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Numerous pharmaceutical and industrial chemicals are classified as endocrine‐disrupting chemicals (EDCs) that interfere with hormonal homeostasis, leading to developmental disorders and other pathologies. The synthetic estrogen 17α‐ethynylestradiol (EE2) is used in oral contraceptives and other hormone therapies. EE2 and other estrogens are inadvertently introduced into aquatic environments through municipal wastewater and agricultural effluents. Exposure of male fish to estrogens increases expression of the egg yolk precursor protein vitellogenin (Vtg), which is used as a molecular marker of exposure to estrogenic EDCs. The mechanisms behind Vtg induction are not fully known, and we hypothesized that it is regulated via DNA methylation. Adult zebrafish were exposed to either dimethyl sulfoxide or 20 ng/L EE2 for 14 days. Messenger RNA (mRNA) expression and DNA methylation were assessed in male zebrafish livers at 0, 0.25, 0.5, 1, 4, 7, and 14 days of exposure; and those of females were assessed at 13 days (n ≥ 4/group/time point). To test the persistence of any changes, we included a recovery group that received EE2 for 7 days and did not receive any for the following 7 days, in the total 14‐day study. Methylation of DNA at the vtg1 promoter was assessed with targeted gene bisulfite sequencing in livers of adult male and female zebrafish. A significant increase in vtg1 mRNA was observed in the EE2‐exposed male fish as early as 6 h. Interestingly, DNA methylation changes were observed at 4 days. Decreases in the overall methylation of the vtg1 promoter in exposed males resulted in levels comparable to those in female controls, suggesting feminization. Importantly, DNA methylation levels in males remained significantly impacted after 7 days post–EE2 removal, unlike mRNA levels. These data identify an epigenetic mark of feminization that may serve as an indicator of not only estrogenic exposure but also previous exposure to EE2. Environ Toxicol Chem 2024;43:1547–1556. © 2024 SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA. [ABSTRACT FROM AUTHOR]

Bromate (⁠ BrO 3 − ⁠) is a water disinfection byproduct (DBP) previously shown to induce nephrotoxicity in vitro and in vivo. We recently showed that inhibitors of DNA methyltransferase 5-aza-2′-deoxycytidine (5-Aza) and histone deacetylase trichostatin A (TSA) increased BrO 3 − nephrotoxicity whereas altering the expression of the cyclin-dependent kinase inhibitor p21. Human embryonic kidney cells (HEK293) and normal rat kidney (NRK) cells were sub-chronically exposed to BrO 3 − or epigenetic inhibitors for 18 days, followed by 9 days of withdrawal. DNA methylation was studied using a modification of bisulfite amplicon sequencing called targeted gene bisulfite sequencing. Basal promoter methylation in the human p21 promoter region was substantially lower than that of the rat DNA. Furthermore, 5-Aza decreased DNA methylation in HEK293 cells at the sis-inducible element at 3 distinct CpG sites located at 691, 855, and 895 bp upstream of transcription start site (TSS). 5-Aza also decreased methylation at the rat p21 promoter about 250 bp upstream of the p21 TSS. In contrast, sub-chronic BrO 3 − exposure failed to alter methylation in human or rat renal cells. BrO 3 − exposure altered histone acetylation in NRK cells at the p21 TSS, but not in HEK293 cells. Interestingly, changes in DNA methylation induced by 5-Aza persisted after its removal; however, TSA- and BrO 3 − -induced histone hyperacetylation returned to basal levels after 3 days of withdrawal. These data demonstrate novel sites within the p21 gene that are epigenetically regulated and further show that significant differences exist in the epigenetic landscape between rat and human p21, especially with regards to toxicant-induced changes in histone acetylation. [ABSTRACT FROM AUTHOR]

Speciation is the result of an accumulation of reproductive barriers between populations, but pinpointing the factors involved is often difficult. However, hybrid zones can form when these barriers are not complete, especially when lineages come into contact in intermediate or modified habitats. We examine a hybrid zone between two closely related riverine turtle species, Sternotherus depressus and S. peltifer, and use dual‐digest RAD sequencing to understand how this hybrid zone formed and elucidate genomic patterns of reproductive isolation. First, the geographical extent and timing of formation of the hybrid zone is established to provide context for understanding the role of extrinsic and intrinsic reproductive isolating mechanisms in this system. The strength of selection on taxon‐specific contributions to maintenance of the hybrid zone is then inferred using a Bayesian genomic cline model. These analyses identify a role for selection inhibiting introgression in some genomic regions at one end of the hybrid zone and promoting introgression in many loci at the other. When selective pressures necessary to generate outliers to the genomic cline are considered with the geographical and temporal context of this hybrid zone, we conclude that habitat‐specific selection probably limits introgression from S. depressus to S. peltifer in the direction of river flow. However, selection is mediating rapid, unidirectional introgression from S. peltifer to S. depressus, which is probably facilitated by anthropogenic habitat alteration. These findings indicate a potentially imminent threat of population‐level genomic extinction for an already imperiled species due to ongoing human‐caused habitat alteration. [ABSTRACT FROM AUTHOR]

The origin and eventual loss of biogeographic barriers can create alternating periods of allopatry and secondary contact, facilitating gene flow among distinct metapopulations and generating reticulate evolutionary histories that are not adequately described by a bifurcating evolutionary tree. One such example may exist in the two-lined salamander (Eurycea bislineata) species complex, where discordance among morphological and molecular datasets has created a "vexing taxonomic challenge." Previous phylogeographic analyses of mitochondrial DNA (mtDNA) suggested that the reorganization of Miocene paleodrainages drove vicariance and dispersal, but the inherent limitations of a single-locus dataset precluded the evaluation of subsequent gene flow. Here, we generate triple-enzyme restriction site-associated DNA sequencing (3RAD) data for > 100 individuals representing all major mtDNA lineages and use a suite of complementary methods to demonstrate that discordance among earlier datasets is best explained by a reticulate evolutionary history influenced by river drainage reorganization. Systematics of such groups should acknowledge these complex histories and relationships that are not strictly hierarchical. [Amphibian; hybridization; introgression; Plethodontidae; stream capture.] [ABSTRACT FROM AUTHOR]

Evolutionary biologists from Darwin forward have dreamed of having data that would elucidate our understanding of evolutionary history and the diversity of life. Sequence capture is a relatively old DNA technology, but its use is growing rapidly due to advances in (i) massively parallel DNA sequencing approaches and instruments, (ii) massively parallel bait construction, (iii) methods to identify target regions and (iv) sample preparation. We give a little historical context to these developments, summarize some of the important advances reported in this special issue and point to further advances that can be made to help fulfill Darwin's dream. [ABSTRACT FROM AUTHOR]

Degraded DNA from suboptimal field sampling is common in molecular ecology. However, its impact on techniques that use restriction site associated next-generation DNA sequencing ( RADSeq, GBS) is unknown. We experimentally examined the effects of in situ DNA degradation on data generation for a modified double-digest RADSeq approach (3 RAD). We generated libraries using genomic DNA serially extracted from the muscle tissue of 8 individual lake whitefish ( Coregonus clupeaformis) following 0-, 12-, 48- and 96-h incubation at room temperature posteuthanasia. This treatment of the tissue resulted in input DNA that ranged in quality from nearly intact to highly sheared. All samples were sequenced as a multiplexed pool on an Illumina MiSeq. Libraries created from low to moderately degraded DNA (12-48 h) performed well. In contrast, the number of RADtags per individual, number of variable sites, and percentage of identical RADtags retained were all dramatically reduced when libraries were made using highly degraded DNA (96-h group). This reduction in performance was largely due to a significant and unexpected loss of raw reads as a result of poor quality scores. Our findings remained consistent after changes in restriction enzymes, modified fold coverage values (2- to 16-fold), and additional read-length trimming. We conclude that starting DNA quality is an important consideration for RADSeq; however, the approach remains robust until genomic DNA is extensively degraded. [ABSTRACT FROM AUTHOR]

