Lee A. Weigt

Comprehensive DNA barcode coverage of North American birds

DNA barcoding seeks to assemble a standardized reference library for DNA-based identification of eukaryotic species. The utility and limitations of this approach need to be tested on well-characterized taxonomic assemblages. Here we provide a comprehensive DNA barcode analysis for North American birds including 643 species representing 93% of the breeding and pelagic avifauna of the USA and Canada. Most (94%) species possess distinct barcode clusters, with average neighbour-joining bootstrap support of 98%. In the remaining 6%, barcode clusters correspond to small sets of closely related species, most of which hybridize regularly. Fifteen (2%) currently recognized species are comprised of two distinct barcode clusters, many of which may represent cryptic species. Intraspecific variation is weakly related to census population size and species age. This study confirms that DNA barcoding can be effectively applied across the geographical and taxonomic expanse of North American birds. The consistent finding of constrained intraspecific mitochondrial variation in this large assemblage of species supports the emerging view that selective sweeps limit mitochondrial diversity.

An Unprecedented Aggregation of Whale Sharks, Rhincodon typus, in Mexican Coastal Waters of the Caribbean Sea

Whale sharks, Rhincodon typus, are often perceived as solitary behemoths that live and feed in the open ocean. To the contrary, evidence is accumulating that they are gregarious and form seasonal aggregations in some coastal waters. One such aggregation occurs annually north of Cabo Catoche, off Isla Holbox on the Yucatán Peninsula of Mexico. Here we report a second, much denser aggregation of whale sharks (dubbed “the Afuera”) that occurs east of the tip of the Yucatán Peninsula in the Caribbean Sea. The 2009 Afuera event comprised the largest aggregation of whale sharks ever reported, with up to 420 whale sharks observed in a single aerial survey, all gathered in an elliptical patch of ocean approximately 18 km2. Plankton studies indicated that the sharks were feeding on dense homogenous patches of fish eggs, which DNA barcoding analysis identified as belonging to little tunny, Euthynnus alletteratus. This contrasts with the annual Cabo Catoche aggregation nearby, where prey consists mostly of copepods and sergestid shrimp. Increased sightings at the Afuera coincide with decreased sightings at Cabo Catoche, and both groups have the same sex ratio, implying that the same animals are likely involved in both aggregations; tagging data support this idea. With two whale shark aggregation areas, high coastal productivity and a previously-unknown scombrid spawning ground, the northeastern Yucatán marine region is a critical habitat that deserves more concerted conservation efforts.

Using DNA barcoding to assess Caribbean reef fish biodiversity: expanding taxonomic and geographic coverage.

This paper represents a DNA barcode data release for 3,400 specimens representing 521 species of fishes from 6 areas across the Caribbean and western central Atlantic regions (FAO Region 31). Merged with our prior published data, the combined efforts result in 3,964 specimens representing 572 species of marine fishes and constitute one of the most comprehensive DNA barcoding “coverages” for a region reported to date. The barcode data are providing new insights into Caribbean shorefish diversity, allowing for more and more accurate DNA-based identifications of larvae, juveniles, and unknown specimens. Examples are given correcting previous work that was erroneous due to database incompleteness.

Using DNA Barcodes to Identify Bird Species Involved in Birdstrikes

Abstract We determined effectiveness of using mitochondrial DNA barcodes (cytochrome c oxidase subunit 1 [CO1]) to identify bird–aircraft collision (birdstrike) cases that lacked sufficient feather evidence for morphological diagnosis. From September through December 2006, 821 samples from birdstrike events occurring in the United States were submitted for DNA analysis. We successfully amplified a CO1 DNA barcode product from 554 (67.5%) of the samples; 267 (32.5%) did not contain viable DNA and depended on morphological methods (microscopy) for Order or Family level identification. We deemed 19 cases inconclusive either because the DNA barcode recovered from the sample did not meet our 98% match criteria when compared to the Barcode of Life Database (BoLD) or because the DNA barcode matched to a set of ≥2 closely related species with overlapping barcodes, preventing complete species identification. Age of the sample (≤6 months) did not affect DNA viability, but initial condition of the sample and the collection method was critical to DNA identification success. The DNA barcoding approach has great potential in aiding in identification of birds (and wildlife) for airfield management practices, particularly in regions of the world that lack the vast research collections and individual expertise for morphologic identifications.

Seven new species within western Atlantic Starksia atlantica, S. lepicoelia, and S. sluiteri (Teleostei, Labrisomidae), with comments on congruence of DNA barcodes and species

Abstract Specimens of Starksia were collected throughout the western Atlantic, and a 650-bp portion of the mitochondrial gene cytochrome oxidase-c subunit I (COl) was sequenced as part of a re-analysis of species diversity of western Central Atlantic shorefishes. A neighbor-joining tree constructed from the sequence data suggests the existence of several cryptic species. Voucher specimens from each genetically distinct lineage and color photographs of vouchers taken prior to dissection and preservation were examined for diagnostic morphological characters. The results suggest that Starksia atlantica, Starksia lepicoelia, and Starksia sluiteri are species complexes, and each comprises three or more species. Seven new species are described. DNA data usually support morphological features, but some incongruence between genetic and morphological data exists. Genetic lineages are only recognized as species if supported by morphology. Genetic lineages within western Atlantic Starksia generally correspond to geography, such that members of each species complex have a very restricted geographical distribution. Increasing geographical coverage of sampling locations will almost certainly increase the number of Starksia species and species complexes recognized in the western Atlantic. Combining molecular and morphological investigations is bringing clarity to the taxonomy of many genera of morphologically similar fishes and increasing the number of currently recognized species. Future phylogenetic studies should help resolve species relationships and shed light on patterns of speciation in western Atlantic Starksia.

