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Despite decades of molecular research, phylogenetic relationships in Palearctic vipers (genus Vipera) still essentially rely on a few loci, such as mitochondrial barcoding genes. Here we examined the diversity and evolution of Vipera with ddRAD-seq data from 33 representative species and subspecies. Phylogenomic analyses of ∼ 1.1 Mb recovered nine major clades corresponding to known species/species complexes which are generally consistent with the mitochondrial phylogeny, albeit with a few deep discrepancies that highlight past hybridization events. The most spectacular case is the Italian-endemic V. walser, which is grouped with the alpine genetic diversity of V. berus in the nuclear tree despite carrying a divergent mitogenome related to the Caucasian V. kaznakovi complex. Clustering analyses of SNPs suggest potential admixture between diverged Iberian taxa (V. aspis zinnikeri and V. seoanei), and confirm that the Anatolian V. pontica corresponds to occasional hybrids between V. (ammodytes) meridionalis and V. kaznakovi. Finally, all analyzed lineages of the V. berus complex (including V. walser and V. barani) form vast areas of admixture and may be delimited as subspecies. Our study sets grounds for future taxonomic and phylogeographic surveys on Palearctic vipers, a group of prime interest for toxinological, ecological, biogeographic and conservation research., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Asian mock vipers of the genus Psammodynastes and African forest snakes of the genus Buhoma are two genera belonging to the snake superfamily Elapoidea. The phylogenetic placements of Psammodynastes and Buhoma within Elapoidea has been extremely unstable which has resulted in their uncertain and debated taxonomy. We used ultraconserved elements and traditional nuclear and mitochondrial markers to infer the phylogenetic relationships of these two genera with other elapoids. Psammodynastes, for which a reference genome has been sequenced, were found, with strong branch support, to be a relatively early diverging split within Elapoidea that is sister to a clade consisting of Elapidae, Micrelapidae and Lamprophiidae. Hence, we allocate Psammodynastes to its own family, Psammodynastidae new family. However, the phylogenetic position of Buhoma could not be resolved with a high degree of confidence. Attempts to identify the possible sources of conflict in the rapid radiation of elapoid snakes suggest that both hybridisation/introgression during the rapid diversification, including possible ghost introgression, as well as incomplete lineage sorting likely have had a confounding role. The usual practice of combining mitochondrial loci with nuclear genomic data appears to mislead phylogeny reconstructions in rapid radiation scenarios, especially in the absence of genome scale data., (© 2024. The Author(s).)

This short essay pretends to make the reader reflect on the concept of biological mass and on the added value that the determination of this molecular property of a protein brings to the interpretation of evolutionary and translational snake venomics research. Starting from the premise that the amino acid sequence is the most distinctive primary molecular characteristics of any protein, the thesis underlying the first part of this essay is that the isotopic distribution of a protein's molecular mass serves to unambiguously differentiate it from any other of an organism's proteome. In the second part of the essay, we discuss examples of collaborative projects among our laboratories, where mass profiling of snake venom PLA2 across conspecific populations played a key role revealing dispersal routes that determined the current phylogeographic pattern of the species., (© 2021 The Author(s).)

