Anticancer potency of Egyptian venom snakes on MCF-7 and HepG2 carcinoma cells.

Breast and hepatic cancers are the leading incidences in the globe occurring of the human sufferings from various cancers. Snake venoms have been reported to provide effective therapeutic agents. The current study investigates the anticancer potency of Egyptian venoms snakes on two cells: breast cancer cells (MCF-7) and hepato-cancer cells (HepG2) (In vitro assay). The examined venoms were more potent on MCF-7 than HepG2 cells. Their inhibition % on MCF-7 ranged from 71.47 to 99.02% with medium inhibition concentrations (IC50s): 3.48, 3.60, 3.70, 4.33, and 4.49 µg/ml for venoms: Echis pyramid (E.H), Cerastes vipera (C.V), Naja haje (N.H), Echis coloratus (E.C), and Cerastes cerastes (C.C), respectively. The values of IC50s on HepG2 were 4.32, 17.77, 59.72, 63.75, and 217.90 µg/ml for toxins: E.C, E.P, C.V, C.C, and N.H, respectively. Some biomarkers were conducted to investigate the apoptotic effects of toxins into the cells. Increasing profiles of lactate dehydrogenase (LDH) activity and levels of glutathione content (GSH) and malodialdhyde (MDA) as well as repairment of DNA indicated such these actions. So, more reliable investigations on these venoms were needed to provide intelligent therapeutic agent for cancer treatment.

Evaluation of venom diversity and antivenom quality from the venom of long-term captive vs recently wild captured Pseudocerastes persicus snake: An In vitro and In vivo study.

Snakebite envenomation is a life-threatening condition and antivenoms are used as the most effective treatment. Venom obtained from snakes in long-term captivity showed some variations in comparison to the venom of the wild snakes. The objective of this study is to compare the venom of the Pseudocerastes persicus under long-term captivity and wild conditions as well as the antivenom obtained from these venoms. We have analyzed venom samples and produced trivalent antivenoms using the venom of long-term captive (LTC) or recently wild-captured (RWC) Pseudocerastes persicus, and RWC Macrovipera lebetina, and Echis carinatus. The HPLC analysis revealed that the RWC snakes' venom had three peaks that were not present in the LTC snake's venom. Further analysis using MALDI-TOF and MS/MS showed that the fraction with a retention time (RT) of 14 min contained a toxin from the Kunitz-type serine protease inhibitor (KUT) class, while the fraction with RT 21 a peptide identified within the snake venom metalloproteinase (SVMP) class. The third peak was identified as a sphingolipid. Interestingly, the in vivo preclinical tests showed no significant differences in the effectiveness of the antivenoms. which could be due to the cross-immunogenicity or cross-reactivity between different toxins in the venom. According to our results, small variations in the venom composition of a species do not lead to a decrease in the efficacy of the polyvalent antivenom.

On the role of TFEC in reptilian coloration.

Reptilian species, particularly snakes and lizards, are emerging models of animal coloration. Here, I focus on the role of the TFEC transcription factor in snake and lizard coloration based on a study on wild-type and piebald ball pythons. Genomic mapping previously identified a TFEC mutation linked to the piebald ball python phenotype. The association of TFEC with skin coloration was further supported by gene-editing experiments in the brown anole lizard. However, novel histological analyses presented here reveal discrepancies between the ball python and the anole TFEC mutants phenotype, cautioning against broad generalizations. Indeed, both wild-type and piebald ball pythons completely lack iridophores, whereas the TFEC anole lizard mutants lose their iridophores compared to the wild-type anole. Based on these findings, I discuss the potential role of the MiT/TFE family in skin pigmentation across vertebrate lineages and advocate the need for developmental analyses and additional gene-editing experiments to explore the reptilian coloration diversity.

The pancreas does not contribute to the non-adrenergic-non-cholinergic stimulation of heart rate in digesting pythons.