Background Urogenital schistosomiasis caused by Schistosoma haematobium is widely distributed across Africa and is increasingly being targeted for control. Genome sequences and population genetic parameters can give insight into the potential for population- or species-level drug resistance. Microsatellite DNA loci are genetic markers in wide use by Schistosoma researchers, but there are few primers available for S. haematobium. Methods We sequenced 1,058,114 random DNA fragments from clonal cercariae collected from a snail infected with a single Schistosoma haematobium miracidium. We assembled and aligned the S. haematobium sequences to the genomes of S. mansoni and S. japonicum, identifying microsatellite DNA loci across all three species and designing primers to amplify the loci in S. haematobium. To validate our primers, we screened 32 randomly selected primer pairs with population samples of S. haematobium. Results We designed >13,790 primer pairs to amplify unique microsatellite loci in S. haematobium, (available at http://www.cebio.org/projetos/schistosoma-haematobium-genome). The three Schistosoma genomes contained similar overall frequencies of microsatellites, but the frequency and length distributions of specific motifs differed among species. We identified 15 primer pairs that amplified consistently and were easily scored. We genotyped these 15 loci in S. haematobium individuals from six locations: Zanzibar had the highest levels of diversity; Malawi, Mauritius, Nigeria, and Senegal were nearly as diverse; but the sample from South Africa was much less diverse. Conclusions About half of the primers in the database of Schistosoma haematobium microsatellite DNA loci should yield amplifiable and easily scored polymorphic markers, thus providing thousands of potential markers. Sequence conservation among S. haematobium, S. japonicum, and S. mansoni is relatively high, thus it should now be possible to identify markers that are universal among Schistosoma species (i.e., using DNA sequences conserved among species), as well as other markers that are specific to species or species-groups (i.e., using DNA sequences that differ among species). Full genome-sequencing of additional species and specimens of S. haematobium, S. japonicum, and S. mansoni is desirable to better characterize differences within and among these species, to develop additional genetic markers, and to examine genes as well as conserved non-coding elements associated with drug resistance. [ABSTRACT FROM AUTHOR]

Ligating adapters with unique synthetic oligonucleotide sequences (sequence tags) onto individual DNA samples before massively parallel sequencing is a popular and efficient way to obtain sequence data from many individual samples. Tag sequences should be numerous and sufficiently different to ensure sequencing, replication, and oligonucleotide synthesis errors do not cause tags to be unrecoverable or confused. However, many design approaches only protect against substitution errors during sequencing and extant tag sets contain too few tag sequences. We developed an open-source software package to validate sequence tags for conformance to two distance metrics and design sequence tags robust to indel and substitution errors. We use this software package to evaluate several commercial and non-commercial sequence tag sets, design several large sets (maxcount = 7,198) of edit metric sequence tags having different lengths and degrees of error correction, and integrate a subset of these edit metric tags to polymerase chain reaction (PCR) primers and sequencing adapters. We validate a subset of these edit metric tagged PCR primers and sequencing adapters by sequencing on several platforms and subsequent comparison to commercially available alternatives. We find that several commonly used sets of sequence tags or design methodologies used to produce sequence tags do not meet the minimum expectations of their underlying distance metric, and we find that PCR primers and sequencing adapters incorporating edit metric sequence tags designed by our software package perform as well as their commercial counterparts. We suggest that researchers evaluate sequence tags prior to use or evaluate tags that they have been using. The sequence tag sets we design improve on extant sets because they are large, valid across the set, and robust to the suite of substitution, insertion, and deletion errors affecting massively parallel sequencing workflows on all currently used platforms. [ABSTRACT FROM AUTHOR]

Abstract: Factors affecting the type and frequency of germline mutations in animals are of significant interest from health and toxicology perspectives. However, studies in this field have been limited by the use of markers with low detection power or uncertain relevance to phenotype. Whole genome sequencing (WGS) is now a potential option to directly determine germline mutation type and frequency in family groups at all loci simultaneously. Medical studies have already capitalized on WGS to identify novel mutations in human families for clinical purposes, such as identifying candidate genes contributing to inherited conditions. However, WGS has not yet been used in any studies of vertebrates that aim to quantify changes in germline mutation frequency as a result of environmental factors. WGS is a promising tool for detecting mutation induction, but it is currently limited by several technical challenges. Perhaps the most pressing issue is sequencing error rates that are currently high in comparison to the intergenerational mutation frequency. Different platforms and depths of coverage currently result in a range of 10–103 false positives for every true mutation. In addition, the cost of WGS is still relatively high, particularly when comparing mutation frequencies among treatment groups with even moderate sample sizes. Despite these challenges, WGS offers the potential for unprecedented insight into germline mutation processes. Refinement of available tools and emergence of new technologies may be able to provide the improved accuracy and reduced costs necessary to make WGS viable in germline mutation studies in the very near future. To streamline studies, researchers may use multiple family triads per treatment group and sequence a targeted (reduced) portion of each genome with high (20–40×) depth of coverage. We are optimistic about the application of WGS for quantifying germline mutations, but caution researchers regarding the resource-intensive nature of the work using existing technology. [Copyright &y& Elsevier]

The diversity of available 2nd and 3rd generation DNA sequencing platforms is increasing rapidly. Costs for these systems range from <$100 000 to more than $1 000 000, with instrument run times ranging from minutes to weeks. Extensive trade-offs exist among these platforms. I summarize the major characteristics of each commercially available platform to enable direct comparisons. In terms of cost per megabase (Mb) of sequence, the Illumina and SOLiD platforms are clearly superior (≤$0.10/Mb vs. >$10/Mb for 454 and some Ion Torrent chips). In terms of cost per nonmultiplexed sample and instrument run time, the Pacific Biosciences and Ion Torrent platforms excel, with the 454 GS Junior and Illumina MiSeq also notable in this regard. All platforms allow multiplexing of samples, but details of library preparation, experimental design and data analysis can constrain the options. The wide range of characteristics among available platforms provides opportunities both to conduct groundbreaking studies and to waste money on scales that were previously infeasible. Thus, careful thought about the desired characteristics of these systems is warranted before purchasing or using any of them. Updated information from this guide will be maintained at: and . [ABSTRACT FROM AUTHOR]

Metallothionein (MT), a protein involved in metal regulation and detoxification, has been used widely as a biomarker of metal exposure. In the present study, a transgenic strain of the free-living soil nematode Caenorhabditis elegans was developed using the C. elegans MT-2 (mtl-2) promoter to control the transcription of green fluorescence protein (GFP) reporter. Response of this transgenic system to Cd, Hg, Cu, Zn, Ni, Pb, and As exposure in aquatic media was tested by quantifying GFP expression after 24 h of exposure. Response in Cd-spiked soil was tested in a similar manner. The mtl-2 transcription also was measured using real-time reverse transcription-polymerase chain reaction to gain a mechanistic understanding of the transgene expression. Green fluorescence protein is induced by Cd, Hg, Cu, and Zn in a time- and concentration-dependent manner; mtl-2 transcription is consistent with the GFP response. The minimum concentrations of Cd, Hg, Cu, and Zn that induce GFP response are 2- to 1,000- fold lower than concentrations affecting traditional endpoints, such as lethality or behavioral change. The system responds to Cd in soil in a similar manner. Neither Ni nor Pb induces GFP, and neither induces mtl-2 transcription. Arsenic does not induce GFP, yet an increase in mtl-2 transcription was found, suggesting that As may interfere with GFP signaling. This mtl-2::GFP transgenic bioassay represents an alternative approach to quantify, both easily and quickly, a surrogate of MT in response to metal exposure (e.g., Cd, Hg, Cu, and Zn) in a variety of environments and potentially may be used for quantitative or semiquantitative biomonitoring of metal contamination in soils and aquatic systems. [ABSTRACT FROM AUTHOR]