Snakebites in Europe are mostly due to bites from Viperidae species of the genus Vipera. This represents a neglected public health hazard with poorly defined incidence, morbidity and mortality. In Europe, fourteen species of "true vipers" (subfamily Viperinae) are present, eleven of which belong to the genus Vipera. Amongst these, the main medically relevant species due to their greater diffusion across Europe and the highest number of registered snakebites are six, namely: Vipera ammodytes, V. aspis, V. berus, V. latastei, V. seoanei and V. ursinii. Generally speaking, viper venom composition is characterised by many different toxin families, like phospholipases A2, snake venom serine proteases, snake venom metalloproteases, cysteine-rich secretory proteins, C-type lectins, disintegrins, haemorrhagic factors and coagulation inhibitors. A suspected snakebite is often associated with severe pain, erythema, oedema and, subsequently, the onset of an ecchymotic area around one or two visible fang marks. In the field, the affected limb should be immobilised and mildly compressed with a bandage, which can then be removed once the patient is being treated in hospital. The clinician should advise the patient to remain calm to reduce blood circulation and, therefore, decrease the spread of the toxins. In the case of pain, an analgesic therapy can be administered, the affected area can be treated with hydrogen peroxide or clean water. However, anti-inflammatory drugs and disinfection with alcohol or alcoholic substances should be avoided. For each patient, clinical chemistry and ECG are always a pre-requisite as well as the evaluation of the tetanus immunisation status and for which immunisation may be provided if needed. The treatment of any clinical complication, due to the envenomation, does not differ from treatments of emergency nature. Antivenom is recommended when signs of systemic envenomation exist or in case of advanced local or systemic progressive symptoms. Recommendations for future work concludes. The aim of this review is to support clinicians for the clinical management of viper envenomation, through taxonomic keys for main species identification, description of venom composition and mode of action of known toxins and provide a standardised clinical protocol and antivenom administration., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Historical data can clarify ecological attributes of fauna in sites that have subsequently been altered by anthropogenic activities. We used the 1960s notebooks of pioneering East African naturalist C.J.P. Ionides to extract quantitative information on captures of 484 snakes of five species (rhinoceros vipers Bitis nasicornis, black mambas Dendroaspis polylepis, Jameson's mambas D. jamesoni, water cobras Naja annulata, and eastern forest cobras N. subfulva). High capture rates suggest high abundances of all species. The relative numbers of each species collected changed over the years and differed seasonally, reflecting targeting by Ionides. Sex ratios and age-class distributions differed among species and were affected by factors such as month of collection and time of day. Habitat use was affected by species, sex and body size: for example, arboreality became less common with increasing body size in the rhinoceros viper and black mamba, and males were found in arboreal sites more often than were females. In both D. jamesoni and D. polylepis, adult males and females were recorded together in September-October, suggesting reproductive activity at this time of year. Although fragmentary, the data from Ionides' notebooks provide a unique glimpse into ecological patterns of snakes within an African landscape half a century ago.

Due to its various structures in bio-compounds, snake venom is the indisputable result of evolutionary stages of molecules with an increasingly complex structure, high specificity, and of great importance for medicine because of their potential. The present study proposed an underpinning examination of venom composition from nine species of venomous snakes using a useful and replicable methodology. The objective was the extension of the evaluation of protein fractions in the field up to 230 kDa to permit possible identification of some fractions that are insufficiently studied. The gel capillary electrophoresis method on the chip was performed using an Agilent 2100 bioassay with the 80 and 230-LabChip Protein kits. Interpretation of electrophoresis was performed using the Protein 2100 expert (Agilent) test software as follows: a) Protein 80 (peak size scale): 1.60, 3.5, 6.50, 15.00, 28.00, 46.00, 63.00, 95.00 kDa; b) Protein 230 (peak size scale): 4.50, 7.00, 15.00, 28.00, 46.00, 63.00, 95.00, 150.00, 240.00 kDa. The screening revealed the presence of compounds with a molecular weight greater than 80 kDa, in the case of Vipera aspis and Vipera xantina palestinae. For V. aspis, a 125 kDa molecular weight pro-coagulant protein was identified, known as being involved in the reduction of plasma clotting time without any direct activity in the fibrinogen coagulation process. The samples examined on the Protein 230-LabChip electrophoresis chip can be considered as a novelty with possible uses in medicine, requiring further approaches by advanced proteomics techniques to confirm the intimate structural features and biological properties of snake venoms.

