Annals of the Missouri Botanical

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Annals of the Missouri Botanical Garden

Volume 87, Number 1 Winter 2000

The Annals, published quarterly, contains papers, primarily in systematic botany, contributed from the Missouri Botanical Garden, St. Louis. Papers originating out- side the Garden will also be accepted. All manuscripts are reviewed by qualified, independent reviewers. Authors should write the Managing Editor for information concerning arrangements for publishing in the ANNALS. Instructions to Authors are printed in the back of the last issue of each volume and are also available online at

www.mobot.org (through MBG Press).

Editorial Committee Victoria C. Hollowell

itor, Missouri Botanical Garden Amy Scheuler McPherson Managing Editor, | Missouri Botanical Garden Diana Gunter Associate Editor, Missouri Botanical Garden Vicki Couture Senior Secretary Barbara Mack Administrative Assistant _ Ihsan A. Al-Shehbaz Missouri Botanical Garden

Gerrit Davidse Missouri Botanical Garden

Roy E. Gereau

Missouri Botanical Garden Peter Goldblatt

Missouri Botanical Garden Gordon McPherson Missouri Botanical Garden P. Mick Richardson

Missouri Botanical Garden

Henk van der Werff Missouri Botanical Garden

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© Missouri Botanical Garden 2000

THE ANNALS OF THE MISSOURI BOTANICAL GARDEN (ISSN 0026-6493) is published quar- terly by the Missouri Botanical Garden, 2345 Tower Grove Avenue, St. Louis, MO 63110. Periodicals postage paid at St. Louis, MO and additional mailing offices. POSTMASTER: Send address changes to ANNALS OF THE MISSOURI BOTANICAL GARDEN, 96 Allen Marketing & Management, P.O. Box 1897, Lawrence, KS 66044-8897

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Volume 87 Number 1 2000

Annals of the Missouri Botanical

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OUR UNKNOWN PLANET: RECENT DISCOVERIES AND THE FUTURE. INTRODUCTION!

P. Mick Richardson?

Recent years have seen many new discoveries in the plant, animal, and other kingdoms. Can we es- timate how many more organisms are out there to be discovered? An international group of experts has been invited to St. Louis to give their thoughts and predictions about this intriguing area of biology. The symposium will consist of an exciting set of presen- tations, ranging from flowering plants in the U.S., Australia, and the tropics, to freshwater fishes, mam- mals, and last, but not least in every sense, extre- mophiles and other bacteria.—F lyer advertising the 45" Annual Systematics Symposium.

In February 1998 I was in London, and purely by chance I was able to attend a talk at the Linnean Society on coelocanths, expertly delivered by Pete Forey. Later that year my wife and I were impressed by a coelocanth specimen in a delightful marine in Tulear, a specimen caught lo- cally off the southwest coast of Madagascar and thought to have been swept southwards from the populations around the Comoro Islands. However, coelocanths have recently been found in Indonesia,

science museum

and one wonders if there are more of them around the world to be discovered. The finding of this sec- ond species of coelocanth was headline news to bi- ologists, as was the discovery of the Wollemi Pine and several new large mammals, discussed in the following pages. However, the regular and contin- ued reporting of new species usually attracts less attention. For example, one of my former students recently published five new species in the brome- liad genus Cryptanthus (Ramirez, 1998). To my mind, such a publication may actually be more im- portant than the discoveries which make headline news because the new species of Cryptanthus are the result of full and detailed revisions of the genus based on field, herbarium, and laboratory studies. The readers can judge for themselves in the follow- ing papers that comprise the published proceedings of the symposium.

1998 Annual Systematics Symposium dif- fered from the usual format in that there were ten speakers rather than the usual seven. This allowed for a greater diversity in composition of the program

! This and the eight articles that follow it are the proceedings of the 45th Annual Systematics Symposium of the T

Missouri Botanical Garden, Our Unknown Planet: October, 1998, at the Missouri Botanical Garden in St. Lo

Recent оте and the Future.

he symposium was held 9-10 s, Missouri, U.S.A

е symposium was supported in part by the National p d Foundation under grant number DEB-9420140. I

thank Peter Raven for helping to select a fine diversity of speakers, Kathy Hurlbert and her e

rt staff for wonderful

help in organizing and administrating the symposium, Barbara Alongi for her fine illustration used for the cover of the

sy mposium brochure, :

? Missouri Botanical Garden, P.O. Box 299, St.

and the symposium speakers for being such a pleasant. group of scientists. Louis, Missouri 63166, U.S.A.

ANN. Missouni Bor. GARD. 87: 1-2. 2000.

Annals of the Missouri Botanical Garden

and allowed a wider variety of organisms to be cov- ered in some detail. The morning session began with Michael Madigan’s talk on prokaryotic organ- isms, a group that constitutes the bulk of evolu- tionary diversity on Earth and which is of increas- ing interest for use in biotechnology and related areas. Think about where systematics would be without the PCR methodology so necessary for cur- rent molecular systematics techniques. Literally, the book on molecular systematics of plants had to be rewritten within six years (Soltis et al., 1992, 1998). Microbes were followed by Richard Brusca’s fascinating talk on arthropod diversification, and this made me think of eating deliciously tasty large crabs caught in the River Jurua in Acre, Brazil. Next was Ebbe Nielsen’s discourse on insects, un- fortunately not included in the published proceed- ings. Our current knowledge of freshwater fishes was the subject of John Lundberg and his col- leagues. If South American fishes are so incom- pletely known, I wonder if the diversity of fishes we ate alongside the aforementioned crabs may have been species new to science. Fortunately, they were photographed before they went into the frying pan, leaving some clues at least to their existence. Last in the animal line of talks was John Kinnon, who told us about new ungulates being dis- covered in Vietnam and his predictions about where future finds will likely be made. The morning session ended with some wonderful video footage and slides from Lynn Margulis, not published here, but see her books Five Kingdoms: An Illustrated Guide to the Phyla of Life on Earth (Margulis & Schwartz, 1998) and Symbiotic Planet: A New Look at Evolution (Margulis, 1998). While not everyone will agree with Margulis's concept of monophyly, there is no denying that she has a a very in- teresting viewpoint to overall ни of biodi- versity, and at the same time she makes an urgent appeal for all biologists and paleontologists to in-

ac-

tegrate their analyses and discuss criteria for es- tablishment of higher taxa

he plant talks came after lunch. lain Prance informed the audience (and now the readers) that the number of angiosperms is currently underesti- mated, and he confidently predicted e there are in fact between 300,000 and 320, He used specific examples of the discovery of me in Mad- agascar and other areas, as well as detailed studies of all genera in areas in Brazil and Brunei, to de- velop his case for an intensification of the rate of collection to confirm his predictions before it is too late. Barbara Briggs's talk on botanical discoveries in Australia contrasted the media attention given to the discovery of a new genus of conifers compared to the uncharismatic discovery of 61 new species in the Restionaceae and allied families. Finally, Barbara Ertter made the surprising announcement that the rate of discovery of new plants in the Unit- ed States and Canada has been constant for the past century and shows no evidence of tapering off.

Mike Donoghue gave a very entertaining and stimulating after-dinner talk, emphasizing that the current age of discovery may be different from ear- lier ones, but it is both richer and more illuminat- ing. It is the duty of all systematists to capture the imagination of other scientists and, even more im- portantly, the public at large.

Literature Cited Margulis, L. 1998. паши |“ lution. rg Books, & V. Schwartz. " 1998, Fi ive Kingdoms: An Il- lustrated Guide to the Phyla of Life on Earth. Freeman,

San Francisco. Ramirez, I. M. 1998. Five new species of wit scarce (Bromeliaceae) and some nomenclatural novelties. Har- 2:

vard T | 3: 215-224.

A New Look at Evo-

Soltis . E. Soltis & J. J. Doyle (Editors). s Noc n matics of Plants. Chapman & Hall, N Ene E., P. S. Soltis & J. J. 1998.

Doyle (Editors). "dene Systematics of Plants П: DNA Sequencing. Kluwer, Boston

EXTREMOPHILIC BACTERIA AND MICROBIAL DIVERSITY!

Michael T. Madigan?

ABSTRACT

rokaryotic microorganisms inhabit *extreme environments"—habitats in which some chemical or physical variable(s) при significantly from that found in habitats that support plant and animal life. Great strides have been

made in t years

have "лије рај, metabolic properties and os evolutionary historie е ге gam s, as all c all known life forms. As our knowledge of bacterial diversity

rs in the isolation and characterization of extremophilic pm and many of them turn out to

Prokaryotes that grow at very high tem- ellular nic din need to be made heat M x and

improves, primarily from the introduction шаша: tools for assessing bacterial phylogeny and diversity and from new nces in isolation and laboratory culture, it is becoming clear that the bulk of evolutionary diversity on Earth does not reside in plants and animals, but instead in the invisible prokaryotic world r great interest in mining the diverse genetic resourc h's smallest cells for use in biotechnology and related areas

words: extremophilic bacteria, evolutionary history, microbial diversity, prokaryotes.

Since the days almost 100 years ago when Robert Koch and his associates isolated the first pure cul- tures of bacteria, microbiologists worldwide have been isolating laboratory cultures of literally thou-

ands of different bacteria. These include, of course, most of the causative agents of infectious diseases, but more important from the standpoint of the web of life on Earth, many of the bacteria that carry out critical chemical reactions that form the “life sup- port” system for plants and animals (Madigan et al., 2000). Despite the diversity of organisms that are already known, it is now clear that microbiologists have only seen the tip of iceberg; most microorgan- isms that exist in nature, in particular the bacteria, have not yet been obtained in laboratory culture! Indeed, with the help of new molecular tools micro- biologists have explored a variety of microbial hab- itats and have detected not only new species of bac- teria, but new genera, families, orders, and even phyla (Barns et al., 1994; Hugenholtz, et al., 1998). Imagine finding a new phylum of plants or animals today! The challenge for microbiologists now is to isolate these organisms, learn about their basic bi- ology, and harness their vast genetic resources for the benefit of mankind.

А NATURAL PICTURE OF THE BACTERIAL WORLD

Great excitement has pervaded the field of mi- crobial diversity in recent years because of the new-found ability of microbiologists to experimen-

tally determine the evolutionary relationships of

bacteria. This giant leap forward emerged from the tools of molecular biology, especially as regards the development of rapid gene sequencing methods and powerful algorithms for the comparative analysis of nucleic acid sequences. But for these advances to impact microbial evolution, a gene or genes that reflected the evolutionary history of an organism had to be identified. Such an evolutionary “Rosetta Stone" had long been sought, but not until the ad- vent of comparative ribosomal RNA sequencing as a rapid and specific means for deducing bacterial phylogenies (Woese, 1987) did microbiologists have the tool they needed to classify bacteria in a nat- ural fashion—the way botanists and zoologists had classified their subjects for over a century using primarily phenotypic characteristics such as bones or leaf arrangements as evolutionary guideposts. Two key concepts have emerged from compara- tive molecular sequencing of ribosomal RNAs: (1) that cells evolved along three major lineages, the Bacteria, the Archaea, and the Eukarya, instead of just two, the prokaryotes and the eukaryotes (Fig. 1); and (2) that the evolutionary difference between a mouse and an elephant (or between Chlorella and Trillium, for the more botanically oriented) pales by comparison to the evolutionary distance between virtually any two common soil bacteria you might want to mention, like Pseudomonas and Bacillus. The first of these conclusions, that prokaryotic life contains two major evolutionary lineages, is slowly but surely becoming mainstream thinking among microbiologists, and is even gaining support

! The d of M. T. Madigan is supported by National Science Foundation grant OPP 980

? Depa

nt of Microbiology and Center for Systematic mus Mailcode 6508, Southern m University, Car- bondale, Illinois 62901-6508, U.S.A. madigan@micro.siu.e

ANN. Missouni Bor. GARD. 87: 3-12. 2000.

Annals of the

Missouri Botanical Garden

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Моште 87, Митбег 1 2000

adigan Extremophilic Bacteria

from macrobiologists as evidenced by the inclusion of this concept in recent biology textbooks (Raven & Johnson, 1999; Raven et al., 1999). However, the second conclusion, that morphologically quite different plants or animals can be extremely closely related in a molecular evolutionary sense, has been for many a harder pill to swallow. If one steps back for a moment and considers that it is not the evo- lution of the mouse and the elephant, or the alga and the flowering plant, as intact entities, that mo- lecular sequencing speaks to, but instead, the evo- lutionary history of the cells that make them up, it is easier to understand why the bulk of evolutionary change has occurred in the prokaryotic world; pro- karyotes have existed for some 3.8 billion years, while the mouse and the elephant have only very recently evolved and diverged.

Prokaryotes ruled the Earth for at least 2 billion years before the modern (organelle-containing) eu- karyotic cell appears in the fossil record. And metazoans (multi-celled plants and animals) have only existed for some half billion years or so. So by the time the stage was set for what botanists and zoologists consider the “evolutionary diversification

29

of plants and animals,” most of cellular evolution had already occurred. Diversification of the mouse and the elephant, for example, was simply a matter of arranging cells in different ways to yield what appears to the eye to be highly divergent organisms. But in terms of their evolutionary history, the mouse and the elephant are virtually identical organisms.

In contrast to higher organisms, prokaryotes have had more evolutionary time to show great genetic divergence. However, unlike metazoans, evolution- ary change in prokaryotes is not manifest in mor- phological variation. For whatever reason(s), bac- teria maintained a very small size and changed relatively little (compared with metazoans) in mor- phology through billions of years of evolutionary history. But that is not to say they did not evolve. Molecular sequencing tells us that they have in- deed evolved but that the product of this evolution- ary change is invisible—instead of big changes in size or shape, evolutionary change in the prokary- otes focused on metabolic diversity and the genetic capacities to explore and eventually colonize every conceivable environment on Earth, including ex- treme environments. Thus we must go to the genes of the prokaryotes to see their true phylogenetic diversification, and with advances in nucleic acid sequencing, this world is now beginning to open up (Madigan et al., 2000)

Using comparative ribosomal RNA sequencing microbiologists can now not only construct natural

relationships of prokaryotes (Fig. 1), but can also use phylogenetic information to construct highly specific nucleic acid probes as a means of identi- fying and tracking specific microorganisms in the environment. А natural application of this technol- ogy has been to take these tools into various ex- treme environments and probe for the diversity of microbial life therein. The fallout from these stud- ies, which historically followed by many years more classical enrichment and isolation approaches, has been an awareness that extreme environments are not a place for “hangers on,” but instead are hab- itats that flourish with microbial life, especially pro-

ave revealed for our understanding of the physi- ochemical limits to life.

EXTREME ENVIRONMENTS AND EXTREMOPHILES

Microbiological examination of extreme environ- ments has revealed many new prokaryotes. By “ex- here, it is meant an environ- at humans would consider extreme ог uninhabitable: extremes of heat or cold, pH, salin- ity, pressure, and even radiation. As previously mentioned, extreme environments are inhabited by diverse populations of microorganisms, most of which have evolved to live only in the presence of

oe

the extreme. These organisms are the “extremo-

Oren, 1999). Several classes of extremophiles are recognized in microbiology, and Е odi cultures of representatives of each class are know

1). Organisms in each class are denoted a de- scriptive term, usually a word with Greek or Latin roots followed by the combining form “phile,” Greek for “loving.” Thus there are thermophiles and hyperthermophiles (organisms growing at high or very high temperatures, respectively), psychrophiles (organisms that grow best at low temperatures), acidophiles and alkaliphiles (organisms optimally adapted to acidic or basic pH values, respectively), barophiles (organisms that grow best under pres- sure), and halophiles (organisms that require NaCl for growth) (Table 1). Instead of trying to be inclu- sive here, as literally hundreds of different species could be included, the organisms listed in Table 1 in each of the ex- tremophile categories. The column of most interest in Table 1 is the one labeled “optimum,” for here

are the current “record holders”

6 Annals of the Missouri Botanical Garden Table 1. Classes and examples of extremophiles*. Descriptive

Extreme term Genus/species Minimum Optimum Maximum Temperature

High Hyperthermophile ^ Pyrolobus fumarii 90°C 106°C 113°

Low Psychrophile Polaromonas vacuolata 0°C 4°C 12°C pH

Low Acidophile Picrophilus oshimae —0.06 0.7 (60°C): 4

High Alkaliphile Natronobacterium grego- 8.5 0 (20% NaCl)! 12

ryt

Pressure Barophile MT41 (Mariana Trench) 500 atm 700 atm (4?C) 21000 atm Salt (NaCl) Halophile Halobacterium salinarum 1596 2596 32% (saturation)

* [n each category the organism listed is the current "record holder" for requiring a particular extreme condition for wth.

» Strain MT41 does not yet have а formal genus and species name and is also a psychrophile. * P. oshimae is also а thermophile, growing optimally at 60°C. 1 М. gregoryi is also an extreme halophile, growing optimally at 20% NaCl.

it becomes clear that these organisms are not mere- ly tolerating their lot, but that they actually do best in their punishing habitats; indeed most actually require their extreme condition(s) in order to repro- duce at all.

Extremophiles are of interest to both basic and applied biology. In a basic sense, these organisms hold many interesting biological secrets, such as the biochemical limits to macromolecular stability and the genetic instructions for constructing mac- romolecules stable to one or another extreme (Ma-

Oren, 1999). But in an applied sense, these organisms have yielded an amazing array of enzymes capable of catalyzing specific biochemical reactions under extreme conditions (Adams & Kel- ly, 1995). Such enzymes have served as grist for industry in applications as diverse as laundry de- tergent additives aree ne and the ge- netic identification of с s (Taq DNA poly-

ase and its use in [pm оа сһаіп reaction, РС

Another important realization that has emerged from the study of extremophiles is that some of these organisms form the cradle of life itself. Many extremophiles, in particular the hyperthermophiles, lie close to the “universal ancestor” of all extant life on Earth (Fig. 1). Thus, an understanding of the basic biology of these organisms is an oppor- tunity for biologists to “look backward in time” so to speak, to a period of early life on Earth. This exciting realization has fueled much research on these organisms in order to understand the nature of primitive life forms, how the first cells “made a living” in Earth’s early days, and how early organ-

isms set the stage for the evolution of modern life forms.

LIFE AT HIGH TEMPERATURE

Although thermophilic bacteria (organisms with growth temperature optima between 45°C and 80°C) have been known for over 80 years, hyperthermo- philic bacteria—organisms with optima above 80°C—have only been recognized more recently

in Brock, 1978), Karl Stetter and co-workers at gensburg (Germany) have proceeded to isolate over 30 genera (> 70 species) of hyperthermophiles. Brock was the first to demonstrate, often using sim- ple but ingenious field experiments, that bacteria were present in boiling hot springs in Yellowstone National Park (Brock, 1978). By contrast, Stetter's group, whose focus has been on isolation and cul- ture, has isolated many of the hyperthermophiles known today, including Pyrolobus fumarii, a re- markable prokaryote that can grow up to 113°C (Ta- ble 1, Fig. 2) (Blóchl et al., 1997).

ermophilic microorganisms can be isolated from virtually any environment that receives inter- mittent heat, such as soil, compost, and the like. But hyperthermophiles thrive only in very hot and constantly hot environments, including hot springs, both terrestrial and undersea (hydrothermal vents), and active sea mounts, where volcanic lava is emit- ted directly onto the sea floor (Stetter, 1999). It is also strongly suspected, and some supportive evi- dence exists, that hyperthermophiles reside deep

Volume 87, Number 1 2000

Madigan 7 Extremophilic Bacteria

Figure 2. ‘Transmission electron micrograph of a cell of Pyrolobus Јитаги, the most thermophilic of all known TO

ind can grow at up to 113°C. Even higher temper- atures are tolerated but do not support growth. Micrograph

courtesy of Reinhard Rachel, Universität Regensburg.

within the earth, living a buried existence and re- lying on geothermal heat for their metabolic activ- ities and reproduction (Stetter, 19¢ The most extreme of known hyperthermophiles, those with temperature optima above 100°C, have come from submarine hydrothermal vents (Stetter, 1996, 1999), and examples include P таги (Blóchl et al., 1997, and Fig. 2) and the methano- gen Methanopyrus kandleri (Kurr et al., 1991). Both of these amazing prokaryotes are members of the Archaea (Fig. 1) and are chemolithotrophs (organ- isms that use inorganic compounds as energy sources), using molecular hydrogen, H,, as their electron donor (energy source), reducing either NO, (P fumarii) or CO, (M. kandleri) as electron acceptors to grow by anaerobic respiration (Madi- gan et al., 2000; Stetter, 1999). Besides requiring substantial heat for growth, these bacteria can sur- vive temperatures substantially above their upper growth temperature limits, making a conventional at 121°C) insufficient for sterilizing cultures of either species! P. fumarii and M. kandleri originated from

autoclave regimen (15 min.

hydrothermal vent chimneys (Blóchl et al., 5 Stetter, 1999). These аге precipitated iron mineral deposits that form as extremely hot water (up to 400°C) containing various minerals emerges from deep-sea hydrothermal vents (note that although this water is superheated, it does not boil because of the hydrostatic pressure of the water column, usually 2000-3000 m, that overlies these vents). Although the water that emerges is too hot for life,

the chimneys, which are often only about 0.5 cm thick, show a temperature gradient from about 300°C inside to 2°C outside. Because prokaryotes are so small, microenvironments differing in tem- perature exist across the chimney wall leading to ideal habitats for various species of heat-loving bacteria.