We assessed colony- and island-level genetic differentiation for the flightless cormorant ( Phalacrocorax harrisi), an endangered Galápagos endemic that has one of the most limited geographical distributions of any seabird, consisting of only two adjacent islands. We screened 223 individuals from both islands and nine colonies at five microsatellite loci, recovering 23 alleles. We found highly significant genetic differentiation throughout the flightless cormorant's range on Fernandina and Isabela Islands (global FST = 0.097; P<0.0003) both between islands (supported by Bayesian analyses, FST and RST values) and within islands (supported only by FST and RST values). An overall pattern of isolation-by-distance was evident throughout the sampled range ( r = 0.4169, one-sided P ≤ 0.02) and partial Mantel tests of this relationship confirmed that ocean is a dispersal barrier ( r = 0.500, one-sided P ≤ 0.003), especially across the 5-km gap between the two islands. The degree of detected genetic differentiation among colonies is surprising, given the flightless cormorant's limited range, and suggests a role for low vagility, behavioural philopatry, or both to limit dispersal where physical barriers are absent. We argue that this population should be managed as at least two genetic populations to better preserve the species-level genetic diversity, but, for demographic reasons, advocate the continued conservation of all breeding colonies. [ABSTRACT FROM AUTHOR]

The feather beta (β keratins of the white leghorn chicken (order Galliformes, Gallus gallus domesticus) are the products of a multigene family that includes claw, feather, feather-like, and scale genes (Presland et al. 1989a). Here we characterize the feather β-keratin genes in additional bird species. We designed primers for polymerase chain reactions (PCR) using sequences available from chicken, cloned the resulting amplicons to isolate individual copies, and sequenced multiple clones from each PCR reaction for which we obtained amplicons of the expected size. Feather β-keratins of 18 species from eight avian orders demonstrate DNA sequence variation within and among taxa, even in the protein-coding regions of the genes. Phylogenies of these data suggest that Galliformes (fowl-like birds), Psittaciformes (parrots), and possibly Falconiformes (birds of prey) existed as separate lineages before duplication of the feather β-keratin gene began in Ciconiiformes (herons, storks, and allies), Gruiformes (cranes, rails, and allies), and Piciformes (woodpeckers and allies). Sequences from single species of Coraciiformes (kingfishers) and Columbiformes (pigeons) are monophyletic and strikingly divergent, suggesting feather β-keratin genes in these birds also diverged after these species last shared a common ancestor with the other taxa investigated. Overall, these data demonstrate considerable variation in this structural protein in the relatively recent history of birds, and raise questions concerning the origin and homology of claw, feather-like, and scale β-keratins of birds and the reptilian β-keratins. [ABSTRACT FROM AUTHOR]

Bacterial resistances to diverse metals and antibiotics are often genetically linked, suggesting that exposure to toxic metals may select for strains resistant to antibiotics and vice versa. To test the hypothesis that resistances to metals and antibiotics are coselected for in environmental microbial assemblages, we investigated the frequency of diverse resistances in freshwater microcosms amended with Cd, Ni, ampicillin or tetracycline. We found that all four toxicants significantly increased the frequency of bacterioplankton resistance to multiple, chemically unrelated metals and antibiotics. An ampicillin-resistant strain of the opportunistic human pathogen Ralstonia mannitolilytica was enriched in microcosms amended with Cd. Frequencies of antibiotic resistance were elevated in microcosms with metal concentrations representative of industry and mining-impacted environments (0.01–1 mM). Metal but not antibiotic amendments decreased microbial diversity, and a weeklong exposure to high concentrations of ampicillin (0.01–10 mg l−1) and tetracycline (0.03–30 mg l−1) decreased microbial abundance only slightly, implying a large reservoir of antibiotic resistance in the studied environment. Our results provide first experimental evidence that the exposure of freshwater environments to individual metals and antibiotics selects for multiresistant microorganisms, including opportunistic human pathogens. [ABSTRACT FROM AUTHOR]

To test the hypothesis that industrial metal contaminants select for microorganisms tolerant to unrelated agents, such as antibiotics, we analyzed metal and antibiotic tolerance patterns in microbial communities in the intake and discharge of ash settling basins (ASBs) of three coal-fired power plants. High-throughput flow-cytometric analyses using cell viability probes were employed to determine tolerances of entire bacterioplankton communities, avoiding bias toward culturable versus nonculturable bacteria. We found that bacterioplankton collected in ASB discharges were significantly more tolerant to metal and antibiotic exposures than bacterioplankton collected in ASB intakes. Optical properties of microorganisms collected in ASB discharges indicated no defensive physiological adaptations such as formation of resting stages or excessive production of exopolymers. Thus, it is likely that the elevated frequency of metal and antibiotic tolerances in bacterioplankton in ASB discharges were caused by shifts in microbial community composition, resulting from the selective pressure imposed by elevated metal concentrations or organic toxicants present in ASBs. [ABSTRACT FROM AUTHOR]

We examined the population genetic structure of the diamondback terrapins (Malaclemys terrapin), within and among estuaries. Based on mark-recapture studies, these estuarine turtles have high site fidelity that is likely to make them vulnerable to local extinctions. We tested if observed site fidelity of adults would be reflected in intraestuarine population genetic structure of six highly polymorphic microsatellite loci (five tetranucleotide and one dinucleotide). No evidence was found for population structuring within the Charleston estuary nor among three different estuaries in South Carolina. We then examined four other terrapin populations from North Carolina to New York, as well as from the Florida Keys and from Texas. With increasing geographical distance, genetic differentiation increased from South Carolina through New York, but overall values were low. The dinucleotide locus contributed significantly more to the genetic differentiation of some population comparisons than any of the other loci. Interestingly, terrapins from South Carolina to New York were much more genetically similar to those from Texas (ρ  = 0.154) than to those from Florida (ρ  = 0.357). We attribute this pattern to extensive translocations of terrapins during the early 20th century to replenish diminished populations and to provide turtle farms with stocks. Terrapins collected in Texas were especially sought for shipment to the northeastern US because of their larger size. Our study indicates no population structure within or among adjacent estuaries. Thus, the mark-recapture information from adult and subadult feeding locations is a poor predictor of population genetic structure. Additionally, it appears that past human activities may have drastically altered the genetics of current populations. Finally, our data suggest that translocation of eggs or head starting of terrapins within estuaries or among adjacent estuaries is acceptable from a genetic standpoint. [ABSTRACT FROM AUTHOR]

We sought to refine genetic management of the endangered Whooping Crane ( Grus americana ) population by developing comprehensive genetic pedigrees for the captive population. Improvements to the studbook were accomplished by addition of pedigree information derived from leg-banding data on wild juvenile and founder similarity coefficients calculated from microsatellite DNA profiles to the original studbook pedigree. Incorporation of pedigrees derived from data on leg-banding of wild juveniles did not greatly alter the previous relatedness structure of the captive population, but incorporation of microsatellite similarity coefficients produced a substantially different view of the population structure. Microsatellite data provided new information on shared founder genotypes and provided a new DNA–based studbook pedigree that will assist in genetic management of the Whooping Crane population. [ABSTRACT FROM AUTHOR]

Presents a study that investigated the genetic variation of Gulf Coast sandhill cranes using mitochondrial DNA sequencing. DNA isolation and amplification; Discussion of the restriction enzyme survey; Information on the morphological measurements to classify sandhill cranes from the Gulf Coast of Texas.

