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What sorts of DNA can be found in an urban environment? Last year I helped supervise Trinity High School students Brenda Tan and Matt Cost in an investigation of New York City apartments, sidewalks, and supermarkets with DNA barcoding. Brenda and Matt spent 4 months collecting and documenting everyday items that might contain DNA, and delivered specimens to Center for Conservation Genetics, American Museum of Natural History for testing; 151 (70%) of 217 items yielded DNA barcodes, including a feather duster (ostrich), a hot dog from a street vendor (cow), a dog biscuit (American bison), and a fly in a shipment of grapefruit from Texas (Oriental latrine fly Chrysomya megacephala, an invasive species in southern U.S.). Among other surprising results, the student investigators found 95 different animal species, 16% of human and pet food items mislabeled, and a genetically distinct mystery cockroach that might be a new subspecies or species. I encourage you to peruse the Rockefeller University DNAHouse site which includes their narrative and Q+A reports, spreadsheets detailing specimens and results, and high-resolution images, including of cockroach!

Following example of 2008 student-led “Sushigate.” Brenda and Matt’s DNAHouse study is capturing wide public interest, including stories in New York Times, New York Post, NPR, NBC TV, and over 230 media sites in 9 languages and 30 countries so far. If high school students can make original discoveries with important regulatory and scientific implications using DNA barcoding, then wide application to food products, products from protected and regulated species, detection of invasive species, and biodiversity surveys, including by interested public, is not far off. The most important for general public is food, and I expect to see growing attention on the part of regulatory agencies, distributers, retailers, and consumers to identifying mislabeled food products using DNA barcodes.

In 16 December 2009 Biol Lett, researchers from University of Guelph, University of British Columbia, and Agriculture and Agri-Food Canada report on COI barcodes for 11,289 individuals representing 1,327 species of Lepidoptera (moths and butterflies) collected in eastern North America. This large collection revealed the same patterns of highly restricted intraspecific variation, uncommon barcode sharing, and overlooked diversity seen in numerous smaller studies: average variation within species was 0.43%, while average among congeneric species was 7.7%, 18-fold higher. Only nine cases (0.7%) of barcode sharing between species were observed, and at the same time, large divergences (>2%) suggesting overlooked taxa were found in 67 (5.1%) of cases (in some cases morphological and ecological differences supporting species status were observed). The survey included multiple individuals per species collected at sites 500 to 2800 km apart, with “no significant increase in genetic distances with geographical separation.” Hebert, deWaard, and Landry conclude “an effective identification system can be constructed for the Lepidoptera fauna of eastern North American without extensive geographical surveys of each species,” and that, given likely similar patterns in most terrestrial and marine fauna, “a comprehensive barcode library for animal life can be assembled rapidly,” with diverse benefits to society and science.

In my view, this study should lay to rest the early and persisting worries of some taxonomists that single gene DNA barcoding would distinguish species only in limited situations. For example, in a 2004 PLoS Biol commentary, Moritz and Cicero cautioned “But to determine when and where [DNA barcoding]…is applicable, we now need to discover the boundary conditions.” The 2009 answer is that there are no major restrictions to wide application of DNA barcoding in animals, taxonomically or geographically, and the one regularly encountered limitation is very young species, which represent a small fraction of recognized taxa even in intensively studied groups. At the same time DNA barcoding speeds taxonomic assessment by flagging genetically distinct forms, many of which are found represent unrecognized species, including species that would likely otherwise remain hidden indefinitely. Together with prior work this study refutes a widely-cited (pre-barcoding) estimate that 23% of animal taxa have shared or overlapping mitochondrial DNA sequences (Funk and Omland Ann Rev Genet 2003); this estimate presumably reflected biases in then-existing databases. In closing, I note that Hebert, deWaard, and Landry offer a new yardstick, namely the fraction of the animal kingdom mapped, lifting our eyes up to the goal of a rapid identification system for all eukaryotic life.