Vertebrates elevate heart rate when metabolism increases during digestion. Part of this tachycardia is due to a non-adrenergic-non-cholinergic (NANC) stimulation of the cardiac pacemaker, and it has been suggested these NANC factors are circulating hormones that are released from either gastrointestinal or endocrine glands. The NANC stimulation is particularly pronounced in species with large metabolic responses to digestion, such as reptiles. To investigate the possibility that the pancreas may release hormones that exert positive chronotropic effects on the digesting Burmese python heart, a species with very large postprandial changes in heart rate and oxygen uptake, we evaluate how pancreatectomy affects postprandial heart rate before and after autonomic blockade of the muscarinic and the beta-adrenergic receptors. We also measured the rates of oxygen consumption and evaluated the short-term control of the heart using the spectral analysis of heart rate variability and the baroreflex sequence method. Digestion caused the ubiquitous tachycardia, but the intrinsic heart rate (revealed after the combination of atropine and propranolol) was not affected by pancreatectomy and therefore hormones, such as glucagon and insulin, do not appear to contribute to the regulation of heart rate during digestion in Burmese pythons.

Distinct regulatory networks control toxin gene expression in elapid and viperid snakes.

BACKGROUND: Venom systems are ideal models to study genetic regulatory mechanisms that underpin evolutionary novelty. Snake venom glands are thought to share a common origin, but there are major distinctions between venom toxins from the medically significant snake families Elapidae and Viperidae, and toxin gene regulatory investigations in elapid snakes have been limited. Here, we used high-throughput RNA-sequencing to profile gene expression and microRNAs between active (milked) and resting (unmilked) venom glands in an elapid (Eastern Brown Snake, Pseudonaja textilis), in addition to comparative genomics, to identify cis- and trans-acting regulation of venom production in an elapid in comparison to viperids (Crotalus viridis and C. tigris). RESULTS: Although there is conservation in high-level mechanistic pathways regulating venom production (unfolded protein response, Notch signaling and cholesterol homeostasis), there are differences in the regulation of histone methylation enzymes, transcription factors, and microRNAs in venom glands from these two snake families. Histone methyltransferases and transcription factor (TF) specificity protein 1 (Sp1) were highly upregulated in the milked elapid venom gland in comparison to the viperids, whereas nuclear factor I (NFI) TFs were upregulated after viperid venom milking. Sp1 and NFI cis-regulatory elements were common to toxin gene promoter regions, but many unique elements were also present between elapid and viperid toxins. The presence of Sp1 binding sites across multiple elapid toxin gene promoter regions that have been experimentally determined to regulate expression, in addition to upregulation of Sp1 after venom milking, suggests this transcription factor is involved in elapid toxin expression. microRNA profiles were distinctive between milked and unmilked venom glands for both snake families, and microRNAs were predicted to target a diversity of toxin transcripts in the elapid P. textilis venom gland, but only snake venom metalloproteinase transcripts in the viperid C. viridis venom gland. These results suggest differences in toxin gene posttranscriptional regulation between the elapid P. textilis and viperid C. viridis. CONCLUSIONS: Our comparative transcriptomic and genomic analyses between toxin genes and isoforms in elapid and viperid snakes suggests independent toxin regulation between these two snake families, demonstrating multiple different regulatory mechanisms underpin a venomous phenotype.

An extremely rare serovar of Salmonella enterica (Yopougon) discovered in a Western Whip Snake (Hierophis viridiflavus) from Montecristo Island, Italy: case report and review.

Reptiles, including snakes, can be asymptomatically infected with multiple pathogen microorganisms, including Salmonella spp., which is considered an important concern for public and animal health. Small and uninhabited isles are quite ecologically different from mainland and represent interesting fields of study, to discover unexpected biological and microbiological aspects of their wild inhabitants. This work reports the presence of the very rare Salmonella enterica serovar Yopougon, isolated in a carcass of a native wild snake (Hierophis viridiflavus) from an Italian uninhabited island of Mediterranean Sea, Montecristo. To our knowledge, S. enterica serovar Yopougon was previously isolated only once 34 years earlier in Ivory Coast, from a human fecal sample. In the present study, we present the genomic characterization of the new isolate, the phylogenetic comparison with the previously isolated S. enterica serovar Yopougon strain of human origin and with other sequences available in public databases. In addition, an extensive review of available data in the literature and from our case history is provided. Our finding represents an example of the ability of some pathogens to travel for very long distances within their hosts and then to infect others, even from different taxa.

Comparative morphology of oral glands in snakes of the family Homalopsidae reveals substantial variation and additional independent origins of salt glands within Serpentes.