AbstractEggs were sampled from 22 wild American alligator nests from the Rockefeller Wildlife Refuge in south-west Louisiana, along with the females guarding the nests. Three nests were sampled in 1995 and 19 were sampled in 1997. Females and offspring from all clutches were genotyped using five polymorphic microsatellite loci and the three nests from 1995 were also genotyped using one allozyme locus. Genotypes of the hatchlings were consistent with the guarding females being the mothers of their respective clutches. Multiple paternity was found in seven of the 22 clutches with one being fathered by three males, and the remaining six clutches having genotypes consistent with two males per clutch. Paternal contributions of multiply sired clutches were skewed. Some males sired hatchlings of more than one of the 22 clutches either as one of two sires of a multiple paternity clutch, as the sole sire of two different clutches, or as the sole sire of one clutch and one of two sires of a multiply sired clutch. There was no significant difference between females that had multiple paternity clutches and those that had singly sired clutches with respect to female total length (P = 0.844) and clutch size (P = 0.861). Also, there was no significant correlation between genetic relatedness of nesting females and pairwise nest distances (r2 = 0.003, F1,208 = 0.623, P = 0.431), indicating that females in this sample that nested close to one another were no more related than any two nesting females chosen at random. Eleven mutations were detected among hatchlings at the five loci over the 22 clutches. Most of these mutations (eight of 11) occurred at Amiµ-17, the only compound microsatellite locus of the five used in this study, corresponding to a mutation rate of 1.7 × 10-3. Finally, most of the mutations (82%) were homoplasious, i.e., mutating to an allelic state already present in this Louisiana population. [ABSTRACT FROM AUTHOR]

The Whooping Crane (Grus americana) is an endangered bird that suffered a severe population bottleneck; only 14 adults survived in 1938. We assessed the genetic effect of this human-caused bottleneck by sequencing 314 base pairs ( bp) of the mitochondrial DNA control region from cranes that lived before, during, and after this bottleneck. The maximum length of DNA amplifiable from museum specimens was negatively correlated with age, and only 10 of 153 specimens yielded the entire 314 bp sequence. Six haplotypes were present among the prebottleneck individuals sequenced, and only one of these persists in the modern population. The most common modern haplotype was in low frequency in the prebottleneck population, which demonstrates the powerful effect of genetic drift in changing allele frequencies in very small populations. By combining all available data, we show that no more than one-third of the prebottleneck haplotypes survived the human-caused population bottleneck. High levels of variation of substitution rates among nucleotide sites prevented us from estimating the prebottleneck population size. Our data will be incorporated into the captive breeding program to allow better management decisions regarding the preservation of current genetic diversity. These data offer the first glimpse into the genetic toll this species has paid for human activities. [ABSTRACT FROM AUTHOR]

Discusses the development of polymerase chain reaction assays of microsatellite loci for Swainson's Warbler, the only species in the genus Limnothlypis and among the rarest wood warblers in North America. Protocols that were used to obtain and score microsatellites; Cross-species amplification experiments that were conducted by testing individuals for amplifiability; Results.

Examines the genetic variation of Michigan elk through the use of protein electrophoresis on cellulose acetate. Differences in allele frequencies; Morphism; Mean heterozygosities and homozygosities; Founding individuals.

Wild animal diets are challenging to quantify, and the various methods for doing so have strengths and weaknesses. Combining multiple methods can allow ecologists to assess their level of confidence in particular results, increase sample size, and investigate diet over varying time scales. The biases of traditional gut content–based methods are mostly well understood. Newer methods may have important biases that can only be worked out through comparison to established ones. We collected data on the diet of wild Plains Hog-nosed Snakes (Heterodon nasicus) using multiple, fundamentally dissimilar methods, combined analytically using a Bayesian framework to describe an ontogenetic dietary shift. Gut contents were the most straightforward, but yielded a small sample size that fell below any reasonable threshold for making generalizations. Stable isotopes indicated an obvious ontogenetic dietary shift, but were labor-intensive, and conclusions are limited by multiple methodological caveats including similarity among prey groups, maternal carryover effects, and uncertainty in trophic enrichment factors. Fecal environmental DNA (eDNA) was intermediate in terms of effort, yielding results congruent with the other two methods, but the interpretation of which would likely have been confounded by contaminants had we not used all three methods in tandem. Several apparent artifacts are discussed. There are some reassuring similarities among methods. There are also several differences. The most complete picture uses data from all methods taken together. Future studies should attempt to compare the biases, expense, and potential drawbacks of these and other methods in greater detail. [ABSTRACT FROM AUTHOR]

Eight polymorphic microsatellite DNA loci were isolated from the flightless cormorant ( Phalacrocorax harrisi) for future population genetic studies. Genetic variability was assessed using at least 38 individuals from two populations. Allele numbers ranged from three to nine per locus. Mean observed heterozygosity varied from 0.27 to 0.78. No locus deviated from Hardy–Weinberg (HW) or linkage equilibria in either population. The high levels of detected polymorphism indicate the utility of these markers for population genetic studies of this Galápagos species. [ABSTRACT FROM AUTHOR]

New study systems and tools are needed to understand how divergence and speciation occur between lineages with gene flow. Migratory birds often exhibit divergence despite seasonal migration, which brings populations into contact with one another. We studied divergence between 2 subspecies of Northern Saw-whet Owl (Aegolius acadicus), in which a sedentary population on the islands of Haida Gwaii, British Columbia (A. a. brooksi), exists in the presence of the other form (A. a. acadicus) during migration but not during the breeding season. Prior research showed fixed mtDNA divergence but left open the question of nuclear gene flow. We used 2,517 ultraconserved element loci to examine the demographic history of this young taxon pair. Although we did not observe fixed single nucleotide polymorphism differences between populations among our genotyped individuals, 100% of the birds were diagnosable and δaδI analyses suggested the demographic model best fitting the data was one of split-bidirectional-migration (i.e. speciation with gene flow). We dated the split between brooksi and acadicus to ~278 Kya, and our analyses suggested gene flow between groups was skewed, with ~0.7 individuals per generation coming from acadicus into brooksi and ~4.4 going the opposite direction. Coupled with an absence of evidence of phenotypic hybrids and the birds' natural history, these data suggest brooksi may be a young biological species arising despite historic gene flow. [ABSTRACT FROM AUTHOR]

Simple Summary: Gastrointestinal tract disorders (GITDs) are a serious problem for both adult and young horses. In many cases, the sources of these disorders are unknown, and a link with alterations of the gastrointestinal (GI) microbiome (i.e., GI dysbiosis) is suspected. New alternatives to conventional amplicon sequencing are needed to decipher the real connection between GITDs and dysbiosis. This study compares the performance of target enrichment by hybridization capture (TEHC), a new approach for microbiome sequencing, with conventional amplicon sequencing for the characterization of the equine fecal microbiome. GITDs are among the most common causes of death in adult and young horses in the United States (US). Previous studies have indicated a connection between GITDs and the equine gut microbiome. However, the low taxonomic resolution of the current microbiome sequencing methods has hampered the identification of specific bacterial changes associated with GITDs in horses. Here, we have compared TEHC, a new approach for 16S rRNA gene selection and sequencing, with conventional 16S rRNA gene amplicon sequencing for the characterization of the equine fecal microbiome. Both sequencing approaches were used to determine the fecal microbiome of four adult horses and one commercial mock microbiome. Our results show that TEHC yielded significantly more operational taxonomic units (OTUs) than conventional 16S amplicon sequencing when the same number of reads were used in the analysis. This translated into a deeper and more accurate characterization of the fecal microbiome when the samples were sequenced with TEHC according to the relative abundance analysis. Alpha and beta diversity metrics corroborated these findings and demonstrated that the microbiome of the fecal samples was significantly richer when sequenced with TEHC compared to 16S amplicon sequencing. Altogether, our study suggests that the TEHC strategy provides a more extensive characterization of the fecal microbiome of horses than the current alternative based on the PCR amplification of a portion of the 16S rRNA gene. [ABSTRACT FROM AUTHOR]

