Phylogeographic History of Endangered Hokuriku Salamander, Hynobius takedai (Amphibia: Caudata).

Knowledge of the phylogeographic history of organisms is valuable for understanding their evolutionary processes. To the best of our knowledge, the phylogeographic structure of Hokuriku salamander, Hynobius takedai, an endangered species, remains unclear. This study aimed to elucidate the phylogeographic history of H. takedai, which is expected to be strongly influenced by paleogeographic events. Phylogenetic analysis based on partial sequences of the mitochondrial DNA cytochrome b gene confirmed the genetic independence of H. takedai, and the divergence time with closely related species was estimated to be from the Late Pliocene to the Early Pleistocene. In the phylogenetic tree, two clades were identified within H. takedai, and their haplotypes were found in samples collected from the west and east of the distribution range. These intraspecific divergences were strongly influenced by geohistorical subdivisions of the current major distribution areas in the Middle Pleistocene. One clade was further subdivided and its formation may have been influenced by sea level changes in the Late Pleistocene.

Bifenthrin induces changes in clinical poisoning symptoms, oxidative stress, DNA damage, histological characteristics, and transcriptome in Chinese giant salamander (Andrias davidianus) larvae.

Bifenthrin (BF) is a broad-spectrum insecticide that has gained widespread use due to its high effectiveness. However, there is limited research on the potential toxic effects of bifenthrin pollution on amphibians. This study aimed to investigate the 50 % lethal concentration (LC50) and safety concentration of Chinese giant salamanders (CGS) exposed to BF (at 0, 6.25,12.5,25 and 50 µg/L BF) for 96 h. Subsequently, CGS were exposed to BF (at 0, 0.04, and 4 µg/L BF) for one week to investigate its toxic effects. Clinical poisoning symptoms, liver pathology, oxidative stress factors, DNA damage, and transcriptome differences were observed and analyzed. The results indicate that exposure to BF at 4 µg/L significantly decreased the adenosine-triphosphate (ATP), superoxide dismutase (SOD), glutathione (GSH), and catalase (CAT) contents in the brain, liver, and kidney of CGS. Additionally, the study found that the malondialdehyde (MDA), reactive oxygen species (ROS), and 8-hydroxydeoxyguanosine (8-OHdG) contents were increased. The liver tissue exhibited significant inflammatory reactions and structural malformations. RNA-seq analysis of the liver showed that BF caused abnormal antioxidant indices of CGS. This affected molecular function genes such as catalytic activity, ATP-dependent activity, metabolic processes, signaling and immune system processes, behavior, and detoxification, which were significantly upregulated, resulting in the differential genes significantly enriched in the calcium signaling pathway, PPARα signaling pathway and NF-kB signaling pathway. The results suggest that BF induces the abnormal production of free radicals, which overwhelms the body's self-defense system, leading to varying degrees of oxidative stress. This can result in oxidative damage, DNA damage, abnormal lipid metabolism, autoimmune diseases, clinical poisoning symptoms, and tissue inflammation. This work provides a theoretical basis for the rational application of bifenthrin and environmental risk assessment, as well as scientific guidance for the conservation of amphibian populations.

Phylogenetic relationships and genetic differentiation of two Salamandrella species as revealed via COI gene from Northeastern China.

Due to traditional classification methods' limitations, some cryptic species remain undiscovered. To better explore the existence of the Schrenck salamander (Salamandrella tridactyla, a cryptic species of Siberian salamander S. keyserlingii) in China, we conducted a molecular phylogenetic analysis to confirm the taxonomic relationship among Salamandrella species and investigate genetic variation. We used complete sequences of the mitochondrial COI gene from 65 specimens collected across a wide range in Northeastern China. Thirty-five haplotypes were obtained from six populations. They showed medium-high haplotype diversity (Hd) and low nucleotide polymorphism (π). The phylogenetic tree and haplotype network analysis revealed that populations from Greater Khingan Ridge (Huma: HM) and Lesser Khingan Ridge (Tieli: TL) belong to S. keyserlingii, while populations from Changbai Mountain (Shangzhi-zhuziying: SZ, Shangzhi-cuijia: SC, Hailin: HL, and Baishan: BS) belong to S. tridactyla. This indicates the monophyly of Salamandrella and each of the two species. There was a substantial level of genetic differentiation between different species and within populations of the same species. This differentiation was significantly related to geographical distance. At last, the mismatch distribution and neutrality analyses indicated that the TL populations have undergone expansion of history. The study supplements the distributional range of Schrenck salamander. And it provides a theoretical basis for species conservation of Salamandrella species.