Atractaspidines are poorly studied, fossorial snakes that are found throughout Africa and western Asia, including the Middle East. We employed concatenated gene-tree analyses and divergence dating approaches to investigate evolutionary relationships and biogeographic patterns of atractaspidines with a multi-locus data set consisting of three mitochondrial (16S, cyt b, and ND4) and two nuclear genes (c-mos and RAG1). We sampled 91 individuals from both atractaspidine genera (Atractaspis and Homoroselaps). Additionally, we used ancestral-state reconstructions to investigate fang and diet evolution within Atractaspidinae and its sister lineage (Aparallactinae). Our results indicated that current classification of atractaspidines underestimates diversity within the group. Diversification occurred predominantly between the Miocene and Pliocene. Ancestral-state reconstructions suggest that snake dentition in these taxa might be highly plastic within relatively short periods of time to facilitate adaptations to dynamic foraging and life-history strategies., Competing Interests: MFB is affiliated with Flora Fauna & Man, Ecological Services Ltd. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Scaling evolutionary trees to time is essential for understanding the origins of clades. Recently developed methods allow including the entire fossil record known for the group of interest and eliminated the need for specifying prior distributions for node ages. Here we apply the fossilized birth-death (FBD) approach to reconstruct the diversification timeline of the viperines (subfamily Viperinae). Viperinae are an Old World snake subfamily comprising 102 species from 13 genera. The fossil record of vipers is fairly rich and well assignable to clades due to the unique vertebral and fang morphology. We use an unprecedented sampling of 83 modern species and 13 genetic markers in combination with 197 fossils representing 28 extinct taxa to reconstruct a time-calibrated phylogeny of the Viperinae. Our results suggest a late Eocene-early Oligocene origin with several diversification events following soon after the group's establishment. The age estimates inferred with the FBD model correspond to those from previous studies that were based on node dating but FBD provides notably narrower credible intervals around the node ages. Viperines comprise two African and an Eurasian clade, but the ancestral origin of the subfamily is ambiguous. The most parsimonious scenarios require two transoceanic dispersals over the Tethys Sea during the Oligocene.

Understanding the influence of geographical events and climate changes on genetic diversity is essential in explaining current patterns of genetic structure and geographic distribution of organisms. We inferred phylogenetic relationships, investigated historical demography, explored the evolutionary history, and clarified intraspecific taxonomy of Protobothrops mucrosquamatus, which is one of the commonest and most wide-ranging Asian pitvipers. A total of 184 samples from 54 localities were sequenced and analyzed for two mitochondrial gene fragments and two nuclear genes. Phylogenetic reconstruction based on mtDNA sequences revealed the existence of a minimum of five geographically structured and well-supported lineages within P. mucrosquamatus. Based on the mtDNA gene tree, and the geographic relationship between populations allied by matrilineal lineages, a complex longitudinal and latitudinal diversification pattern was uncovered in P. mucrosquamatus. The estimated date of the origin of the species (about 5.3 Ma) and divergence of the intraspecific lineages match the rapid uplifting of Qinghai-Xizang Plateau, and is also consistent with those of some other co-distributed Asian pitvipers. Formation of the two island lineages (Taiwan and Hainan) was generally congruent with the first isolation of the islands, but the two lineages showed different relationships with the continental Asian populations in comparison with some other pitvipers. Population historical demographic analyses, based on three methods, showed that all lineages have experienced slight population expansion in and around the Dali Glacial. Tests of intraspecific taxonomy indicated that no cryptic taxon is present within this widely distributed snake., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Thrombotic microangiopathy (TMA), which includes the spectrum of hemolytic uremic syndrome and thrombotic thrombocytopenic purpura, is an uncommon complication of hump-nosed pit viper envenomation. We describe 4 cases of TMA following hump-nosed pit viper (Hypnale spp) bites in Sri Lanka. The first case is a typical TMA that spontaneously resolved with supportive treatments. The second and third cases are related to hemolytic uremic syndrome complicated with acute kidney injury that required hemodialysis. The fourth case is thrombotic thrombocytopenic purpura associated with acute kidney injury that required hemodialysis and therapeutic plasma exchange. For each patient we describe the circumstances of the bite, clinical features, laboratory findings, and management., (Copyright © 2018 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.)

This case report documents envenomation by an arboreal white-lipped green pit viper (Trimeresurus albolabris), a species found in South and Southeast Asia that causes the majority of venomous snakebites among Southeast Asian pit vipers. Clinical features vary from asymptomatic to serious coagulopathy that may progress into life-threatening or fatal hemorrhage. The proven life-threatening cases described in published literature, however, are sparse. Practically, no specific antivenom targeted to pit viper bites is available in Nepal. We report a case (managed with several non-evidence-based interventions) of noticeable coagulopathic envenomation due to confirmed T albolabris bite in Nepal. This is the first known reported case of such a bite in Nepal. This case highlights the urgent need to improve diagnosis, monitoring, and supportive care for bite victims and to study the effectiveness of Thai pit viper antivenoms for the treatment of T albolabris envenomations., (Copyright © 2018 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.)