We performed whole-genome multi-locus sequence typing for 2554 genes in a large and heterogenous panel of 180 Listeria monocytogenes strains having diverse geographical and temporal origins. The subtyping data was used for characterizing genetic variation and evaluating patterns of linkage disequilibrium in the pan-genome of L. monocytogenes. Our analysis revealed the presence of strong linkage disequilibrium in L. monocytogenes, with ~99% of genes showing significant non-random associations with a large majority of other genes in the genome. Twenty-seven loci having lower levels of association with other genes were considered to be potential "hot spots" for horizontal gene transfer (i.e., recombination via conjugation, transduction, and/or transformation). The patterns of linkage disequilibrium in L. monocytogenes suggest limited exchange of foreign genetic material in the genome and can be used as a tool for identifying new recombinant strains. This can help understand processes contributing to the diversification and evolution of this pathogenic bacteria, thereby facilitating development of effective control measures. [ABSTRACT FROM AUTHOR]

Determining how genetic diversity is structured between populations that span the divergence continuum from populations to biological species is key to understanding the generation and maintenance of biodiversity. We investigated genetic divergence and gene flow in eight lineages of birds with a trans‐Beringian distribution, where Asian and North American populations have likely been split and reunited through multiple Pleistocene glacial cycles. Our study transects the speciation process, including eight pairwise comparisons in three orders (ducks, shorebirds and passerines) at population, subspecies and species levels. Using ultraconserved elements (UCEs), we found that these lineages represent conditions from slightly differentiated populations to full biological species. Although allopatric speciation is considered the predominant mode of divergence in birds, all of our best divergence models included gene flow, supporting speciation with gene flow as the predominant mode in Beringia. In our eight lineages, three were best described by a split‐migration model (divergence with gene flow), three best fit a secondary contact scenario (isolation followed by gene flow), and two showed support for both models. The lineages were not evenly distributed across a divergence space defined by gene flow (M) and differentiation (FST), instead forming two discontinuous groups: one with relatively shallow divergence, no fixed single nucleotide polymorphisms (SNPs), and high rates of gene flow between populations; and the second with relatively deeply divergent lineages, multiple fixed SNPs, and low gene flow. Our results highlight the important role that gene flow plays in avian divergence in Beringia. [ABSTRACT FROM AUTHOR]

The western mosquitofish, Gambusia affinis, is a freshwater poecilid fish native to the southeastern United States but with a global distribution due to widespread human introduction. Gambusia affinis has been used as a model species for a broad range of evolutionary and ecological studies. We sequenced the genome of a male G. affinis to facilitate genetic studies in diverse fields including invasion biology and comparative genetics. We generated Illumina short read data from paired-end libraries and in vitro proximity-ligation libraries. We obtained 54.9×coverage, N50 contig length of 17.6 kb, and N50 scaffold length of 6.65 Mb. Compared to two other species in the Poeciliidae family, G. affinis has slightly fewer genes that have shorter total, exon, and intron length on average. Using a set of universal single-copy orthologs in fish genomes, we found 95.5% of these genes were complete in the G. affinis assembly. The number of transposable elements in the G. affinis assembly is similar to those of closely related species. The high-quality genome sequence and annotations we report will be valuable resources for scientists to map the genetic architecture of traits of interest in this species. [ABSTRACT FROM AUTHOR]

Accurate and consistent delimitation of species and their relationships provides a necessary framework for comparative studies, understanding evolutionary relationships, and informing conservation management. Despite the ever-increasing availability of genomic data, evolutionary dynamics can still render some relationships exceedingly difficult to resolve, including underlying speciation events that are rapid, recent, or confounded by post-speciation introgression. Here we present an empirical study of musk turtles ( Sternotherus ), which illustrates approaches to resolve difficult nodes in the Tree of Life that robust species-tree methods fail to resolve. We sequence 4430 RAD-loci from 205 individuals. Independent coalescent-based analyses, corroborated with morphology and geography, strongly support the recognition of cryptic species within Sternotherus , but with conflicting or weak support for some intraspecific relationships. To resolve species-tree conflict, we use a likelihood-based approach to test support for alternative demographic models behind alternative speciation scenarios and argue that demographic model testing has an important role for resolving systematic relationships in recent, rapid radiations. Species-tree and demographic modeling strongly support the elevation of two nominal subspecies in Sternotherus to species and the recognition of a previously cryptic species ( S. intermedius sp. nov.) described within. The evolutionary and taxonomic history of Sternotherus is discussed in the context of these new species and novel and well-supported systematic hypotheses. [ABSTRACT FROM AUTHOR]

The blacklegged tick (Ixodes scapularis (Journal of the Academy of Natural Sciences of Philadelphia, 1821, 2, 59)) is a vector of Borrelia burgdorferi sensu stricto (s.s.) (International Journal of Systematic Bacteriology, 1984, 34, 496), the causative bacterial agent of Lyme disease, part of a slow‐moving epidemic of Lyme borreliosis spreading across the northern hemisphere. Well‐known geographical differences in the vectorial capacity of these ticks are associated with genetic variation. Despite the need for detailed genetic information in this disease system, previous phylogeographical studies of these ticks have been restricted to relatively few populations or few genetic loci. Here we present the most comprehensive phylogeographical study of genome‐wide markers in I. scapularis, conducted by using 3RAD (triple‐enzyme restriction‐site associated sequencing) and surveying 353 ticks from 33 counties throughout the species' range. We found limited genetic variation among populations from the Northeast and Upper Midwest, where Lyme disease is most common, and higher genetic variation among populations from the South. We identify five spatially associated genetic clusters of I. scapularis. In regions where Lyme disease is increasing in frequency, the I. scapularis populations genetically group with ticks from historically highly Lyme‐endemic regions. Finally, we identify 10 variable DNA sites that contribute the most to population differentiation. These variable sites cluster on one of the chromosome‐scale scaffolds for I. scapularis and are within identified genes. Our findings illuminate the need for additional research to identify loci causing variation in the vectorial capacity of I. scapularis and where additional tick sampling would be most valuable to further understand disease trends caused by pathogens transmitted by I. scapularis. [ABSTRACT FROM AUTHOR]

Multiple paternity occurs in most species and animal groups that have been studied. Because mating involves fitness costs to individual females, theory predicts that polyandrous females gain greater fitness benefits than costs, allowing the behavior to be maintained. Genetic, rather than material, benefits often occur in species where males provide females with little more than sperm and seminal fluid. We compared fitness correlates of single- and double-sire clutches from female marbled salamanders ( Ambystoma opacum) at the egg, hatchling, and metamorph stages of offspring development. Because clutches were collected from experimental breeding groups, strict paternity exclusion of offspring using microsatellite data allowed us to categorize each clutch as having either one or two fathers. Early offspring viability and size of hatchlings were not different between single- and multiple-paternity clutches. Larvae from the two clutch types were allowed to develop together in field enclosures until metamorphosis. Although there was no difference in size at metamorphosis, survival to metamorphosis was significantly higher in multiple-paternity clutches (44% vs. 40%) suggesting a benefit for females. The results were consistent with genetic benefits, although maternal effects could not be ruled out. The data did not support predictions of the genetic bet-hedging and good sperm hypotheses for genetic benefits of polyandry. [ABSTRACT FROM AUTHOR]