Discovery of ex situ individuals of Andrias sligoi, an extremely endangered species and one of the largest amphibians worldwide.

The South China giant salamander, Andrias sligoi, is one of the largest extant amphibian species worldwide. It was recently distinguished from another Chinese species, the Chinese giant salamander, Andrias davidianus, which is considered Critically Endangered according to the International Union for Conservation of Nature (IUCN) Red List. It appears too late to save this extremely rare and large amphibian in situ. Another extant species of the same genus, Andrias japonicus, inhabits Japan. However, the introduction of Chinese giant salamanders into some areas of Japan has resulted in hybridization between the Japanese and Chinese species. During our genetic screening of giant salamanders in Japan, we unexpectedly discovered four individuals of the South China giant salamander: two were adult males in captivity, and one had recently died. The last individual was a preserved specimen. In this study, we report these extremely rare individuals of A. sligoi in Japan and discuss the taxonomic and conservational implications of these introduced individuals.

Discerning structure versus speciation in phylogeographic analysis of Seepage Salamanders (Desmognathus aeneus) using demography, environment, geography, and phenotype.

Numerous mechanisms can drive speciation, including isolation by adaptation, distance, and environment. These forces can promote genetic and phenotypic differentiation of local populations, the formation of phylogeographic lineages, and ultimately, completed speciation. However, conceptually similar mechanisms may also result in stabilizing rather than diversifying selection, leading to lineage integration and the long-term persistence of population structure within genetically cohesive species. Processes that drive the formation and maintenance of geographic genetic diversity while facilitating high rates of migration and limiting phenotypic differentiation may thereby result in population genetic structure that is not accompanied by reproductive isolation. We suggest that this framework can be applied more broadly to address the classic dilemma of "structure" versus "species" when evaluating phylogeographic diversity, unifying population genetics, species delimitation, and the underlying study of speciation. We demonstrate one such instance in the Seepage Salamander (Desmognathus aeneus) from the southeastern United States. Recent studies estimated up to 6.3% mitochondrial divergence and four phylogenomic lineages with broad admixture across geographic hybrid zones, which could potentially represent distinct species supported by our species-delimitation analyses. However, while limited dispersal promotes substantial isolation by distance, microhabitat specificity appears to yield stabilizing selection on a single, uniform, ecologically mediated phenotype. As a result, climatic cycles promote recurrent contact between lineages and repeated instances of high migration through time. Subsequent hybridization is apparently not counteracted by adaptive differentiation limiting introgression, leaving a single unified species with deeply divergent phylogeographic lineages that nonetheless do not appear to represent incipient species.

Reassessment of species delimitation using nuclear markers in three lentic-breeding salamanders from the Chugoku District of Japan (Amphibia: Caudata: Hynobiidae).

Hynobius akiensis sensu lato has recently been split into three species based on short sequence analyses of cyt-b gene of mtDNA and without data of nuclear DNA, and strange sympatric distribution in some areas has been indicated in two species. We analyzed nuclear DNA marker (SNPs) and complete sequence of cyt-b in H. akiensis sensu lato to reassess species delimitation and genetic introgression among species. As a result, we found two lineages with discordant mitochondrial and nuclear DNA in some areas. Of H. akiensis sensu lato, each of the two contains the type locality of two species recently reported (H. sumidai and H. geiyoensis), and the use of these names has been previously advocated. However, their sympatric distribution was rejected based on nuclear DNA data, which we consider is more reliable than mtDNA. We thus clarify geographic boundary of these two species and revise the species delimitations.