Using diffusible iodine-based contrast-enhanced computed tomography (diceCT), we examined the morphology of the oral glands of 12 species of the family Homalopsidae. Snakes of this family exhibit substantial interspecific morphological variation in their oral glands. Particular variables are the venom glands, ranging from large (e.g., Subsessor bocourti) to small (e.g., Erpeton tentaculatum). The supra- and infralabial glands are more uniform in morphology, being the second most developed in almost all the sampled species. Premaxillary glands distinct from the supralabial glands were observed in five species (Myron richardsonii, Bitia hydroides, Cantoria violacea, Fordonia leucobalia, and Gerarda prevostiana), in addition to Cerberus rynchops, the only species in which this condition was previously documented associated with the excretion of salt. In the three species of the saltwater group of homalopsids (C. violacea, F. leucobalia, and G. prevostiana), the premaxillary glands also extend posteriorly, occupying a large area above the supralabial gland, a condition not observed in any other species of snake studied thus far. Character evolution analyses indicate that premaxillary glands differentiated from the supralabial gland and evolved independently three or four times in the family, always in lineages that invaded marine habitats. Our results suggest that the differentiated premaxillary glands are likely salt glands, as is the case in C. rynchops. If corroborated, this increases to six or seven the number of independent evolutionary origins of salt glands in snakes that have undergone an evolutionary transition to marine life.

In-depth immunorecognition and neutralization analyses of Micrurus mipartitus and M. dumerilii venoms and toxins by a commercial antivenom.

In Colombia, the Micrurus genus comprises 30 species, including M. mipartitus and M. dumerilii, which are of major clinical relevance due to their wide geographical distribution and the number of snakebites inflicted by them. These neurotoxic envenomations are characterized by neuromuscular paralysis attributed to venom components such as three-finger toxins (3FTx) and phospholipases (PLA2). Additionally, there is limited information available on the neutralizing coverage of commercially available antivenoms, underscoring the need to perform studies to assess the cross-neutralizing ability of these life-saving products. Therefore, we present an in-depth immunorecognition analysis by the anticoral-INS antivenom from Colombia on the M. mipartitus and M. dumerilii venoms. The antivenom cross-recognized the whole venoms and their components with different intensities. For instance, the antivenom showed better recognition on PLA2s than on 3FTxs in both venoms. Moreover, at doses tested, the antivenom totally neutralized the lethal effect of M. dumerilii venom; however, it did not neutralize this effect induced by M. mipartitus venom and its main toxic components from the southwestern region of the department of Antioquia. Furthermore, the anticoral-INS antivenom displayed better cross-immunorecognition of PLA2-predominant Micrurus venoms than of 3FTx-predominant Micrurus venoms. This highlights the need to include venoms from both types of venom patterns in the immunization mixture to produce antivenoms against coral snakes. Finally, our results suggest the need for further research to optimize the composition of immunizing mixtures for antivenom production and improve their efficacy against coral snake envenomation in Colombia and the Americas.

Ecological factors and parity mode correlate with genome size variation in squamate reptiles.

BACKGROUND: Evidence of correlation between genome size, the nuclear haploid DNA content of a cell, environmental factors and life-history traits have been reported in many animal species. Genome size, however, spans over three orders of magnitude across taxa and such a correlation does not seem to follow a universal pattern. In squamate reptiles, the second most species-rich order of vertebrates, there are currently no studies investigating drivers of genome size variability. We run a series of phylogenetic generalized least-squares models on 227 species of squamates to test for possible relationships between genome size and ecological factors including latitudinal distribution, bioclimatic variables and microhabitat use. We also tested whether genome size variation can be associated with parity mode, a highly variable life history trait in this order of reptiles. RESULTS: The best-fitting model showed that the interaction between microhabitat use and parity mode mainly accounted for genome size variation. Larger genome sizes were found in live-bearing species that live in rock/sand ecosystems and in egg-laying arboreal taxa. On the other hand, smaller genomes were found in fossorial live-bearing species. CONCLUSIONS: Environmental factors and species parity mode appear to be among the main parameters explaining genome size variation in squamates. Our results suggest that genome size may favour adaptation of some species to certain environments or could otherwise result from the interaction between environmental factors and parity mode. Integration of genome size and genome sequencing data could help understand the role of differential genome content in the evolutionary process of genome size variation in squamates.

Morphological and molecular phylogenetic characterization of Sarcocystis kani sp. nov. and other novel, closely related Sarcocystis spp. infecting small mammals and colubrid snakes in Asia.

