Development Of Dna Markers And The Genetic Analysis Of Seven Populations Of The Freshwater Mussel Lampsilis Radiata Siliquoidea Bivalvia Unionidae In North Central Minnesota PDF Download

Identification of freshwater mussel species and subpopulations is challenging due to the plasticity of morphological characters. Molecular analyses are a powerful alternative for systematics and for biological surveys and restoration efforts. Yet most molecular methods require sacrificing animals for tissue sampling, an approach that cannot be applied to threatened and endangered species. This study demonstrated, through field trials with 3 endemic yet abundant Lake Waccamaw mussel species, that hemolymph extraction produced no mortality in animals in their native environment. The authors developed extraction methods that make hemolymph a reliable source for DNA analysis that, despite lower DNA yield, shows percent success rate in polymerase chain reaction (PCR) assays that rivals that of body tissues from sacrificed animals. The authors followed sampled and control mussels in field enclosures in Lake Waccamaw for 8 weeks and observed no mortality of either treatment group. A small, statistically not significant reduction in growth (5 - 8%) was observed in the hemolymph-removal group. The authors report the development and application of PCR assays from two mitochondrial DNA (mtDNA) regions, followed with digestion by restriction enzymes that reveal species-diagnostic DNA restriction fragment length polymorphisms (RFLP). The results are rapid and inexpensive PCRRFLP assays to discriminate among sibling species in North Carolina that are difficult and sometimes misidentified in the field, even by experienced biologists. Their regional sampling and phylogenetic analysis of mtDNAs indicates that the major taxa in Lake Waccamaw are not strictly endemic to the Lake. Samples collected in the Lake showed mtDNA sequences that were identical or very similar to sequences obtained from other rivers within the Yadkin-PeeDee and Lumber drainages. Lake Waccamaw Elliptio wacamawensis shared haplotypes with, and/or fell within a clade of, mussels identified as E. congaraea and E. wacamawensis from the Waccamaw River. The same was true of Lampsilis fullerkati specimens, which were in a clade intermingled with L. radiata and L. radiata radiata specimens from the Waccamaw and Yadkin-PeeDee rivers. Additional sampling, morphological and molecular analysis, particularly of type specimens, is needed to further resolve taxonomic relationships and biogeographic ranges. A potential cryptic species, misidentified as E. complanata, and evidence for a cryptic phylogenetic species within L. radiata radiata both appeared in their collection. Together, their results validate the use of hemolymph extraction, followed by PCR-RFLP assays and targeted DNA sequencing, as a rapid, inexpensive, and reliable identification method for field biologists at NCDOT and other agencies who routinely survey NC freshwater mussel populations.