Amended diagnosis, mitochondrial genome, and phylogenetic position of Sphyranura euryceae (Neodermata, Monogenea, Polystomatidae), a parasite of the Oklahoma salamander.

Polystomatidae is a monogenean family whose representatives infect mainly (semi)-aquatic tetrapods. Species of Sphyranura Wright, 1879 exhibit ectoparasitism on salamander hosts, with molecular work supporting their inclusion within Polystomatidae, at an early diverging, yet unresolved, position in the clade of otherwise endoparasitic polystomatid parasites of batrachian hosts. Records of representatives of Sphyranura are scarce with genetic data only available for S. oligorchis Alvey, 1933. Based on detailed morphological examination and comparison with type material, we identified worms belonging to Sphyranura infecting Oklahoma salamander (Eurycea tynerensis) as S. euryceae Hughes & Moore, 1943. Along with an amended diagnosis of Sphyranura, we provide the first molecular data for S. euryceae in the form of a mitochondrial genome and nuclear (18S, 28S rRNA) markers. Close morphological similarity between the two species of Sphyranura is reflected in low genetic divergence. Mitochondrial level comparison reveals instances of tRNA gene rearrangements in polystomatids. Although the phylogenetic reconstruction supports Sphyranura as early branching in the lineage of polystomatid monogeneans infecting batrachians, certain nodes remain unresolved.

Title: Diagnostic modifié, génome mitochondrial et position phylogénétique de Sphyranura euryceae (Neodermata, Monogenea, Polystomatidae), un parasite de la salamandre de l'Oklahoma. Abstract: Les Polystomatidae sont une famille de monogènes dont les représentants infectent principalement les tétrapodes (semi)-aquatiques. Les espèces de Sphyranura Wright, 1879 présentent un ectoparasitisme sur les hôtes salamandres, et des travaux moléculaires soutiennent leur inclusion dans les Polystomatidae, à une position divergente précoce mais non résolue dans le clade des Polystomatidae endoparasites d'hôtes batraciens. Les signalements des représentants de Sphyranura sont rares et les données génétiques ne sont disponibles que pour S. oligorchis Alvey, 1933. Sur la base d'un examen morphologique détaillé et d'une comparaison avec le matériel type, nous avons identifié les vers appartenant à Sphyranura infectant la salamandre de l'Oklahoma (Eurycea tynerensis) comme S. euryceae Hughes & Moore, 1943. Parallèlement à un diagnostic modifié de Sphyranura, nous fournissons les premières données moléculaires pour S. euryceae sous la forme d'un génome mitochondrial et de marqueurs nucléaires (ARNr 18S, 28S). La similitude morphologique étroite entre les deux espèces de Sphyranura se traduit par une faible divergence génétique. La comparaison au niveau mitochondrial révèle des cas de réarrangements des gènes des ARNt chez les Polystomatidae. Bien que la reconstruction phylogénétique soutienne Sphyranura comme un rameau précoce dans la lignée des monogènes Polystomatidae infectant les batraciens, certains nœuds restent non résolus.

Development and validation of an environmental DNA assay to detect federally threatened groundwater salamanders in central Texas.