Using nucleic acid probe technology several morphological types of bacteria have been detected in hydrothermal vent chimney walls (Harmsen et al., 1997), suggesting that these compact thermal gradients may contain many different microbial populations in addition to those already isolated. And for my botanical friends reading this paper, I would be remiss if I did not point out that P. fumarii and M. kandleri are good examples of primary pro- ducers totally divorced from sunlight, a capacity widespread in the microbial world. Besides growing at almost unbelievably high temperatures, P fu- marii and M. kandleri are also autotrophs, capable of growing in a simple anaerobic mineral salts me- dium supplied with СО, and H,; neither sunlight nor a key product of photosynthesis, O,, is required for either organism. Indeed, it has been hypothe- sized that long before the process of photosynthesis evolved, anaerobic H,-based chemolithotrophy was the major means by which new organic material was synthesized on Earth (Madigan et al., 2

For an organism to grow at high temperatures, especially as high as those of the hyperthermo- philes discussed here, all cellular components, in- cluding proteins, nucleic acids, and lipids, must be heat stable (Adams & Kelly, 1995; Ladenstein & Antranikian, 1998; Wiegel & Adams, 1998; van de Vossenberg et al., 1998a). The thermostability of enzymes from various hyperthermophiles, referred

een documented, and О,

to as extremozymes, has some have been found to remain active up to 1 (Adams & Kelly, 1995). The structural features that dictate thermal stability in proteins are not well understood, but a small number of noncovalent fea- tures seem characteristic of thermostable proteins. These include a highly apolar core, which appar- ently makes the inside of the protein “sticky” and thus more resistant to unfolding, a small surface-

tends to remove options for flexibility and thus in- troduce rigidity to the molecule, and extensive ionic bonding across the protein’s surface that helps the compacted protein resist unfolding at high temper- ature (Ladenstein & Anthranikian, 1998). In ad- dition to these intrinsic stability factors, special proteins called chaperonins are synthesized by hy- perthermophiles. Chaperonins function to bind heat

Annals of the Missouri Botanical Garden

denatured proteins and refold them into their active form. The thermosome is a type of chaperonin that is widespread among hyperthermophiles capable of growth above 100°C, like P. fumarii and M. kan- dleri (Stetter, 1999).

everal factors may combine to prevent DNA from melting in hyperthermophiles. However, the two most important features appear to be the en- zyme reverse DNA gyrase, which catalyzes the pos- itive supercoiling of closed circular DNA (by con- trast, nonhyperthermophiles contain DNA gyrase, an enzyme that supercoils DNA in a negative twist- ed fashion), and various types of DNA binding pro- teins, including histone-like proteins (Madigan & Oren, 1999; Pereira & Reeve, 1998). For various physicochemical reasons, positively supercoiled DNA is more resistant to thermal denaturation than is negatively supercoiled DNA. And the fact that reverse gyrase seems to be the only protein thus far found universally among hyperthermophiles (re- gardless of their metabolic pattern) (Madigan & Oren, 1999) points to an important role for it in the heat stability of DNA.

Several hyperthermophiles contain DNA binding proteins that appear to play a role in maintaining DNA in a double-stranded form at high tempera- ture. Some of these proteins are structurally related to the core histones of eukaryotic cells and function to wind and compact the DNA into nucleosome-like structures (Pereira & Reeve, 1998). Others have no structural relationship to histones but when bound to DNA alter its structure in such a way as to sig- nificantly raise its melting temperature (Madigan & Oren, 1999). It is likely that the combination of positive supercoiling of DNA along with proteins that prevent DNA melting is a major solution to the maintenance and integrity of DNA in hyperther- mophiles.

Heat can also affect membrane stability. As all biologists know, in organisms living at moderate temperatures cell membranes are constructed along the typical “lipid bilayer” model: hydrophobic res- idues (fatty acids) inside oppose each other and retain an affinity for one another while hydrophilic residues (such as glycerol phosphate) lie at the sur- face of the environment and the cytoplasm, respec- tively, maintaining contact with the aqueous phase. If one applies sufficient heat to such a membrane architecture the two leaflets of the membrane will pull apart, leading to membrane damage and cy- toplasmic leakage. To prevent this from occurring at very high temperatures, hyperthermophiles have evolved a novel membrane structure. Instead of forming a membrane as a lipid bilayer, as just dis- cussed, some hyperthermophiles chemically bond

the opposing hydrophobic residues from each layer of the membrane together (van de Vossenberg et al., 1998a). This forms a lipid monolayer instead of a bilayer, and prevents the membrane from melting at high temperature. Although the precise chemis- try of lipid monolayer membranes can vary some- what from species to species, they are common among hyperthermophiles and are likely an impor- tant evolutionary response to life at high tempera- ture.

LIFE AT Low TEMPERATURES

How about life at the other end of the thermom- eter? Cold environments on Earth are actually much more common than hot ones. For example, the oceans, which make up over one half the Earth's surface, maintain an average temperature of about 2?C. And vast land masses are intermittently cold and in some cases permanently cold, or even frozen. However, cold temperatures are no barrier to microbial life, as various microorganisms flourish in cold environments, even in ice (Horikoshi & Grant, 1998; Madigan & Marrs, 1997). Many mi- croorganisms have been isolated capable of growth at refrigerator temperatures (4—8°C). These are usu- ally psychrotolerant, meaning that although they are capable of growth in the cold, they grow better at warmer temperatures, usually 25-35°С. True psy- chrophiles, defined as microorganisms that grow best

15°C or lower, are usually only present in per- manently cold environments like the Arctic, or in particular, the Antarctic (Horikoshi & Grant, 1998).

А variety of microorganisms including algae and diatoms have been found in Antarctic sea ice— ocean water that remains frozen for much of the year. Sea ice is the habitat for one well-character- ized bacterium, Polaromonas vacuolata, the genus name indicating its affinity for cold temperatures ). Polaromonas vacuolata grows

—

Irgens et al., optimally at 4°C and finds temperatures above 12° too warm for growth (Table 1). Other psychrophiles are known, but because some of them appear to be very sensitive to warming, great care must be taken in their isolation and culture to prevent killing them off at temperatures as low as room temperature. An understanding of the biochemistry and mo- lecular biology of psychrophilic bacteria is in a much earlier stage than that of the hyperthermo- philes. From what is known about the biochemistry of psychrophiles, it appears that their proteins func- tion optimally at low temperatures because they are constructed in such a way so as to maximize flex- ibility; this is essentially the opposite strategy from that of hyperthermophiles (see earlier). Moreover,

Моште 87, Митбег 1 2000

Madigan Extremophilic Bacteria

proteins from psychrophiles are typically more po- lar and less hydrophobic than proteins from hy- perthermophiles, a fact that undoubtedly also as- sists in their relative flexibility.

Besides keeping their enzymes functional, psy- chrophiles have other biological problems to con- tend with, transport of nutrients across the mem- brane being chief among them. However, just as margarine, with its higher content of unsaturated fats, can stay softer than butter at cold tempera- tures, psychrophiles regulate the chemical compo- sition of their membranes, including in particular the length and degree of unsaturation of fatty acids, to keep them sufficiently fluid to allow for transport processes, even at temperatures below freezing (Horikoshi & Grant, 1998). Applications of en- zymes from psychrophiles include the cold food in- dustry, where enzymes that work at refrigerator tem- peratures are sometimes desirable, as well as producers of cold-water laundry detergents (see more on this below).

LIFE IN BATTERY ACID OR SODA

Many extremophiles have evolved to grow best

t extremes of pH: these are the acidophiles and b. alkaliphiles (Horikoshi & Grant, 1998). Al- though extremely acidic or alkaline (below pH 3 or above pH 10) habitats are rare on earth, in such environments one can find a variety of microorgan- isms thriving in chemistry the equivalent of battery acid or soda-lime. Highly acidic environments can result naturally from geochemical activities, such as from the oxidation of SO, and H,S produced in hydrothermal vents and hot springs, and from the metabolic activities of certain acidophiles them- selves. For example, the iron sulfide-oxidizing bac- terium Thiobacillus ferrooxidans can generate acid by oxidizing Fe?* to Fe?*, the latter of which pre- cipitates out as Fe(OH), (Fe** + ЗЊО — Fe(OH), + 3H*), or by oxidizing H8. to S0,- (НЗ + 20, = 502 + H^). Thiobacillus ferrooxidans is par- ticularly active in surface coal mining operations where exposure to oxygen of pyrite (FeS,) in the coal seam triggers acid production from the meta- bolic activities of this and related bacteria. Runoff from these habitats can often have a pH of less than 2, fueling conditions for further acidophile activity.

The most acidophilic of all bacteria known thus far is Picrophilus oshimae, whose pH optimum for growth is just 0.7 (Schleper et al., 1995) (Table 1). Picrophilus oshimae is also a thermophile (temper- ature optimum, 60°C) so this organism must be sta- ble to both hot and acidic conditions. Cultures of P. oshimae were isolated from an extremely acidic

(< pH 1) solfatara in Italy, and the organism has clearly evolved to require these highly acidic con- ditions for its very existence.

Interestingly, however, acid-loving extremo- philes, even those as extreme as P. oshimae, cannot tolerate great acidity inside their cells, where it would destroy such important molecules as DNA. They thus survive by keeping the acid out. The internal pH of P. oshimae is about pH 5, and it is the cytoplasmic membrane of this organism that keeps protons from passively entering the cell. However, studies of the P. oshimae membrane have shown that it can only retain its integrity in acidic solutions; above an external pH of about 4 the P. oshimae membrane spontaneously disintegrates. Major unanswered questions concerning the metab- olism of P. oshimae and other extreme acidophiles concern how they generate a proton motive force during respiration and related issues of bioenerget- ics involving membrane-mediated proton translo- cation (van de Vossenberg et al., 1998b).

Various acid-tolerant enzymes from acidophiles, primarily ones located on the cell surface or ones excreted from the cell into the acidic milieu, have been studied and potential industrial applications identified. These are primarily as animal-feed sup- plements where the enzymes function to break down inexpensive grains to more nutritionally ben- eficial forms directly in the animal’s stomach. Such enzymes have been widely used in the poultry in- dustry and have been shown to reduce feed costs and the time necessary to get birds to market.

Extreme alkaliphiles live in soils laden with soda (natron) or in soda lakes where the pH can rise to as high as 12. Natronobacterium gregoryi (Table 1), for example, was isolated from Lake Magadi, a soda lake located in the Rift Valley of Africa; N. gregoryi grows optimally at a pH of about 10 (Table 1) (Hor- ikoshi & Grant, 1998). In the opposite scenario from the acidophiles, alkaliphiles have to contend with the problems associated with high pH. Above a pH of 8 or so, certain biomolecules, notably RNA, break down. Consequently, like acidophiles, alka- liphiles must maintain their cytoplasm nearer to neutrality than their environment. Nevertheless, any proteins found in the cell wall or in the mem- brane that make contact with the environment must be stable to high pH. Indeed, many such enzymes have been studied and a number have found in- dustrial applications, especially in the laundry de- tergent industry. Detergents that are “enzyme en- riched” contain proteases and lipases (enzymes that degrade proteins or fats, respectively, in clothing stains) that function at the high pH of soapy solu- tions (Horikoshi & Grant, 1998). In addition, alkali-

Annals of the Missouri Botanical Garden

active enzymes from thermophiles and psychro- philes have been discovered and commercialized to better target detergent additives to hot water or cold water applications, respectively.

Besides keeping their cytoplasm near neutrality, alkaliphiles have other biological problems to con- tend with. For example, consider the problem of membrane-mediated bioenergetics—protons ex- truded to the external surface of the membrane en- ter a sea of hydroxyl ions. Nevertheless, biochem- ical studies of this problem have shown that a proton motive force is indeed formed by extreme alkaliphiles and drives some of the energy-requir- ing reactions in the cell, such as motility and trans- port. Sometimes in ATP synthesis, an ion gradient of Ма“, rather than H+, drives this key bioenergetic process in extreme alkaliphiles шалк ры & Grant,

1998). This is probably not surpri о considers that many (but not all) extreme alkali- philes are also extreme halophiles (see below), re- quiring high salt as well as high pH for metabolism and reproduction.

ing when one

LIFE IN A BRINE

Another remarkable group of extremophiles are the halophiles—organisms adapted to grow best in salty solutions (Oren, 1999; Ventosa et al., 1998). And for extreme о like Halobacterium, a

“salty solution” me e from 25% NaCl up to saturation (32% NaCl) (Table 1). Halophilic microorganisms abound in hypersaline lakes suc

the Dead Sea, the Great Salt Lake, and solar salt evaporation ponds. Such lakes are often colored red by the dense microbial communities of pigmented halophiles such as Halobacterium (Javor, 1989).

, and under- ground salt deposits. To date, a very large number of halophilic bacteria have been grown in culture including members of all domains of life, including the Eukarya (Kamekura, 1998). chaeal halophiles as exemplified by Halobacterium species remain the most halophilic organisms

owever, the ar-

own.

Halophiles are able to live in salty conditions by preventing dehydration of their cytoplasm. They do this by either producing large amounts of an inter- nal organic solute or by concentrating an organic or inorganic solute from their environment (Hori- koshi & Grant, 1998; Oren, 1999). Th patible solutes” is often used to describe organic osmolytes, of which there are several types, but not all halophiles employ such solutes (Madigan & Oren, 1999; Oren, 1999). For example, as its os-

e term “com-

molyte, the archaeon Halobacterium (Table 1) con- centrates large amounts of potassium (K+, as KCl) from its environment. Dissolved KCl in the cyto- plasm of Halobacterium cells is present at a con- centration equal to or slightly above that of the dis- solved NaCl outside, and in this way cells maintain the tendency for water to enter and thereby prevent dehydration. As would be expected from such a salty cytoplasm, enzymes that function inside of cells of Halobacterium have evolved to require this large dose of K* for catalytic activity. By contrast, membrane or cell wall-positioned proteins in Hal- obacterium require Ма’ and are typically stable only in the presence of high Ма’ (Madigan & Oren, 999)

=

Extreme halophiles are sources of а variety of biomolecules that can function under salty condi- tions. Applications of salt-active enzymes include those that can break down viscous materials pres- ent in oil wells (oil is often found in geographic strata that contain salt) as well as enzymes that can carry out desirable transformations in highly salted foods. In addition, some halophiles that produce organic compatible solutes have been commercial- ized for the production of these solutes as skin care

supplements (Madigan & Oren, 1999).

OTHER EXTREMOPHILES

Extremophilic microorganisms adapted to high pressure or which show no deleterious effects from exposure to high levels of radiation are also known. Barophiles are microorganisms that grow best under pressure greater than 1 atmosphere. Extreme bar- ophiles are the most interesting in this regard as they actually require pressure, and in some cases, extreme pressure, for growth (Table 1). Strain

T41, for example, a bacterium isolated from ma- rine sediments in the Mariana Trench near the Phil- ippines (a depth of greater than 10,000 m), requires at least 500 atmospheres of pressure in order to grow and grows optimally at 700 atmospheres (and at a temperature of 4°C because strain МТАЛ is also a psychrophile). Because laboratory culture of ex- treme barophiles is rather difficult, comparatively little is known about their important biomolecules. However, although probably all macromolecules in extreme barophiles need to be biochemically tai- lored to high pressure to some extent, experiments with moderately barophilic bacteria, some of which can be grown without pressure, have pointed to nu- trient transport proteins in the cytoplasmic mem- brane as key cell components requiring structural modifications in order to function at high pressure

(Horikoshi & Grant, 1998).

Volume 87, Number 1 2000

Маадап Extremophilic Bacteria

The bacterium Deinococcus radiodurans is an amazingly radiation-resistant microorganism (Mur- ray, 1992). This remarkable organism can survive 30,000 Grays of ionizing radiation, sufficient to lit- erally shatter its chromosome into hundreds of pieces (by contrast, a human can be killed by ex- posure to as little as 5 Grays). A powerful DNA repair machinery exists in cells of D. radiodurans that is able to piece the shattered chromosome back together and yield viable cells. Because of its re- markable radiation resistance, Deinococcus has been proposed as a cleanup agent for the biore- mediation of toxic materials in contaminated soils that are also radioactive from the leakage of radio- active materials; these conditions exist primarily at nuclear weapons production sites.

EXTREMOPHILES IN THE EVOLUTION OF LIFE

A focus of research on extremophiles has cen- tered on the hyperthermophiles. As discussed ear- lier, there is good reason to believe that at least some hyperthermophiles have evolved relatively lit- tle from their ancestors present on earth over 3.5 billion years ago (Figs. 1, 2). If true, an understand- ing of the biology of hyperthermophiles may yield a glimpse of what life was like eons ago. In this connection the genomes of several hyperthermo- philes have been sequenced (Madigan & Oren, 1999), and the large number of genes they contain that lack counterparts in other organisms suggests that their biological secrets have at this point only been partially revealed. As if living in boiling water isn’t enough, just imagine what other tricks hy- perthermophiles might be able to perform!

As previously mentioned, the excitement in mi- crobial diversity these days comes from the fact that the evolutionary history of the prokaryotes can now be experimentally determined. Microbiologists no longer have to propose bacterial phylogenies based on speculation or “educated guesses” of what type of microbe likely preceded another; the phylogenies themselves are etched in the sequences of mole- cules, and all one has to do is read them. Moreover, the application of molecular phylogenetic methods to natural environments (Barns et al., ; Hu- genholtz et al., 1998) has given us the exciting news that the diversity of the microbial world is enormous—indeed it is beyond our wildest expec- tations. Thus, in the final analysis bacterial diver- sity will likely dwarf that of all of the rest of biology, perhaps by several orders of magnitude. But only continued and expanded research into the diversity of microbial life in all environments, extreme and

otherwise, will yield the data needed to confirm this.

It may indeed be humbling to many biologists to think that prokaryotes dominate living diversity. But within the rich genetic resources of the pro- karyotes undoubtedly lies more benefit for human- kind than we will extract from any other group of organisms. Antibiotics, fermentation, and biotech- nology are only the beginning. The best is yet to come.

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tending the upper temperature "for life to 113?C. Extre- mophiles 1: 14—21.

Brock, T. D. 1978. Thermophilic Microorganisms and Life at High Temperatures. Springer, New York.

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UNRAVELING THE HISTORY OF ARTHROPOD BIODIVERSIFICATION!

Richard C. Brusca?

ABSTRACT

Current views of arthropod phylogeny are assessed in light of recent research in morphological and molecular phylogenetics, m biology. neurobiology, and po Recent fossil discoveries and molecular clock

data inform us that were already

pod diversification began in the Pre he аан а iose metazoan re im on earth.

cambrian, and suggest that by the Cambrian the arthropods “The c ombination of metamerism and jointed appendages

(with intrinsic musculature), and the evolutionary potential of homeotic genes, has profoundly affected arthropod evolution

avors a monophyletic Arthropoda. Accumulating evidence

and created many Wap aces ‘al homoplasies. | supports a hypothesis that in

ects and modern crustaceans ато

а phylogenetic sister group, and that they, and perhaps

also trilobites, chelicerates, p myriapods, all could have evolved out of an ancient crustacean stem line. Two implic ations

of this hypothesis are that Crus Key words:

stacea comprise a paraphyletic taxon and insects may be viewed as “flying crustaceans.” Arthropoda, arthropod evolution, Crustacea, insects, Met

PREFACE. THE CHALLENGE OF UNRAVELING METAZOAN PHYLOGENY

Despite great progress made in zoology during the 20th century, there remain many fundamental, unanswered questions concerning metazoan evolu- nd relationships of

e part, this stems from covering unambiguous йоген enc from ancient lineages. Recent molecular and paleonto- logical studies suggest that major splits among the Metazoa occurred in the Precambrian, some per- haps nearly a billion years ago (Wray et al., 1996; Ayala et al., 1998; Seilacher et al., 1998; Li et al., 1998). In part, it may also be because the field of comparative morphology has lost popularity (and employment opportunities). And finally, the emerg- ing field of molecular phylogenetics is still so new that every year sees improvements in the data an- alyzed and the phylogenetic inference methods used. For example, prior to 1997 most molecular analyses were based on small numbers of taxa and short sequences of a single gene, usually the in- herently problematic 188 rDNA gene. Recently, however, new (and larger) molecular data sets have been developed based on other conserved nuclear genes, mitochondrial gene order, and gene dupli- cation data. Because it is unlikely that a single gene will recover the full phylogeny of Metazoa, the future will no doubt see analyses of multiple gene sets.