Incorporation of DNA barcoding into a large-scale biomonitoring program: opportunities and pitfalls

Abstract Taxonomic identification of benthic macroinvertebrates is critical to protocols used to assess the biological integrity of aquatic ecosystems. The time, expense, and inherent error rate of species-level morphological identifications has necessitated use of genus- or family-level identifications in most large, statewide bioassessment programs. Use of coarse-scale taxonomy can obscure signal about biological condition, particularly if the range of species tolerances is large within genera or families. We hypothesized that integration of deoxyribonucleic acid (DNA) barcodes (partial cytochrome c oxidase subunit I sequences) into bioassessment protocols would provide greater discriminatory ability than genus-level identifications and that this increased specificity could lead to more sensitive assessments of water quality and habitat. Analysis of DNA barcodes from larval specimens of Ephemeroptera, Plecoptera, and Trichoptera (EPT) taxa collected as part of Marylands Biological Stream Survey (MBSS) revealed ∼2 to 3× as many DNA-barcode groups or molecular operational taxonomic units (mOTUs) as morphologically identified genera. As expected, geographic distributions for several mOTUs were tighter than for the parent genus, but few mOTUs showed closer associations with water-quality variables or physical-habitat features than did the genus in which they belonged. The need for improved protocols for the consistent generation of DNA barcodes is discussed.

Invasive lionfish preying on critically endangered reef fish

Caribbean coral reef ecosystems are at the forefront of a global decline and are now facing a new threat: elimination of vulnerable species by the invasive lionfish (Pterois spp.). In addition to being threatened by habitat destruction and pollution, the critically endangered social wrasse (Halichoeres socialis), endemic to Belize’s inner barrier reef, has a combination of biological traits (small size, schooling, and hovering behavior) that makes it a target for the invasive lionfish. Based on stomach content analyses, this small fish comprises almost half of the lionfish diet at the inner barrier reef in Belize. The combination of lionfish predation, limited range, and ongoing habitat destruction makes the social wrasse the most threatened coral reef fish in the world. Other species with small range and similar traits occur elsewhere in the Caribbean and face similar risks.

DNA barcoding fishes.

This chapter is an overview of the techniques for DNA barcoding of fishes from field collection to DNA sequence analysis. Recommendations for modifications of field protocols and best tissue sampling practices are made. A variety of DNA extraction protocols is provided, including high-throughput robot-assisted methods. A pair of well-tested forward and reverse primers for PCR amplification and sequencing are presented. These primers have been successfully used for DNA barcode on a wide array of marine fish taxa and also work well in most freshwater and cartilaginous fishes. Recipes and cycling protocols for both PCR amplification and sequencing and cleanup methods for the reaction products are provided. A method for the consistent production of high-quality DNA barcodes from DNA sequence data is given and stringent guidelines for judging the quality of raw sequence data are laid out.

A New Species of Soapfish (Teleostei: Serranidae: Rypticus), with Redescription of R. subbifrenatus and Comments on the Use of DNA Barcoding in Systematic Studies

A new species of Rypticus is described from the Bahamas, Bermuda, Florida, and the Caribbean Sea. The species previously has been confused with the spotted soapfish, R. subbifrenatus Gill 1861, with which it shares a similar pattern of dark spotting on the body. The new species differs from R. subbifrenatus in having yellow pigment on the pectoral fin and distal portions of the soft dorsal, caudal, and anal fins in life (pale in preservative); a different configuration of dark spots on the head; usually dark spots on the belly and caudal fin; almost always four dorsal-fin spines; and modally 25 total dorsal-fin elements, 15 pectoral-fin rays, and 23 total caudal-fin rays. The lower jaw typically extends further anteriorly beyond the upper jaw in the new species than in R. subbifrenatus, and the caudal peduncle is usually narrower. The new Rypticus typically inhabits deeper waters than R. subbifrenatus, and is commonly found on vertical slopes and walls vs. shallow, flat areas. The new species likely would have continued to go unnoticed without examination of genetic data, as there was little reason to look further at R. subbifrenatus until DNA barcoding revealed two distinct genetic lineages within the species. The value of DNA barcoding data in systematic studies and the need for increased support of taxonomy are highlighted. A neotype for Rypticus subbifrenatus is designated.

The Northern Snakehead Channa argus (Anabantomorpha: Channidae), a non-indigenous fish species in the Potomac River, U.S.A

ABSTRACT Mitochondrial sequence variation was examined in the northern snakehead, Channa argus (Cantor, 1842), a species of fish native to Asia and recently collected in the eastern United States. There are seven unique haplotypes in 29 specimens studied, with no haplotype shared between areas of introduction. One haplotype was shared by 15 individuals from the Potomac River system (both males and females <480 mm total length), indicating possible breeding within the system. A single large adult male from the Potomac River system has a unique haplotype. All Crofton, Maryland, specimens had one haplotype. Two haplotypes were found in Meadow Lake, FDR Park, Philadelphia, Pennsylvania. Unique haplotypes were found in Wheaton, Maryland, and Shrewsbury, Massachusetts. These results support the conclusion that there were several independent introductions of the northern snake-head into these waters, and that no two introductions came from the same maternal source.