The molecular detection of DNA fragments that are shed into the environment (eDNA) has become an increasingly applied tool used to inventory biological communities and to perform targeted species surveys. This method is particularly useful in habitats where it is difficult or not practical to visually detect or trap the target organisms. Central Texas Eurycea salamanders inhabit both surface and subterranean aquatic environments. Subterranean surveys are challenging or infeasible, and the detection of salamander eDNA in water samples is an appealing survey technique for these situations. Here, we develop and validate an eDNA assay using quantitative PCR for E. chisholmensis, E. naufragia, and E. tonkawae. These three species are federally threatened and constitute the Septentriomolge clade that occurs in the northern segment of the Edwards Aquifer. First, we validated the specificity of the assay in silico and with DNA extracted from tissue samples of both target Septentriomolge and non-target amphibians that overlap in distribution. Then, we evaluated the sensitivity of the assay in two controls, one with salamander-positive water and one at field sites known to be occupied by Septentriomolge. For the salamander-positive control, the estimated probability of eDNA occurrence (ψ) was 0.981 (SE = 0.019), and the estimated probability of detecting eDNA in a qPCR replicate (p) was 0.981 (SE = 0.011). For the field control, the estimated probability of eDNA occurring at a site (ψ) was 0.938 (95% CRI: 0.714-0.998). The estimated probability of collecting eDNA in a water sample (θ) was positively correlated with salamander relative density and ranged from 0.371 (95% CRI: 0.201-0.561) to 0.999 (95% CRI: 0.850- > 0.999) among sampled sites. Therefore, sites with low salamander density require more water samples for eDNA evaluation, and we determined that our site with the lowest estimated θ would require seven water samples for the cumulative collection probability to exceed 0.95. The estimated probability of detecting eDNA in a qPCR replicate (p) was 0.882 (95% CRI: 0.807-0.936), and our assay required two qPCR replicates for the cumulative detection probability to exceed 0.95. In complementary visual encounter surveys, the estimated probability of salamanders occurring at a known-occupied site was 0.905 (SE = 0.096), and the estimated probability of detecting salamanders in a visual encounter survey was 0.925 (SE = 0.052). We additionally discuss future research needed to refine this method and understand its limitations before practical application and incorporation into formal survey protocols for these taxa.

Molecular cloning, tissues distribution, and function analysis of thioredoxin-like protein-1 (TXNL1) in Chinese giant salamanders Andrias davidianus.

Thioredoxin-like protein-1 (TXNL1) is the member of thioredoxin superfamily, a family of thiol oxidoreductases. TXNL1 plays an important role in scavenging ROS and the maintenance of cellular redox balance. However, its physiological functions in Andrias davidianus have not been well understood. In the present study, the full-length cDNA encoding thioredoxin-like protein-1 (AdTXNL1) of A. davidianus was cloned, the mRNA tissue distribution was analyzed, and the function was characterized. The Adtxnl1 cDNA contained an open reading frame (ORF) of 870 bp encoding a polypeptide of 289 amino acids with the N-terminal TRX domain, a Cys34-Ala35-Pro36-Cys37 (CAPC) motif, and the C-terminal proteasome-interacting thioredoxin domain (PITH). The mRNA of AdTXNL1 was expressed in a wide range of tissues, with the highest level in the liver. The transcript level of AdTXNL1 was significantly up-regulated post Aeromonas hydrophila challenge in liver tissue. Moreover, the recombinant AdTXNL1 protein was produced and purified, and used to investigate the antioxidant activity. In the insulin disulfide reduction assay, rAdTXNL1 exhibited strong antioxidant capability. Altogether, the thioredoxin-like protein-1 may be involved in reduction/oxidation (redox) balance and as an important immunological gene in A. davidianus.

Encouraging news for in situ conservation: Translocation of salamander larvae has limited impacts on their skin microbiota.

The key role of symbiotic skin bacteria communities in amphibian resistance to emerging pathogens is well recognized, but factors leading to their dysbiosis are not fully understood. In particular, the potential effects of population translocations on the composition and diversity of hosts' skin microbiota have received little attention, although such transfers are widely carried out as a strategy for amphibian conservation. To characterize the potential reorganization of the microbiota over such a sudden environmental change, we conducted a common-garden experiment simulating reciprocal translocations of yellow-spotted salamander larvae across three lakes. We sequenced skin microbiota samples collected before and 15 days after the transfer. Using a database of antifungal isolates, we identified symbionts with known function against the pathogen Batrachochytrium dendrobatidis, a major driver of amphibian declines. Our results indicate an important reorganization of bacterial assemblages throughout ontogeny, with strong changes in composition, diversity and structure of the skin microbiota in both control and translocated individuals over the 15 days of monitoring. Unexpectedly, the diversity and community structure of the microbiota were not significantly affected by the translocation event, thus suggesting a strong resilience of skin bacterial communities to environmental change-at least across the time-window studied here. A few phylotypes were more abundant in the microbiota of translocated larvae, but no differences were found among pathogen-inhibiting symbionts. Taken together, our results support amphibian translocations as a promising strategy for this endangered animal class, with limited impact on their skin microbiota.