Emerging molecular studies have corroborated many, and challenged some, paradigms of metazoan phylogeny. For example, whereas some molecular studies have supported the long-held close rela- tionship between annelids and arthropods (Wheeler et al., 1993), recent studies have not done so (Lake, 1990; Halanych et al., 1995; Eernisse, 1997; Agui- naldo et al., 1997). Furthermore, the discovery of new animal phyla, and thus new fundamental body plans, continues to occur. The first edition of Lin- naeus's (1735) Systema Naturae listed 14 groups that we now recognize as distinct animal phyla. To- day, we recognize 34 animal phyla. Three former phyla have recently been sunk: Pentastomids are now placed within the Arthropoda (allied with the Maxillopoda), and vestimentiferans and pogonoph- orans are now regarded as annelids (probably high- ly modified polychaetes) (McHugh, 1997; Brusca & Brusca, in press).

Most of the large-bodied animal groups were dis- covered by the end of the 19th century. We are now on a track of discovery of microscopic metazoa, and three new animal phyla have been discovered just since 1956: Gnathostomulida (1956), Loricifera 1983), and Cycliophora (1995). There is a corre- lation between the discovery of new animal phyla and their body sizes: phyla described in the period of 1901-1920 have maximum body lengths of 3- 10 mm; phyla described in the period of 1941— 1960 have maximum body lengths of just 1 mm;

P mm

! This paper е from reviews by Wendy Moore, Lisa Nagy, and an anonymous reviewer. SF-PEET m ag B-9521649) to the author. Special thanks go to the inde СТА Peter Raven

ported in part b by for encouraging "bs ы. of this

This work was sup-

? Columbia University, Biosphere 2 uda: P.O. Box 689, Oracle, Arizona 85623, U.S.A.

ANN. MissounRi Bor.

GARD. 87: 13-25. 2000.

Annals of the Missouri Botanical Garden

phyla described in the 1980s and 1990s have max-

imum body lengths of less than 0.5 mm. Most of

the small-bodied phyla are meiofaunal, although cycliophorans live as commensals on the mouth ap- pendages of various marine crustaceans (Funch & Kristensen, 1997). The discovery of these minute animals presents challenges to those of us interest- ed in animal phylogeny. They are so small that a great deal of their anatomy is reduced or otherwise altered. We know almost nothing about their de- velopmental biology, and they are so rare that mo- lecular biologists have not yet extracted gene se- quences from them. I predict that the discovery of new microscopic phyla will continue for another half-century.

The challenges of unraveling animal phylogeny are not unique to molecular biology, small animals, or new phyla. Biology has a long history of skir- mishing over phylogenetic issues at all levels. The evolutionary history of the Arthropoda has been one of the most challenging issues biologists struggled with throughout the 20th century. What follows is an update (as of mid 1998) on what we know about arthropod evolutionary history.

ARTHROPOD EVOLUTION: BACKGROUND

There are five clearly distinguished groups of ar- thropods: trilobites (extinct since the end of the Pa- leozoic; ~ 4000 described species); Chelicerifor- mes (horseshoe crabs, eurypterids, arachnids, pycnogonids; ~ 75,000 described living species); crustaceans (crabs, shrimp, isopods, and their kin; ~ 50,000 described living species); hexapods (in- sects and their kin; 878,000 to 1.5 million de- scribed living species); and myriapods (centipedes, millipedes, and their kin; ~ 14,000 described liv- ing species). And, there are two close allies of the arthropods, tardigrades (water bears) and опу- chophorans (Peripatus and their kin). The close re- lationship between the Tardigrada and the Arthrop- oda has never been seriously questioned (Brusca & Brusca, 1990), and recent molecular work contin- ues to support a sister-group relationship between these two phyla (Garey et al., 1996). There are now 1.02 to 1.64 million described arthropods, known from virtually all environments on earth. Estimates of undescribed arthropod species range from 3 to 100 million. The arthropods (Table 1) comprise about 85% of all described metazoan species.

The arthropods also encompass an unparalleled range of structural and taxonomic diversity, have a rich fossil record, and have become favored ani- mals of evolutionary developmental biology. Arthro- pods were among the earliest animals to evolve.

Table 1.

Fossil record of major arthropod groups.

Tardigrades: Middle Cambrian to present Onychophora: Middle Cambrian to present Trilobita: Early Cambrian to Permian

Xiphosura: Early Ordovician/Silurian to present Eurypterida: Early Ordovician through mid-Permian Arachnida: Upper Silurian to present Pycnogonida: Devonian to present

Crustacea: Early Cambrian (or Vendian) to present Hexapoda: Lower Devonian to present

Myriapoda: Upper Silurian to present

Recent work (Waggoner, 1996) suggests that even the Ediacaran (Vendian) fauna, of the latest Pre- cambrian, included early arthropod taxa, perhaps true Crustacea.

Ever since Darwin, biologists have asked the question, “How has the incredibly successful di- versification of arthropods come about?” Why are there so many arthropods? Is there something “spe- cial” about these animals? What is the phyloge- netic history of the Arthropoda? Specifically, are the arthropods monophyletic and what are the re- lationships of the major arthropod groups to one another? There have been four great challenges to biologists in answering these questions. (1) Until the last decade of the 20th century, there had been a lack of hypotheses on arthropod evolution based on principles of explicit phylogenetic inference. (2) We have a very incomplete understanding of ar- thropod development, though this is improving quickly with the advent of molecular developmental biology. (3) There has been a paucity of compre- hensive studies based on fossils from the earliest ages of arthropod evolution (late Precambrian and early Paleozoic). (4) It is apparent that high levels of homoplasy exist among the arthropods. In just the past 10 years, major discoveries have begun to address each of these challenges, as discussed be- ow.

Work by the great comparative biologist Robert Snodgrass in the 1930s established a benchmark in arthropod biodiversity research. Table 2 shows a classification of the arthropods at that time, and it is this classification that one still finds in most modern biology textbooks. The Snodgrass classifi- cation embraces three important hypotheses:

(1) Arthropods comprise a monophyletic taxon.

(2) Myriapods and hexapods form a sister group, a taxon called Atelocerata (= Tracheata, or Uni- ramia of some authors). The Atelocerata have been united by several seemingly powerful at- tributes:

Volume 87, Number 1 2000

Brusca Arthropod Biodiversification

Table 2

Classification of the arthropods and their allies sensu Snodgrass (1938).

Phylum Arthropoda Subphylum Trilobita Subphylum Chelicerata Class Merostomata Subclass Xiphosura. Horseshoe crabs Subclass | Class Arachnida. Land spiders, mites, etc. Class Pycnogonida. Sea spiders ge ibn Mandibulata Class Crustacea. Crabs, shrimps, isopods, etc. Class heata (= Atelocerata) Subclass He Superorder Protura. Proturans

xapoda

Superorder Insecta. Insects Subclass Myriapoda Chilopoda. Centipedes Diplopoda. Millipedes Symphyla. Symphylans

Superorder Superorder Superorder

Superorder Рапгорода. Pauropodans

Eurypterida. Eurypterids; extinct Paleozoic arthropods

(a) A tracheal риу system.

(b) Uniramous legs.

(c) Use of Malpighian tubules for excretion.

(d) Loss of the second head appendages—the second antennae (as the name Atelocerata im- plies). Vestiges of the anlagen of this appendage can be seen during the embryogeny of some insects (e.g., Sharov, 1953; Brukmoser, 1965).

(3) The Crustacea and the Tracheata form a sister

the a name that Snod-

grass himself coined.

group, Mandibulata

For a brief period of time in the mid-century the concept of a polyphyletic Arthropoda, championed mainly by S. Manton and D. Anderson, enjoyed some popularity (Manton, 1973, 1977; Manton & Anderson, 1979; Anderson, 1979), and Anderson (1996) still maintains this view. The Mantonian view of arthropods placed the myriapods, hexapods, and onychophorans in a separate lineage (Manton’s phylum “Uniramia”) with an origin apart from the rest of the arthropods. However, this idea, based on flawed phylogenetic argumentation and an inade- quate embryological foundation, did not long sur-

ive the rigors of scientific testing and modern methods of phylogenetic inference (see below). In addition to phylogenetic analyses, studies of Perm- sae а insects (Kukalová-Peck, 1991a, b, 1992; Kukalová-Peck & Brauckmann, 1990) have hoan that early pterygotes probably possessed polyramous appendages, further under- mining the Manton-Anderson Uniramia hypothesis. Additional support for arthropod monophyly has come from studies of compound eyes using a mono-

ian

clonal antibody raised against a specific glycopro- tein (3G6), to crystalline cones, eucones, and other elements in a variety of insect and crustacean ret- inas (Edwards & Meyer, 1990).

It was not until the i 1980s that Snodgrass’s long-standing view of arthropod relationships began to be seriously questioned with: (1) the appearance of explicit morphological and molecular phyloge- netic analyses, (2) the discovery of the amazing po- tential of homeobox genes in arthropod develop- ment and evolution, (3) the emergence of molecular-based evolutionary developmental biol- ogy, and (4) the discovery of exquisite new Cam- brian preservations from Sweden, China, and else-

ere.

MORPHOLOGICAL PHYLOGENETIC STUDIES OF ARTHROPODS

Morphological phylogenetic studies of the arthro- pods are summarized in Table 3. Overall, these analyses suggest three important conclusions:

(1) The arthropods are a monophyletic taxon.

(2) The relationship of the Crustacea to the insects and myriapods is ambiguous; that is, Snod- grass's Mandibulata is a taxon of questionable validity.

(3) The monophyly of the Atelocerata (insects + myriapods) is also questionable.

Waggoner (1996) included in his analysis a num- ber of arthropod-like fossils belonging to the “Ven- dian fauna," from the latest Precambrian (= Edi- acara Period) that had generally been regarded as

16 Annals of the Missouri Botanical Garden Table 3. Morphological views of monophyly within the arthropods. Arthropods Mandibulates Tracheata

Year Author(s) monophyletic monophyletic monophyletic 1990 Brusca & Brusca Yes Yes n.a. 1991 k Yes No n.a. 1992 Eernisse et al. Yes n.a. n.a. 1993 Backeljau et al. Yes n.a. n.a. 1993 Wheeler et al Yes Yes Yes 1994 Wills et al Yes Yes Yes 1995 Wills et al Yes No No 1996 Nielsen et al. Yes Yes Yes 1996 Waggoner Yes No No 1997 Emerson & Schram Yes No No 1998 Strausfelc Yes No No

“problematica.” He also included 21 Cambrian ar- thropods, and various modern taxa. He concluded that: (a) the Arthropoda are monophyletic, (b) the Ediacaran arthropod-like fossils are, in fact, true arthropods, and (c) the anomalocarids (and their kin) fall out very close to the base, and are probably the most primitive known arthropods. Anomalocar- ids were giant predatory arthropods (arguably, true Crustacea) that reached a meter in length. They are known from both the Precambrian and the Cam- brian, and they were probably the m poe of that time (Briggs, 1994; Chen et al.,

he most recent phylogenetic us ji = pods was based on anatomical features of the сеп- tral nervous system (Strausfeld, 1998). Strausfeld used 100 conserved neural characters in the brains of a variety of segmented invertebrates to recon- struct phylogenetic relationships among the arthro- pods. His analysis suggested that insects and crus- taceans comprise a sister group, that the myriapods are a polyphyletic group (1.е., chilopods and dip- opods are not sister taxa), and that pycnogonids are true chelicerates. The most important neuronal synapomorphies of Crustacea—Insecta are elements of the optic lobes and mid-brain, particularly fea- tures of the midline neuropils and neuropils asso- ciated with the compound eyes. This analysis cor- roborated earlier neurological descriptive work by Strausfeld et al. (1995), which also concluded that insects are closer to crustaceans than to any other arthropod group.

All arthropod central nervous systems use the same fundamental p plan of construction (Whitington et al., ; Thomas et al. 4; Strausfeld, 1998). ы. a fundamental distinc tion between the early embryonic development of the myriapod nervous system and that of insects + crustaceans was recognized some time ago. Whi- tington et al. (1991) found that in insects and crus-

taceans longitudinal connectives are pioneered by segmental neurons, whereas in the centipede Eth- mostigmus rubipres longitudinal connectives are pi- oneered from neurons in the brain that send their axons posteriorly to set up the parallel connectives. This difference between centipede and insect-crus- tacean ventral nervous systems is compounded by the fact that the pattern of segmental neurons in centipedes is quite different from that found in in- sects and crustaceans; centipede ganglia receive contributions from more widely distributed neu- rons, and there are more neurons in the centipede ventral cord when segmental axons are laid down. Comparisons of early neuronal outgrowth during embryonic development of the brain and thoracic ganglia also suggest a close affinity Quar crus- taceans and insects (Harzsch et al., : anos et al., 1995; Whitington et al., T Paulus (1979) argued for arthropod monophyly on the basis of shared characters in the organization of photo- receptors and their satellite cells in compound and single-lens eyes. He further noted that insect and crustacean ommatidia, with their developmentally fixed numbers of cells, share more fine structural characters than either do with the chilopod om- matidia (which comprise an indeterminate number of elements).

MOLECULAR PHYLOGENETIC STUDIES OF ARTHROPODS

Molecular о studies of the Arthropo- e 4. Field et al. (1988) sequenced a short У вс of 185 rRNA but used

da are summarized in

representatives of just 10 phyla (only 4 of which were arthropods). Despite its limitations, the Fie

et al. work was pioneering. It was the first molec- ular phylogenetic study to test the monophyly of the arthropods, which it supported, and the work ini-

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Вгизса Arthropod Biodiversification

2000 Table 4. Molecular views of monophyly within the arthropods. Myriapoda + Arthropods Crustacea + Hexapoda Year Author(s) monophyletic Hexapoda — (Tracheata) Data 1988 Field et al. Yes Yes No 18S rRNA 1989 Patterson Yes Yes No 185 rRNA 1990 Lake No No Yes 185 rRNA 1990 Field et al. Yes Yes No 188 rRNA 1991 Turbeville et al. Yes Yes No 185 rRNA 1992 Ballard et al. Yes Yes No 125 rRNA (mitochondrial) 1992 Winnepenninckx et al. Yes Yes n.a. 85 rDNA 1993 Van « ге: Yes Yes f 185 rDNA 1993 Wheeler et al. Yes Yes No 18S rDNA + ubiquitin 1995 Winnepenninckx et al. Yes Yes n.a. 18S rDNA 1995 Friedrich & Tautz Yes Yes No 18S + 28S rDNA 1996 Garey Yes Yes n.a. 18S rDNA 1997 Regier 8 Shultz Yes Yes No EF-la + POLII 1997 Eernisse Yes Yes No 185 rDNA 1997 Spears & Abele Yes Yes No 185 rDNA

tiated a stream of follow-up studies, continuing to

e 188 rRNA sequences, and later the 185 rDNA gene itself. Each subsequent study has tended to use more taxa and longer nucleotide sequences for its data base, but until very recently most also con- tinued to rely on the 18S gene. Problems associated with the 185 gene, use of short gene sequences, and single-gene phylogenetic inferences are well known and need not be repeated here. Further- more, although there are now over 300 metazoan 185 sequences available, most published phyloge- nies have been based on fewer than 20 sequences (Eernisse, 1997). This is despite studies that sug- gest a minimum o taxa are needed to ac- curately identify the root node of a large clade (Le- cointre et al., 1993a, b; Sanderson, 1996; Hillis, 1996). In spite of methodological and sampling problems, recent molecular studies are beginning to converge on some similar conclusions. However, as Spears and Abele (1997) pointed out, “. . . in the crusade for understanding relationships among crustacean and other arthropod lineages, the rDNA data represent but a relic, and not the Holy Grail itself.”

The most recent 18S sequence data suggest that insects share fewer similarities with the myriapods than they do with the Crustacea. Spears and Abele (1997) analyzed 31 18S sequences, and their re- sults suggested that neither crustaceans nor insects were monophyletic. When they removed the “prob- lematic” long-branched crustacean taxa (Remipe- dia, Cephalocarida, Mystacocarida), a myriapod + chelicerate clade emerged first, with insects as the

sister group to a paraphyletic Crustacea. The Spears and Abele analysis also strongly supported malacostracan monophyly. Eernisse (1997) ana- lyzed 103 sequences and concluded that (1) the Arthropoda are monophyletic, but only if the tar- digrades are included [probably another 185 arti- fact], and (2) hexapods are more closely related to crustaceans than they are to myriapods. Regier and Shultz (1997) made a complete and welcome break with the 18S gene, using sequences from two other nuclear genes, the elongation factor (EF-la) gene and the A polymerase II (POLII) gene. These trees were robust and mostly in agreement with the 18S work, concluding that: (1) Arthropods are monophyletic, (2) Crustacea are paraphyletic, and 3) insects are not the sister group of the myriapods, but arose from within the Crustacea.

Recent work by Boore et al. (1995) examined not gene sequences, but the linear arrangement of mi- tochondrial genes. This new type of data corrobo- rates the gene sequence work and recognizes a mi- tochondrial gene arrangement that is unique to the crustaceans and insects alone.

In summary, the majority opinion from the mo- lecular research, and the most recent opinions from both the morphological and molecular work, rec- ognize four key features in arthropod phylogeny:

~

(1) Arthropods are monophyletic.

(2) Neither the Mandibulata nor the Atelocerata are natural groups.

3) Crustaceans and insects constitute а sister group, exclusive of the myriapods.

~

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(4) Crustacea are likely to constitute a paraphyletic taxon

These last three conclusions are in conflict with 150 years of morphology-based thinking. Thus, two profound implications of these new studies are that the morphological attributes linking insects to myr- iapods might all be convergences (e.g., uniramous legs, tracheal system, Malpighian tubules), and that insects are actu ying crustaceans” (in the same sense that birds are flying reptiles).

EMERGING VIEWS FROM DEVELOPMENTAL STUDIES

The unique combination of segmentation and jointed appendages has allowed arthropods to de- velop modes of locomotion and feeding, and body region specialization, unavailable to other metazoan phyla. We now know that the fates of these seg- mental units and their appendages are under the ultimate orchestration of homeotic genes. These genes select the critical developmental pathways to be followed by cells during morphogenesis. Ho- meobox genes determine such basic body architec- ture as the dorso-ventral and the anterior-posterior body axes, where body appendages form, and the general types of appendages that form (Averof & Patel, 1997; Panganiban et al., 1997; Shubin et al., 1997). Homeobox genes can either suppress limb development, or modify it to create alternative ap- pendage morphologies. A growing body of evidence suggests that these unique genes have probably played major roles in the evolution of new body plans among arthropods and the Metazoa in general (Davidson et al., 1995; Williams & Nagy, 1995; Panganiban et al., 1995).

The degree to which homeobox genes have been conserved is remarkable, and most of them proba- bly date back at least to the Cambrian. For exam- ple, homologues of the Pax-6 gene seem to dictate where eyes will develop in all animal phyla. Pax- 6 is so similar in protostomes (insects) and deu- terostomes (mammals) that the genes can be ex- perimentally interchanged and still function correctly. Homeobox genes modulate the expression of dozens of interacting, downstream, developmen- tal genes whose products drive morphogenesis. The profound evolutionary potential of homeobox genes lies in this hierarchical nature. Variation in the out- put of these multigene networks can arise at many levels, simply by tinkering with the relative timing of gene expression—an evolutionary process we know as heterochrony. To understand the profound potential of homeobox genes to drive evolutionary change, consider that within the Drosophila genome 85-170 different genes might be regulated by the

product of a single homeobox gene, the Ultrabi- thorax (Ubx) gene (Carroll, 1995 A good example of the evolutionary potential of homeobox genes is seen in the abdominal limbs of insects. Abdominal limbs (*prolegs") occur on lar- vae of various insects in several orders, and they are ubiquitous in the order Lepidoptera, i.e., cat- erpillars. Abdominal limbs were almost certainly present in adult insect ancestors. Hence prolegs may have reappeared in such groups as the Lepi- doptera through something as simple as the de-re- pression of an ancestral limb development program (1.е., they represent an atavism). We now know that proleg formation is initiated by a change in the reg- ulation and expression of the BX-C gene complex e., the Bithorax complex, which includes the Hox s Ubx, abdA, and abdB) during embryogenesis (Carroll, 1995 Molecular and developmental biology also seem to have broken the deadlock on the arguments over origins of uniramous and biramous limbs (e.g., Po- padic et al., 1996; Panganiban et al., 1995, 1997; Shubin et al., 1997; Emerson & Schram, 1997). We now know that limb branching is a second-order phenomenon, probably orchestrated largely by the homeobox gene Distal-less (DII). This single gene initiates development of unbranched limbs in in- sects and branched limbs in crustaceans. Antibod- ies that recognize ОИ proteins show expression at the tips of insect limbs and also in biramous crus- 1995). Branched

limbs are formed when the gene is expressed ec-

tacean limbs (Panganiban et al.,

topically in Drosophila (Diaz-Benjumea et al., 1994). In fact, ОИ occurs in many animal phyla, where it is expressed at the tips of ectodermal body outgrowths in such different structures as the limbs of vertebrates, parapodia and antennae of poly- chaete worms, tube feet of echinoderms, siphons of tunicates, and appendages of arthropods. Further- more, recent work suggests that whether an arthro- pod mandible is *whole-limb" (i.e., built of many segments) or "gnathobasic" (i.e., built of only the basalmost segments) also depends on the expres- sion of the gene Distal-less. Thus, Dil is expressed in the whole limb (or multisegmented) jaws of myr- iapods, but not in the gnathobasic jaws of crusta- ceans and insects—still further testimony to the probable sister-group relationship of insects and Crustacea.