Snake venom serves as a tool of defense against threat and helps in prey digestion. It consists of a mixture of enzymes, such as phospholipase A2, metalloproteases, and l-amino acid oxidase, and toxins, including neurotoxins and cytotoxins. Beside their toxicity, venom components possess many pharmacological effects and have been used to design drugs and as biomarkers of diseases. Viperidae is one family of venomous snakes that is found nearly worldwide. However, three main vipers exist in the Middle Eastern region: Montivipera bornmuelleri , Macrovipera lebetina , and Vipera (Daboia) palaestinae . The venoms of these vipers have been the subject of many studies and are considered as a promising source of bioactive molecules. In this review, we present an overview of these three vipers, with a special focus on their venom composition as well as their biological activities, and we discuss further frameworks for the exploration of each venom.

A proteomic and transcriptomic comparative analysis of the venoms of three Atheris species (A. squamigera, A. nitschei and A. chlorechis) was carried out by size exclusion liquid chromatography, gel electrophoresis, mass spectrometry, and mRNA sequencing. The improved proteomic profiling utilised in this work was combined with transcript studies, advancing our insights into venom composition, protein distribution and inter-species variation among the three bush vipers. Crude venoms of all three samples contained at least 10-20 protein components, ranging in size from ≤ 3 to > 98 kDa. Both approaches yielded converging overall information, pointing to phospholipases, disintegrins, serine proteases and metalloproteases as the major toxin classes, which are likely to explain the local and systemic symptoms observed in envenomation by Atheris genus. Being considered as the main factors involved in the distinct venom-induced pathologies, these identified snake venom proteins are of particular interest in terms of understanding their physiological and biological function as well as for their contribution in potential medical treatments.

Medically relevant cases of snakebite in Europe are predominately caused by European vipers of the genus Vipera. Systemic envenoming by European vipers can cause severe pathology in humans and different clinical manifestations are associated with different members of this genus. The most representative vipers in Europe are V. aspis and V. berus and neurological symptoms have been reported in humans envenomed by the former but not by the latter species. In this study we determined the toxicological profile of V. aspis and V. berus venoms in vivo in mice and we tested the effectiveness of two antivenoms, commonly used as antidotes, in counteracting the specific activities of the two venoms. We found that V. aspis, but not V. berus, is neurotoxic and that this effect is due to the degeneration of peripheral nerve terminals at the NMJ and is not neutralized by the two tested antisera. Differently, V. berus causes a haemorrhagic effect, which is efficiently contrasted by the same antivenoms. These results indicate that the effectiveness of different antisera is strongly influenced by the variable composition of the venoms and reinforce the arguments supporting the use polyvalent antivenoms.

Northern and western mountains of Iran are among the most important biodiversity and endemism hot spots for reptiles in the Middle East. Among herpetofauna, the montivipers represent an emblematic and fragmented endemic group for which estimating their level of genetic differentiation and defining conservation priorities is urgently needed. Here, we present the most comprehensive phylogenetic study on the Montivipera raddei species group comprising all 5 known taxa, among which 3 are endemic to Iran. Based on 2 mitochondrial genes, phylogenetic and phylogeographic analyses revealed 3 major lineages each presenting very contrasting distribution areas. The Iranian montivipers are highly structured in clades showing low genetic diversity and corresponding to high altitude summits. Molecular dating revealed the role of Quaternary paleo-climatic oscillations and altitudinal movements of montivipers in shaping genetic diversity and differentiation of these sky-island taxa. In addition, the best scenario of historical biogeography allowed identifying 3 possible refugial areas in Iran most likely arising by vicariance. Based on our mitochondrial results and pending additional data, we recognize 3 candidate species among the M. raddei complex: M. raddei, Montivipera latifii, and Montivipera kuhrangica that are coherent with their geographical distribution. We propose that the most appropriate evolutionary significant units for conservation of the montivipers are represented by 13 units among which 6 are recognized as high priority. Finally, we suggest some recommendations to the IUCN as well as to the Iranian conservation policies with respect to conservation prioritization.