We investigated the morphology and phylogenetic relationships of novel and previously recognized Sarcocystis spp. infecting small mammals and colubrid snakes in Asia. The nuclear 18S rRNA and mitochondrial cox1 of Sarcocystis sp.1 from mangrove snakes (Boiga dendrophila) in Thailand and Sarcocystis sp.2 from a ricefield rat (Rattus argentiventer) in Sumatra were partially sequenced. Sporocysts of Sarcocystis sp.1 induced development of sarcocysts in experimentally infected rats, which showed a unique ultrastructure that was observed previously by S.P. Kan in rats from Malaysia; therefore, we describe this species as Sarcocystis kani sp. nov. Its integration into the 18S rRNA phylogeny of Sarcocystis spp. cycling between small mammals and colubrid snakes helped clarify relationships among the so-called S. zuoi-complex of molecularly cryptic species: Sarcocystis kani sp. nov., S. sp.2, S. attenuati, S. scandentiborneensis, and S. zuoi were all included in this clade. Tree topology was resolved into dichotomies congruent with the morphological disparities between the taxa. However, cox1 gene sequencing (including newly sequenced S. singaporensis and S. zamani) revealed that Sarcocystis kani, S. attenuati, and S. scandentiborneensis were identical suggesting a recent, common ancestry. To identify other distinctive features, lineage-specific molecular patterns within both genes were examined revealing that all 18S rRNA sequences of the S. zuoi - complex possess a unique, 7-nt long motif in helix 38 of domain V7 that was different in S. clethrionomyelaphis which branched off basally from the complex. Three-dimensional homology modelling of COX1 protein structure identified amino acid substitutions within the barcode area specific for the S. zuoi-complex and substantial divergence in structurally important amino acids between Sarcocystis species of snakes as definitive hosts and other lineages of the Sarcocystidae. We discuss the utility of selected genes for species delimitation of the Sarcocystis spp. under investigation, which probably evolved during recent radiations of their intermediate and definitive hosts.

Extinction debt and functional traits mediate community saturation over large spatiotemporal scales.

Determining if ecological communities are saturated (have a limit to the number of species they can support) has important implications for understanding community assembly, species invasions, and climate change. However, previous studies have generally been limited to short time frames that overlook extinction debt and have not explicitly considered how functional trait diversity may mediate patterns of community saturation. Here, we combine data from biodiversity surveys with functional and phylogenetic data to explore if the colonisation events after the Great American Biotic Interchange (closure of the Panamanian Isthmus) resulted in increases in species richness of communities of the snake family Dipsadidae. We determined the number and the direction of dispersal events between Central and South America by estimating ancestral areas based on a Bayesian time-calibrated phylogenetic analysis. We then evaluated whether variation in community saturation was mediated by the functional similarity of six traits for the resident and colonizing snakes and/or local environmental conditions. We found that colonised communities did not support more species than those that were not colonised. Moreover, we did not find an association between the functional diversity across sites and whether they were colonised by members from the lineages dispersing across the Isthmus or not. Instead, variation in species richness was predicted best by covariates such as time since colonisation and local environment. Taken together, our results suggest that snake communities of the Dipsadidae across the neotropics are saturated. Moreover, our research highlights two important factors to consider in studies of community saturation: extinction debt and the functional differences and similarities in species' ecological roles.

Determinar si las comunidades ecológicas están saturadas (si tienen un límite en el número de especies que pueden albergar) tiene importantes implicaciones para entender el ensamblaje de comunidades, las invasiones de especies y el cambio climático. Sin embargo, los estudios previos en esta área se han limitado generalmente a marcos temporales cortos, ignorando el concepto de deuda de extinción y no considerando explícitamente cómo la diversidad de rasgos funcionales puede mediar en los patrones de saturación de las comunidades. En este trabajo combinamos datos publicados de muestreos de campo con datos funcionales y filogenéticos para explorar si los eventos de colonización después del Gran Intercambio Biótico Americano (ocurrido con el cierre del istmo de Panamá) resultaron en aumentos en la riqueza de especies de las comunidades de la familia de serpientes Dipsadidae. Determinamos el número y la dirección de los eventos de dispersión entre América Central y América del Sur mediante la estimación de áreas ancestrales basada en un análisis filogenético Bayesiano calibrado en el tiempo. Luego evaluamos si la variación en la saturación de las comunidades estaba mediada por la similitud funcional de seis rasgos para las serpientes residentes y colonizadoras y/o por las condiciones ambientales locales. Encontramos que las comunidades colonizadas no contienen más especies que aquellas que no fueron colonizadas. Además, no encontramos ninguna relación entre la diversidad funcional de los sitios considerados y el hecho de que estuvieran colonizados o no por miembros de los linajes que se dispersaron a través del Istmo. En cambio, la variación en la riqueza de especies se predijo mejor por covariantes como el tiempo transcurrido desde la colonización y el clima local. En conjunto, nuestros resultados sugieren que las comunidades de Dipsadidae a lo largo del neotrópico están saturadas. Además, nuestra investigación destaca dos factores importantes a considerar en los estudios de saturación de comunidades: la existencia de una deuda de extinción y las diferencias y similitudes funcionales en los papeles ecológicos de las especies.