THE PALEONTOLOGICAL DATA

Recent work has shown the fossil record of ar- thropods dates back to the early Cambrian, or per- haps the late Precambrian. And, by the mid-Paleo-

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Brusca 19 Arthropod Biodiversification

Table 5

Some important Precambrian and Cambrian arthropod Lagerstdtten faunas.

Age

Name

Principal location

Orsten fauna Upper Cambrian (~ Burgess Shale fauna Chengjiang fauna Ediacaran fauna

510 MYA) Middle Cambrian (~520 MYA) Lower Cambrian (~530 MYA) 560-600 MYA)

Latest Precambrian (~

Southern Sweden British Columbia Southern China Ediacara Hills, Aust.

zoic, all five arthropod lineages were in existence and had already undergone substantial radiation. Arthropods are also the first land animals for which we have a geological record (Labandeira et al., 1988; Kukalová-Peck, 1990), and by the Late 51- lurian the first terrestrial scorpions and myriapods were already present. In fact, both terrestrial and marine myriapods have been reported from this pe- riod (Almond, 1985; Hahn et al., 1986; Labandeira et al., 1988), although molecular data suggest that myriapods might have arisen as early as the Cam- brian (Friedrich & Tautz, 1995). The first centipede fossils occur in the Upper Silurian (— 414 MYA) and, along with trigonotarbid arachnids, constitute the earliest known land animals (Jeram et al., 1990). The first millipede fossils occur in Devonian deposits (Almond, 1985; Robison, 1990); they are similar to the extant genus Craterstigmus (Shear et al., 1984) and are contemporaneous with the first terrestrial mites, pseudoscorpions, and scorpions (Størmer, 1969, 1977; Shear et al., 1987), as well as the first hexapods (Greenslade, 1988). The ear- liest known fossil hexapods are bristletails and col- lembolans from 390-million-year-old Gaspé mud- stone (Labandeira et al., 1988). Some good records of these early creatures now exist, and the presence of these predatory arthropods suggests that complex terrestrial ecosystems were in place at least as early as the late Silurian. Perhaps the most important ancient arthropod fossils are those in which even the soft parts of the animal were preserved—the so-called ancient Lagerstátten (Table 5).

These ancient fossils have pushed the age of or- igin for the arthropods back to at least 600 MYA, and they provide us with critically important data on early arthropod anatomy and evolution. These extraordinary faunas are now telling us that Crus- tacea probably predate the appearance of trilobites in the fossil record, running counter to a long-held belief that trilobites were the most ancient arthro-

pods. The recently exploited Chengjiang fauna of

south China is Lower Cambrian, about 10 million years older than the Middle Cambrian Burgess Shale fauna (Chen et al., 1994). The Chengjiang fauna is very well preserved and includes at least 100 species of animals, many without hard skele-

tons, including the first known members of many modern groups. However, it is the arthropods that dominate this fauna, including trilobites and. bra- doriid “crustaceans” (and also tardigrades and on- ychophorans). The largest of the Chengjiang ani- mals is Anomalocaris, also known from Ediacaran and Middle Cambrian deposits (Briggs, 1994). The Chengjiang fauna is very similar to that of the Bur- gess Shale, and it demonstrates that the arthropods were already far advanced by this early date.

The spectacular recent discovery by Klaus Müll- er and Dieter Walossek (Müller, 1983, 1992; Müller & Walossek, 1985; Müller et al., 1995; Walossek & Müller, 1992, 1997) of microscopic arthropods from the Upper Cambrian Orsten deposits of Swe- den, has brought to light a rich fauna of minute crustaceans, crustacean larvae, and various crus- tacean-like arthropods. Among them, for example, is Skara, a cephalocarid-, or mystacocarid-like crustacean for which both naupliar larvae and adults have been recovered (the nauplius larvae are only a couple hundred microns long; adults are about 1 mm in length) (Müller & Walossek, 1986). Skara, and many other Orsten Crustacea, were sure- ly meiofaunal animals not unlike modern marine meiofaunal crustaceans.

Fossils from this Cambrian site in Sweden have been collected since the days of Linnaeus, who ac- tually described the first fossils from this area in 1757 (trilobites and conodonts). However, a brand- new kind of collecting began with Müller and Wal- ossek's work in the 1980s. This new Orsten material is all microscopic, three-dimensional fossils. The Orsten arthropods show little or no signs of decom- position. They preserve details less than 1 micro- meter in size (e.g., cuticular pores, the bristles on setae). Dozens of Orsten microcrustacea have so far been described. The recovery of these three-dimen- sionally preserved animals and the developmental series that have been found (with successive larval, juvenile, and adult instars—in animals less than 1 mm in length) have provided us with information on the detailed anatomy of body segments and ap- pendages of many ancient stem-arthropods. The Or- sten fauna shows that Cambrian Crustacea had all the attributes of modern crustaceans, such as com-

Annals of the Missouri Botanical Garden

pound eyes, a head shield, naupliar larvae (with locomotory first antennae), and biramous append- ages on the second and third head somites (the sec- ond antennae and mandibles).

Taken together, this recent paleontological work corroborates Whitington’s observations long ago about the Burgess Shale fauna, that during Cam- brian times the non-trilobite arthropods were both morphologically more varied and more numerous than were the trilobites (despite popular belief). We also now know that arthropods have probably been the dominant animals in terms of species diversity since the Cambrian. Arthropods comprise over one- third of all species described from Lower Cambrian strata.

Briggs and Fortey (1989) cladistically analyzed 23 of the Cambrian arthropod taxa, plus 5 extant groups. Their tree placed the Crustacea at the very base, as a paraphyletic sequence of taxa, and placed the trilobites and chelicerates near the top of the tree. The most recent molecular work does not conflict with this tree, in viewing the Crustacea as a paraphyletic group from which the other major arthropod clades emerged.

THE PENTASTOMIDA

Pentastomids are obligatory parasites of verte- brate respiratory systems. There are about 100 de- scribed species, all of which infest various tetra- pods, including two cosmopolitan species that infest humans. The blood-sucking adults inhabit re- spiratory tracts of their hosts, where they anchor themselves by means of their hooklike head ар- pendages. For years it was believed that pentasto- mids were allied with the onychophorans as ver- miform, pre-arthropod creatures. However, several recent molecular studies (using 185 gene sequenc- es) have revealed the pentastomids to be highly modified crustaceans (Abele et al., 1989, 1992; 1996). Corroborative independent work over the past few years has come from cla-

Garey et al.,

distic analyses of sperm and larval morphology, nervous system anatomy, and cuticular fine struc- ture (Wingstrand, 1972, 1978; Storch, 1984; Storch & Jamieson, 1992). Furthermore, Miiller and Wal- ossek’s work on the Swedish Orsten fauna proves that the pentastomids (and also the tardigrades) had appeared at least by the Upper Cambrian, long be- fore the land vertebrates had even evolved (Miiller & Walossek, 1988; Walossek & Miiller, 1994). So, we must ask what the original hosts of these para- sitic crustaceans might have been. Walossek, Miill- er, and even Stephen Jay Gould have noted that Conodont fossils are common in all the Cambrian

localities that have yielded pentastomids, and thus the conodonts (also long a mystery, but now widely fish-like vertebrates) might have been the original hosts of the pentasto-

regarded as parts of early mids.

THE ONYCHOPHORA

As with pentastomids, onychophorans, too, were part of the amazing, early-Cambrian, explosive ma- rine diversification. They have been found in Bur- gess Shale-type faunas at several localities, in Cam- brian deposits from China and Siberia, and in the Swedish Orsten fauna (Xianguang & Weiguo, 1988; Xianguang & Junyuan, 1989; Ramskóld & Hou, 199 nd, we now know that Conway Morris's original reconstruction of Hallucigenia (from the Burgess Shale) had the animal turned upside-down. Ramskóld and Hou (1991) recently turned Hallu- cigenia over and found a second pair of legs, con- cluding it was an onychophoran with long dorsal spines. And there is now an onychophoran known from the Chengjiang deposits of China with side plates and spines (Ramskóld & Hou, 1991). Ay- sheaia (also from the Burgess Shale) was originally described by Walcott as an annelid, but it, too, is now regarded as an early marine onychophoran.

CONCLUSIONS

Let us now return to our two fundamental ques- tions regarding arthropod evolution: Why are there so many arthropods, and what is the phylogenetic history of the arthropods? As to the first question, I propose six over-arching scenarios, each complex in its own right

(1) The numerical superiority of arthropods is not

a recent event. Recent fossil discoveries, and molecular clock data, inform us that arthropod diversification began very early in the history of the Metazoa, in the Precambrian, and by the Cambrian the arthropods were probably already the most speciose metazoan phylum on earth. Arthropods have been on a powerful phyloge- 600 MY. They have had a great deal of time to PEN and with the exception of the trilobites and the eu- rypterids, all the major lineages have survived and continue to radiate.

netic trajectory for well over

S

Their great size range, especially on the smaller end of the scale, adapts arthropods for a great variety of ecological niches. The Cambrian Or- sten. deposits tell us that a whole fauna of in- terstitial/meiofaunal arthropods already existed as early as the mid-Cambrian, and this habitat

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Brusca 21 Arthropod Biodiversification

has continued to be rich in adaptive radiation

and specialized species ever since. Similar small-body-size niches are filled in a great many specialized environments today. We find high diversities of minute arthropods in habitats such as marine sediments, coral reefs, among the fronds of algae, on mosses and other prim- itive plants, and on the bodies of every kind of animal imaginable. There are even arthropod faunas that live strictly on the gills of other crustaceans (mites and small crustaceans). Small insects and mites have exploited virtually every terrestrial microhabitat available.

(3) The close relationship and coevolution with flowering plants (on land) and algae (in aquatic

een a powerful force in

—

environments) have the radiation of the arthropods. It is not just the insects that have been on a coevolutionary tra- jectory with plants—many crustaceans utilize algae as both a living substrate and a food source and show strong evidence of coevolu-

tion.

(4) The arthropods (insects) were the first flying an- imals, and the ability to fly led them into niches other invertebrates simply could not penetrate.

(5) Metamerism (the serially repeated body seg- ments and appendages of arthropods) provides an enormous amount of easily manipulated body plan material upon which evolutionary processes can act. Given the great age, sheer diversity, and our emerging knowledge of reg- ulatory genes in these animals, a high level of homoplasy is no longer surprising.

(6) The potential for major changes in body plans due to variations in homeobox genes, and the downstream genes they regulate, is just begin- ning to be realized, but this potential is clearly enormous. There seems little doubt that chang- es in homeotic genes over time have profoundly affected arthropod evolution. Considering the number and position of limbs in arthropods, and the flexibility of homeobox and regulatory switches, it is little wonder that arthropod an-

atomical diversity seems so endless.

As to the second question—what is the phylo- genetic history of the Arthropods—it seems the plasticity of the arthropod body and homeobox gene expression may have produced an even higher level of homoplasy than once thought. As a result, some traditional morphological classifications are in con- flict with molecular classifications. All the evidence suggests that the arthropods are monophyletic. However, fossil data, recent comparative neuroan- atomical research, and molecular data all suggest

that Crustacea are a paraphyletic group, and that the Crustacea and Insecta are very closely related to one another, but not to the Myriapoda. In fact, the insects appear to have arisen from within a crustacean stem line. Further, recent molecular and fossil data are beginning to suggest that the trilo- bites, chelicerates, insects, myriapods, and recent crustaceans all might have emerged from crusta- cean stem-line ancestors. This view of a paraphy- letic Crustacea spinning off a series of other major arthropod lineages might explain why morpholo- gists have been unable to come to agreement on the sister-group relationships of the major arthropod lineages. Resolution of this conflict will come, | predict, within the next two decades, with further understanding of the genetic regulation of devel- opmental processes, examination of new nuclear and mitochondrial genes (and use of multiple gene data sets in phylogenetic analyses), and as more cladistic analyses include fossil species, particu- larly the growing series of Chengjiang, Orsten, and related arthropods.

A SPECULATION

The realization that insects might have arisen out of an ancestral crustacean stem line leads to many new implications concerning arthropod evolution. For example, given this scenario, one could search about among the Crustacea for a likely ancestor to the insects and in doing so recognize the presence of a “fixed” 19-segmented body plan in insects and certain crustaceans (or more likely a 20-segmented plan in each, Kukalová-Peck, 1991а, b; Scholtz et al., 1994; Scholtz, 1995). All insects are fixed on this body plan. Of all the crustacean higher taxa, this body plan consistently occurs only in the sub- class Eumalacostraca—the crabs, shrimps, isopods, and their kin. Thus, if the insects did evolve from a crustacean ancestor, one might spec ulate that they could have evolved from a Examining the Eumalacostraca for a piile i insect ancestry, there is only one group that is truly ter- restrial, has evolved gas-exchange tracheae (grant- ed, probably convergently to those of insects), has reduced/lost one pair of antennae (antennae one re- duced in oniscideans, antennae two reduced hexapods), and has strictly uniramous walking legs (as do the insects)—the terrestrial isopods (Isopo- da: Oniscidea). Could it be that insects are not only flying crustaceans, but flying isopods?

The concept of a Eumalacostraca—Insecta sister- group relationship finds strong support in the com- parative anatomy of arthropod central nervous sys- tems. Development of the compound eye follows

„пала

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similar morphogenetic events in insects and eu- malacostracans (Hafner & Tokarski, in press). In addition, the optic lobes of pterygote insects and eumalacostracans are distinguished by nested reti- notopic neuropils, each of which represents the whole eye. In these two taxa, these neuropils com- prise an anatomically distinct lamina, medulla, and lobula complex (Strausfeld, 1996). The presence of these structures in pterygote insects and eumala- costracans was viewed as a homology indicating a sister-group relationship between these taxa Osorio and Bacon (1994) and Nilsson and Osorio (1997). Further, eumalacostracans that have so far been examined also possess a distinctive form of neuron, called a bushy T-cell, which was first rec- ognized in insects on the basis of its characteristic dendritic “tree” situated near the inner face of the medulla (Strausfeld, 1976). Bushy T-cells in insects and eumalacostracan crustaceans send their axons to large tangential dendrites that extend across sub- stantial areas of the retinotopic mosaic. In those pterygote orders investigated, bushy T-cells com- prise part of an evolutionarily conserved subset of retinotopic elements that contribute to elementary motion detector circuits (Strausfeld & Lee, 1991; Douglass & Strausfeld, 1995, 1996). The presence of these cell types in eumalacostracan crustaceans and pterygote insects implies that either identical circuits have independently in the two groups, or the circuit for motion detection evolved in a common ancestor to insects and crustaceans has been maintained basically unchanged through- out the history of both groups. That the latter is more likely is suggested by the presence of small field retinotopic neurons that arise from the inner layer of the medulla of the apterygote Thermobia and extend into the lateral lobe of the protocere- brum (Strausfeld, 1998).

All crustacean nervous systems so far examined possess the architectonic and positional equivalent of a fan-shaped body, the neurons of which extend laterally into the рое с) lobes, as they do іп insects (Strausfeld, 1998). However, except in iso-

insects. Further, in pterygote insects and isopods (but not in decapods or apterygotes) the fan-shaped body is supplied by a bridge of neuropil that lies posteriorly in the brain and connects the left and right protocerebral hemispheres. Strausfeld (1998) concluded that, while fan-shaped bodies are syna-

morphic to insects and crustaceans, the proto- cerebral bridge may have evolved independently in insects and isopods.

Many morphological features are in conflict with

a close malacostracan-insect relationship, includ- ing differences in tagmata arrangement and loca- tions of the gonopores. The fossil record also does not support an isopod + insect sister-group rela- tionship. The oldest known isopod fossils are only 300 million years in age (Phreatoicidea: Hesslerella, Carboniferous) (Brusca & Wilson, 1991). However, a recent analysis of phreatoicidean phylogeny sug- gests the isopods might have had their origin con- siderably earlier than this (Wilson & Keable, in press), and further examination of this unconven- tional idea may be warranted.

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E

SO MANY FISHES, SO LITTLE Ай Kottelat,” Ger ith,’ Melanie

TIME: AN OVERVIEW OF Stiassny,? and Anthony C. Gille

RECENT ICHTHYOLOGICAL

DISCOVERY IN

CONTINENTAL WATERS’

ABSTRACT

Although freshwaters contribute only about 0.01% to Earth’s water supply, their fishes now number more than 10,000 species and thus account for at least 40% of all fish species. The continental fish faunas differ greatly in taxonomic composition and species richness, our state of knowledge of them, and the rate of discovery of unknown kinds. The ichthyofaunas of North America (about 1050 species), Europe (about 360), and Australia-New Guinea (about 500), are the most thoroughly documented, but new species continue to be described based on discovery of previously unseen orms and species-level taxonomic splits of known species. The ichthyofaunas of tropical Asia (perhaps > 3000), Africa (perhaps > 3000 species), and South and Central America (perhaps >> 5000 species), are species-rich yet inc omple tely known. Tropical freshwaters are the hot spots of recent and likely future ichthyological discoveries. Especially in the species that signal new generic-level taxa are common, and new family-level groups are found

occasionally. Everywhere ongoing phylogenetic studies oft gg r | unsuspected relationships. These аге imes of exciting disco advancement of knowledge in freshwater ichthyology. New discoveries beckon us to seek the many remaining unknowns in the diversity of life on our planet. These are also times of rapid and destructive

change i in freshwater habitats around the pe se threats alert us to the increasing potential for permanent loss and i icd of ve of our planet's rich aquatic biota.

Key words: Actinopterygii, Characiformes, Craniata, Cypriniformes, freshwater, Gymnotiformes, ichthyofaunas, ich- ыл. Otophysi, з finned fish, Siluriformes

The species of craniate animals (hagfishes + ver- tebrates) are probably the most thoroughly docu- mented of all the large clades in the tree of life. Yet new species of living craniates are described

i ichthyologists. Characterized in this way, the total mber of living fish species is about 25,000 and accounts for roughly 50% of the extant species rich- ness of the Craniata, while the other half are tetra- pods (Nelson, 1994). Thus delimited, fishes are not a сна не group because their subgroups аге along a phylogenetic "ladder" below the asd. (Fig. 1). The vast majority of fish species, owever, belong to one clade: the Actinopterygii or ray-finned fishes. There are at least 23,700 living

frequently, and most of these are fishes. In a recent tally, Eschmeyer (1998) found that about 200 fish species are described annually

Fishes are water-dwelling craniates with perma- nent gills borne on the walls of pharyngeal arches

or pouches. Fishes have median fins supported by cartilaginous or bony rays, and most have paired

actinopterygian species (Nelson, 1994). The species richness of the ray-finned fishes strongly contrasts

fins, but never limbs bearing digits (Bond, 1996; with the e present species poverty of the other higher Helfman et al., 1997). Uniquely, fishes are studied fish clades (Fig. 1).

! We applaud and and acknowledge Peter Raven and P. Mick Richardson for creating the 45th Annual System- atics Symposium at the Missouri Botanical Garden, and for inviting us to participate. We ч оиг тапу usse s who provided helpful Sc and suggestions on drafts of this manuscript: Alberto Akama, na C rnandes, Carl Ferraris, John Friel, lan Pos ui Hopkins, Michel Jegá, Dave Johnson, Luiz Миљан, Lucinda ме Dade, Steven Norris, Lynn Parenti, Мапо C. C. de Pinna, Lácia Rapp Py-Daniel, јен = sis, Ole Seehausen, Monica Toledo-Piza, Guy Teugels, P. J. Unmeck, Richard Vari, Stan Weitzman, and anonymous reviewers. Editors Amy Mc- Pherson and Victoria Hollowell at the MBG Press offered us patient and wise assistance өн чаша ће MER: process. Permission roduce ursi ade or photographs was granted by the authors or creators noted in figure captions, and the publishers of: American Museum Novitates; Copeia; Ichthyological Е xploration of Freshwaters; Occasional Papers of the Museum of Zoology, ay of Michigan; Proceedings of the Academy of Natural Sciences, Philadelphia; and Revue Suisse de Zoologie.

? Department a enis and Evolutionary о The University of ta Tucson, Arizona 85721, U.S.A. осће 12, Саве > postale 57, < 19-2952 Cornol, Switzer lan

S. Museum of Natural History, New Yo rk, New York 10024, U.S • Department of Zoology, The Natural History Museum, Cromwell Road, London SW7 5BD, United ETE

ANN. Missouni Bor. GARD. 87: 26—62. 2000

Моште 87, Митбег 1

Lundberg et al. 27 Ichthyological Discovery

| Hagfishes 25 | | | Lampreys | 99 F ISharks, rays 1,200 ! ! |S IRay-finned fishes «423,700! Н E | Coelacanths 1(2?) | S | | |Lungfishes — — — 7 | Tetrapods 24,000

Figure l. The phylogenetic tree of the major groups of living craniate animals and estimated number of extant species in each (based on Nelson, 1994, and Compagno, pers. comm.).

