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Background Snake venoms are trophic adaptations that represent an ideal model to examine the evolutionary factors that shape polymorphic traits under strong natural selection. Venom compositional variation is substantial within and among venomous snake species. However, the forces shaping this phenotypic complexity, as well as the potential integrated roles of biotic and abiotic factors, have received little attention. Here, we investigate geographic variation in venom composition in a wide-ranging rattlesnake ( Crotalus viridis viridis ) and contextualize this variation by investigating dietary, phylogenetic, and environmental variables that covary with venom. Results Using shotgun proteomics, venom biochemical profiling, and lethality assays, we identify 2 distinct divergent phenotypes that characterize major axes of venom variation in this species: a myotoxin-rich phenotype and a snake venom metalloprotease (SVMP)-rich phenotype. We find that dietary availability and temperature-related abiotic factors are correlated with geographic trends in venom composition. Conclusions Our findings highlight the potential for snake venoms to vary extensively within species, for this variation to be driven by biotic and abiotic factors, and for the importance of integrating biotic and abiotic variation for understanding complex trait evolution. Links between venom variation and variation in biotic and abiotic factors indicate that venom variation likely results from substantial geographic variation in selection regimes that determine the efficacy of venom phenotypes across populations and snake species. Our results highlight the cascading influence of abiotic factors on biotic factors that ultimately shape venom phenotype, providing evidence for a central role of local selection as a key driver of venom variation.

Hybridization is increasingly understood as common throughout and beyond the speciation process, rather than an anomaly. Sympatric taxa are expected to exhibit strong reproductive isolation, and although hybridization may occur, it often results in inviable offspring. We investigated hybridization among three ranid frogs (Rana areolata, R. palustris, and R. sphenocephala) in eastern Oklahoma, where their distributions and breeding phenology overlap. Using micro‐CT scans of cranial morphology, genomic SNP data, and phenological records, we confirmed two putative hybrids collected in the field—remarkable given the phylogenetic distance between these non‐sister taxa. Genomic data show split ancestry from parental populations, representing one R. areolata × R. palustris and one R. areolata × R. sphenocephala. Cranial morphology indicates hybrids exhibit intermediate phenotypes, and our assessment identified a third likely hybrid, a specimen collected two decades earlier from the same area. Both confirmed hybrids were R. areolata backcrosses, but minimal introgression throughout the dataset suggests hybrid fitness may be lower than that of parental populations. Hybridization appears facilitated by overlapping breeding strategies and ecological factors leading to misdirected amplexus. This study provides the first documentation of natural hybridization in R. areolata, a species of conservation concern throughout its range. Our findings emphasize the utility of high‐resolution morphological data (micro‐CT) in complementing genomic approaches for hybrid diagnosis and suggest cranial morphology may be an effective method for hybrid identification in similar systems. Understanding this atypical three‐species hybridization has important implications for conservation, as hybrid fitness and introgression can influence population dynamics and genetic integrity. We investigated natural hybridization among three rapid frogs in eastern Oklahoma. Using genomic SNP data and cranial morphometrics, we confirmed the presence of hybrids exhibiting intermediate traits. Despite overlapping breeding phenologies, minimal introgression suggests hybrid fitness may be considerably lower than that of parental populations.

Background: Although antivenom is the standard treatment for snakebite envenoming, its efficacy may be impacted by geographic variation in venom composition, emphasizing the need for region-specific antivenom development. Methods: We report a case of snakebite envenoming, in which the patient was bitten on the hand by a captive Malayan pit viper (Calloselasma rhodostoma) with typical clinical manifestations following. Antivenom (produced in Thailand) was administered at 33 and 39 h post-bite. Venom from the causative individual snake was collected for compositional analysis via SDS-PAGE. Enzymatic activity of the venom was evaluated through the degradation of casein and phospholipid substrates, along with the assessment of enzymatic inhibition by two regionally specific antivenoms produced in Vietnam (AV. Cr. VN.) and Thailand (AV. Cr. TL.). Results: The patient showed good recovery, with complete normalization by day 7. SDS-PAGE profiling of the venom revealed five major enzymes, with SVSP, SVMP and PLA2 being the most abundant (16.7%, 40.11% and 26.11%, respectively). Antivenom inhibition tests revealed remaining casein percentages of 67.43% (AV. Cr. VN) and 59.35% (AV. Cr. TL). Blood agar assays indicated that phospholipase activity was reduced to 21.01% by AV. Cr. VN. and 23.30% by AV. Cr. TL. Conclusions: Our results show that the Vietnamese antivenom generated greater inhibitory activity against proteinases compared to the Thai product, underscoring the importance of using regionally specific antivenoms that are more effective against the venom profiles of locality-matched snake populations.