Malaysian and Chinese King Cobra Venom Cytotoxicity in Melanoma and Neonatal Foreskin Fibroblasts Is Mediated by Age and Geography.

Snake venoms constitute a complex, rapidly evolving trait, whose composition varies between and within populations depending on geographical location, age and preys (diets). These factors have determined the adaptive evolution for predatory success and link venom heterogeneity with prey specificity. Moreover, understanding the evolutionary drivers of animal venoms has streamlined the biodiscovery of venom-derived compounds as drug candidates in biomedicine and biotechnology. The king cobra (Ophiophagus hannah; Cantor, 1836) is distributed in diverse habitats, forming independent populations, which confer differing scale markings, including between hatchlings and adults. Furthermore, king cobra venoms possess unique cytotoxic properties that are used as a defensive trait, but their toxins may also have utility as promising anticancer-agent candidates. However, the impact of geographical distribution and age on these potential venom applications has been typically neglected. In this study, we hypothesised that ontogenetic venom variation accompanies the morphological distinction between hatchlings and adults. We used non-transformed neonatal foreskin (NFF) fibroblasts to examine and compare the variability of venom cytotoxicity between adult captive breeding pairs from Malaysian and Chinese lineages, along with that of their progeny upon hatching. In parallel, we assessed the anticancer potential of these venoms in human-melanoma-patient-derived cells (MM96L). We found that in a geographical distribution and gender-independent manner, venoms from hatchlings were significantly less cytotoxic than those from adults (NFF; ~Log EC50: 0.5-0.6 vs. 0.2-0.35 mg/mL). This is consistent with neonates occupying a semifossorial habitat, while adults inhabit more above-ground habitats and are therefore more conspicuous to potential predators. We also observed that Malaysian venoms exhibited a slightly higher cytotoxicity than those from the Chinese cobra cohorts (NFF; Log EC50: 0.1-0.3 vs. 0.3-0.4 mg/mL), which is consistent with Malaysian king cobras being more strongly aposematically marked. These variations are therefore suggestive of differential anti-predator strategies associated with the occupation of distinct niches. However, all cobra venoms were similarly cytotoxic in both melanoma cells and fibroblasts, limiting their potential medical applications in their native forms.

Comparisons of the Oral Microbiota from Seven Species of Wild Venomous Snakes in Taiwan Using the High-Throughput Amplicon Sequencing of the Full-Length 16S rRNA Gene.

A venomous snake's oral cavity may harbor pathogenic microorganisms that cause secondary infection at the wound site after being bitten. We collected oral samples from 37 individuals belonging to seven species of wild venomous snakes in Taiwan, including Naja atra (Na), Bungarus multicinctus (Bm), Protobothrops mucrosquamatus (Pm), Trimeresurus stejnegeri (Ts), Daboia siamensis (Ds), Deinagkistrodon acutus (Da), and alpine Trimeresurus gracilis (Tg). Bacterial species were identified using full-length 16S rRNA amplicon sequencing analysis, and this is the first study using this technique to investigate the oral microbiota of multiple Taiwanese snake species. Up to 1064 bacterial species were identified from the snake's oral cavities, with 24 pathogenic and 24 non-pathogenic species among the most abundant ones. The most abundant oral bacterial species detected in our study were different from those found in previous studies, which varied by snake species, collection sites, sampling tissues, culture dependence, and analysis methods. Multivariate analysis revealed that the oral bacterial species compositions in Na, Bm, and Pm each were significantly different from the other species, whereas those among Ts, Ds, Da, and Tg showed fewer differences. Herein, we reveal the microbial diversity in multiple species of wild snakes and provide potential therapeutic implications regarding empiric antibiotic selection for wildlife medicine and snakebite management.