The large-scale ecological distribution of fishes is strongly bimodal: 5846 of fish species are marine, 41% (about 10,000 species) live principally in freshwater, and only about 160 (~ 1%) regularly migrate between salt and fresh water (Cohen, 1970; McDowell, 1988). This diversity of freshwater fishes causes some pause in light of the impressive fact that freshwater makes up a tiny amount, only about 0.01%, of Earth's water supply. Thus, Horn (1972 calculated that freshwaters hold a far greater “den- sity" of fish species than the oceans—greater by 7500 times! But, of course, most marine species live in a relatively small volume of seawater in the productive photic zone, especially around coral

МУ

reefs.

In this paper we focus on the diversity of fresh- water fishes, those living in continental lakes, riv- ers, streams, and swamps. In addition to their great species numbers, freshwater fishes are interesting to us because of their tremendous variety in form, function, and habit. Freshwater fishes provide evo- lutionary biologists with some . the best — of natural selection and adaptation, e.g., gupp (Endler, 1983) and а (Bell & Foster, 1994), divergence and speciation, e.g., Laurentian Great Lakes whitefishes (Smith & Todd, 1984) and North American Great Basin fishes (Hubbs & Mill- er, 1948; G. R. Smith et al., in press), reticulate evolution, e.g., Catostomidae (Uyeno & Smith,

1972), species flocks (African cichlids, Stiassny, this paper), and Lake Titicaca Orestias (Parenti, 1984a; Costa, 1997), and the historical develop- ment of biotas on scales from regional, e.g., post- glacial age North America (Bailey & Smith, 1981) to continental, e.g., North America (Patterson, 1981; Grande, 1994) and Africa-South America (Lundberg, 1993).

Here we are particularly interested in the ex- panding knowledge of freshwater fishes: the

amounts, natures, and sources of recent ichthyolog- ical discoveries and prospects for the future. Our first concern is with the new species of freshwater fishes that are being found and described. In ad- dition, we call attention to discoveries of other im- portant aspects of fish diversity: new phylogenetic lines and unexpected relationships, newly foun extraordinary characteristics of phenotypes and natural histories, and new understanding of the pro- cesses that control rates of speciation and extinc- tion. The major sources of previously unseen fishes are explorations in biotically uncharted waters, es- pecially in remote tropical areas, and in difficult- to-sample aquatic habitats such as deep river chan- nels, cataracts, and leaf litter. Yet survey work at long-visited sites, such as the African Great Lakes and Amazon margins, continues to yield many new species. Unrecognized and even formerly unseen species are routinely found in the course of thor- ough taxonomic studies, especially in revisions of species-rich, freshwater groups of ostariophysans, atherinomorphs, and percomorphs. Another source of recent species descriptions involves taxonomic concepts and practice. Application of species cri- teria that emphasize diagnosability or distinctness of populations over traditional geographically wide- spread, apes Auges results in an increase of recognized speci

ollowing some : additional background informa- tion aimed mainly at the non-specialist, we take a regional approach to showcasing freshwater fish di- versity and recent discoveries. Based on our indi- vidual expertise, authorship responsibility follows geography: Smith—North America, Lundb South and Central America, tropical Asia, Stiassny—Africa, Gill—Australia and New Guinea.

erg—

FRESHWATER FISH FAUNAS: TAXONOMIC COMPOSITION

Most higher fish clades include species that sometimes or permanently live in freshwater. All lamprey species reproduce in freshwater; some re- main there whereas others migrate to the sea to feed. Modern lungfishes are restricted to freshwater. Several species of sharks, sawfishes, and rays reg- ularly enter freshwater, and the Potamotrygonidae are a moderately diverse clade of sting rays con- fined to South American rivers and lakes. Scores of actinopterygians live in freshwater. Familiar ex- amples of these are sturgeons, gars, carps, pira- nhas, tetras, electric eels, catfishes, pikes, trouts, guppies, sticklebacks, black basses, sunfishes, darters, cichlids, bettas, and gouramies. If biotic

Annals of the Missouri Botanical Garden

success is measured by species numbers and breadth of overall distribution, the dominant grou of freshwater fishes is the Otophysi (Ostariophysi in

(carps, minnows, barbs, suckers, loaches) of North America, Europe, Asia and Africa, presently with roughly 2700 species; (2) Characiformes (tetras, pi- ranhas), now with at least 1300 species, distributed today in the Neotropics and Africa, and formerly extending to Europe and the Arabian Peninsula; (3) Gymnotiformes (electric eel and knifefishes) of South America with over 90 described species; (4) Siluriformes (catfishes) currently with more than

0 species and an overall distribution including all continents, even Antarctica as recorded by fos- sils from Seymour Island (Grande & Eastman,

BIOGEOGRAPHY AND PHYSIOLOGICAL ECOLOGY OF FISHES

Freshwater fishes play important roles in a va- riety of biogeographic studies. Some are endemic markers that delimit regional biotas. Fish species are censused in determinations of regional and global patterns of species richness or biodiversity. Each clade of fishes has its singular biogeographic history. Many freshwater fish clades are compo- nents of biotas that share with others common vi- cariant or dispersal-based biogeographic histories. Many fish species and higher taxa serve as biotic indicators of former connections among regions from watersheds to continents.

Monophyletic groups of strictly freshwater fishes have special significance in continental biogeogra- phy because they require freshwater for dispersal, and thus their distributions are correlated with the evolution of topography and watersheds. The degree to which fishes are physiologically and behaviorally restricted to freshwater varies widely. Species and higher taxa of fishes are commonly grouped ac- cording to their observed present and presumed historical habitat and physiological salinity toler- ances (Myers, 1938; Darlington, 1957). So-called “primary” freshwater fish groups, such as most oto- physans, spend their entire lives in freshwater hab- itats and are physiologically incapable of coping with seawater. It is important to know the рћујо- genetic level at which subgroups within a clade show strict primary freshwater distributions (Pat- terson, 1975; Lundberg, 1993)

"Secondary" freshwater fish taxa, such as cypri- nodontiforms and cichlids, are usually limited to freshwater, especially for reproduction, and have

North America Europe 360 1,050 9 | ЧЩ Tropical EMD = Asia > 3,000

SA ING

Australia & New Guinea 500

South & Central America »5,000

Africa «3,000 Figure 2. Estimated number of living fish species in (ће major freshwater faunas. Base map created by Xerox Corporation's Palo Alto Research Center, Map Viewer.

their greatest diversity and abundance there, but occasional individuals or member species may be found regularly in coastal saltwater habitats. Sec- ondary freshwater fishes might, therefore, disperse through the sea. "Peripheral" fishes are marine groups that include species with individuals that may move sporadically into freshwater or species that are permanent freshwater residents; common examples include herrings, anchovies, needlefishes, puffers, pipefishes, drums, gobioids, and soles. All continental ichthyofaunas include peripheral groups with persistent freshwater populations, and the freshwater faunas of many islands comprise en- tirely peripheral fishes. These are generally regard- ed as "marine dispersants" but, unless or even if their closest intragroup relatives are known to be resident marine taxa, peripheral fishes should not automatically be ignored in comparing interconti- nental faunas.

DISTINCTNESS OF FRESHWATER СНТНУ‹

ГА ОМА

At the scale of continents (and Alfred Wallace’s Zoogeographic Realms) ichthyofaunas differ greatly in their taxonomic composition and species rich- ness (Fig. 2). Australia and Europe each have hun- dreds of fish species, North America has somewhat more than 1000, tropical Asia and Africa each have about 3000, and there may be more than 5000 fish species in the American tropics. Especially at the species level, there are also major differences in thoroughness of our knowledge of continental fish faunas. Higher-level taxonomic differences are mostly caused by the long and distant isolation of the freshwaters of most continents. Differences in species diversity probably have more to do with factors that control both terms of biological diver- sification: speciation and extinction. Differences in the extent of knowledge of species result from dif- ferent histories of exploration and study, as well as species richness. It is not surprising that we know

Volume 87, Number 1 Lundberg et al. 29 2000 Ichthyological Discovery

Figure Fish icons of North America. —A. Petromyzon marinus, sea lamprey. —B. Polyodon ie ли p —C. Lei 25165 05565, _ gar. —D. Amia NO bowfin E. Lavinia exilicauda, hitch. nus pla- tyrhynchus, mountain suc —G. Noturus furiosus, Carolina шайган, —Н. Aphredoderus nm pirate perch, —1. Percopsis omiscomaycus, trout-perch. . Spe leti poulsoni, Alabama cavefish. —K. Umbra limi, central mud- minnow. —L. Esox темин, musk БШ ge. —M. randria bimaculata, spottail кае —N. кези notatus, blackstripe topminnow. —O. Goodea atripinnis, blac kfin goodea. —P. Percina caprodes, logperch. —Q. Archoplites interruptus, Sacramento perch. —R. Elassoma evergladei, Everglades pygmy sunfish. Drawn n Tum Шу,

more about the less diverse European, North Amer- ican, and Australian faunas, than the rich African, tropical Asian, and South American assemblages.

NORTH AMERICA

The North American freshwater fish fauna oc- cupies the Nearctic Realm: from Canada and Alas- ka, south to the Transvolcanic Axis south of the Mexican Plateau (Fig. 2). Over 1050 species (e.g., Fig. 3) in about 175 genera and 32 primarily fresh- water families (plus a few freshwater species be-

longing to 35 genera in 24 mostly marine families) occupy rivers and lakes currently or recently drain- Caribbean, and

ing to the Arctic, eastern Pacific,

western Atlantic. The numerically dominant fami- lies are: Cyprinidae (305 species), Percidae (172), Poeciliidae (75), Catostomidae (68), Ictaluridae (48), Goodeidae (40), Fundulidae (37), Centrarchi- dae (32), Atherinidae (35), Cottidae (27), and Cich- lidae (21). These 11 families make up about 80% of the species in the fauna.

Historical biogeography of the fauna was sum-

30

Annals of the Missouri Botanical Garden

marized in the comprehensive treatise Systematics, Historical Ecology, and North American Freshwater Fishes edited by Mayden (1992). Regional bioge- ography was thoroughly analyzed in Zoogeography of North American Freshwater Fishes edited by Ho- cutt and Wiley (1986). Systematic accounts and distributional records have been documented in about four dozen regional, provincial, or state fish monographs and the Atlas of North American Fresh- water Fishes (Lee et al., 1980). Nine families and 128 genera are now endemic to North America, but some (e.g., Amiidae, Hiodontidae) were more wide- spread in the past. The principal intercontinental relationships of most of the fauna (e.g., Cyprinidae, Catostomidae, Salmonidae, Esocidae, Percidae) are with Eurasia, whereas the Characidae, Pimelodi- dae, Cyprinodontidae, Fundulidae, Profundulidae, Poeciliidae, and Cichlidae have neotropical rela- tionships (Patterson, 1981; Grande, 1994).

The combined fauna of Atlantic and Gulf of Mex- ico drainages is several times as diverse as the Pa- cific drainage fauna because its land area is larger and has been geologically and climatically more stable during the Cenozoic. The Mississippi Basin is the center of North American freshwater fish di- versity; its fauna is the largest with 375 species. The Eastern Highlands, mostly in the Mississippi drainage, have the most distinctive fishes with 57 species (Mayden, 1985, 1987). Diversity decreases with distance away from the Mississippi Basin heartland. The vast northern glaciated areas are de- pauperate because of long winters and slow post- glacial recolonization (Fig. 4; see also C. L. Smith,

5). Western and southern faunas are depauper- ate because they are mountainous, arid, and iso- lated by barriers. The Great Basin (Hubbs & Miller, 1948; Hubbs et al., 1974) and Colorado River fau- nas, in western North America, have the lowest di- versity, but highest endemism (Miller, 1959), over 50%, indicating a high extinction rate (G. R. Smith et al., in press).

Freshwaters of the United States were thoroughly explored in the 19th century and the first half of the 20th century; North America now has few un- discovered species except in Mexico and Central America. Recently described additions to the North American fauna are mostly allopatric, divergent lo- cal populations of darters, minnows, and suckers that are elevated to species status based on the phylogenetic and evolutionary species concepts (Mayden & Wood, 1995). An unusual recent dis- covery was Scaphirhynchus suttkusi Williams & Clemmer, a new species of sturgeon from the Mo- bile Basin. Molecular data have also revealed di- verse populations of salmons, trouts, and whitefish-

50 150°, a 40 440 э}. Е з0 20 420

ZA во-1 49-79 25-48

LJ 10-24 СО 1-9 Figure 4. Fish species diversity gradients displayed as nu

cies richness from 80 to 105; 49 to 79; 25 to 48; 10 to 24; 9

es (Bernatchez & Wilson, 1998), some of which are sympatric but ecologically different. Some of these forms fit species concepts based on inferred genetic and ecological differences between populations that spawn in different times and places. With emphasis on historical processes of origin, however, G. R. Smith et al. (1995) defined species as lineages sep- arated from each other by genetically based mor- phological, reproductive, ecological, or behavioral barriers sufficient to confer long-term historical in- dependence.

Monographs on groups of the North American freshwater fish fauna have been produced recently. The most magnificent are The Evolutionary Biology of the Threespine Stickleback, edited by Bell and Foster (1994), and Native Trout of Western North America, by Behnke (1992). Two of North America’s famous relictual fishes and their fossil relatives, paddlefishes (Polyodontidae) and bowfins (Ami- idae), are RT treated by Grande and Bemis (1991, so notable as popular as well as мен а valuable treatments аге The Handbook of Darters by Page (1983) and The Amer- ican Darters by Kuehne and Barbour (1983). The fauna has also benefited from outstanding ecologi- cal and evolutionary treatises, such as that by Mat- thews and Heins (1987). The field eagerly awaits

Моште 87, Митбег 1 2000

Lundberg et al. 31 Ichthyological Discovery

the appearance of the Freshwater Fishes of Mexico, in preparation by Robert R. Miller

Because North American fish species are well described, the most exciting recent discoveries are of evolutionary relationships and patterns. New in- sights on relationships are the result of research in molecular and morphological phylogeny, an area of systematics substantively pioneered by North American ichthyologists (Wiley, 1981; Burr & May- den, 1992). Systematic summaries of most of the important groups can be found in Mayden (1992). The higher-level systematics of representative fish taxa is treated in Stiassny et al. ineteen of the 21 richest North American families have been the object of substantial phylogenetic work (Burr & Mayden, 1992: 32). More than 250 phy- logenetic papers on North American fishes have been published, but much more cladistic work is needed (Burr & Mayden, 1992: 59). Unexpected recent discoveries include the close relationship of the live-bearing Goodeidae (Fig. 30) of the Mexi- can Plateau to the egg-laying Crenichthys and Em- petrichthys of the Great Basin (Parenti, 1981; Webb, 1998) and the relationship of the pygmy sunfishes, Elassoma (Fig. 3R), to the sticklebacks (Johnson & Springer, 1997). Other relationships, based on DNA sequences, include spinedace (Hubbs & Miller, 1960) and creek chubs among cyprinids (Simons & Mayden, 1997), and Pacific trouts and salmons among salmonids (Stearley & Smith, 1993; Philips & Oakley, 1997). In addition, studies of phylogeny have revealed the frequent influence of introgres- sion on evolutionary patterns of North American freshwater fishes (G. R. Smith, 1992; Dowling & Secor, 1997).

New evolutionary analyses relate patterns in fish diversity through time to the geological control of rates of evolution and extinction. In tectonically fragmented and volcanically disturbed areas, high extinction rates control diversity. Comparison of fossil and Recent North American fishes suggests that they have not evolved substantially in response to Pleistocene environmental changes. These re- sults are consistent with those based on discovery of early to middle Cenozoic fishes closely related and similar to extant species in North America and South America (Cavender, 1986; Wilson & Wil- liams, 1992; Lundberg, 1998). Molecular and mor- phological data in their geological context indicate that anagenic change and speciation are much slower than geological and climatic changes (G. R. Smith et al., in press). These results suggest that the current (latest of the past two dozen or so cy- cles) post-glacial species assemblages in glaciated

regions, such as the Great Lakes basin, are not

Б 5 N Q л © о о 1 1 1 J

E o

T T T T T Џ

MEAN NUMBER OF DORSAL SPINES

Џ ct i. 70 80 90 100 110 TIME ( х 1000 YEARS) ure 5. Short-term fluctuations within a 100,000-yr. trend in stickleback spines (from Bell et al., 1985) den onstrating limited significance of ка Баев over the long course of phenotypic evolution

likely to be ecological communities with fine-tuned interspecific interactions.

Studies of rates of fish evolution were pioneered by Hubbs and Miller (1948) in the Great Basin. They documented rapid evolution based on ob- served changes attributed to post-pluvial isolation (in the past 10,000 years) of populations assumed to have been uniform when waters were connected in pluvial times (e.g., Kocher & Stepien, 1997). Current studies negate the assumption of geneti- cally uniform species (G. R. Smith et al., in press). Studies of Pliocene and Pleistocene morphological changes in the Great Basin show that the early, rapid responses to environmental change such as those documented by Hubbs and Miller (1948, 1974) do not usually lead to new species (Bell et al., 1985; G. R. Smith et al., in press). The isolation of small populations in rapidly changing environ- ments promotes rapid changes, i over the lon term these appear to be short-term fluctuations within slow trends (Fig. 5; Bell & "Haglund, 1982; Bell et al.,

Fossil TM suggest that the modern North American families probably date back more than 65 million years to the Cretaceous, most genera to the Miocene, and species mostly to the Pliocene or early Pleistocene. Species formation averages slow- er than one branch per million years per clade in rivers (depending on the family) but may be more rapid in lakes (Echelle & Kornfield, 1984). Molec- ular evolution (Kocher & Stepien, 1997) may be stochastically constant within 10-25% error; for certain mitochondrial genes, it varies from 0.596 sequence divergence per million years in salmonids to about 196/m.y. in cyprinids and cyprinodontoids Fig. 6; G. R. Smith et al., in press). The a estimates suggest that adaptive evolutionary re- sponses to the current global ecological crisis are unlikely.

—

Annals of the Missouri Botanical Garden

32 8 ‚25 c Cyprinidae 5 .20- o $ 15 Cyprinodontoidei 8 ло 5 Salmonid 5 .05- опіаае © Р) Ч Т Т Т 0

5 10 Million years before present

sure 6. Estimated rates of mitochondrial gene se- quence divergence vary from 0.5% to 1% per million years among salmonids, cyprinids, and id E dE Fossil discoveries enable eed. of these rates when synapomorphies of earliest 1 fossils can be used to identify the internode segment or pi of the lineage in

which the fossil fish w ember. The age of the fossil can then be e to е а minimum sana of bis age in millions of years—the deno the equation; The оме sequence eae eg eke

xa for which an age estimate is available Же becomes ifa num cud in the rate equation. The rates are then

sed to e e the ages of other vicariance barriers and ages of

Prospects for future ichthyological discoveries in North America include some new species in Mexico and important advances in our knowledge of pat- terns and processes of fish diversification.

SOUTH AND CENTRAL AMERICA

The vast Neotropical ichthyofauna, estimated to contain between 5000 and 8000 species (Schaefer, 1998; Vari & Malabarba, 1998), inhabits the fresh- waters of South and Central America (Fig. 2), with a handful of cichlids, pimelodid catfishes, and characids extending north into Mexico or the south- ernmost U.S. The great majority of Neotropical fishes belong to one of five dominant groups: characi- forms, siluriforms, gymnotiforms, cyprinodontiforms, or cichlids.

Within the Neotropics the Amazon Basin con- tains Earth's most diverse riverine fish fauna that certainly far exceeds 1000 species. The Orinoco, Paraná, and other large, tropical rivers flowing to the Atlantic are also species-rich. Fish diversity drops sharply in the watersheds emptying into the Caribbean and Pacific, and southward into temper- ate South America where the taxonomic composi- tion also changes markedly. Central American idees contain roughly 300 species (Bussing,

. In this region the San Juan Basin of Nica- ragua and Costa Rica has the most diverse fauna with about 54 species (Bussing, 1985).

Neotropical cichlids and gymnotiforms are en- demic, monophyletic clades. On the other hand, Neotropical characiforms (Vari & Malabarba, 1998), siluriforms (de Pinna, 1998), and cyprino-

dontiforms (Costa, 1998) each contain several sep- arately monophyletic subgroups with incompletely known extralimital relationships. Some Neotropical fish clades have their closest relatives of today in African freshwaters (Lundberg, 1993; Vari & Mal- abarba, 1998; de Pinna, 1998): lungfish, arapaima, ctenoluciid + erythrinid and some characid char- aciforms, doradoid catfishes, aplocheiloid and poe- ciliid cyprinodontiforms, cichlids, and nandids. A few of the ed cyprinodontiforms are most closely related to American taxa (Parenti, 1981). Many ibd fishes, including individ- ual species and some small clades, have their prox- imate relatives and presumed ancestry in coastal marine waters, e.g., river sting rays, various her- rings and anchovies, drums, soles, needlefishes, toadfishes, and a puffer.