What lives in soil? In August 2009 Pesq Agropec Bras (open access) an international cohort of 10 researchers from Canada, France, US, Taiwan, and Russia examine prospects for speeding assessment of soil animal diversity with COI DNA barcoding. As test sets, Rougerie and colleagues explore taxonomic and sequence diversity in earthworms and collembolans (springtails). These two groups comprise a similar number of named species (earthworms, 6000; springtails, 7900), but these totals likely underestimate true diversity, particularly for springtails, which are tiny (0.2 – 6.0 mm), challenging collecting and morphology.

For earthworms, the researchers analyzed COI sequences from 457 specimens collected in 13 countries around the world (including North America, South America, Caribbean, Europe, Middle East, Southeast Asia, and Australia); these represented at least 49 genera in 8 families. 87 species were identified by morphology, representing about 1/2 of specimens; the remainder, mostly those from Philippines and Brazil, could not be identified to species. Applying a threshold approach, the researchers found 192 (10% cutoff) and 211 (4% cutoff) genetic clusters, including two or more divergent clusters in 13 (15%) of named species. None of species showed sequence sharing or overlaps.

For springtails, 695 specimens from a similar global distribution of sites were analyzed, representing 88 genera in 16 familes; only 44 species were formally identified with morphologic characters, consistent with the “difficult and poorly known taxonomy of these organisms.” Of note, the authors report that “a specific protocol was developed for [collembolans] so that voucher specimens could be recollected after DNA extraction and thus be used for further morphological examination.” Sequence comparisons showed a “typical…bimodal distribution of intra- versus interspecific divergences such as the one also reported in earthworms.” Applying distance thresholds as above gave 215 (10% cutoff) and 227 (4% cutoff) genetic clusters. In conclusion, efficient assessment of soil animal diversity calls out for DNA barcoding.

On a separate note, soil animals may be useful for understanding mitochondrial evolution. Even factoring in species diversity, these animals appear to have enormous population sizes, given densities up to 4 x 10³ earthworms/m² and 1.8 x 10⁶ collembolans/m³, yet show typical bimodal pattern of intra- << inter-specific variation noted above. Along these lines, in November 2009 Nature Nick Lane explores whys of mitochondrial evolution and speciation, including a possible “radically new picture of mitochondrial genes being tightly regulated by selection.” Stay tuned!

In current Aquatic Conserv Marine Freshwater Ecosys, researchers from Muséum national d’Histoire naturelle, France, report on 80 years of taxonomic confusion that has contributed to near extinction for a once abundant north Atlantic skate. Iglésias and colleagues found that two forms, lumped together in 1926 as European common skate (Dipturus batis, Linnaeus 1798), in fact represent distinct species with morphologic, genetic (in mitochondrial genome), and life history differences.

As the researchers report, this taxonomic oversight obscured the disappearance of one species, the flapper skate (D. cf. intermedia) because it was confused with the less threatened  blue skate (D. cf. flossada).  Iglesias and colleagues marketplace survey revealed additional sources of confusion. They analyzed 4,110 skates landed over a 2 year period from 103 fishing cruises in four main French ports by 41 different French commercial trawlers, and found that five skate species (included the two named above) from two genera are variously lumped together under just two marketplace names, the aforementioned “European common skate (D. batis)” and “longnose skate (D. oxyrinchus);” according to their analysis the latter species, formerly common, is also locally extirpated, and most specimens with this name represent other species.

For newly rediscovered blue and flapper skates, the researchers report 20 diagnostic substitutions in approximately 2600-nucleotide segment spanning 12s and 16s RNA. Other than the 10 mitochondrial sequences included in this report, I find only two other D. batis sequences in GenBank (and none as yet under either of two resurrected names). It is remarkable that so little genetic information has been collected for such recently abundant, commercially important (annual landings in 1000’s of tons), and now threatened species. To aid standardized application of molecular identification techniques, I hope the authors will also analyze COI barcode sequences for their specimens. Then I look forward to school children aiding conservation and helping find new species by DNA barcoding specimens from their local fish markets!