Snakes of the cosmopolitan family Viperidae comprise around 329 venomous species showing a striking heterogeneity in species richness among lineages. While the subfamily Azemiopinae comprises only two species, 70% of all viper species are arranged in the subfamily Crotalinae or the "pit vipers". The radiation of the pit vipers was marked by the evolution of the heat-sensing pits, which has been suggested to be a key innovation for the successful diversification of the group. Additionally, only crotalines were able to successfully colonize the New World. Here, we present the most complete molecular phylogeny for the family to date that comprises sequences from nuclear and mitochondrial genes representing 79% of all living vipers. We also investigated the time of divergence between lineages, using six fossils to calibrate the tree, and explored the hypothesis that crotalines have undergone an explosive radiation. Our phylogenetic analyses retrieved high support values for the monophyly of the family Viperidae, subfamilies Viperinae and Crotalinae, and 22 out of 27 genera, as well as well-supported intergeneric relationships throughout the family. We were able to recover a strongly supported sister clade to the New World pit vipers that comprises Gloydius, Ovophis, Protobothrops and Trimeresurus gracilis. Our results agree in many aspects with other studies focusing on the phylogenetics of vipers, but we recover new relationships as well. Despite the addition of new sequences we were not able to resolve some of the poor supported relationships previously suggested. Time of divergence estimates suggested that vipers started to radiate around the late Paleocene to middle Eocene with subfamilies most likely dating back to the Eocene. The invasion of the New World might have taken place sometime close to the Oligocene/Miocene boundary. Diversification analyses suggested a shift in speciation rates during the radiation of a sub-clade of pit vipers where speciation rates rapidly increased but slowed down toward the present. Thus, the evolution of the loreal pits alone does not seem to explain their explosive speciation rates. We suggest that climatic and geological changes in Asia and the invasion of the New World may have also contributed to the speciation shift found in vipers., (Copyright © 2016 Elsevier Inc. All rights reserved.)

In the present study, we recognized progressively high immunological cross-reactivity between Pseudocerastes persicus fieldi (Pf) venom and six other medically important Egyptian snake venoms belonging to families Viperidae and Elapidae. Antibodies with a range of bonding strengths were shown to be involved in such cross-reactivity. Two strategies have been tried to access specificity; (i) using affinity purified species-specific anti-Pf antivenom antibodies, (ii) conducting the assay in the presence of ammonium thiocyanate (NH4SCN). The discrimination power of the prepared species-specific antivenom was demonstrated by its ability to detect Pf venom over a range of Pf concentrations (2.5 ng-2.5 μg) in a variety of body fluids. The assay could distinguish circulating Pf antigens from other viper antigens in the whole blood of experimentally envenomed mice. What seems promising in our work is the use of the chaotrope, NH4SCN, which renders the reaction medium more favorable for the specific homologous antigen-antibody interactions, primarily via preventing lower avid antibodies to share and, to a bit lesser extent, by decreasing non-specific absorbance signals frequently encountered with ELISA assays. The ELISA described herein may be useful for clinicians for identification of snake bites inflicted by Pf snake species. Balancing between specificity and sensitivity has to be considered for best results., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The Near and Middle East is a hotspot of biodiversity, but the region remains underexplored at the level of genetic biodiversity. Here, we present an extensive molecular phylogeny of the viperid snake genus Montivipera, including all known taxa. Based on nuclear and mitochondrial data, we present novel insights into the phylogeny of the genus and review the status of its constituent species. Maximum likelihood methods revealed a montane origin of Montivipera at 12.3Mya. We then analyzed factors of mountain viper diversity. Our data support substantial changes in effective population size through Plio-Pleistocene periods. We conclude that climatic oscillations were drivers of allopatric speciation, and that mountain systems of the Near and Middle East have strongly influenced the evolution and survival of taxa, because climatic and topographical heterogeneities induced by mountains have played a crucial role as filters for dispersal and as multiple refugia. The wide diversity of montane microhabitats enabled mountain vipers to retain their ecological niche during climatic pessima. In consequence the varied geological and topographical conditions between refugia favoured genetic isolation and created patterns of species richness resulting in the formation of neoendemic taxa. Our data support high concordance between geographic distributions of Montivipera haplotypes with putative plant refugia., (Copyright © 2016. Published by Elsevier Inc.)

