Análise preliminar revela baixos níveis de diversidade na estrutura filogeográfica da cascavel mexicana Crotalus polystictus (Serpentes: Viperidae).

Se investigaron las relaciones matrilineales entre poblaciones de la serpiente de cascabel Mexicana cabeza de lanza  (Crotalus polystictus), esta especie se distribuye en valles que presentan elevaciones altas de la meseta del sur de México. Se analizó un fragmento mitocondrial de la ATPasa 8 y 6, los genes (589 pares de bases) revelaron bajos niveles de diversidad genética, con poco polimorfismo nucleótico entre la muestra geográfica analizada. La poca divergencia (1.0%) intraespecífica encontrada en los genes de la ATPasa 8 y 6 en C. polystictus contrastan con los fuertes porcentajes de divergencia (~1.0–14.1%) que han sido observados dentro de otros linajes de serpientes de cascabel mexicanas que habitan elevaciones altas. La variación intraespecífica es observadacomúnmente en especies de cascabel que se distribuyen en elevaciones menores y que presentan una distribución amplia (por ejemplo, C. tigris). Proponemos que la baja diversidad genética encontrada en C. polystictus comparada con la diversidad registrada en otras serpientes de cascabel que habitan elevaciones altas, se debe a diferencias ecológicas que han dado como resultado una respuesta evolutiva diferente en esta especie a los eventos climáticos del Pleistoceno. Nuestros resultados de una aparente baja diversidad genética en C. polystictus son un fuerte soporte para hacer un llamadoa la importancia de realizar iniciativas de conservación para proteger praderas con elevaciones altas en el centro de México.Foram investigadas as relações matrilineares entre as populações da cascavel mexicana Crotalus polystictus, uma espécie que habita vales de altitude do planalto do sul do México. Os genes (589 pares de bases) de um fragmento mitocondrial da ATPase 8 e 6 revelaram níveis relativamente baixos de diversidade genética, com poucos polimorfismos de nucleotídeos entre a amostra geográfica analisada. A baixa divergência da sequência intraespecífica (1.0%) encontrada nos genes da ATPase 8 e 6 de C. polystictus contrasta com asfortes divergências (~1.0–14.1%) observadas em outras linhagens de cascavel que habitam grandes altitudes, sendo que a variação intra-específica é observada comumente em espécies de cascavel que estão distribuídas em altitudes mais baixas e apresentam uma ampla distribuição (por exemplo, C.tigris). Propomos que a baixa diversidade genética encontrada em C. polystictus, comparada com a de outras cascavéis que habitam altitudes elevadas, pode refletir diferenças ecológicas que resultaram em uma resposta evolutiva diferente aos eventos climáticos do Pleistoceno. Nossos resultados deuma baixa diversidade genética aparente em C. polystictus destacam a importância de iniciativas de conservação para proteger os campos de altitude da região central do México.We investigated matrilineal relationships among populations of the Mexican lance-headed rattlesnake (Crotalus polystictus), a pitviper inhabiting high-elevation valleys of the densely populated southern Mexican Plateau. A fragment of the mitochondrial ATPase 8 and 6 genes (589 base pairs) revealed comparatively low levels of genetic diversity, with few nucleotide polymorphisms across the portion of the geographic distribution sampled. The shallow intraspecific sequence divergence (1.0%) in C. polystictus ATPase 8 and 6 genes contrasts with deepdivergences (~1.0–14.1%) observed within other montane rattlesnake lineages from the Mexican highlands, and is more typical of intraspecific variation observed in lowland rattlesnake species with similar distributional extents (e.g., C. tigris). We posit that the low genetic diversity in C. polystictus relative to that of other highland rattlesnakes may reflect ecological differences resulting in a different evolutionary response to Pleistocene climatic events. Our finding of apparently low genetic diversity in C. polystictus highlights the importance of conservation initiatives to protect high elevation grasslands in central Mexico

Rapid range expansion in the Great Plains narrow-mouthed toad (Gastrophryne olivacea) and a revised taxonomy for North American microhylids

We investigated genetic variation within the Great Plains narrow-mouthed toad, Gastrophryne olivacea, across its geographic range in the United States and Mexico. An analysis of mitochondrial DNA (mtDNA) from 105 frogs revealed remarkably low levels of genetic diversity in individuals inhabiting the central United States and northern Mexico. We found that this widespread matrilineal lineage is divergent (ca. 2% in mtDNA) from haplotypes that originate from the western United States and western coast of Mexico. Using a dataset that included all five species of Gastrophryne and both species of the closely related genus Hypopachus, we investigated the phylogenetic placement of G. olivacea. This analysis recovered strong support that G. olivacea, the tropically distributed G. elegans, and the temperately distributed G. carolinensisconstitute a monophyletic assemblage. However, the placement of G. pictiventris and G. usta render Gastrophryne paraphyletic with respect to Hypopachus. To complement our mitochondrial analysis, we examined a small fragment of nuclear DNA and recovered consistent patterns. In light of our findings we recommend (1) the resurrection of the nomen G. mazatlanensis Taylor (1943) for the disjunct western clade of G. olivaceaand (2) the tentative placement of G. pictiventris and G. usta in Hypopachus. To explore possible scenarios leading to low levels of genetic diversity in G. olivacea, we used mismatch distributions and Bayesian Skyline plots to examine historic population expansion and demography. Collectively these analyses suggest that G. olivacea rapidly expanded in effective population size and geographic range during the late Pleistocene or early Holocene. This hypothesis is consistent with fossil data from northern localities and contemporary observations that suggest ongoing northern expansion. Given our findings, we suspect that the rapid range expansion of G. olivacea may have been facilitated by ecological associations with open habitats and seasonal water bodies