It is scarcely surprising that no comprehensive treatise yet exists for the Neotropical fish fauna. However, two recent publications have immensely advanced access to information about the fauna. The massive Catalog of Fishes (Eschmeyer, 1998) provides the most thorough listing ever retrieved of the binomials applied to all fishes and also contains an extensive bibliography of descriptive ichthyolo- gy. The electronic version of the Catalog has great- ly facilitated an estimate of the historical account- ing of published description of Neotropical fishes. The symposium volume Phylogeny and. Classifica- tion of Neotropical Fishes (Malabarba et al., 1998) contains 28 papers that summarize much up-to- date systematic knowledge of higher Neotropical axa.

c

a The discovery of Neotropical fish species became an active enterprise by about 1825 and it continues at a high rate. The latest ca. 50-year trends in spe- cies description for the Neotropical siluriforms, gymnotiforms, characiforms, cyprinodontoids, and cichlids are shown in Figure 7 (data from Eschmey- er, 1998). Overall about 1400 species were de- scribed during this period, and the vast majority of these are considered valid. Using the recent esti- mates for total Neotropical fish species richness, this amount of discovery and description could rep- resent about 25% of the whole ichthyofauna.

The levels of recent discovery and estimates of total species richness for the five major groups are truly i impressive. The siluriforms are the richest of

species (Nelson, 1994). 491 Neotropical catfishes were described, and more than half of these were published in the last 20 years. Several known catfish species await descrip- tion, and certainly other unrecognized and unseen species will turn up in natural history collections

Volume 87, Number 1 2000

Lundberg et al. Ichthyological Discovery

<< Characiformes m æ< Silurifor ^. noti < Сутпо{ Ног = 4 165 Se >= = = я Cic < ч a чч o аф << << <a 150 < < << < < < < << << - = = ок. + 135 «ec a < o «v 4 < c «< E ~ о а сас «< ~ а җы ~ 120 а ee o - о и a а. И = «ьс «асс абс аА ё a ee чада: с «йч © 105 < «і сас сс ойыс аф [7] << «< ae «адаг седи чв Ф << ас сана с ен А аф --— анас «адас андаг фр б << aime age т a 90 я << to tee Em o n << < а €—X abe ес А o = pres > LEM eee або te — Ф 75 с a$ = 2 m —_ <_< << L сан abe abe адаг адас [5] > dn БЕ б ьс «ee abe «бе M ан Ф << << << << « d с або «бо ес и а. ы LJ << 4 ~ I ee tee або ве лес tee << «v | а сас афс чо с << ate — = — << = сана ee аа ed — «ча о 4 cee mee ~ ас сас саас афс са афс эы о a бе бе әдә со | ос 45 | << ЧЕ чс ада аф ол ас о о ве бе йэ 4 до ә саас dM NE o b ete eee << бе ве Фи abe «she ee ee а eee ee << бе ве 4 -— ado p de Lm orn A^ | ———— «abe a —- ад 4 бе n 4 4 = & o ate = a =й abe abe eo ——— «ae «ш ae а & Ф ate ate 15 | бе a аа --— abo = ee —— чё —— = = == & «ae ate aio an чш -— «e ди а - «до де dde 4 XR & 4 -— <= «alte m ебә o © m 4 --—- ле с 4 = = 4 =e ate @ 4 ate ate ote 46 4 + а & = = = 4 4 = а 4 = & 46 T 46 T TE & & 1952 1955 1958 1961 1964 1967 1970 1973 1976 1979 1982 1985 1988 1991 1994 1997 3 - Year periods ending in date shown Figure 7. Neotropical fish species "d "sc ds dry 1950-1997 for characiforms, siluriforms, gymnotiforms, cyprino-

dontiforms, and cichlids based on data

and new field samples. Thus, any estimate of the total species richness of Neotropical catfishes is now imprecise, but close to or more than 2000 spe- cies should be expected Mago-Leccia (1994) NE 94. valid species of electric fishes; Campos da Paz and Albert (1998) estimated the total to exceed 100 species. In just the last 25 years, descriptions of 31 new gymnoti- form species and 13 new genera were published, and we are aware of an additional 32 undescribed species. As with catfishes, it is difficult to estimate the total species richness of Neotropical characi- forms because of their taxonomic complexity and the high rate of discovery of undescribed species. Nelson (1994) estimated an overall total of 1300 characiform species, and Vari (1998) predicted that far greater than 2 exist overall. Of these only about 225 are African. Between 1950 and 1997, over 450 characiform species were described in the Neotropics, and although annual rates of descrip- tion vary, there is no indication of reduction. Ex- perts working with characiform taxonomy are aware of many dozens of undescribed forms. South and Central American cyprinodontiforms are estimated at about 375 species (Huber, 1998;

n Eschmeyer (1998). Each whole fish i

partial fish icons equal one or two desc dcin each column covers a three-year peri

icon represents three descriptions, iod.

Nelson, 1994). From the Neotropics 203 cyprino- dont species have been described since 1950, and of these over 80% (166) were published after 1974. These numbers would be higher if new species in shared and closely related genera from Mexico, the U.S., and the West Indies were included. Neotrop- ical cichlids are not as species diverse as African cichlids, but their numbers are high and increasing. Kullander's current (1998) estimate is that the total will be about 450 Neotropical cichlids, of which more than 100 are undescribed. About 30% (134) of the estimated total have been described since 1974.

The continuing high level of discovery of fish species in the Neotropics may be explained by the combination of an increased amount of field and taxonomic work applied to a very large, taxonomi- cally complex, and undersampled fauna (see also Vari & Malabarba, 1998). We could cite dozens of recent examples of ichthyological discovery coming out of small and incidental to large, focused field exploration throughout the Neotropics. However, the recent history of ichthyological exploration in Venezuela illustrates the great magnitude of dis- covery, on one large, hydrographically complex re-

Annals of the Missouri Botanical Garden

gional scale, that is still possible in many areas of South America. About 30 years ago Mago-Leccia (1970) published the first comprehensive list of Ve- nezuelan freshwater fishes, which included 474 species. In 1997 Taphorn et al. were able to revise the Venezuelan list upward to 1065 species (a 125% increase) based largely on the combined re- sults of much survey work carried out during the intervening approximately 25 years.

The number of taxonomists involved in describ- ing Neotropical fishes has increased over the years, although we may have reached a plateau. Using characiform taxonomy as an example, for the 10 quarter-century periods between 1758 and 1990 the numbers of authors of species descriptions were 4, 5, 9, 18, 12, 31, 28, 38, 64, 57, and in the 7 years since 1990 already 54 authors have been in- volved in describing characiforms. Between 1970 and 1997, 228 authors contributed to papers de- scribing approximately 960 species of ee

characiforms, siluriforms, gymnotiforms, cyprino- dontiforms, and cichlids. As expected, рана productivity of species descriptions varies tremen-

ously: the range of descriptions across author is 1-102, with a strong mode of 1 description per au- thor, and a median of 2. Seven highly productive individuals were sole or co-authors of at least 40 species descriptions and, remarkably, the total con- tribution of these 7 persons accounts for over half (524) of the 960 recent descriptions.

A general pattern of biodiversity is that species richness increases with sample size, e.g., with area, time, specimens, taxonomic scope, etc. The effect of taxonomic scope on Neotropical fish diversity is evident at different ranks. For example, differences in the amounts of species discovery among catfish families are correlated with the species richness of those families (Fig. 8). The same relationship ap- pears to hold at the genus level with the very large genera Corydoras, Aspidoras, Trichomycterus, and Hypostomus accounting for over 3096 of all Neo- tropical siluriform species described during the last ca. 25 years. Among characiforms, too, the large genera Leporinus, Hyphessobrycon, Cyphocharax, Curimata, Astyanax, and Creagrutus account for about 3096 of the species described in the same

riod.

Exemplifying the great yield of new species that can be found by in-depth study of relatively small- bodied characiforms, Vari and Harold (1998) have raised the species number of Creagrutus (east of the Andes) from 15 to 56, a 37396 increase. Re- visionary work at the species level seems more of- ten than not to produce a net increase in the

ber of Neotropical fish species. Kullander (1980).

150- m ~ ш г = .96

Lo ш о 2100 2 a ul a Ca * 504 d Pi та Тг Ф sc Au t Do Di 100 200 300 400 500 600 CURRENT TOTAL SPECIES Figure 8. Relationship between species richness of

Neotropical catfish families and numbers of new species described during the last ca. 25 yr. based on data from Eschmeyer ipu Ар = Азрг єй йе, At = Astroble- pidae, Au = Auchenipteridae (inc luding Ageneiosidae and Centromoc Ме), Са = Callic

5 & d. E Ф

|

Trichomycteridae. (One fossil but no extant species of Ме- matogenyidae were described in this period.)

for example, reported that by 1977, 31 nominal species of the cichlid genus Apistogramma had been described, of which he judged only 20 to be valid. In his 1980 revision, Kullander added 18 new Apistogramma; he recognized 36 overall and estimated that the genus contains > 40 species. By 1997 an additional 17 species of Apistogramma had been described, about a 50% increase in the 17 years following the initial revision. On the other hand, revisionary work may sometimes suggest con- specificity that might yield a net reduction in the number of recognized forms. In a revision of the widely distributed catfish genus Rhamdia, with about 100 nominal species, Silfvergrip (1996) pro- posed that only 11 species are valid (the author described 3 of these in the wor

Whereas the numbers of new species are high and the common taxa are becoming more so, the qualities of recent ichthyological discoveries in the Neotropics are perhaps more impressive than quan- tity. Neotropical fishes are famous for their diversity of diet and trophic apparati. New species with strik- ingly distinct feeding specializations are common. Among characiforms the most unusual recently found dietary specialist is the serrasalmine Ossub- tus xinguense (Fig. 9A; Jégu, 1992), a species from turbulent cataracts that feeds on aquatic macro- phytes, especially Potemogeton. During its devel-

Volume 87, Number 1 0

Lundberg et al. 35 Ichthyological Discovery

New dietary specialists from the Neotropics.

Figure 9. —A. Ossubtus xinguense, left, mouth; right, an adult specimen about 153 mm long (re- produced with permission from Jegu, 1992). —B. Magos-

drawings of teeth and

ternarchus duccis, left, anterior third of adult specimen skeleton of teeth and jaws (re- 1996). —

. Sternarc AI sp. (Copy right John С. Lundbe TE and hn P. Sullivar

about 235 mm long; right, produc ‘ed with permission from Lundberg et al.,

opment the snout, jaws, and gill chamber of Ossub- tus undergo a metamorphosis, twisting ventrally so that the mouth and blade-like teeth are directed at the plants attached to the rocky substrate. An in- teresting twist was added to this case with the dis- covery of a parasitic isopod living in the gill cham- ber of the fish that also develops a contorted morphology, presumably in response to its host’s cursive ontogeny (Thatcher, 1995).

Most gymnotiforms have generalized diets of

small invertebrates or fishes. However, the recently described genus Magosternarchus (Fig. 9B; Lund- berg et al., 1996) includes two species of Amazon Basin apteronotids that, with greatly enlarged jaws and teeth, feed on the tails of other electric fishes. Another “new” specialist among electric knife fish- es is Rhabdolichops zareti (Lundberg & Mago-Lec- cia, 1986), from deep channels of the Orinoco, that feeds mostly on allochthonous zooplankton washed into the rivers mainstem channels from the pro-

1987).

Providing an example of an odd newly discovered

ductive marginal savannas (Lundberg et al.,

structure possibly related to electrolocation of prey, an undescribed species of the apteronotid Sternar- chogiton bears a probe-like, somewhat protrusible organ on its lower jaw (Fig. 9C)

The evolution of small to miniature-size species is commonplace among Neotropical fishes (Weitz- man & Vari, 1988). It is no surprise that very small fishes are discovered relatively late in survey work; 40 of the 85 little species listed by Weitzman and Vari were described in the latest 25-year period, more than twice that of the previous quarter cen- tury, and several more have been discovered re- cently that fall below or approach their size crite- rion of 26 mm. In the Neotropics the evolution of reduced size is most common among characiforms, siluriforms, and cyprinodontiforms, and there are a

andful of remarkably small engraulids (Amazon- sprattus scintilla, Roberts, 1984) and gobioids (Mi- crophylipnus species, Myers, 1927). Weitzman and Vari (1988) pointed out that miniature species of most taxa live in slow-flowing or still waters. The most strikingly modified small and miniature cat- fishes, however, are found in swiftly flowing and often deep, river channels. Examples of these in- clude the aspredinid Micromyzon akamai (Fig. 10A; Friel & Lundberg, 1996), a blind, nearly pig- mentless, heavily armored species that matures at about 12 mm, making it one of the smallest known catfish species. Bathycetopsis oliveirai (Fig. 10B; Lundberg & Rapp Py-Daniel, 1994) is a deep-water ca. 35-mm cetopsid catfish that has completely lost its eyes and pigment but has enormously hypertro- phied olfactory organs. There are two new groups of very small, microphthalmic, channel-dwelling pi- melodids: Horiomyzon retropinnatus (Fig. 10C; Stewart, 1986), with wing-like pectoral fins, and an undescribed clade with at least four species bearing thickened bones and skin studded with sensory or- gans.

At the other end of the size spectrum, there are occasional discoveries of large to very large fish species. Toledo-Piza (1997; Toledo-Piza et al., in press) has determined that the well-known fang- toothed cynodontine Hydrolycus, long considered monotypic, contains at least four species of which two are undescribed. Lundberg and Akama (1999, and in prep.) have identified an unrecognized spe- cies of “goliath” catfish, Brachyplatystoma (Fig. 10D), in the central and lower Amazon. This im- portant species is now known to reach close to a meter in length, and it is among the most common in the commercial and artesanal fisheries where it is confused with B. filamentosum.

Annals of the MESE Botanical Garden

кк 10. fishes

long ^ rode ed wi ith pe 996

Recent discoveries of very small and large the Amazon. —A myzon akamai, rmission n from

14 mm Friel & Lundberg,

~ X

S N N © = т ® LI = S — 5 = = 5 & = = z ~ = = ge = % О

produced with permission from Stewart, . —D. Bra- chyplatystoma sp., specimen about 500 mm long. (Сору- right John С. Lundberg.)

-

Novel aspects of reproductive biology are being discovered in Neotropical fishes. In addition to the continuing discovery of new genera and species & Menezes, 1998; Malabarba, 1998), recent investigations of the glandulocaudine and cheirodontine Characidae have turned up a

see Weitzman

new mode of reproduction for characiforms. These with males developing a variety of specialized, court- ship-related features of the caudal fins such as bent fin rays with hooks and fleshy tissues that appear to be glandular (Fig. 11A; Burns et al., 1995, 1997; Malabarba & Weitzman, 1999). Almost all male glandulocaudines develop a cutaneous caudal gland thought to produce a pheromone, and the gland is surrounded by hypertrophied scales and in some species putative *pumping"

fishes are convergently sexually dimorphic,

muscles. Oddly, although the males lack any obvious intromittent organ, many of these species are internal fertilizers.

The known limits of sexually dimorphic charac- teristics are being stretched by some Neotropical fishes. Recent collections made during the high- water season near Manaus have turned up breeding associations of electric fishes and morphologically intermediate individuals showing two cases of ex- treme sex dimorphism that have misled taxonomic decisions. The nominal species Aperonotus anas is

‘igure 11.

E p sexual dimorphism found in | X

fishes. —A. Left, Xenurobrycon

Fig Neotropical male о characin dal gland; right, 1 in male Mimagoniates T (from Weitzman & Fink, 1985;

mac ropus, with a cau-

odified scales surrounding caudal gland

‚ 15.4 mm long,

ermissio n to

офисе images granted by Smith- The

NMNH. Division: of Fishes). —B.

sonian Institution,

а ия exodon. (Copyright John undberg and John P. Sullivan). —C. Neblinichthys pilo-

sus, male about 50 mm long (reproduced with permission from Ferraris et al., 1986).

based on a large, hypermorphically long-snouted male of A. hasemani (Cox Fernandes, 1998; Cox Fernandes & Lundberg, 1999). The nominal, mono- typic Oedemognathus exodon is based on large males of Sternarchogiton nattereri that develop strong externalized teeth, perhaps for use in combat over mates (Fig. 11B; Cox Fernandes & Lundberg, in prep.). Ferraris et al. (1986) described a bizarre catfish, Neblinichthys pilosus (Fig. 11C), in which males develop on their snouts a cartoon-like bush of elongate odontodes, i.e., cu-

loricariid armored

taneous teeth.

Among discoveries during the last 25 years, the greatest scientific interest attaches to previously unseen fishes that represent new phylogenetic lines

Volume 87, Number 1 2000

Lundberg et al. 37 Ichthyological Discovery

at or above the family level. These discoveries have been most common among catfishes and often result in broad changes in higher classification. The lor- ic — Scoloplax dicra (Fig. 12A, B; Bailey & Bas- kin, 1976) was first described as a new subfamily of es 'ariidae, but was soon elevated to family rank by Isbrücker (1980). Three more Scoloplax species were described by Schaefer et al. (1989), and Schaefer (1990) determined that this clade is the likely sister taxon of the vast assemblage of Loricariidae + Astroblepidae.

е Trichomycteridae presently include about 200 species (perhaps over 100 in the large genus Trichomycterus) arranged in eight or nine subfami- lies (de Pinna, 1998). The most widely known tri- chomycterids are the *candirás? or “parasitic cat- fishes” that feed within the gill chambers of larger fishes on blood and epithelia. (Candirás finally earned their notoriety for supposed inadvertent en- try into the urethrae of humans with the first med- ically verified case of such behavior in. Brazil in 1998.) Of more scientific interest, however, is the recent explosion of systematic knowledge of the perplexing diversity of trichomycterids. Although 16 of the 27 trichomycterid species described be- tween 1975 and 1997 belong to Trichomycterus, the remaining 11 (4096) are placed in new or previ- ously rare, poorly known genera and subfamilies. The highly derived Trichogenys (Fig. 12C, D; Brit- ski & Ortega, 1983) has been placed in its own subfamily by Isbrücker (1986). Two scribed genera, Copionodon (Fig. 12E, F) and Gla-

newly de-

phyropoma, form a small, plesiomorphic clade of three species that de Pinna (1992) placed as the sister taxon to all other trichomycterids. The sub- families Sarcoglanidinae and Glanapteryginae, known from very few specimens, contain perhaps the most extraordinarily spec alized and diminutive catfishes in the world (Fig. 12G, n addition to major range extensions, in just the last decade the ranks of the sarcoglanidines were increased by four new monotypic genera (e.g., Fig. 12K, L; de Pinna, 1989; de Pinna & Starnes, 1990; Costa & Bock- mann, 1994b; Costa, 1994), and two species and one genus were added to the glanapterygines (de Pinna, 1988, 1989; Costa & Bockmann, 1994a). It is no surprise that this remarkable amplification of trichomycterid diversity at the genus and family- group levels has caused a significant revision of our phylogenetic understanding (de Pinna, 1992, 1998; Stiassny & de Pinna, 1994). This case emphasizes the major impact that taxon sampling can have in Neotropical fish systematics (see also Schaefer,

Discoveries of new species and new character

evidence in known clades often lead to altered phy- logenetic arrangements and classifications. Exam- ples include the catfishes Helogenes, discovered to be closely related to the Cetopsidae (de Pinna & Vari, 1995), and Hypophthalmus (Fig. 13A), which is now confidently placed within Pimelodinae in- stead of in isolation in its own family (Howes, 1983; Lundberg et al., 1991). The first examinations (only within the last three years) of skeletal specimens of the monotypic, по Зао Francisco endemic Cono- rhynchos conirostris (Fig. 13B) have revealed that this supposed pimelodid, described in 1840, does not belong to any diagnosed family group of silur- iforms (Bockman, de Ferraris, comm., pers. obs.). There are noteworthy recent higher-level discoveries in other Neotropical groups as well. Stiassny (1991) has found synapomorphies that unite New World cichlids as the monophyletic sister taxon of some African cichlids. On the other hand, there is increasing evidence of multiple sis-

inna pers.

ter-taxon relationships between African and South American characiform and cyprinodontiform clades (Parenti, 1981; Vari, 1995; Buckup, 1998; Costa, 1998)

Thus, at every level knowledge of Neotropical fishes continues to grow at remarkable rates: myr- iad species, their intra- and intercontinental rela- tionships, and their richness and novelty of char- acteristics and natural history. Prospects for ichthyological discovery in South and Central America continue to be as high as ever.