Middle America is one of the most biodiverse regions in the world, harboring an exceptional number of rare and endemic species. This is especially true of Middle American cloud forests, where montane specialists occupy restricted, high-elevation ranges making them attractive candidates for investigating historical biogeography and speciation. One such highland-restricted species, the black speckled palm-pitviper (Bothriechis nigroviridis), occupies the Central, Tilarán, and Talamanca Cordilleras in Costa Rica and Panama. In this study, we investigate the genetic and morphological variation among populations of B. nigroviridis by inferring a multilocus phylogeny (21 individuals) and analyzing meristic scale characters with a principal component analysis (64 individuals). We find B. nigroviridis sensu stricto to be composed of two deeply divergent lineages, one with a restricted range in the northern and central Cordillera Talamanca and the other ranging throughout the Central, Tilarán, and Talamanca Cordilleras. Furthermore, these two lineages are morphologically distinct, with previously unrecognized differences in several characters allowing us to name and diagnose a new species B. nubestris sp. nov. We also examine the genetic and morphological variation within B. nigroviridis and discuss biogeographic hypotheses that may have led to the diversification of Bothriechis lineages.

The molecular events underlying the evolution of the Snake Venom Metalloproteinase (SVMP) family from an A Disintegrin And Metalloproteinase (ADAM) ancestor remain poorly understood. Comparative genomics may provide decisive information to reconstruct the evolutionary history of this multi-locus toxin family. Here, we report the genomic organization of Echis ocellatus genes encoding SVMPs from the PII and PI classes. Comparisons between them and between these genes and the genomic structures of Anolis carolinensis ADAM28 and E. ocellatus PIII-SVMP EOC00089 suggest that insertions and deletions of intronic regions played key roles along the evolutionary pathway that shaped the current diversity within the multi-locus SVMP gene family. In particular, our data suggest that emergence of EOC00028-like PI-SVMP from an ancestral PII(e/d)-type SVMP involved splicing site mutations that abolished both the 3' splice AG acceptor site of intron 12* and the 5' splice GT donor site of intron 13*, and resulted in the intronization of exon 13* and the consequent destruction of the structural integrity of the PII-SVMP characteristic disintegrin domain.

Sarafotoxins (SRTX) are endothelin-like peptides extracted from the venom of snakes belonging to the Atractaspididae family. A recent in vivo study on anesthetized and ventilated animals showed that sarafotoxin-b (SRTX-b), extracted from the venom of Atractaspis engaddensis, decreases cardiac output by inducing left ventricular dysfunction while sarafotoxin-m (SRTX-m), extracted from the venom of Atractaspis microlepidota microlepidota, induces right ventricular dysfunction with increased airway pressure. The aim of the present experimental study was to compare the respiratory effects of SRTX-m and SRTX-b. Male Wistar rats were anesthetized, tracheotomized and mechanically ventilated. They received either a 1 LD50 IV bolus of SRTX-b (n = 5) or 1 LD50 of SRTX-m (n = 5). The low-frequency forced oscillation technique was used to measure respiratory impedance. Airway resistance (Raw), parenchymal damping (G) and elastance (H) were determined from impedance data, before and 5 min after SRTX injection. SRTX-m and SRTX-b injections induced acute hypoxia and metabolic acidosis with an increased anion gap. Both toxins markedly increased Raw, G and H, but with a much greater effect of SRTX-b on H, which may have been due to pulmonary edema in addition to bronchoconstriction. Therefore, despite their structural analogy, these two toxins exert different effects on respiratory function. These results emphasize the role of the C-terminal extension in the in vivo effect of these toxins.

The mitochondrial genome sequence of Ussuri mamushi is analyzed and presented publically for the first time. The genome is 17,208 bp in length and contains 13 protein-coding genes, 2 ribosomal RNA genes, 22 transfer RNA genes and 2 control regions. The overall base composition is A (32.4%), C (28.4%), T (26.3%) and G (12.9%). The base compositions clearly presented the A-C skew, which is the most obvious in the protein-coding genes. Mitochondrial genome analyses based on MP, ML, NJ and Bayesian analyses yielded identical phylogenetic trees, indicating a close phylogenetic affinity of the 13 Crotalinae species. It appeared that no less than two major phyletic lineages were present in Crotalinae. The main clades within the Crotalinae include Protobothrops. A clade (G. brevicaudus, G. ussuriensis, G. intermedius, and G. sahatilis) with the Ovophis as the sister taxon to Protobothrops was supported by bootstrap values of 88%. The four Gloydius species formed a paraphyletic group with the high bootstrap value (100%) in all examinations.

