Massive Molecular Parallel Evolution of the HSP90AA1 Gene between High-elevation Anurans

HSP90AA1 is part of the heat shock protein 90 gene family and has important functions against heat stress. We report a case of molecular level parallel evolution of the HSP90AA1 gene in high elevation amphibians. IISP90AA1 gene sequences of four high-elevation anurans, Bufo gargarizans, Nanorana parkeri, Rana kukunoris, and Scutiger boulengeri, were compared along with five of their low-elevation relatives. A total of 16 amino-acid sites were identified as parallel evolution between N parkeri and R. kukunoris. We generated both model based (Zhang and Kumar's test) and empirical data based (parallel/divergence plotting) null distributions for non-parallel evolution, and both methods clearly determined that the observed number of parallel substitutions were significantly more than the null expectation. Furthermore, on the HSP90AA1 gene tree, N. parkeri and R. kukunoris formed a strongly supported Glade that was away from their respective relatives. This study provides a clear case of molecular parallel evolution, which may have significant implications in understanding the genetic mechanisms of high-elevation adaptation

Tail Waving Speed Affects Territorial Response in the Toad-headed Agama Phrynocephalus vlangalii

Territorial response affects a resident's access to resources and mates, but the response level is likely flexible due to the trade-off between benefits and costs. Residents have to decide the response based on their own quality, resource benefits and intruder threat. Here we examined the association between territorial response and intruder threat using a newly developed 3D animation in male Phrynocephalus vlangalii. Three intruder stimuli (fast tail waving plus tail coil, slow tail waving plus tail coil and tail coil only) were animated based on display characters of real male P. vlangalii, and played to the resident in the field using a within subject design. We found that residents who faced fast tail waving plus tail coil displays more often emerged from their burrows compared with those faced slow tail waving plus tail coil displays and tail coil only displays, suggesting high speed tail waving display likely encodes high threat and triggers high territorial response. We also found that residents who faced slow tail waving plus tail coil displays opted to stay in the burrow compared with those faced tail coil only displays, suggesting slow tail waving display likely encodes low threat and functions in social conflict alleviation. Taken together, our study casts important insight on the association between intruder tail waving speed and resident's territorial response. and provides some evidence that intruder tail waving speed is positively associated with its threat level

Big mountains but small barriers: Population genetic structure of the Chinese wood frog (Rana chensinensis) in the Tsinling and Daba Mountain region of northern China

<p>Abstract</p> <p>Background</p> <p>Amphibians in general are poor dispersers and highly philopatric, and landscape features often have important impacts on their population genetic structure and dispersal patterns. Numerous studies have suggested that genetic differentiation among amphibian populations are particularly pronounced for populations separated by mountain ridges. The Tsinling Mountain range of northern China is a major mountain chain that forms the boundary between the Oriental and Palearctic zoogeographic realms. We studied the population structure of the Chinese wood frog (<it>Rana chensinensis</it>) to test whether the Tsinling Mountains and the nearby Daba Mountains impose major barriers to gene flow.</p> <p>Results</p> <p>Using 13 polymorphic microsatellite DNA loci, 523 individuals from 12 breeding sites with geographical distances ranging from 2.6 to 422.8 kilometers were examined. Substantial genetic diversity was detected at all sites with an average of 25.5 alleles per locus and an expected heterozygosity ranging from 0.504 to 0.855, and two peripheral populations revealed significantly lower genetic diversity than the central populations. In addition, the genetic differentiation among the central populations was statistically significant, with pairwise <it>F</it>_{<it>ST </it>}values ranging from 0.0175 to 0.1625 with an average of 0.0878. Furthermore, hierarchical AMOVA analysis attributed most genetic variation to the within-population component, and the between-population variation can largely be explained by isolation-by-distance. None of the putative barriers detected from genetic data coincided with the location of the Tsinling Mountains.</p> <p>Conclusion</p> <p>The Tsinling and Daba Mountains revealed no significant impact on the population genetic structure of <it>R. chensinensis</it>. High population connectivity and extensive juvenile dispersal may account for the significant, but moderate differentiation between populations. Chinese wood frogs are able to use streams as breeding sites at high elevations, which may significantly contribute to the diminishing barrier effect of mountain ridges. Additionally, a significant decrease in genetic diversity in the peripheral populations supports Mayr's central-peripheral population hypothesis.</p

The prevalence of genome replacement in unisexual salamanders of the genus Ambystoma (Amphibia, Caudata) revealed by nuclear gene genealogy