Chinese Giant Salamander Iridovirus 025L Is a Viral Essential Gene.

Ranavirus is a large nucleocytoplasmic DNA virus. Chinese giant salamander iridovirus (CGSIV) belongs to the ranavirus genus, and its replication involves a series of essential viral genes. Viral PCNA is a gene closely associated with viral replication. CGSIV-025L also encodes PCNA-like genes. We have described the function of CGSIV-025L in virus replication. The promoter of CGSIV-025L is activated during viral infection, and it is an early (E) gene that can be effectively transcribed after viral infection. CGSIV-025L overexpression promoted viral replication and viral DNA replication. siRNA interfered with CGSIV-025L expression and attenuated viral replication and viral DNA replication. The Δ025L-CGSIV strain with the deletion of CGSIV-025L could not replicate normally and could be rescued by the replenishment of 025L. CGSIV-025L was proven to be an essential gene for CGSIV by overexpression, interference, and deletion mutation experiments. CGSIV-025L was found to interact with CGSIV-062L by yeast two-hybrid, CoIP, and GST pulldown. Thus, the current study demonstrated that CGSIV-025L is an essential gene of CGSIV, which may be involved in viral infection by participating in viral DNA replication and interacting with replication-related proteins.

An efficient environmental DNA detection method for rare species: a case study of a small salamander (Hynobius boulengeri).

Loss of biodiversity is a serious concern, and amphibians are particularly threatened. Most small salamanders in Japan are endangered. Distributional information is fundamental to the conservation of these rare species; however, small salamanders are generally difficult to locate or catch. Environmental DNA analysis is an effective survey method for monitoring such rare species. The conventional polymerase chain reaction (PCR) method, which combines PCR amplification with subsequent electrophoresis, and the real-time PCR method, which uses fluorescent material, are commonly used for this purpose. In this study, a comparison of these two detection methods was conducted using a rare salamander species, Hynobius boulengeri, as a model case. We compared three points: (i) detection sensitivity, (ii) influence of environmental factors related to detection, and (iii) time and financial costs of the two methods. To perform this comparison, we developed a real-time PCR detection assay, conducted field surveys, and compared the time and financial costs of conventional and real-time PCR methods. The comparison showed no statistical difference in the detection sensitivity from field samples, and the effects of environmental factors tended to be similar. In addition, the financial cost was lower for the conventional PCR method while the time cost was lower for the real-time PCR method. Therefore, selecting eDNA detection methods based on objectives, time, and financial costs will promote efficient monitoring and contribute to the conservation of rare species.

A small noncoding RNA links ribosome recovery and translation control to dedifferentiation during salamander limb regeneration.

Building a blastema from the stump is a key step of salamander limb regeneration. Stump-derived cells temporarily suspend their identity as they contribute to the blastema by a process generally referred to as dedifferentiation. Here, we provide evidence for a mechanism that involves an active inhibition of protein synthesis during blastema formation and growth. Relieving this inhibition results in a higher number of cycling cells and enhances the pace of limb regeneration. By small RNA profiling and fate mapping of skeletal muscle progeny as a cellular model for dedifferentiation, we find that the downregulation of miR-10b-5p is critical for rebooting the translation machinery. miR-10b-5p targets ribosomal mRNAs, and its artificial upregulation causes decreased blastema cell proliferation, reduction in transcripts that encode ribosomal subunits, diminished nascent protein synthesis, and retardation of limb regeneration. Taken together, our data identify a link between miRNA regulation, ribosome biogenesis, and protein synthesis during newt limb regeneration.

Digest: Salamanders reveal novel trajectories of amphibian MHC evolution.

