United States Environmental Protection Agency Environmental Monitoring and Support Laboratory Cincinnati OH 45268 Research and Development EPA/600/S4-86/032 Jan. 1987 SEPA Project Summary Taxonomy of Ceriodaphnia (Crustacea: Cladocera) in U.S. Environmental Protection Agency Cultures Dorothy B. Berner This study investigated the taxonomy of three groups of the cladoceran genus Ceriodaphnia in cultures being used by the U.S. Environmental Protection Agency. One taxonomic group was identified as C. reticulata. The second group was identi- fied as C. dub/a. The third group was tax- onomically nearly identical to C. dubla but was determined to be a hitherto undescrib- ed phenotypic variant of C. dubla, and is designated as C, dubla, toothed-pecten variety. Specimens of this form have been found in populations of C. dubla collected west of the Mississippi River. Similarities in the general morphology, postabdomens, and ephippia of C. reti- culata and C. dubia suggest that they are evolutionary closely related and might be able to hybridize and produce offspring having an ovate-toothed pecten like that of the C. dubia variant. Experiments designed to test this possibility were in- conclusive although two successful inter- specific matings were observed. It is sug- gested that the relationship between these two Ceriodaphnia could be further eluci- dated by study of more extensive field samples, and by interspecific breeding experiments that include hatching of hybrid young from ephippia and study of their taxonomy and fertility. This Project Summary was developed by EPA's Environmental Monitoring and Support Laboratory, Cincinnati, OH, to announce key findings of the research pro- ject that is fully documented in a separate report of the same title (see Project Report ordering information at back). Introduction The U.S. Environmental Protection Agency (EPA) Environmental Research Laboratory-Duluth, the Environmental Monitoring and Support Laboratory- Cincinnati, and other EPA and private laboratories have been exploring the suitability of Ceriodaphnia for short-term (7 day) toxicity testing for several years. Initial Ceriodaphnia stock cultures established during 1969 in the Duluth laboratory with animals obtained from fish ponds at EPA's Fish Toxicology Station, Newtown, OH, were identified as Cerio- daphnia reticulata. In 1982-83, subtle dif- ferences in the appearance of the cultured animals suggested that the stocks com- prised more than one species. Subsequent microscopic examination revealed that their cultures contained two, possibly three, species of Ceriodaphnia. Two were tentatively identified as C. reticulata and C. dubia (or C. affinis because a problem in synonomy exists), but the third was unidentifiable and appeared to have characteristics of both the other two. The goals of the study reported herein were to verify the identification of Cerio- daphnia in culture at the EMSL-Cincinnati Facility, Newtown and at other EPA laboratories, and to try to determine, by taxonomic comparison and by interspe- cific matings, if the unidentifiable animals were hybrids of the two former species, a morph of one of those species, or a new species. Materials and Methods EPA's Ceriodaphnia were compared with specimens from other sources, including ------- specimens from the U.S. Natural History Museum (Smithsonian), Washington, DC, the British Museum of Natural History, London; and the Lilljeborg Collection, Uppsala, Sweden, using optical and scan- ning electron microscopy. Conclusions 1. Nearly all Ceriodaphnia cultures at the EPA Newtown Facility and Duluth lab- oratories, and other cultures derived from them, were C. reticulata or C. dub/a. 2. A third Ceriodaphnia found in a few EPA cultures, particularly those from EPA's Athens, GA, laboratory, was a morpholog- ical variant of C. dubia, which it resembles almost completely. The morph was rever- sibly altered to the dubia form when the animals were cultured in reconstituted, rather than well water. The males are indis- tinguishable from C. dubia males, regardless of culture medium. Specimens of this variant have been found in natural populations of C. dubia west of the Mississippi River. It is designated in the full report as C. dubia, toothed-pecten variety. 3. Experiments attempting to hybridize C. reticulata and C. dubia were incon- clusive, although two successful inter- specific matings occurred. Taxonomically, these species appear to be closely enough related that males might mistake a female of the other species as their own. To ascer- tain whether hybridization is possible, more experiments of the kind attempted in this study would have to be carried out. Lastly, the morphology of such hybrids should be compared with specimens from field populations in which C. reticulata and C. dubia coexist, to see if hybrid forms occur naturally. 4. Comparison of the EPA C. dubia with N. American and European populations designated C. dubia or C. affinis revealed no significant differences among them. This study therefore supports earlier con- clusions that the two names are synon- omous, and that C. dubia takes prece- dence over C. affinis. It is likely, however, that other species of Ceriodaphnia exist that have characteristics similar to that of dubia, that should not be used to identify animals found in natural populations. Recommendations As an outcome of this study, the follow- ing recommendations were made: 1. There needs to be a nationally organ- ized and funded program for sampling freshwaters, accompanied by support for systematic studies, development of refer- ence collections, and the publication of adequate taxonomic keys. Difficulties en- countered by EPA personnel in identifying the species in their cultures reflect the in- adequacy of the keys commonly used in N. America to identify zooplankton. Fur- thermore, original species' descriptions are frequently in Latin, German, or French, are not readily available, and may not be ap- plicable to N. America because they are of taxa from other continents. As the need to monitor freshwaters increases, so does the need for contemporary, comprehen- sive reference collections and taxonomic information. 2. It is suggested that some of the key characteristics in C. dubia are under nutri- tional control. This could possibly be tested by rearing them in defined medium and feeding them algae also grown in de- fined medium. This would make it possible to manipulate the presence and concentra- tions of micronutrients, which seem likely candidates for such control. Such studies might contribute to the knowledge of some of the factors affecting polymorph- ism in the Daphniidae. Dorothy B. Berner is with Temple University, Philadelphia. PA 19122. William J. Horning, II, is the EPA Project Officer (see below). The complete report, entitled "Taxonomy of Ceriodaphnia (Crustacea:Cladocera) in U.S. Environmental Protection Agency Cultures." 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