Limitations of Climatic Data for Inferring Species Boundaries: Insights from Speckled Rattlesnakes

Phenotypes, DNA, and measures of ecological differences are widely used in species delimitation. Although rarely defined in such studies, ecological divergence is almost always approximated using multivariate climatic data associated with sets of specimens (i.e., the "climatic niche"); the justification for this approach is that species-specific climatic envelopes act as surrogates for physiological tolerances. Using identical statistical procedures, we evaluated the usefulness and validity of the climate-as-proxy assumption by comparing performance of genetic (nDNA SNPs and mitochondrial DNA), phenotypic, and climatic data for objective species delimitation in the speckled rattlesnake (Crotalus mitchellii) complex. Ordination and clustering patterns were largely congruent among intrinsic (heritable) traits (nDNA, mtDNA, phenotype), and discordance is explained by biological processes (e.g., ontogeny, hybridization). In contrast, climatic data did not produce biologically meaningful clusters that were congruent with any intrinsic dataset, but rather corresponded to regional differences in atmospheric circulation and climate, indicating an absence of inherent taxonomic signal in these data. Surrogating climate for physiological tolerances adds artificial weight to evidence of species boundaries, as these data are irrelevant for that purpose. Based on the evidence from congruent clustering of intrinsic datasets, we recommend that three subspecies of C. mitchellii be recognized as species: C. angelensis, C. mitchellii, and C. Pyrrhus

Rapid Microsatellite Identification from Illumina Paired-End Genomic Sequencing in Two Birds and a Snake

Identification of microsatellites, or simple sequence repeats (SSRs), can be a time-consuming and costly investment requiring enrichment, cloning, and sequencing of candidate loci. Recently, however, high throughput sequencing (with or without prior enrichment for specific SSR loci) has been utilized to identify SSR loci. The direct “Seq-to-SSR” approach has an advantage over enrichment-based strategies in that it does not require a priori selection of particular motifs, or prior knowledge of genomic SSR content. It has been more expensive per SSR locus recovered, however, particularly for genomes with few SSR loci, such as bird genomes. The longer but relatively more expensive 454 reads have been preferred over less expensive Illumina reads. Here, we use Illumina paired-end sequence data to identify potentially amplifiable SSR loci (PALs) from a snake (the Burmese python, Python molurus bivittatus), and directly compare these results to those from 454 data. We also compare the python results to results from Illumina sequencing of two bird genomes (Gunnison Sage-grouse, Centrocercus minimus, and Clark's Nutcracker, Nucifraga columbiana), which have considerably fewer SSRs than the python. We show that direct Illumina Seq-to-SSR can identify and characterize thousands of potentially amplifiable SSR loci for as little as $10 per sample – a fraction of the cost of 454 sequencing. Given that Illumina Seq-to-SSR is effective, inexpensive, and reliable even for species such as birds that have few SSR loci, it seems that there are now few situations for which prior hybridization is justifiable

Rediscovery of the Endangered Carchi Andean Toad, \u3cem\u3eRhaebo colomai\u3c/em\u3e (Hoogmoed, 1985), in Ecuador, with Comments on Its Conservation Status and Extinction Risk

Since 1984 there have been no records of Rhaebo colomai (Hoogmoed, 1985) within the territory of Ecuador. This species was known from 2 localities in the province of Carchi, northwestern Ecuador, and the department of Nariño, southwestern Colombia, which were reported in 1979 and 2015, respectively. We report the recent sightings of R. colomai at 3 new localities in Ecuador and discuss and evaluate this species’ extinction risk and conservation status

Report from the First Snake Genomics and Integrative Biology Meeting

This report summarizes the proceedings of the 1st Snake Genomics and Integrative Biology Meeting held in Vail, CO USA, 5-8 October 2011. The meeting had over twenty registered participants, and was conducted as a single session of presentations. Goals of the meeting included coordination of genomic data collection and fostering collaborative interactions among researchers using snakes as model systems.Organismic and Evolutionary Biolog