The Azemiops snakes are pit-less and phylogenetically located at the Crotalinae and Viperinae divergence. cDNAs encoding five Azemiops venom phospholipase (sPLA2) molecules were cloned and sequenced; their signal-peptides were similar to those of crotalid sPLA2s. Based on their calculated pI-values and residue-49 substitutions, they were designated as Af-E6, Af-N49a, Af-N49a1, Af-N49a2, and Af-N49b, respectively. The first three isoforms, comprising 3-4% of the venom proteins, were purified by reversed-phase HPLC. Af-E6 is catalytically active and has >80% sequence-similarity to other Glu(6)-PLA2 (a pitviper venom-marker). Results of phylogenetic analyses reveal that acidic Af-N49a and Af-N49a1 are rather unique and loosely linked with crotalid PLA2s, while Af-N49b is related to the viperid PLA2s with Ser(1) substitution. Notably, the Asn(49)-substitutions in these molecules imply catalytic-independent mechanisms. The 3D-models of Af-E6 and Af-N49a have surface electropotential maps similar to each other and to those of antiplatelet PLA2s, while the Af-N49b model is similar to basic and myotoxic sPLA2 molecules. From Azemiops feae and four other Viperidae, we cloned five novel Cys-rich secretory proteins (CRISPs). Azemiops CRISP and natriuretic-peptide precursors share more sequence similarities with those of crotalid venoms than with viperid venoms, further supporting the theory that Azemiops are sister taxons to pit vipers, especially Tropedolaemus., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Background: Studying patterns of introgression can illuminate the role of hybridization in speciation, and help guide decisions relevant to the conservation of rare taxa. Vipera magnifica and Vipera orlovi are small vipers that have high conservation status due to their rarity and restricted distributions in an area of the Caucasus region where two other related species are present - V. kaznakovi and V. renardi. Despite numerous observations of hybridization between different species of small vipers, and the potential of a hybrid origin for V. magnifica and V. orlovi based on their distribution with respect to V. kaznakovi and V. renardi, hypotheses of a hybrid origin have not been formally tested. Here we generate genomic-scale data by performing next generation sequencing of double digest restriction-site associated DNA libraries, and use these multilocus data to test whether these two species are of hybrid origin., Results: We generated over nine hundred loci for 38 specimens of six taxa, and analysed the dataset using Bayesian clustering and multivariate methods, as well as Patterson D-statistics, which can distinguish between incomplete lineage sorting and introgression as explanations for shared polymorphism. The results demonstrate a pattern of historical admixture in the two purported hybrids that is consistent with past gene flow from V. renardi into V. kaznakovi. The average admixture proportion in individuals was low (6.39 %) in the case of V. magnifica, but was higher in V. orlovi (19.02 %). We also show that the specific individual samples used in D-statistic tests can have a significant impact on inferences regarding the magnitude of introgression, suggesting the importance of including multiple individuals in these analyses., Conclusions: Our results support the conclusion that both V. orlovi and V. magnifica had formed through a hybridization event between V. kaznakovi and V. renardi. Given a low proportion of admixture and absence of clear ecological and morphological differences V. magnifica should be treated as a marginal population of V. kaznakovi. Further studies that include analyses of ecological segregation of V. orlovi from parental taxa and search for evolutionary consequences of hybridisation would clarify if V. orlovi is a distinct hybrid species. Until this we recommend preserving the current taxonomy and protection status of V. orlovi.

Despite the public health significance and potential applications to medical research, the evolutionary history of the Asian Lance-headed pitvipers (genus Protobothrops) remains inadequately studied. Previous research generally focused on a few selected species with no comprehensive molecular study of Protobothrops. Here, we conduct the first study to infer the phylogenetic relationships of all currently recognized species in this genus based on four mitochondrial DNA fragments and four nuclear genes. Bayesian Inference and Maximum Likelihood analyses show that within Protobothrops there are four strongly supported clades forming distinct subgroups. The first subgroup, which is sister to the other three, consists of three species, P. himalayanus, P. kaulbacki, and P. sieversorum. The second subgroup contains only P. mangshanensis. The final two subgroups, which are sister groups, include the other four and six Protobothrops species. Although our findings provide additional information on the phylogenetic relationships of the genus Protobothrops, the placement of P. dabieshanensis and P. elegans remains problematic. In addition, our molecular results indicate that P. trungkhanhensis, currently considered endemic to Vietnam, should be added to the species known from China. Our ancestral area estimation indicated that Protobothrops likely originated in southwestern China. This study improves our understanding of the evolutionary relationships among species of Asian Lance-headed pitvipers, providing a greater framework for future studies.