Dynamic gap crossing in Dendrelaphis, the sister taxon of flying snakes.

Arboreal animals commonly use dynamic gap-crossing behaviors such as jumping. In snakes, however, most species studied to date only employ the quasi-static cantilever crawl, which involves a whole-body reach. One exception is the paradise tree snake (Chrysopelea paradisi), which exhibits kinematic changes as gap distance increases, culminating in dynamic behaviors that are kinematically indistinguishable from those used to launch glides. Because Chrysopelea uses dynamic behaviors when bridging gaps without gliding, we hypothesized that such dynamic behaviors evolved ancestrally to Chrysopelea. To test this predicted occurrence of dynamic behaviors in closely related taxa, we studied gap bridging locomotion in the genus Dendrelaphis, which is the sister lineage of Chysopelea. We recorded 20 snakes from two species (D. punctulatus and D. calligastra) crossing gaps of increasing size, and analyzed their 3D kinematics. We found that, like C. paradisi, both species of Dendrelaphis modulate their use of dynamic behaviors in response to gap distance, but Dendrelaphis exhibit greater inter-individual variation. Although all three species displayed the use of looped movements, the highly stereotyped J-loop movement of Chrysopelea was not observed in Dendrelaphis. These results support the hypothesis that Chrysopelea may have co-opted and refined an ancestral behavior for crossing gaps for the novel function of launching a glide. Overall, these data demonstrate the importance of gap distance in governing behavior and kinematics during arboreal gap crossing.

Comparing morphological and secretory aspects of cephalic glands among the New World coral snakes brings novel insights on their biological roles.

Oral and other cephalic glands have been surveyed by several studies with distinct purposes. Despite the wide diversity and medical relevance of the New World coral snakes, studies focusing on understanding the biological roles of the glands within this group are still scarce. Specifically, the venom glands of some coral snakes were previously investigated but all other cephalic glands remain uncharacterized. In this sense, performing morphological and molecular analysis of these glands may help better understand their biological role. Here, we studied the morphology of the venom, infralabial, rictal, and harderian glands of thirteen species of Micrurus and Micruroides euryxanthus. We also performed a molecular characterization of these glands from selected species of Micrurus using transcriptomic and proteomic approaches. We described substantial morphological variation in the cephalic glands of New World coral snakes and structural evidence for protein-secreting cells in the inferior rictal glands. Our molecular analysis revealed that the venom glands, as expected, are majorly devoted to toxin production, however, the infralabial and inferior rictal glands also expressed some toxin genes at low to medium levels, despite the marked morphological differences. On the other hand, the harderian glands were dominated by the expression of lipocalins, but do not produce toxins. Our integrative analysis, including the prediction of biological processes and pathways, helped decipher some important traits of cephalic glands and better understand their biology.

Diagnosis of endoparasite species and subtypes of Cryptosporidium spp. with one health importance, in feces from captive snakes in Rio de Janeiro, Brazil.

In captivity, snakes may present chronic infections with high mortality, such as those caused by Cryptosporidium serpentis, or they may be pseudoparasitized by species that present zoonotic potential. Thus, the aim of this study was to evaluate the frequency of helminths and protozoa in the feces of captive snakes, characterize the species and subtypes of Cryptosporidium spp. and correlate the parasites detected with other information obtained from these animals. Feces were collected from 189 snakes kept at the Vital Brazil Institute, Rio de Janeiro, including samples from Bothrops jararaca, Bothrops jararacussu, Bothrops moojeni, Bothrops atrox, Bothrops leucurus, Crotalus durissus and Lachesis muta. All the samples were subjected to microscopy techniques and the polymerase chain reaction (PCR) in association with sequencing, to identify Cryptosporidium spp.. Forms of parasites infecting the snakes were identified through microscopy in 50.8% of the samples. Helminths were detected more often than protozoa in the feces of these animals, mainly comprising eggs resembling Kalicephalus sp. and oocysts of Eimeria sp.. Pseudoparasites such as Syphacia sp., Aspiculuris sp. and Hymenolepis nana were also detected. Through correlating the results obtained from parasitological staining techniques and PCR, the total frequency of Cryptosporidium sp. was found to be 19%. The species C. tyzzeri and C. parvum were identified. Characterization using the target gp60 showed subtypes with high potential for zoonotic transmission, especially IIaA15G2R1 and IIaA14G2R1 of C. parvum and IXbA8 of C. tyzzeri. This study highlighted the need for more intensive health management in the Institute's serpentarium and, especially, in its bioterium where rodents are reared as a food source for these snakes.