Report from the First Snake Genomics and Integrative Biology Meeting

This report summarizes the proceedings of the 1st Snake Genomics and Integrative Biology Meeting held in Vail, CO USA, 5-8 October 2011. The meeting had over twenty registered participants, and was conducted as a single session of presentations. Goals of the meeting included coordination of genomic data collection and fostering collaborative interactions among researchers using snakes as model systems

Rediscovery of the Endangered Carchi Andean Toad, Rhaebo colomai (Hoogmoed, 1985), in Ecuador, with comments on its conservation status and extinction risk

Since 1984 there have been no records of Rhaebo colomai (Hoogmoed, 1985) within the territory of Ecuador. This species was known from 2 localities in the province of Carchi, northwestern Ecuador, and the department of Nari��o, southwestern Colombia, which were reported in 1979 and 2015, respectively. We report the recent sightings of R. colomai at 3 new localities in Ecuador and discuss and evaluate this species��� extinction risk and conservation status

Microsatellite discovery in an insular amphibian (Grandisonia alternans) with comments on cross-species utility and the accuracy of locus identification from unassembled Illumina data

The Seychelles archipelago is unique among isolated oceanic islands because it features an endemic radiation of caecilian amphibians (Gymnophiona). In order to develop population genetics resources for this system, we identified microsatellite loci using unassembled Illumina MiSeq data generated from a genomic library of Grandisonia alternans, a species that occurs on multiple islands in the archipelago. Applying a recently described method (PALFINDER) we identified 8001 microsatellite loci that were potentially informative for population genetics analyses. Of these markers, we screened 60 loci using five individuals, directly sequenced several amplicons to confirm their identity, and then used eight loci to score allele sizes in 64 G. alternans individuals originating from five islands. A number of these individuals were sampled using non-lethal methods, demonstrating the efficacy of non-destructive molecular sampling in amphibian research. Although two loci satisfied our criteria as diploid, neutrally evolving loci with the statistical power to detect population structure, our success in identifying reliable loci was very low. Additionally, we discovered some issues with primer redundancy and differences between Illumina and Sanger sequences that suggest some Illumina-inferred loci are invalid. We investigated cross-species utility for eight loci and found most could be successfully amplified, sequenced and aligned across other species and genera of caecilians from the Seychelles. Thus, our study in part supported the validity of using PALFINDER with unassembled reads for microsatellite discovery within and across species, but importantly identified major limitations to applying this approach to small datasets (ca. 1 million reads) and loci with small tandem repeat sizes

Elevated AKR1C3 expression promotes prostate cancer cell survival and prostate cell-mediated endothelial cell tube formation: implications for prostate cancer progressioan

<p>Abstract</p> <p>Background</p> <p>Aldo-keto reductase (AKR) 1C family member 3 (AKR1C3), one of four identified human AKR1C enzymes, catalyzes steroid, prostaglandin, and xenobiotic metabolism. In the prostate, AKR1C3 is up-regulated in localized and advanced prostate adenocarcinoma, and is associated with prostate cancer (PCa) aggressiveness. Here we propose a novel pathological function of AKR1C3 in tumor angiogenesis and its potential role in promoting PCa progression.</p> <p>Methods</p> <p>To recapitulate elevated AKR1C3 expression in cancerous prostate, the human PCa PC-3 cell line was stably transfected with an AKR1C3 expression construct to establish PC3-AKR1C3 transfectants. Microarray and bioinformatics analysis were performed to identify AKR1C3-mediated pathways of activation and their potential biological consequences in PC-3 cells. Western blot analysis, reverse transcription-polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and an <it>in vitro </it>Matrigel angiogenesis assays were applied to validate the pro-angiogenic activity of PC3-AKR1C3 transfectants identified by bioinformatics analysis.</p> <p>Results</p> <p>Microarray and bioinformatics analysis suggested that overexpression of AKR1C3 in PC-3 cells modulates estrogen and androgen metabolism, activates insulin-like growth factor (IGF)-1 and Akt signaling pathways, as well as promotes tumor angiogenesis and aggressiveness. Levels of IGF-1 receptor (IGF-1R) and Akt activation as well as vascular endothelial growth factor (VEGF) expression and secretion were significantly elevated in PC3-AKR1C3 transfectants in comparison to PC3-mock transfectants. PC3-AKR1C3 transfectants also promoted endothelial cell (EC) tube formation on Matrigel as compared to the AKR1C3-negative parental PC-3 cells and PC3-mock transfectants. Pre-treatment of PC3-AKR1C3 transfectants with a selective IGF-1R kinase inhibitor (AG1024) or a non-selective phosphoinositide 3-kinases (PI3K) inhibitor (LY294002) abolished ability of the cells to promote EC tube formation.</p> <p>Conclusions</p> <p>Bioinformatics analysis followed by functional genomics demonstrated that AKR1C3 overexpression promotes angiogenesis and aggressiveness of PC-3 cells. These results also suggest that AKR1C3-mediated tumor angiogenesis is regulated by estrogen and androgen metabolism with subsequent IGF-1R and Akt activation followed by VEGF expression in PCa cells.</p