How does amphibian MHC diversity fit in the landscape of vertebrate evolution? Mimnias et al. (2022) address this gap in the field of MHC evolution by focusing on the lesser described MHC class I in salamanders. These findings contribute to understanding MHC diversity and the susceptibility of amphibians to pathogens, which could lead to future research on a major threat to amphibian biodiversity, chytrid fungi.

Whole-Mount In Situ Hybridization (WISH) for Salamander Embryos and Larvae.

Whole-mount in situ hybridization (WISH) is widely used to visualize transcribed gene sequences (mRNA) in developing embryos, larvae, and other nucleotide probe permeable tissue samples. This methodology involves the hybridization of an antisense nucleotide probe to the target mRNA, followed by chromogen or fluorescence-based detection. Here we describe a protocol for the spatiotemporal analysis of mRNA transcripts in axolotl embryos/larvae using digoxigenin-labeled riboprobes, anti-digoxigenin alkaline phosphatase, Fab fragments antibody, and NBT/BCIP chromogen detection.

Speciation Hypotheses from Phylogeographic Delimitation Yield an Integrative Taxonomy for Seal Salamanders (Desmognathus monticola).

Significant advances have been made in species delimitation and numerous methods can test precisely defined models of speciation, though the synthesis of phylogeography and taxonomy is still sometimes incomplete. Emerging consensus treats distinct genealogical clusters in genome-scale data as strong initial evidence of speciation in most cases, a hypothesis that must therefore be falsified under an explicit evolutionary model. We can now test speciation hypotheses linking trait differentiation to specific mechanisms of divergence with increasingly large data sets. Integrative taxonomy can, therefore, reflect an understanding of how each axis of variation relates to underlying speciation processes, with nomenclature for distinct evolutionary lineages. We illustrate this approach here with Seal Salamanders (Desmognathus monticola) and introduce a new unsupervised machine-learning approach for species delimitation. Plethodontid salamanders are renowned for their morphological conservatism despite extensive phylogeographic divergence. We discover 2 geographic genetic clusters, for which demographic and spatial models of ecology and gene flow provide robust support for ecogeographic speciation despite limited phenotypic divergence. These data are integrated under evolutionary mechanisms (e.g., spatially localized gene flow with reduced migration) and reflected in emergent properties expected under models of reinforcement (e.g., ethological isolation and selection against hybrids). Their genetic divergence is prima facie evidence for species-level distinctiveness, supported by speciation models and divergence along axes such as behavior, geography, and climate that suggest an ecological basis with subsequent reinforcement through prezygotic isolation. As data sets grow more comprehensive, species-delimitation models can be tested, rejected, or corroborated as explicit speciation hypotheses, providing for reciprocal illumination of evolutionary processes and integrative taxonomies. [Desmognathus; integrative taxonomy; machine learning; species delimitation.].

Morphological differentiation in giant salamanders, Andrias japonicus, A. davidianus, and their hybrids (Urodela, Cryptobranchidae), and its taxonomic implications.

For a long time, it has been debated whether the two giant salamanders, Andrias japonicus from Japan and A. davidianus from China, are conspecific or heterospecific. Morphological information about their diagnostic characteristics has been limited, without considering sexual dimorphism and/or body size variation. Recently, A. davidianus, which was introduced into Japan sometime in the past, has been found to hybridize with A. japonicus in situ. Taxonomic identification of individuals involved in this unusual breeding is made based on mitochondrial and nuclear DNA analyses. This identification method is time-consuming and costly. Thus, developing easier methods of identification, such as utilizing external morphological characteristics, is urgently needed. In this study, we verify previous descriptions showing that A. davidianus has a longer relative tail length than A. japonicus, and the tubercles on the lower jaw and throat were present in both sexes of A. davidianus. In addition, many head characteristics were found to be relatively larger in A. davidianus than in A. japonicus, which were new distinguishing characters. These morphological differences help support the idea that these are heterospecific lineages. In hybrids, relative values of head width and tail length were larger than those of A. japonicus, and the tubercles on their lower jaw and throat were present as in A. davidianus, suggesting that the hybrids and A. davidianus are distinguishable from A. japonicus.