Sequence capture and restriction site associated DNA sequencing (RAD-Seq) are two genomic enrichment strategies for applying next-generation sequencing technologies to systematics studies.At shallowtimescales, such as within species, RAD-Seq has been widely adopted among researchers, although there has been little discussion of the potential limitations and benefits of RAD-Seq and sequence capture. We discuss a series of issues that may impact the utility of sequence capture and RAD-Seq data for shallow systematics in non-model species.We review prior studies that used both methods, and investigate differences between the methods by re-analyzing existing RAD-Seq and sequence capture data sets from a Neotropical bird (Xenops minutus). We suggest that the strengths of RAD-Seq data sets for shallow systematics are the wide dispersion of markers across the genome, the relative ease and cost of laboratory work, the deep coverage and read overlap at recovered loci, and the high overall information that results. Sequence capture's benefits include flexibility and repeatability in the genomic regions targeted, success using low-quality samples, more straightforward read orthology assessment, and higher per-locus information content. The utility of a method in systematics, however, rests not only on its performance within a study, but on the comparability of data sets and inferences with those of prior work. In RADSeq data sets, comparability is compromised by low overlap of orthologous markers across species and the sensitivity of genetic diversity in a data set to an interaction between the level of natural heterozygosity in the samples examined and the parameters used for orthology assessment. In contrast, sequence capture of conserved genomic regions permits interrogation of the same loci across divergent species, which is preferable for maintaining comparability among data sets and studies for the purpose of drawing general conclusions about the impact of historical processes across biotas.We argue that sequence capture should be given greater attention as a method of obtaining data for studies in shallow systematics and comparative phylogeography. [ABSTRACT FROM AUTHOR]

Rapid evolutionary radiations are expected to require large amounts of sequence data to resolve. To resolve these types of relationships many systematists believe that it will be necessary to collect data by next-generation sequencing (NGS) and use multispecies coalescent ("species tree") methods. Ultraconserved element (UCE) sequence capture is becoming a popular method to leverage the high throughput of NGS to address problems in vertebrate phylogenetics. Herewe examine the performance of UCE data for gallopheasants (true pheasants and allies), a clade that underwent a rapid radiation 10-15 Ma. Relationships among gallopheasant genera have been difficult to establish. We used this rapid radiation to assess the performance of species tree methods, using ∼600 kilobases of DNA sequence data from ∼1500 UCEs. We also integrated information from traditional markers (nuclear intron data from 15 loci and three mitochondrial gene regions). Species tree methods exhibited troubling behavior. Two methods [Maximum Pseudolikelihood for Estimating Species Trees (MP-EST) and Accurate Species TRee ALgorithm (ASTRAL)] appeared to perform optimally when the set of input gene trees was limited to the most variable UCEs, though ASTRAL appeared to be more robust than MP-EST to input trees generated using less variable UCEs. In contrast, the rooted triplet consensus method implemented in Triplec performed better when the largest set of input gene trees was used. We also found that all three species tree methods exhibited a surprising degree of dependence on the program used to estimate input gene trees, suggesting that the details of likelihood calculations (e.g., numerical optimization) are important for loci with limited phylogenetic information. As an alternative to summary species tree methods we explored the performance of SuperMatrix Rooted Triple - Maximum Likelihood (SMRT-ML), a concatenation method that is consistent even when gene trees exhibit topological differences due to the multispecies coalescent. We found that SMRT-ML performed well for UCE data. Our results suggest that UCE data have excellent prospects for the resolution of difficult evolutionary radiations, though specific attention may need to be given to the details of the methods used to estimate species trees. [ABSTRACT FROM AUTHOR]

Aflatoxins are a group of potent foodborne toxicants naturally occurring in maize and groundnuts. Differential speciesspecific sensitivity to aflatoxins has been documented but cannot be fully explained by the differences in metabolismof these toxicants among animal species. Commensal microbial communities (microbiota) are critical to human and animal health, but few studies have assessed interactions between xenobiotic toxins and those microbiota, and its potential effects to humans and animals. Here, an exploratory dosing experiment was conducted to explore effects of Aflatoxin B1 (AFB1) on the gut microbiota in a commonly used rat model. Male F344 rats were randomly divided into groups and treated with different concentrations of AFB1. Microbial communities in fecal samples were assessed using 16S rRNA sequence analysis. We found that samples from the control group had a phylogenetically diverse community, and that increasing AFB1 doses decreased this diversity but increased evenness of community composition. In addition, the gut microbiota from different samples was clustered according to their dosing regimens. There is no community shift at the phylum level but some lactic acid bacteria were significantly depleted by AFB1. These findings suggested that AFB1 could modify the gut microbiota in a dose-dependent manner. [ABSTRACT FROM AUTHOR]

Since the recognition that allopatric speciation can be induced by large-scale reconfigurations of the landscape that isolate formerly continuous populations, such as the separation of continents by plate tectonics, the uplift of mountains or the formation of large rivers, landscape change has been viewed as a primary driver of biological diversification. This process is referred to in biogeography as vicariance. In the most species-rich region of the world, the Neotropics, the sundering of populations associated with the Andean uplift is ascribed this principal role in speciation. An alternative model posits that rather than being directly linked to landscape change, allopatric speciation is initiated to a greater extent by dispersal events, with the principal drivers of speciation being organism-specific abilities to persist and disperse in the landscape. Landscape change is not a necessity for speciation in this model. Here we show that spatial and temporal patterns of genetic differentiation in Neotropical birds are highly discordant across lineages and are not reconcilable with a model linking speciation solely to landscape change. Instead, the strongest predictors of speciation are the amount of time a lineage has persisted in the landscape and the ability of birds to move through the landscape matrix. These results, augmented by the observation that most species-level diversity originated after episodes of major Andean uplift in the Neogene period, suggest that dispersal and differentiation on a matrix previously shaped by large-scale landscape events was a major driver of avian speciation in lowland Neotropical rainforests. [ABSTRACT FROM AUTHOR]

Background: Deer mice (Peromyscus maniculatus) are the principal reservoir hosts of Sin Nombre virus (SNV), the cause of the great majority of hantavirus cardiopulmonary syndrome (HCPS) cases in North America. SNV, like all hantaviruses with their reservoirs, causes persistent infection without pathology in deer mice and appear to elicit a regulatory T cell response. Deer mice are also susceptible to Andes virus (ANDV), which causes the great majority of HCPS cases in South America, but they clear infection by 56 days post infection without signs of disease. Results: We examined lymph node cell responses of deer mice infected with ANDV to determine expression profiles upon in vitro recall challenge with viral antigen. Because the deer mouse genome is currently unannotated, we developed a bioinformatics pipeline to use known lab mouse (Mus musculus) cDNAs to predict genes within the deer mouse genome and design primers for quantitative PCR (http://dna.publichealth.uga.edu/BlastPrimer/ BlastPrimer.php). Of 94 genes examined, 20 were elevated, the plurality of which were Th2-specific, whereas 12 were downregulated. Other expressed genes represented Th1, regulatory T cells and follicular helper T cells, and B cells, but not Th17 cells, indicating that many cellular phenotypes participate in the host response to Andes virus. Conclusions: The ability to examine expression levels of nearly any gene from deer mice should allow direct comparison of infection with SNV or ANDV to determine the immunological pathways used for clearance of hantavirus infection in a reservoir host species [ABSTRACT FROM AUTHOR]