EUROPE

Europe west of the Ural Mountains comprises roughly one-third of the Palearctic Realm (Fig. 2). The major groups accounting for most (ca. 8096) of the fish species are Cyprinidae (129), Salmonidae 54), Coregonidae (43), Gobiidae (31), Cobitidae 21), Petromyzontidae (11), Clupeidae (11), and Percidae (11). The “classical” checklists of Euro- pean freshwater fishes are outdated and reflect a state of knowledge and species concepts of 30—40 years ago. Maitland (1976) listed 215 species for Europe west of the Urals and 170 species in Europe

йл адар

exclusive of the former U.S.S.R. In а recent review Kottelat (1997) recognized 358 species in Europe (excluding the former U.S.S.R.), an increase of

Northern Europe, much of it glaciated during the Pleistocene, has low fish diversity, usually with only 2 or 3 native species in any watershed. The number of species increases to the south. North of the Alps from France to Russia, the fauna of about 30 spe- cies is relatively uniform. The Danube basin, with

38 Annals of the Missouri Botanical Garden

(251 iu

А NS АА

Figure 12. Recent discoveries of phylogenetic importance among South American sc oloplac ‘id and trichomycterid à , B. The sc oloplac ча Scoloplax dicra, scale bars | mm (reproduced with permission from Bailey & Baskin, 1976). —C, p. The trichogenine Trichogenys longipinnis (from Britski & Ortega, 1983). —E, F. The горот Copionodon pecten, about 60 mm long (reproduced with permission from de Pinna, 1992). —G, H. The glanapterygine E an киша, about 55 mm long (reproduced with permission from de Pinna, 1989). — 1, J. The ТИ ine амтигта camposi, about 38 mm long (reproduced with permission from de Pinna, 1988). —К, L. The sarcoglanidine Stauroglanis gouldingi, about 23 mm long (reproduced with permission from de Pinna, 1989).

el T

Volume 87, Number 1 2000

Lundberg et al. 39 Ichthyological Discovery

Figure 13. among long-known South

а fimbriatus. — В. Соп

Recent discoveries of phylogenetic impor-

бн ‘an catfishes. —A.

orhynchos conirostris. 2

(Copyrights John G. Lundberg and jue. P. Sullivan

about 90 species, has the most diverse fauna on the continent. The fish faunas of the southern European peninsulas contain relatively high numbers of en- demic species but are otherwise species-poor as- semblages: 87 species in the Balkan Peninsula, 29 in the river basins of the Italian Peninsula, and 26 in the Iberian Peninsula. The only endemic Euro- pean fish family is the cyprinodontiform Valenci- idae, with two species. This fauna has its closest relationships with Asia, North America, and north- ernmost Africa.

The cornerstones of European fish systematics were provided in the 16th century in Rondelet's Libri de piscibus (1554) and Gesner’s Nomenclator aquatilium (1560) and Fischbuch (1563). Modern European ichthyology started with Peter Artedi's Ichthyologia (1738), whose system and nomencla- ture were adopted by Carl von Linné in 1758.

Having thus enjoyed 450 years of study (about double that of any other continental fish fauna), one would expect that our modern systematic knowl- edge of European fishes is definitive. This is not the case, and even the most basic question, “How many species?," has no easy answer. That approx- imately 2000 names have been applied to Euro- pean fish species suggests that their taxonomy is not simple (Kottelat, 1997). As with North America, the discovery of previously unknown fish species in European waters is now uncommon. How then do we account for great increase in the number of valid European fish species proposed by Kottelat (1997)?

First, there have been some recent discoveries of a few previously unknown species. Twenty spe- cies were discovered in European waters between

gure 14. Valid European cyprinid species previous-

ly dnd by копу us . Scardinius acarnanicus, 152 m long. —B. Scardinius graecus, 140 mm long. (Copy- rights Maurice Какы t.)

1978 and 1998, and some 10 known species are still unnamed. These are cyprinids, cobitids, and gobiids of small to very small size (3 to 10 cm). Most were found in southern Europe on the Iberian, Italian, and Balkan peninsulas. In 1998, two new cyprinids and one new cobitid were described from the Iberian Peninsula.

Second, several species described by earlier au- thors and considered invalid by later authors, have since been demonstrated to be distinct. This also mainly concerns species from southern Europe. For example, the species of the cyprinid genus Scar- dinius have all been treated as synonyms of S. er- ythrophthalmus. It appears that at least five species should be recognized, some of which are conspic- uously distinct from 5. ү (Fig. 14) (Iliadou et al., 1996; Kottelat, 1 )

Another interesting case is the E Carassius gibelio, originally described from Germany in 1782. This species is traditionally considered as either the wild form of the goldfish, a feral goldfish, or a hybrid. These theories ignore the Japanese litera- ture on morphology and karyotypes of goldfishes that documents the existence of at least five species of Carassius in east Asia, all of them distinct from C. gibelio (see Hosoya, in Nakabo, 1994, for a sum-

ary). They ignore the fact that C. gibelio was ap- pret recorded in European waters as early as 9 (Kentmann’s Codex; Hertel, 1978), long be- E goldfish were first imported to Europe in 1611 or 1691, and maybe even before it was imported from China to Japan (between 1502 and 1748). The hybrid theory does not seem to have support. So given our current knowledge, there is no alternative but to recognize Carassius gibelio as a distinct spe-

cies. Third, application of the Phylogenetic Species

Annals of the Missouri Botanical Garden

Concept (PSC) has played a role in increasing the number of recognized ee (Kottelat, 1997). In this view, a species is “an irreducible (basal) clus- ter of organisms diagnosably distinct from other such clusters, and within which there is a parental pattern of ancestry and descent” (Cracraft, 1989: 34-35). Most earlier work on European fishes ei- ther used some sort of implicit Biological Species Concept or, more frequently, was not explicit re- garding concepts, definitions, and criteria. Incon- sistencies in application of criteria are common. Detailed studies of geographic variation and sys- tematics reveal that fish diversity often has been underestimated, and there is no theoretical or prac- tical advantage in underestimating biodiversity. Subspecies of earlier authors that satisfy species criteria under the PSC must be treated as species. Further, some traditionally recognized subspecies have been found to be nothing more than arbitrarily delimited sections of clines, and these entities must be abandoned because there is no sense in recog- nizing arbitrarily defined units.

e systematics of salmonids and coregonids has been notoriously difficult because of real or per- ceived high degrees of character plasticity and fail- ure to find consistent morphological differences among populations. Most earlier treatments avoided dealing critically with the systematics of salmonids and coregonids, and this resulted in a much un- derestimated number of species. Some “classical” lists merely include a single coregonid taxon, *Cor- egonus sp." The species-level systematics of these families is still far from resolved, but a pragmatic handling of their taxonomy is possible. Using a few simple concepts and definitions, Kottelat (1997) tentatively recognized 44 monophyletic evolution- ary units within the genus Coregonus in Europe that can be recognized as species (cf. 7 species recog- nized by Blanc et al., 1971), and 27 within the genus Salmo (cf. 5 recognized by Stearley & Smith,

). Careful comparisons often show constant differences in morphology and in color patterns, and this is now supported by molecular data (e.g., Bernatchez & Dodson, 1994; Bernatchez et al., 1992). The distinctive endemic species of Salmo provide examples (Fig. 15). Among these, S. trutta (Rhine basin) and S. rhodanensis (Rhóne basin) oc- cur in creeks only a few kilometers apart along the divide between the two basins in western Switzer- land. These species have always been empirically treated as different on the basis of coloration and morphology, a fact corroborated by molecular data.

Some may object that it will be impossible to handle the large number of names that application

of the PSC will produce. This is, however, not the

Fig e 15.

c of three endemic and one wide-

pean Salmo. —A. Salmo carpio, 33 mm long, endemic to Lake Garda, Italy. — . Salmo peristerus, 236 mm long, endemic to the Lake B евра basin, Greece-Albania-FYROM. —С. Salmo rho- danensis, 305 mm long, endemic to the Rhóne basin. — D. Salmo trutta, 235 mm long, spec imen from Rhine ba- sin, Switzerland, caught about 5 km from C. (Copyrights Maurice ud )

point; if they are different species we have to han-

le them as species, give them names as species, and manage them as species. The problem is not the names, but with our perceived limitations. Ar- tificially squeezing all trout species into the same pigeonhole risks confusing critical information about species distinctions. If details of ecology of one species are mixed with the migration data of another and the reproduction of a third one under a single name, it is not surprising that patterns of species diversity are blurred.

Salmonoids are notorious also for the existence in many lakes of several sympatric stocks that have been о considered as different “forms” of a single species. In many cases, the “forms” are not only MADE M different, but have differ- ent habitats, prey on different organisms, have dif- ferent spawning seasons and spawning grounds, and are genetically distinct (but few have been in- vestigated on this last aspect). Under any species concept, they are different species, and several of i thus are groups of

these “polymorphic species”

Volume 87, Number 1

Lundberg et al. 41 PAM D Discovery

species or possibly species flocks. Presently, the best example is probably the three or four species of Salvelinus of Lake Thingvalla in Iceland. Euro- pean examples are the Salmo of Lough Melvin (Ire- land) and Lake Ohrid (Albania and the former Yu- goslavian Republic of Macedonia) and the Coregonus of Lake Konstanz and Lake Thun.

AFRICA

According to current estimates, Africa (Fig. 2) harbors some 2850 species arrayed, depending upon taxonomic convention, within 40—50 families. While perhaps not as rich in species as other trop- ical regions, what Africa lacks in terms of raw num- bers it makes up for in an ichthyofauna that in- cludes some striking examples of evolutionary phenomena ranging from relictual “living fossils” and strange morphological isolates, to stunningly diverse species flocks exhibiting unparalleled spe- ciation rates and adaptive radiations. It is notewor- thy that African fish diversity is marked by an in- teresting preponderance of archaic, phylogenetically isolated taxa. Fishes such as bichirs (Polypteridae), lungfishes (Protopteridae), the butterflyfish (Panto- dontidae), weakly electric elephantfishes (Mormy- ridae), the denticle herring (Denticipitidae), the snake mudhead (Phractolaemidae), and knerids (Kneriidae) are among the many examples found today only in African freshwaters. Perhaps more fa- miliar, and certainly more numerous, are members of the two major freshwater fish radiations: the oto- characiforms, and siluriforms

—

physan cyprinids, and the percomorph cichlids. In Africa an inter- esting taxonomic dichotomy between riverine and lacustrine communities exists. The former are dom- inated by otophysans: cyprinids, characiforms, and a few dominant catfish families (Clariidae, Mochok- idae, and Claroteidae), along with mormyrids and cyprinodontiforms (particularly in rivers of west and central Africa). On the other hand, Africa's many lacustrine ecosystems, particularly those of the Great Rift Valley, are almost completely dominated by remarkable adaptive radiations of cichlid spe- cles.

ven the enormous size of the African conti- nent, an ichthyological tally of 2850 species, al- though unquestionably an underestimate of actual species richness, is remarkably low. By way of par- tial explanation, the common vision of Africa’s in- land waters as characterized by the great Nile, Ni- ger, Congo (Zaire), and Zambezi Rivers is misleading. In fact, over 90% of Africa’s rivers are less than 9 km in length and many flow only sea- sonally. Surprisingly, Africa has more arid and

semi-arid area than any other continent on the planet, and as a result the great majority of Africa’s sh biodiversity is concentrated in the tropical, moister (and historically forested) regions.

Roberts (1975), building on foundations laid by Boulenger (1905) and Pellegrin (1911, 1921), di-

vided Africa into 10 so-called ichthyofaunal prov- inces reflecting broad areas of endemicity. Green- wood (1983) has likened the state-of-the-art to bioaccountancy rather than biogeographic analysis, and Leveque (1997) noted recently that the bioge- ography of African freshwater fishes remains poorly

nown. The problems are manifold: new taxa are constantly being discovered, distribution patterns for most taxa are poorly known, and range exten- sions often encompassing thousands of kilometers are far from uncommon, but perhaps most impor- tantly there is a profound lack of adequate hypoth- eses for the phylogenetic relationships of the spe- cies involved. Despite these shortcomings, the ichthyofaunal provinces as currently recognized (see Leveque, 1997, for an up-to-date summary) are a useful first-pass approach to subdividing the ich- thyofauna into meaningful biogeographic units. Un- questionably, the fauna is phylogenetically richest and most varied in the rivers of tropical west and central Africa, with numbers attenuating in the provinces to the north and south of the tropical band. However, in terms of raw species numbers, it is the cichlids of the lakes of east Africa that dominate the continent's inland waters.

Overall African fishes exhibit a very high level of endemism; almost 100% at the species level and upward of 40% at the familial, no doubt a reflection of the long-term stability and isolation of the Afri- can continent. To the extent that extracontinental affinities are known, it seems that the dominant re- lationship is with Asia (e.g., Notopteridae, Cypri- nidae, Bagridae, Schilbeidae, Clariidae, Channidae, Anabantidae, Mastacembelidae, Scatophagidae, pellonuline clupeids, and etropline cichlids), al- though links with South America (e.g., Lepidosiren- idae, arapaimid osteoglossomorphs, some characi- forms, siluriforms, cyprinodontiforms, and cichlids) and Europe (some cyprinid genera) are also evident in a few clades. Unfortunately, our current under- standing of the phylogenetic relationships of most African clades is insufficient to enable historical interpretation of many distributional data.

While the first African fishes to reach European collections were assembled by the French naturalist Michel Adanson during his stay in Senegal (1749— 1753), much early exploration in Africa was cen- tered on the Nile, a river that has played a major

42 Annals of the Missouri Botanical Garden

Number of species described

0 тиз т == Т LI Li т Тт 1758- 1779- 1799- 1819- 1839- 1859- 1879- 1899- 1919- 1939- 1959. 1979- 1768 1788 1808 1828 1848 1868 1888 1908 1928 1948 1968 1988

Species discovery plotted for three African fish clades (Citharinidae, Mormyridae, and Cichlidae). Black bars in ndic rate the number of species described in 10-year increments from 1748 until 1998. Data compiled mainly from Eschmeyer (1998).

role in the history of Mediterranean civilization (see 1992) for most of west Africa and Skelton (1993) Beadle, 1974, for an excellent discussion of early for southern Africa (including the Zambezi prov- exploration of Africa's inland waters). Паре! (1994) ince), have comprehensive coverage. Additionally, recounted an amusing anecdote in which it is much up-to-date regional information is to be found claimed that on Geoffroy Saint-Hilaire's return from in Teugels et al. (1994), and references to major the Napoleonic Campaigns in Egypt (1799) his fish — revisional studies are to be found in Leveque collections were deposited at the Paris Museum. (1997). As was noted earlier, the great majority of hen Cuvier examined for the first time the ex- African species are members of just two groups: the traordinary bichir (Polypterus), he is reputed to — otophysan cyprinids (ca. 475 species in 23 genera), have declared, “This alone is justification for the — characiforms (characids and citharinids with ca. Egyptian Campaign." Whatever the importance of 208 species in 39 genera), and clariid (ca. 74 spe- nilotic fishes, the real beginnings of modern African cies in 12 genera), clarioteid (ca. 98 species in 18 ichthyology are perhaps best marked by the pub- genera), and mochokid (ca. 167 species in 10 gen- lication of Cuvier and Valenciennes’ multi-volume era) catfishes; and the percomorph cichlids (ca. 870 Histoire naturelle des poissons (1828-1849), in species in 143 genera). Together these seven fam- which about 140 African fishes were described. ilies comprise more than two-thirds of the conti- Some 50 years later when Boulenger published his — nent's ichthyofauna. four-volume Catalog (1909-1916) he was able to Despite almost 200 years of scientific study, record over 1400 species. And by 1991, with the there remains a remarkable amount to learn about publication of the last in an extremely useful series — African fishes, and many surprises no doubt re- of checklists (Базе! et al., 1984, 1986, 1991), the main. At the most basic level, monitoring species tally had risen to about 2850. Although Boulengers ^ numbers (Fig. 16) illustrates the cumulative dis- Catalog remains the only pan-African faunal study, covery rates for three typical African fish clades numerous regional accounts have since been pub- and, as elsewhere in tropical freshwaters, species lished. Two in particular, Leveque et al. (1990, discovery continues, sometimes at an extraordinary

Volume 87, Number 1 2000

Lundberg et al. 43 а Discovery

Ole Seehausen

Figure 17. with permission; иын Ole Seehausen.

rate. For example, recent sampling in east Africa’s Lake Victoria has revealed the existence of a pre- viously unknown assemblage of over 120 stenotopic rock-dwelling cichlid species (Fig. 17) (Seehausen, 1996; Seehausen et al., 1998). The quite unex- pected discovery of this extraordinary radiation not only has major implications for our understanding of the evolutionary dynamics of Africa's cichlid flocks (Kaufman et al., 1997; Turner, 1997) and their conservation (Kaufman, 1992; Kaufman & Ochumba, 1993; Seehausen et al., 1997), but it also illustrates that even in supposedly well-sampled lo- cales there still remain many unknowns.

Equally illustrative is a recent study of the Cross River drainage of Cameroon and Nigeria. Although the Cross was considered reasonably well sampled when Teugels et al. (1992) thoroughly reviewed re- cently collected and historical material in muse- ums, they found that previous figures underesti-

^cent disc оуегу of an unknown assemblage

Lake Victoria. (Reproduced

of rock-dwelling cichlids in

mated true diversity by more than 70% (Stiassny, 1996). Even after that comprehensive study new taxa are turning up. For example, Stiassny, Schliew- en, and Freihof (in prep.) are describing an enig- matic new genus of cichlid fish recently collected in the Cross (Fig. 18). This taxon, which is highly distinctive in appearance, displays a fascinating combination of character states that render its phy- logenetic placement problematical and may result in significant changes to current views of relation- ships among tilapiine cichlids (a major clade of Af- cichlids of considerable economic

rican impor-

ance). Another recent discovery altering our view

=

of the evolutionary dynamics of tilapiine cichlids is that of the Lake Bermin species flock (Fig. 19). ake Bermin, a small Cameroonian crater lake of little more than 0.5 km? has recently been deter- mined to be home to the continent’s first species

—

flock of substrate-spawning tilapiines. The flock in-

Annals of the Missouri Botanical Garden

Teugels « et al.

ЖЕ

85

Estimated Number of species 8

ANN bs zie

00:

m

Before 1992 study

After 1992 study

~

е 18. о estimates of fish diversity in the Cross River basin of West 992).

cludes the smallest known tilapia, Tilapia snyderae, a species that attains sexual maturity at only about 25 mm. The discovery of endemic cichlid flocks in small crater lakes in the Cameroonian highlands provides a marvelous series of models for the study of evolutionary diversification and speciation mech- anisms (Trewavas et al., 1992). For example, Schliewen et al. (1994) rec- ognized the Lake Bermin and Lake Barombi Mbo cichlid flocks as providing the most compelling ex-

1972; Stiassny et al.,

Africa.

Data compiled from

amples of probable sympatric speciation known

among vertebrates

The cichlid radiations of the “living laboratories of evolution

likened to

east African lakes, es pro-

vide some of the most extraordinary examples of vertebrate speciation, adaptive radiation, and eco- morphological diversification known on the planet and have intrigued biologists since their initial dis- covery at the end of the last century (e.g., Moore, 1903; Fryer & Iles, 1972; Fryer, 1976; Greenwood,

Range of interspecific variation in LPJ morphology among

Lake Bermin tilapias

T.snyderae 7

А > DEL

T T thysi

2 117 у ^, ^ / т hi { у tte S T bemim nel

Figure 19.

The Lake Bermin (Cameroon) Tilapia species flock. Data and illustrations after Stiassny et al. (1992).

Volume 87, Number 1 2000

Lundberg et al. 45 Ichthyological Discovery

а Whee ли ы В. brevicephalus

CLERK B. peer

Volume-percentage

т УУУ

Analysis of gut contents of Lake Тапа Barbus species

Figure 20.

1981; Coulter et al., 1986; Galis, 1998; Stiassny & Meyer, 1 e recent explosion in molecular sequencing techniques and analytical methods has resulted in a plethora of studies aimed at unravel- ling phylogenetic, biogeographic, and speciation mechanisms among these fishes (e.g., Sturmbauer & Meyer, 1992; Meyer, 1993; Kocher et al., 1993; 1994; Kornfield & Parker, 1997;

1996, and references therein). The

Schliewen et al., Verheyen et al., results have been stimulating to the field generally, have added significantly to our understanding of evolutionary modes and speciation mechanisms of the lake cichlids, and have served to highlight the important roles for both morphological and molec- ular analyses.

Another recent discovery of considerable interest and evolutionary implication is found in Ethiopia’s Lake Tana, the highland source of the Blue Nile. In an elegant series of taxonomic and ecomorphol- ogical studies of trophic diversification, Nagelkerke and his colleagues have established the existence of an ecologically and reproductively segregated ra- diation of 14 species of the cyprinid genus Barbus (Fig. 20; Nagelkerke et al., 1994; 1997). Since the demise of the Lake Lanao cyprinid radiation in the Philippines, the Lake Tana flock becomes the planet’s only remaining lacustrine cyp- rinid radiation.

Lest the impression remains that new discovery is limited to Africa’s lakes, a few riverine examples serve to redress the balance. Recent work that com-

agelkerke,

The Lake Tana (Ethiopia) Barbus species floc

k. Data and illustrations after Nagelkerke (1997).

bines neurobiology, behavior, and morphology has revealed remarkably rich communities of sympatric mormyrid taxa living in small rainforest rivers of west and central Africa (Hopkins, 1981; Moller, 1995). This is especially well illustrated for species in the genus Brienomyrus from Gabon (Fig. 21), where 11 new species have been discovered by re- cording distinctive electric organ discharges (Al- ves-Gomes & Hopkins, 1997) and detailed mor- phological study (Teugels & Hopkins,

Teugels & Hopkins, in prep.). Mormyrids ереси electric pulses many times per second for purposes of active electrolocation, and as a consequence spe- cies-typical and sex-specific signals are broadcast nearly continuously. The diversity of wave patterns among sympatric species suggests that individuals exploit differences in waveform, polarity, and du- ration to identify the species and sex of signalers. Playback experiments using computer-synthesized EODs have confirmed their importance in repro- ductive isolation and sex recognition (Hopkins & Bass, 1981). There are even individual differences in EODs that are sufficient to permit the tracking their in small streams monitored daily (Friedman & Hopkins, 1996), thus enabling non-intrusive ecological mon-

of individuals and movements

itoring of populations.