First report of envenomation symptoms caused by the Eastern Montpellier snake, Malpolon insignitus (St.-Hilaire, 1827).

Snakebite is a global health problem with disastrous consequences. Accordingly, venoms plus their function and clinical aspects were intensively studied for several snakes, primarily with proteroglyphous and solenoglyphous dentition. Much less is known about the venoms from most opisthoglyphous (rear-fanged) snakes and it is often questionable which symptoms they may cause. Here, we provide the first description of the envenomation symptoms caused by a bite of the Eastern Montpellier snake Malpolon insignitus from Croatia. The clinical manifestations elicited by the bite were only local, and included bleeding, pulsatility and paresthesia, followed by sweating and signs of inflammation. The observed features subsided quickly, and the patient recovered fully with no long-term illness. Therefore, we conclude that M. insignitus does not represent a substantial (i.e. life threatening or morbidity-causing) threat. However, as related species in the Malpolon genus are known to be capable to deliver neurotoxic envenoming, we recommend to nevertheless carefully interact with M. insignitus and recommend that future studies should unveil its venom composition.

Molecular phylogeny reveals cryptic diversity in Sibynophis from China (Serpentes: Sibynophiidae).

The elucidation of species diversity and distribution is critical within the fields of evolution, genetics, and conservation. The genus Sibynophis contains rare snakes that have historically received little attention. In this study, we conducted comprehensive sampling and use both mitochondrial and nuclear genetic markers to explore Sibynophis species diversity within China. Our findings revealed that S. c. miyiensis should be considered synonymous with S. c. grahami, and S. c. grahami should be gave a specific rank as S. grahami. In addition, we discovered S. triangularis was new to China and Myanmar. Based on the specimens and molecular phylogeny results, we redefined the species distribution boundaries of each Chinese species.

Speciation rates are unrelated to the formation of population structure in Malagasy gemsnakes.

Speciation rates vary substantially across the tree of life. These rates should be linked to the rate at which population structure forms if a continuum between micro and macroevolutionary patterns exists. Previous studies examining the link between speciation rates and the degree of population formation in clades have been shown to be either correlated or uncorrelated depending on the group, but no study has yet examined the relationship between speciation rates and population structure in a young group that is constrained spatially to a single-island system. We examine this correlation in 109 gemsnakes (Pseudoxyrhophiidae) endemic to Madagascar and originating in the early Miocene, which helps control for extinction variation across time and space. We find no relationship between rates of speciation and the formation rates of population structure over space in 33 species of gemsnakes. Rates of speciation show low variation, yet population structure varies widely across species, indicating that speciation rates and population structure are disconnected. We suspect this is largely due to the persistence of some lineages not susceptible to extinction. Importantly, we discuss how delimiting populations versus species may contribute to problems understanding the continuum between shallow and deep evolutionary processes.

Genomic evidence sheds new light on phylogeny of Rhabdophis nuchalis (sensu lato) complex (Serpentes: Natricidae).

Stable taxonomy and robust phylogeny are essential for the evolution and conservation of organisms. The Rhabdophis nuchalis (sensu lato) complex presently contains three species (R. nuchalis, R. chiwen, R. pentasupralabialis). Although several studies have explored the diversity and phylogeography of this group, certain issues related to systematics and taxonomy remain unresolved. Here, based on genome-wide data, including single nucleotide polymorphisms (SNPs) generated from ddRAD-seq and mitochondrial DNA (mtDNA), we re-evaluated the phylogenetic relationships and cryptic diversity of this species group. Our results are generally consistent with previous studies but provide some new insights. Phylogenetic relationship reconstruction based on SNPs and mtDNA revealed that three species in the R. nuchalis (sensu lato) complex did not form a monophyly but each species is well supported as monophyletic lineage in SNP-based analyses. Population structure analyses showed genetic admixture between several species pairs. Additionally, the population in eastern Yunnan, China, was identified as a potential cryptic species and thus described as a new species based on morphological data. From our results and previous studies, we redefined the distribution boundary for each species in the R. nuchalis (sensu lato) species complex.