The prevalence of female multiple mating in natural populations is important for many questions in mating system evolution. Several statistical techniques use genetic data to estimate the number of fathers that contribute gametes to broods, but they have not been widely compared to assess the magnitude of differences in their performance. With a combination of new data and reanalysis of previously published data, we compared five analytical approaches: (1) allele-counting, (2) parental reconstruction in GERUD, (3) a Bayesian probability model to estimate the frequency of multiple mating (FMM), (4) computer simulations based on population allele frequencies in HAPLOTYPES and (5) Bayesian parental reconstruction in PARENTAGE. The results show that choice of analysis technique can significantly affect estimates of sire number. Estimates from GERUD conformed exactly to results obtained from strict exclusion of potential sires in an experimental context. However, estimates yielded by HAPLOTYPES and PARENTAGE sometimes exceeded the numbers from GERUD by as much as 120 and 55%, respectively. We recommend GERUD over these other approaches for most purposes because of its accuracy and consistency in this analysis. Our novel genetic data set allowed us to investigate the extent and frequency of multiple paternity in a marbled salamander (Ambystoma opacum) population in South Carolina, USA. A. opacum contrasted with other salamander species by having relatively low levels of multiple paternity (only 31-54% compared with 71-96%). Although A. opacum had the lowest level of multiple paternity under all analytical approaches used here, the magnitude of differences among species varied. [ABSTRACT FROM AUTHOR]

Information describing the possible impacts of manufactured nanoparticles on human health and ecological receptors is limited. The objective of the present study was to evaluate the potential toxicological effects of manufactured zinc oxide nanoparticles (ZnO-NPs; 1.5 nm) compared to aqueous zinc chloride (ZnCl2) in the free-living nematode Caenorhabditis elegans. Toxicity of both types of Zn was investigated using the ecologically relevant endpoints of lethality, behavior, reproduction, and transgene expression in a mtl-2::GFP (gene encoding green fluorescence protein fused onto the metallothionein-2 gene promoter) transgenic strain of C. elegans. Zinc oxide nanoparticles showed no significant difference from ZnCl2 regarding either lethality or reproduction in C. elegans, as indicated by their median lethal concentrations (LC50s; p = 0.29, n = 3) and median effective concentrations (EC50s; Z = 0.835, p = 0.797). Also, no significant difference was found in EC50s for behavioral change between ZnO-NPs (635 mg Zn/L; 95% confidence interval [CI], 477-844 mg Zn/L) and ZnCl2 (546 mg Zn/L; 95% CI, 447-666 mg Zn/L) (Z = 0.907, p = 0.834). Zinc oxide nanoparticles induced transgene expression in the mtl-2::GFP transgenic C. elegans in a manner similar to that of ZnCl2, suggesting that intracellular biotransformation of the nanoparticles might have occurred or the nanoparticles have dissolved to Zn2+ to enact toxicity. These findings demonstrate that manufactured ZnO-NPs have toxicity to the nematode C. elegans similar to that of aqueous ZnCl2. [ABSTRACT FROM AUTHOR]

The article discusses research on the genetic population structure of the Black-throated Blue Warbler or Dendroica caeruslescens which breeds in the forests in eastern North America. The researchers analyzed 333 base pairs of the mitochondrial control region from 287 birds from 14 populations in Michigan and central Appalachian forests. Results of their study revealed relatively high haplotype diversity, low nucleotide diversity, and limited but significant phylogeographic structure in the birds' breeding range and populations.

Background: Genome elucidation is now in high gear for many organisms, and whilst genetic maps have been developed for a broad array of species, surprisingly, no such maps exist for a crocodilian, or indeed any other non-avian member of the Class Reptilia. Genetic linkage maps are essential tools for the mapping and dissection of complex quantitative trait loci (QTL), and in order to permit systematic genome scans for the identification of genes affecting economically important traits in farmed crocodilians, a comprehensive genetic linage map will be necessary. Results: A first-generation genetic linkage map for the saltwater crocodile (Crocodylus porosus) was constructed using 203 microsatellite markers amplified across a two-generation pedigree comprising ten full-sib families from a commercial population at Darwin Crocodile Farm, Northern Territory, Australia. Linkage analyses identified fourteen linkage groups comprising a total of 180 loci, with 23 loci remaining unlinked. Markers were ordered within linkage groups employing a heuristic approach using CRIMAP v3.0 software. The estimated female and male recombination map lengths were 1824.1 and 319.0 centimorgans (cM) respectively, revealing an uncommonly large disparity in recombination map lengths between sexes (ratio of 5.7:1). Conclusion: We have generated the first genetic linkage map for a crocodilian, or indeed any other non-avian reptile. The uncommonly large disparity in recombination map lengths confirms previous preliminary evidence of major differences in sex-specific recombination rates in a species that exhibits temperature-dependent sex determination (TSD). However, at this point the reason for this disparity in saltwater crocodiles remains unclear. This map will be a valuable resource for crocodilian researchers, facilitating the systematic genome scans necessary for identifying genes affecting complex traits of economic importance in the crocodile industry. In addition, since many of the markers placed on this genetic map have been evaluated in up to 18 other extant species of crocodilian, this map will be of intrinsic value to comparative mapping efforts aimed at understanding genome content and organization among crocodilians, as well as the molecular evolution of reptilian and other amniote genomes. As researchers continue to work towards elucidation of the crocodilian genome, this first generation map lays the groundwork for more detailed mapping investigations, as well as providing a valuable scaffold for future genome sequence assembly. [ABSTRACT FROM AUTHOR]

Little is known about nest-site fidelity in wild Alligator mississippiensis (American Alligator). As part of a long-term study on multiple paternity and mate fidelity that required capture and marking of nesting female alligators, we documented several instances of nest-site reuse by individual females. In one case, the female nested at the same site seven years after initial capture, demonstrating long-term nest-site fidelity. Another female used one nest site in 2000, 2002, and 2004. Herein, we provide data on minimum nesting intervals for recaptured alligators, including three cases of females nesting in two consecutive years. Despite tremendous habitat alteration and storm surge from Hurricane Rita in 2005 and a catastrophic drought precluding nesting in 2006, we found female alligators in 2007 at nests within 20 m and 170 m of their nest sites from seven years and three years, respectively, prior to those events. [ABSTRACT FROM AUTHOR]

Factors underlying the evolution of female-biased sexual size dimorphism in mammals are poorly understood. In an effort to better understand these factors we tested whether larger female southern flying squirrels, Glaucomys volans, gained reproductive advantages (larger litters or more male mates) and direct resource benefits, such as larger home ranges or access to more food (i.e. mast-producing trees). As dimorphism can vary with age in precocial breeding species, we compared females during their first reproduction and during a subsequent breeding attempt. Females were not significantly larger or heavier than males at first reproduction, but became about 7% heavier and 22% larger than males at subsequent breeding. Larger females produced larger litters and had home ranges containing a greater proportion of upland hardwood trees. Female body size was not associated with either multiple male mating or home range size, but females with larger home ranges had higher indexes of body condition. Females in precocial breeding flying squirrels initiate reproduction before sexual size dimorphism is evident, and thus, may be allocating resources to both reproduction and growth simultaneously, or delaying growth entirely. Larger females produce more pups and have access to more food resources. Thus, selection for increased female size may partly explain how female-biased sexual size dimorphism is maintained in this species. [Copyright &y& Elsevier]

We assessed the effects of range disjunction, migratory habit, coloniality, and habitat structure on the genetic differentiation of North American Burrowing Owl (Athene cunicularia) populations. Burrowing Owls in North America comprise two forms or subspecies: A. c. floridana in Florida, separated by ∼1,500 km from the western form, A. c. hypugaea, which ranges from Texas to California and north to southern Canada. Burrowing Owls tend to be loosely colonial, and both the Florida populations and southerly populations of A. c. hypugaea from California to Texas are nonmigratory. To assess genetic structure, we examined 201 individuals from nine western and six Florida populations at seven highly variable microsatellite DNA loci. Mean gene diversity (Hexp) was higher in the west than in Florida (0.539 and 0.341, respectively; P < 0.05). Populations within subspecies were essentially panmictic (A. c. floridana: θ = 0.038, ρ = 0.014; A. c. hypugaea: θ = 0.014, ρ = 0.009) and even genetic differentiation across subspecies was modest (θ = 0.051, ρ = 0.014). Nevertheless, the western and Florida forms were easily distinguished by any of several criteria, such as allelic absences in Florida, assignment tests, and well-supported branches on the inferred phylogenetic tree. Genetic differentiation was at least twice as great in resident Florida (θ = 0.038) and California (θ = 0.021) populations as in migratory western populations (θ = 0.012), though 95% confidence intervals of theta estimates overlapped. We found no evidence of a genetic bottleneck that would result in evolutionary disequilibrium within subspecies. In the west, high observed heterozygosity values and evidence of gene flow suggest that population declines and patchy habitat, which currently imperil this species throughout much of its range, have not led to inbreeding or biologically meaningful genetic differentiation among the sampled populations. [ABSTRACT FROM AUTHOR]