Electric catfishes (family Malapteruridae) are en- demic to Africa and are one of the continent's ich- thyological icons. They owe their common name to their peculiar ability to produce stunning discharg-

46

Annals of the Missouri Botanical Garden

Stomatorhinus —

| ¢

) Boulengeromyrus кповртел

l Mormyrops zancltrostris NĚ

} Isichthys henryi

үү ——$%

Г ... Polimyrus marche:

. Marcusentus тооги

Ivindomyrus opdenboschi Г б

ү Paramormyrops gabonensis а

| Petrocephalus simus

| Petrocephalus stuhimani

Marcusenius сопсерћа 5

i Brienomyrus kingsieyae

> Bnenomyrus sp. 7

а. „Л Brienomyrus longicaudatus тб V 47 ~ —— M S. ү SE

А Brienomyrus sp 1 лава |

) ~ Brienomyrus sp 2 E v

р А Brienomyrus sp. 3

Bnenomyrus sp. 6

0 5 10

e 2l.

Mormyrid diversity in the Ivindo River (Gabon). Species-specific

Electric Organ Discharges (EODs)

Mei in middle columns. (With permission and modified after Hopkins, 1981.)

es, up to 350V in large specimens. The first species was formally described 200 years ago as Silurus (now Malapterurus) electricus, and for much of the time since then a single species was recognized. However, electric catfishes are widely distributed across Africa, and careful revisionary work (Norris, in prep.) has recovered a complex of upward of 20 species previously unrecognized in museum collec- tions and cataloged simply under the name M. elec- tricus. Interestingly, the diversity of electric catfish species matches that of other groups conforming to the basic division of ichthyofaunal provinces of the continent. Beyond taxonomic diversity, Norris has uncovered a large amount of ecological and ana- tomical variety. For example, some taxa are highly depressed bottom-dwellers, others are fusiform and presumably active swimmers. By far the most as- tonishing discovery in this family is a clade of dwarf forms in which females can be gravid at sizes as small as 6 cm, a size range in which specimens of other species show little gonadal differentiation. Most of the dwarf catfishes have reduced cephalic

and lateral line systems, elongate adipose fins, and a highly unusual three-chambered swim bladder. The function of the three-chambered bladder is as yet unclear, but Norris has suggested that it may play a role in the acoustic biology of these fishes, either in sound reception or production, or both.

There is little doubt that a similar taxonomic complexity will be revealed when other supposedly widespread "species," such as the enigmatic char- acoid Hepsetus odoe, are subject to the same sort of critical review. New species, and even whole new communities, abound in Africa, and their discovery and documentation will continue to require effort and render important results. But without doubt the greater scientific challenge will be the unraveling of the phylogenetic relationships of the African ich- thyofe auna. Such information is critical to our un- derstanding of the evolutionary history of the fauna, the geologic history and relationships of the conti- nent, and increasingly, as an aid in informing con- servation work. In all these aspects, this is a task that has hardly begun.

Volume 87, Number 1 2000

Lundberg et al. 47 Ichthyological Discovery

The Malagasy region has long been recognized as a biotic entity distinct from that of mainlan Africa. The region includes the Grande Ile of Mad- agascar and its offshore islands, the volcanic Mas- carenes and Comores, and the granitic Seychelles. As noted by Roberts (1975) the region, which is something of an enigma to biogeographers, harbors a highly endemic ichthyofauna. More recently Stiassny and Raminosoa (1994) provided a review of the fauna and, in addition to recognizing many as occupying phyloge- netically basal positions in their respective clades, they noted the enigmatic absence of a great many of the families currently represented in Africa and India. Because of its singular nature, the Malagasy ichthyofauna is not treated in detail in this essay. For more information on the Malagasy ichthyofauna see Teugels et al. (1985), Stiassny and Raminosoa (1994), and Benstead et al. (2000)

of the Malagasy endemics

TROPICAL ASIA

As treated here Tropical Asia approximately equals the Oriental Realm, extending from the In- dus basin eastward to South China and to the Mo- luccas (ca. *Wallace's Line") in Indonesia (Fig. 2). The dominant groups among primary and secondary freshwater families are Cyprinidae (about 1000 species), Balitoridae (about 300), Cobitidae (about 100), Bagridae (about 100), Osphronemidae (85): and among peripheral families, Gobiidae (about 300). This pattern applies principally to the main- land areas and in the insular parts to the major river basins draining onto the continental shelf. On oceanic islands and on the mainland in streams not draining to the continental shelf, the proportion of primary freshwater fishes is lower, and there is an increase in representation of peripheral families. In the Moluccas, Sulawesi, and most Philippine is- lands, there are no primary freshwater fishes, and gobies constitute about half of the inland fish fauna.

While recognizing an element of arbitrary taxo- nomic convention in doing so, fish families are of- ten used as a rough guide to higher-level taxonomic diversity of faunas. A distinguishing feature of the Tropical Asian inland fish fauna is its high number of families. One-hundred-twenty-one families have been recorded from inland waters, either as per- manent, temporary, or occasional residents (cf. 45— 50 in Africa and about 55 in South America). Thir- ty-four of these families are primary or secondary division freshwater fishes, and 18 are endemic to Southeast or tropical Asia. The remaining 87 fam- ilies are peripheral freshwater fishes, most repre- sented by few species. The high number of periph-

eral families in Asian freshwater is explained by the existence in adjacent seas of the highest fish family diversity in the world. The northern tropical Asian ichthyofauna has some affinities with that of northern temperate Asia, with which it shares some cyprinid and cobitoid lineages, but on the whole, tropical Asia shares several lineages with Africa

—

g., Notopteridae, several cyprinid lineages, Bag- ridae, Schilbeidae, Aplocheilidae, Nandidae, Ana- bantidae, Channidae, Mastacembelidae).

An estimated total number of species for tropical Asian freshwater fishes is 3000. More precise fig- ures have been compiled for some parts of South- east Asia (Irrawaddy, Salween, Mekong, Red River basins and intermediate areas, Hainan, Malay Pen- insula, the Philippines and Indonesia eastward to the Moluccas), where 2100 valid species are pres- ently known (Kottelat, in prep.). However, consid- erable differences exist in our ichthyological knowl- edge among regions and countries because of incomplete surveying and variable ichthyological practices among countries.

The Indian ichthyofauna remains in need of in- depth systematic study. Recent work on Sri Lankan and southern Indian fishes indicates that many spe- cies thought to be widely distributed and conspe- cific with species originally described from north- eastern India are in fact assemblages of allopatric species (Pethiyagoda, 1991; Pethiyagoda, pers. comm.; MK, pers. obs.). In Kerala (South India) 10-20% of the fishes in any basin of reasonable size are likely to be undescribed (Pethiyagoda & Kottelat, 1994). Habitat deterioration in this region poses a serious threat to most species, including those that are poorly known or unknown.

The development of ichthyology in China has been largely independent of that in the rest of Asia, but many scientific names applied to China's fresh- water fishes are taken from adjacent faunas without adequate comparison. Survey work is very incom- plete, especially in the hilly areas in the south. Species diversity is underestimated especially for small-sized fishes. For example, the loach identified as Schistura fasciolata is probably an assemblage of at least half a dozen species. Unfortunately, eco- nomic changes in recent years have made illegal electric fishing widely available everywhere. This poses a serious threat to the survival of most en- demic hill stream fish communities. In 1996 one of us (MK) observed in western Sichuan that most small streams surveyed were without fish.

Knowledge of the ichthyofauna of tropical Asia is still in an exploration and discovery phase. The fish fauna of Laos jumped from 216 species re- corded in the scientific literature as of early 199

48

Annals of the Missouri Botanical Garden

to some 370 known by mid-1997 (Kottelat, 1998). This represents an increase of about 80% in about 11 weeks of fieldwork. Some 85 of these species were unnamed, though

of the family Balitoridae that inhabit hill streams and have a relatively small distribution area, often restricted to part of a single river basin. Similar figures can be recorded for most Asian countries (Kottelat & Whitten, 1996b). The situation is likely similar (but much more acute) in Vietnam, where, as in China, ichthyology has been largely isolated.

Survey efforts in little-known areas and poorly sampled habitats continue to yield new discoveries. Kottelat et al. (1993) listed 964 species known in 1 rom western Indonesian inland waters. Ап addendum published in 1996 added 79 species de- scribed in the interval, and others have been dis- covered but not yet reported. Changes in systematic or nomenclatural status affected 56 other species (Kottelat & Whitten, 1996a). Fieldwork in reason- ably remote areas easily results in the discovery of 1 or 2 new species per actual day of fieldwork, but also in the discovery of an almost equal number of identification problems involving earlier known

cies. “Sundaland” (Java, Borneo, Sumatra, Malay Pen- insula) has a rich fauna (total ca. 1200 species). This is due to its historical fragmentation into smaller isolated basins and very diverse habitats ranging from coastal mangroves to high altitudes. One unusual habitat type has been discovered to harbor a very specialized and diverse fauna. This is the peat swamp forest, characterized by a peat- like soil (not made of sphagnum, but of tree roots) with a morphology somewhat similar to northern peat bogs, culminating into peat domes with a cen- tral “lake.” The black water of these oligotrophic lakes is darkly tinted by tannin and has a very low pH (3 to 5). Once thought to be depauperate, there are now more than 100 species known exclusively from these habitats. Most are small fishes: fighting fishes (Betta), licorice gouramies (Parosphromenus), and numerous diminutive cyprinids. A particularly interesting discovery in the peat swamp is Bihun- ichthys monopteroides (Fig. 22), а chaudhuriid earthworm eel (Kottelat & Lim, 1994). This family was considered endemic to Inle Lake in Myanmar, ut it is now known from about a dozen species with an overall wide range in Asia. Bihunichthys monopteroides is the sais chaudhuriid (maxi- mum known size of 36 mm SL) with extreme re- duction or loss of scales, ег line, fins, and a number of cranial bones. It has fossorial habits in soils around the swamps.

A diminutive, fossorial earthworm eel, Bi- blackwater a Ng; drawing by Kelvin Lim).

Figure 22. hunichthys топоріе ur 5, from an swamp (photo by Peter

Asian

In tropical Asia, as in Africa and South America, discovery of miniature je is fairly 3B). Danionella pelluci-

common. Many are cyprinids (Fig. 2 da from Myanmar and Boraras micros from north- east Thailand are adult at about 12 mm SL, and females of the loach Kottelatlimia katik are ripe at about 13 mm SL. In mangroves the smallest known species is an undescribed goby, Gobiopterus from Singapore, that reaches about 10 mm Minia- tures provide one of the most striking examples of discovery of a new fish clade, the family Sundasa- Wesen (Fig. 23C), described based on two species

1981 by Roberts. These are tiny (25 mm SL) and cae fishes with many reduced structures, and, in places, they are extremely abundant. Four additional species have been found on Borneo since their first discovery, and Siebert (1997) provided evidence that the group is related to herrings (Clu- peidae).

At the other end of the size scale, occasional large-sized species are still being described. Some of these were reported by early explorers, but it is only in recent years that they have been carefully diagnosed and described. In Asia an impressive ex- ample is Himantura chaophraya (Fig. 23A), the Mekong sting ray known to science since 1880 but just named in 1990 (Monkolprasit & Roberts,

his immense fish can attain a disc width of 2 m, an estimated total length of 4 m, and a weight of 600 kg

Probably the most interesting communities of Southeast Asian fishes discovered in recent years are the species flocks of the lakes of central Sula- wesi, Indonesia. A group of five tectonic lakes (Mal- ili lakes) and the connecting streams host a fauna

Volume 87, Number 1 2000

Lundberg et al. 49 sil eres Discovery

Figure 23. Recent discoveries of very large and small бі in hs Asia. —A. Freshwater sting гау, Himan- tura chaophraya. —B. A tiny, undescribed cyprinid, 9.9 mm long, from Bo . Sumatra, Malay Peninsula. — One of the Mig puer и EUN Sundasalanx аек hynchus, 23.8 mm long. (Copyrights Maurice Kottelat.)

consisting of 26 known native species, all but 4 endemic to this system. Within this lake system, two units can be distinguished, Lake Matano and lakes Towuti, Mahalona, and Wawontoa; only 2 en- demic fish species are shared by these two units.

e fish species include 3 endemic Hemiramphi- dae (halfbeaks), 6 endemic Gobiidae (gobies demic Oryziidae (ricefishes),

‚ 3 en-

м

4 endemic Telmath- erinidae (sailfin silversides); also present are two endemic genera and 4 non-endemic species (one each in the families Telmatherinidae, Gobiidae, Synbranchidae, Anabantidae). The lakes also con- tain at least 4 endemic crabs, a dozen endemic shrimps, about 60 endemic molluscs, and endemic ostracods, sponges, other invertebrates, and mac- po (Kottelat, 1990b, с, 1991; Larson & Kot- telat, 1992; Bouchet, 1995).

The family Telmatherinidae includes five genera, two endemic to the lakes, one with eight species endemic to the lakes plus one riverine species oc- curring also in an adjacent basin, one monotypic genus endemic to southwestern Sulawesi, and an-

Endemic lake s of Sulawesi.

Figure 2 : courting pair ой Telma reni атотае. —B. scale-eating Telmatherina prognatha. (Copyrights Maurice

Kottelat.)

Fin-ray and

other monotypic genus from баби | an island off Irian Jaya (the Indonesian рам

(Aarn et al., 19

matherinids are

New Guinea) Ше the bis of all tel- “pelagic,” the adults live close to the bottom and are substrate spawners. They are strongly sexually dimorphic (Fig. 24A), and males of several species exhibit color polymorphism. One species (Telmatherina prognatha, Fig. 24B) appar- ently feeds on fin-rays and scales of other species, and another (T. sarasinorum) preys on eggs of other spe The и lakes of central Sulawesi, lakes Poso and Lindu, have less diverse fish faunas. Lake Lin- du apparently hosts a single native species, Xeno- poecilus sarasinorum while Lake

three endemic

(Adrianichthyidae), Poso is inhabited by two endemic Gobiidae, Adrianichthyidae, and two endemic

Oryziidae (Kottelat, 1990a). The Adrianichthyidae

are noteworthy for their breeding behavior. AI- though originally recorded as live-bearers, it now

appears (Fig. 25A) that the female carries a clutch of eggs with her pelvic fins, in a ventral depression of the belly, for about 10 days until they hatch. Adrianichthyidae are closely related to Oryzii- dae, which are known from India to Japan and Ti-

Annals of the Missouri Botanical Garden

Figure 25. Recent a 'overies of oddly specialized . The

fishes i in n tropical ris e adrianichthyid Xenopoe- cilius оорћоги: fertil n ized clutch of eggs. —B. Th cave-dwelling balitorid loach Schistura Oedipus from Thai- land. welling cyprinid Puntius sp. from Java. (Copyrights Maurice Kottelat.)

~ у

mor, in fresh and brackish waters. Interestingly, Adrianichthyidae are known on Sulawesi from that part of the island (the western half) that plate tec- tonics suggest was earlier connected to elements now constituting part of Southeast Asia. On the oth- er hand, the distribution of telmatherinid species (except one) on the eastern half of Sulawesi corre- sponds to a plate earlier connected with the New Guinean one, on which Misool is located, where the family is also represented by an endemic genus (Kottelat, 1991

Tropical Asia contains several of the most exten-

Мм

sive karst areas of the world, and it is not surprising that at least 31 species (some undescribed) of cave fishes are known from this region and many more should be expected. Some of the largest karsts (Laos, Vietnam) have barely been explored speleo- logically. Sixteen species of cave fishes are known m China, seven from Thailand, four from India (three actually from wells), two from Indonesia, and one each from Malaysia and Laos. Most of these belong to genera known from surface waters, but five cannot be linked with known surface species and are treated as distinct genera. Noteworthy is the cyprinid genus Sinocyclocheilus, which has evolved independently at least seven hypogean spe- cies in China (Guizhou, Yunnan, and Guangxi prov- inces). The epigean species of Sinocyclocheilus are usually found associated with springs or in streams under overhanging rocky shores, which make them “pre-adapted” to colonize cave habitats. The known cave fishes in Asia belong to the families Balitori-

dae (13 species, e.g., Fig. 25B), Cyprinidae (12, e.g., Fig. 25C), Cobitidae (2), Synbranchidae (2), Gobiidae (1), and Clariidae (1).

AUSTRALIA AND NEW GUINEA

Australia and New Guinea are perhaps better known for their diverse marine fish faunas than for their freshwater ones. Nevertheless, freshwater fish- es of Australia and New Guinea are distinctive, in- teresting, and have been the subject of significant ichthyological discovery over the past 20 or so years.

With an area roughly equal to the contiguous ‚ Australia (Fig. 2) is generally characterized by lower topographical relief, lower

continental U.S

rainfall, and a much less extensive system of river drainages. However, largely as a consequence of the latitudinal spread of the country (from about 10°40’S to about 43?40'S), and of the influence of the Great Dividing Range on climatic conditions, Australia possesses a wide variety of freshwater habitats. These include artesian springs and ephemeral desert lakes and streams (Great Austra- lian Bight, Lake Eyre, and other internal drainag- es); tropical streams that undergo extensive flooding during summer monsoonal rains (northwestern Aus- rainforest

=

tralia and Gulf of Carpentaria drainages); streams (northeast coast); coastal streams and sand- dune lakes (east coast); and alpine lakes (Tasman- ia). The Murray-Darling River System is by far the largest in Australia, extending southwestward about 1900 km from the interior of southern Queensland to the Southern Ocean.

Stretching around 1600 km, New Guinea is the world’s second-largest island (Fig. 2). Biogeograph- ically and geologically, New Guinea is divided into more-or-less northern and southern provinces, sep- arated by an extensive mountain chain along the long axis of the island. This mountainous topogra- phy, in combination with high rainfall, results in numerous drainage systems, and a large array of freshwater habitats that include: short coastal streams, large lowland rivers, coastal swamps and floodplain lakes, alpine streams and lakes, and large highland rivers.

Australia and New Guinea are much more than just adjacent land masses, as they have been con- nected throughout most of their history. The Sahul Shelf, beneath the shallow Arafura Sea and Torres Strait that now separate the two areas, was emer- gent until as recently as about 6000 years ago dur- ing the latest glacial lowering of sea level, and southern New Guinean streams were confluent with those of the adjacent Australian coast.

Volume 87, Number 1 2000

undberg et al. 51 Ichthyological Discovery

Australian freshwater fishes were reviewed by Allen (1989), who recorded 187 species and sub- species; subsequent collecting and research have taken this count to around 215. The dominant freshwater taxa in the country are m gobioid fam- ilies Eleotrididae and Gobiidae (about 50 species), the superfamily Galaxioidea, an austral group of salmoniform fishes related to the northern smelts (26 species), Terapontidae, an Indo-West Pacific group of perch-like fishes (22 species), Percichthyi- dae, a family of southern Australian and southern South American perch-like fishes (21 species), Me- lanotaeniidae, an Australian-New Guinean endem- ic family (but, see below) of atherinomorph fishes (16 species and subspecies), Plotosidae, an Indo- West Pacific catfish family (about 15 species), and Atherinidae, a worldwide atherinomorph family (15 species and subspecies). Other distinctive compo- nents include the Queensland lungfish, Neocerato- dus forsteri (Neoceratodontidae), and the bony- tongue (osteoglossid) species, Scleropages jardinii and 5. leichardti.

Freshwater fishes of New Guinea were reviewed by Allen (1991), who listed 320 species, including some estuarine forms; subsequent collecting and research have taken this count to about 350. The most diverse freshwater taxa are eleotridids and go- biids (about 115 species), Melanotaeniidae (53 spe- cies), Ariidae, a circumtropical catfish family (21 species), Terapontidae (16 species), Chandidae (— Ambassidae), an Indo-West Pacific family of small, perch-like fishes (15 species), Pseudomugilidae, a family of atherinomorph fishes endemic to Australia and New Guinea (13 species), (about 13 species). Most of these taxa are also spe- cies-rich in northern Australia, emphasizing the historical link between the areas; indeed, about 50 species from southern New Guinea also occur in

and Plotosidae

northern Australia, and many of these are restricted to the two areas.

The amount of recent ichthyological activity in Australia and New Guinea can be gauged conser- vatively from the number of recently described spe- cies and subspecies. Since 1970 about 70 Austra- lian (33% of the total) and about 125 New Guinean (36% of the total) freshwater fish species and sub- species were described or are awaiting description.

Increased understanding of freshwater fish sys- tematics and distributions in the region has largely stemmed from the application of modern collecting and systematic techniques by professional ichthy- ologists. However, no less significant has been the input from amateur ichthyologists and aquarists. In particular, during the last three decades a profound