<p>Abstract</p> <p>Background</p> <p>Unisexual salamanders of the genus <it>Ambystoma </it>exemplify the most ancient lineage of unisexual vertebrates and demonstrate an extremely flexible reproductive system. Unisexual <it>Ambystoma </it>interact with and incorporate genomes from two to four sexual species (<it>A. laterale</it>, <it>A. jeffersonianum</it>,<it>A. texanum</it>, and <it>A. tigrinum</it>), to generate more than 20 genome compositions or biotypes. Unisexual ploidy levels range from diploid to pentaploid, but all contain at least one <it>A. laterale </it>(L) genome. Replacement of nuclear genomes might be responsible for the evolutionary longevity of unisexual <it>Ambystoma </it>but direct evidence for the prevalence of genome replacement in natural populations is absent. Two major puzzling questions have remained unanswered over the last few decades: 1) is genome replacement a common reproductive method in various unisexual populations and, 2) is there an ancient "L" genome that persists in various unisexual genome compositions.</p> <p>Results</p> <p>We examined 194 unisexual and 89 <it>A. laterale </it>specimens from 97 localities throughout their range and constructed a genealogy of the "L" genomes using a nuclear DNA marker (L-G1C12) to answer the above questions. Six L-G1C12 haplotypes (A-F) were shared by individuals in various <it>A. laterale </it>and unisexual populations. The general geographical distribution of the haplotypes in unisexual populations conformed to those found in <it>A. laterale</it>, indicating that "L" genomes in unisexuals are obtained from sympatric or nearby populations of <it>A. laterale</it>.</p> <p>Conclusion</p> <p>Our data demonstrate that genome replacement frequently occurs in unisexual <it>Ambystoma </it>across their range, and support previous speculations that genome replacement is an important reproductive mechanism that can enhance their evolutionary longevity. Our results show that there is no ancient "L" genome in the unisexual lineages, and no particular "L" genome is favored in any unisexual individual. The presence of an "L" genome in all unisexuals implies that it is important to the maintenance of unisexuals. Nuclear gene genealogy is a powerful tool to examine the historical interaction between sperm-dependent unisexuals and their sexual sperm donors. This methodology could be applicable to many other unisexual lineages to improve our understanding of their reproduction and their ability to persist.</p

Species delineation using Bayesian model-based assignment tests: a case study using Chinese toad-headed agamas (genus Phrynocephalus)

<p>Abstract</p> <p>Background</p> <p>Species are fundamental units in biology, yet much debate exists surrounding how we should delineate species in nature. Species discovery now requires the use of separate, corroborating datasets to quantify independently evolving lineages and test species criteria. However, the complexity of the speciation process has ushered in a need to infuse studies with new tools capable of aiding in species delineation. We suggest that model-based assignment tests are one such tool. This method circumvents constraints with traditional population genetic analyses and provides a novel means of describing cryptic and complex diversity in natural systems. Using toad-headed agamas of the <it>Phrynocephalus vlangalii </it>complex as a case study, we apply model-based assignment tests to microsatellite DNA data to test whether <it>P. putjatia</it>, a controversial species that closely resembles <it>P. vlangalii </it>morphologically, represents a valid species. Mitochondrial DNA and geographic data are also included to corroborate the assignment test results.</p> <p>Results</p> <p>Assignment tests revealed two distinct nuclear DNA clusters with 95% (230/243) of the individuals being assigned to one of the clusters with > 90% probability. The nuclear genomes of the two clusters remained distinct in sympatry, particularly at three syntopic sites, suggesting the existence of reproductive isolation between the identified clusters. In addition, a mitochondrial ND2 gene tree revealed two deeply diverged clades, which were largely congruent with the two nuclear DNA clusters, with a few exceptions. Historical mitochondrial introgression events between the two groups might explain the disagreement between the mitochondrial and nuclear DNA data. The nuclear DNA clusters and mitochondrial clades corresponded nicely to the hypothesized distributions of <it>P. vlangalii </it>and <it>P. putjatia</it>.</p> <p>Conclusions</p> <p>These results demonstrate that assignment tests based on microsatellite DNA data can be powerful tools for distinguishing closely related species and support the validity of <it>P. putjatia</it>. Assignment tests have the potential to play a significant role in elucidating biodiversity in the era of DNA data. Nonetheless, important limitations do exist and multiple independent datasets should be used to corroborate results from assignment programs.</p

A new and efficient method for purification of poly-γ- glutamic acid from high-viscosity fermentation broth

Purpose: To devise an efficient strategy for the separation and recovery of high-quality γ-PGA by investigation of the physical properties, pigment properties and microfiltration mode of high-viscosity fermentation broth.Methods: The bacterial strain, Bacillus subtilis 115, was used in this study. The viscosity of the fermentation broth was determined by digital viscometer with spindle SP-2 at 25 oC. The concentrations of glucose and L-glutamate were analyzed with a biosensor equipped with both glucose oxidase and Lglutamate oxidase electrodes. The pigment in the fermentation liquid was scanned with a UV spectrophotometer at wavelength range of 200 - 500 nm and was removed using activated carbon. Measurement of IR spectrum was performed using an IR spectrophotometer with KBr pellet. Results: The results showed that the γ-PGA yield was 35 g/L. The viscosity of the fermentation broth was 1600 mPa.s at the end of the batch fermentation. After 3-fold dilution, the viscosity was reduced to one-fortieth of the original value at 65 °C for 30 min., which allowed effective removal of Bacillus subtilis 115 from the broth. Maximum UV absorption of the pigment was occurred at 260 nm. The pigment was removed by shaking with 0.6 % activated carbon powder at 50 rpm for 20 min, resulting in 88 % decolorization. Concentration with hollow-fiber membrane (MWCO 500,000) resulted in complete removal of residual glucose and glutamic acid from the aqueous solution of γ-PGA. The molecular weight of the γ-PGA was 1095 kDa, and its UV scanning spectrum showed an absorption peak at 216 nm. The decomposition temperature (Td) of the γ-PGA was 312.92 oC. Its IR spectrum was consistent with the presence of carboxyl, hydroxyl, carbonyl and amide groups.Conclusion: An efficient method for the extraction and purification of high-quality γ-PGA from highviscosity fermentation broth.Keywords: Bacillus subtilis 115, γ-Polyglutamic acid, De-pigmentation, Activated carbon, Ultra-filtration, High-viscosity fermentation brot