The evolution of reproductive modes and life cycles in amphibians.

Amphibians have undergone important evolutionary transitions in reproductive modes and life-cycles. We compare large-scale macroevolutionary patterns in these transitions across the three major amphibian clades: frogs, salamanders, and caecilians. We analyse matching reproductive and phylogenetic data for 4025 species. We find that having aquatic larvae is ancestral for all three groups and is retained by many extant species (33-44%). The most frequent transitions in each group are to relatively uncommon states: live-bearing in caecilians, paedomorphosis in salamanders, and semi-terrestriality in frogs. All three groups show transitions to more terrestrial reproductive modes, but only in caecilians have these evolved sequentially from most-to-least aquatic. Diversification rates are largely independent of reproductive modes. However, in salamanders direct development accelerates diversification whereas paedomorphosis decreases it. Overall, we find a widespread retention of ancestral modes, decoupling of trait transition rates from patterns of species richness, and the general independence of reproductive modes and diversification.

Environmentally Associated Variation in Dispersal Distance Affects Inbreeding Risk in a Stream Salamander.

AbstractAvoiding inbreeding is considered a key driver of dispersal evolution, and dispersal distances should be especially important in mediating inbreeding risk because the likelihood of mating with relatives decreases with dispersal distance. However, a lack of direct data on dispersal distances has limited empirical tests of this prediction, particularly in the context of the multiple selective forces that can influence dispersal. Using the headwater stream salamander Gyrinophilus porphyriticus, we tested whether spatial variation in environmental conditions leads to differences in dispersal distances, resulting in spatial variation in the effect of dispersal on inbreeding risk. Using capture-recapture and population genomic data from five streams, we found that dispersal distances were greater in downstream reaches than upstream reaches. Inbreeding risk trended lower for dispersers than nondispersers in downstream reaches but not in upstream reaches. Furthermore, stream reaches did not differ in spatial patterns of individual relatedness, indicating that variation in inbreeding risk was in fact due to differences in dispersal distances. These results demonstrate that environmentally associated variation in dispersal distances can cause the inbreeding consequences of dispersal to vary at fine spatial scales. They also show that selective pressures other than inbreeding avoidance maintain phenotypic variation in dispersal, underscoring the importance of addressing alternative hypotheses in dispersal research.

A new, narrowly endemic species of swamp-dwelling dusky salamander (Plethodontidae: Desmognathus) from the Gulf Coastal Plain of Mississippi and Alabama.

We describe a new, narrowly endemic species of swamp-dwelling dusky salamander (Plethodontidae: Desmognathus pascagoula sp. nov.) from the Gulf Coastal Plain of southeastern Mississippi and southwestern Alabama based on linear morphometrics, mitochondrial DNA, and single nucleotide polymorphisms from 881 loci produced using genotype-by-sequencing. Some populations of the new species were historically referred to as D. auriculatus, a polyphyletic assemblage of at least three species in the Atlantic and Gulf Coastal Plain from Texas to North Carolina. Populations of D. auriculatus from the Gulf Coastal Plain in Louisiana and Mississippi were recently described as D. valentinei. The new species includes populations that were tentatively referred to D. valentinei, but we find it is morphologically, genetically, and geographically distinct. It is smaller, has a more defined dorsal color pattern, more irregular whitish portholes in up to three rows on the lateral surfaces of the body and tail, and a brighter orange or yellowish orange postocular stripe. At present, the new species is known from only six extant populations in the lower Pascagoula, Escatawpa, and Mobile drainages. The latter represents a distinct phylogeographic lineage. We also refer a historical collection from the northeastern side of the Mobile-Tensaw River Delta to this species, suggesting a much broader range in the past. We suspect that more populations remain to be discovered in the area, and their potential species-level distinctiveness should be tested further. This discovery increases knowledge of the biodiversity in the southeastern United States Coastal Plain, a candidate region meeting the global criteria for a biodiversity hotspot, and underscores the amount of cryptic diversity likely remaining to be discovered and described in Nearctic salamanders.