Summary: Finding, characterizing and monitoring reservoirs for antimicrobial resistance (AMR) is vital to protecting public health. Hybridization capture baits are an accurate, sensitive and cost‐effective technique used to enrich and characterize DNA sequences of interest, including antimicrobial resistance genes (ARGs), in complex environmental samples. We demonstrate the continued utility of a set of 19 933 hybridization capture baits designed from the Comprehensive Antibiotic Resistance Database (CARD)v1.1.2 and Pathogenicity Island Database (PAIDB)v2.0, targeting 3565 unique nucleotide sequences that confer resistance. We demonstrate the efficiency of our bait set on a custom‐made resistance mock community and complex environmental samples to increase the proportion of on‐target reads as much as >200‐fold. However, keeping pace with newly discovered ARGs poses a challenge when studying AMR, because novel ARGs are continually being identified and would not be included in bait sets designed prior to discovery. We provide imperative information on how our bait set performs against CARDv3.3.1, as well as a generalizable approach for deciding when and how to update hybridization capture bait sets. This research encapsulates the full life cycle of baits for hybridization capture of the resistome from design and validation (both in silico and in vitro) to utilization and forecasting updates and retirement. [ABSTRACT FROM AUTHOR]

We collected fish and mammals in several radioactively contaminated locations in the Chornobyl Exclusion Zone and analyzed them for 137Cs content. Frequency distributions were built for populations of channel catfish, yellow-necked mice and bank voles. We combined our data with similar data from several other studies to demonstrate the relationship between the standard deviations and means of 137Cs of fish and mammal populations. The frequency distributions of 137Cs in populations of fish and mammals are not normal, as indicated by the strong relationship between standard deviation and mean. Distributions for mammals are more skewed than those for fish. Fish and mammals probably use their environments in fundamentally different ways. The highest concentrations and thus greatest risks are therefore confined to relatively few individuals in each population. [Copyright &y& Elsevier]

Triatominae are the largest radiation of haematophagous species within the true bugs (Hemiptera) and the sole vectors of Trypanosoma cruzi Chagas, the causative agent of Chagas disease, a neglected human disease that affects millions. We used a combined ultraconserved element (UCE) and ribosomal dataset from ethanol‐preserved and pinned specimens in a museomics approach to elucidate the phylogenetic relationships among triatomines, including taxa of four of the five tribes with substantial representation of the Old World fauna. We conclude that: (i) Triatominae are monophyletic and Opisthacidius Berg is their predatory sister taxon; (ii) the three large genera (Rhodnius Stål, Triatoma Laporte and Panstrongylus Berg) are not monophyletic; (iii) Triatomini fall into nine well‐supported clades, only two of which are identical in composition to previously recognized groups; and (iv) the Old World clade is deeply nested within Triatomini. Ancestral character state reconstructions of specific character traits provide insights into the evolutionary history of Triatominae. [ABSTRACT FROM AUTHOR]

A reliable and standardized classification of Listeria monocytogenes is important for accurate strain identification during outbreak investigations. Current whole-genome sequencing (WGS)-based approaches for strain characterization are either difficult to standardize, rendering them less suitable for data exchange, or are not freely available. Thus, we developed a portable and open-source tool, Haplo-ST, to improve standardization and provide maximum discriminatory potential to WGS data tied to a multilocus sequence typing (MLST) framework. Haplo-ST performs whole-genome MLST (wgMLST) for L. monocytogenes while allowing for data exchangeability worldwide. This tool takes in (i) raw WGS reads as input, (ii) cleans the raw data according to user-specified parameters, (iii) assembles genes across loci by mapping to genes from reference strains, and (iv) assigns allelic profiles to assembled genes and provides a wgMLST subtyping for each isolate. Data exchangeability relies on the tool assigning allelic profiles based on a centralized nomenclature defined by the widely used BIGSdb-Lm database. Tests of Haplo-ST's performance with simulated reads from L. monocytogenes reference strains demonstrated high sensitivity (97.5%), and coverage depths of ≥20X were found to be sufficient for wgMLST profiling. We then used Haplo-ST to characterize and differentiate between two groups of L. monocytogenes isolates derived from the natural environment and poultry processing plants. Phylogenetic reconstruction identified lineages within each group, and no lineage specificity was observed with isolate phenotypes (transient versus persistent) or origins. Genetic differentiation analyses between isolate groups identified 21 significantly differentiated loci, potentially enriched for adaptation and persistence of L. monocytogenes within poultry processing plants. [ABSTRACT FROM AUTHOR]

• The Eastern red bat (Lasiurus borealis) and big brown bat (Eptesicus fuscus) are common across the United States corn belt. • We examined the diets of both species to determine the agricultural pests they consume. • We provide new insight into the diversity of bat diets by using the recently developed ANML DNA primers. • 92% of big brown bat samples and 94% of red bat samples contained agricultural pest genera. • Individual guano sample for both bat species contained nearly two pest genera each. To meet growing global food demand, producers are actively searching for ways tocrop yield. One way to boost yield is to reduce arthropod damage. However, insecticides are becoming less effective and can have detrimental effects on human health and the environment. Learning how to harness natural pest control is one way to reduce arthropod damage to crops and avoid adverse effects of insecticide use. Therefore, we examined the diets of two bat species that are likely to provide pest control services across the United States corn belt: the eastern red bat (Lasiurus borealis) and the big brown bat (Eptesicus fuscus). We also show that primer choice can impact the diversity of taxa detected and resecent adavances in primer design can improve diet detection studies. Using new ANML primers to isolate prey DNA in fecal material, we found a more diverse diet than previously reported for both bat species. Big brown bats consumed an average of 16.2 species from 8.5 genera. Red bats consumed an average of 28.6 species from 15.8 genera. We found pest genera in 92 % of big brown bat samples and 94 % of red bat samples. For both bat species, approximately two pest genera were detected in each sample. Corn rootworms (Diabrotica), Cloverworms (Hypena), plant bugs (Lygus), wireworms (Melanotus), seed corn beetles (Stenolophus), and armyworms (Spodoptera) were the most frequently consumed agricultural pests. We confirm earlier studies that document beetles (Coleoptera) and moths (Lepidoptera) as the most common prey items of big brown and red bats, respectively. However, we also document a diverse diet for both bat species that includes many true bugs (Hemiptera). Because pest control services of big brown and red bats are not completely overlapped, we conclude that a diverse bat community is essential to maximize pest control services by bats. [ABSTRACT FROM AUTHOR]

Premise of the study: Ten microsatellite loci were isolated and characterized for the neotropical epiphytic orchid Epidendrum firmum to examine levels of genetic diversity and genetic structure at multiple spatial scales. Methods and Results: We screened loci in 12-25 individuals from each of two populations in Costa Rica and identified 10 polymorphic loci. The number of alleles per locus ranged from one to 15 while observed heterozygosity for polymorphic loci ranged from 0.360 to 0.960. Conclusions: Primers for these informative genetic markers will be useful for quantifying genetic diversity, spatial genetic structure, and gene fl ow in E. firmum. [ABSTRACT FROM AUTHOR]