A new species of viperine viperid snake is described, Bitis harenna sp. nov. The new species is a member of the subgenus Macrocerastes based on it having three scales separating the nasal and rostral shields, and on the combination of 'divisions' of dorsal scale rows on the upper flanks and 'fusions' of rows on the lower flanks. Bitis harenna sp. nov. is distinguished from other members of the subgenus by its unique colour pattern, posterior parietal flange on the lateral wall of the braincase, and possibly by differences in scalation and head proportions. Only a single museum specimen is known, a female collected from 'Dodola' in Ethiopia probably in the late 1960s and previously identified as a possibly unusually coloured and patterned B. parviocula. A live, presumably male, specimen very closely resembling the holotype of Bitis harenna sp. nov. was photographed on the Harenna escarpment of the Bale Mountains National Park, Ethiopia in 2013, providing secure occurrence data and evidence that the holotype is not a uniquely aberrant specimen. A revised key to the species of Bitis in Ethiopia is presented. Aspects of body scalation are compared among species of the subgenus Macrocerastes and between species of Macrocerastes and Bitis, and several systematic characters are highlighted and clarified.

Crude venom from three venomous snakes, Crotalus adamanteus, Crotalus vegrandis and Bitis nasicornis was fractionated by gel filtration chromatography, and selected fractions screened for in-vitro insulinotropic activity using clonal pancreatic BRIN-BD11 cells. Nineteen fractions stimulated insulin secretion and the structural identity of bioactive compounds responsible was probed using MALDI-ToF MS and N-terminal Edman degradation sequencing. Partial N-terminal sequences were determined and their homology to existing sequences identified using BLAST searching. The main insulinotropic peptide families identified were made up of snake venom serine proteinases, phospholipases A2 (PLA2) and disintegrins. Snake venom constituents may have therapeutic potential for diabetes, with each of the three viper venoms tested providing insulinotropic compounds from a range of different toxin families., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

phylin is a package for the r programming environment which offers different methods to spatially interpolate genetic information from phylogeographic data. These interpolations can be used to predict the spatial occurrence of different lineages within a phylogeny using a modified method of kriging, which allows the usage of a genetic distance matrix to derive a model of spatial dependence. phylin improves the available methods to generate interpolated surfaces from a phylogenetic trees by assessing the autocorrelation structure of the genetic information, interpolating the genetic data based on a statistical model, estimating the uncertainty of the predictions and identifying lineage occurrence and contact zones probability without projection of pairwise genetic distances into mid-points between sample locations. The package also includes methods to plot interpolation surfaces and provide summary tables from the generated data and models. We provide an example of the usefulness of this tool by inferring the spatial occurrence of distinct historical evolutionary lineages of the Lataste's viper (Vipera latastei Boscá, 1878) in the Iberian Peninsula and identifying potential contact areas. The maps of phylogenetic patterns obtained with these methods provide a spatial context to test hypotheses related to processes underlying the geographic distribution of genetic diversity and to inform conservation planning., (© 2014 John Wiley & Sons Ltd.)

The phylogeny and historical demography of small Eurasian vipers of the Vipera ursinii and V. renardi complexes were studied using mitochondrial DNA sequences analysed with Bayesian inference, Maximum Likelihood and Maximum Parsimony approaches, and mismatch distributions. Diversification in the group resulted from an initial dispersion in the later Pliocene - Pleistocene in two directions: north-westwards via the Balkans (V. ursinii complex) and north-eastwards from Asia Minor via the Caucasus (V. renardi complex). An independent, comparatively recent transition occurred from montane habitats to lowland grasslands in different mitochondrial lineages during the Late Pleistocene, when representatives of the both complexes had reached lowland steppes to the north. Effective population size showed clear signs of rapid growth in eastern V. renardi, triggered by colonization of vast lowland steppes, but in western V. ursinii complex grew during the Last Glaciation and experienced stabilization in Holocene. Expansion and population growth in lowland lineages of V. renardi was not strongly affected by Pleistocene climatic oscillations, when cold, dry conditions could have favoured species living in open grasslands. The high diversity of closely related haplotypes in the Caucasus and Tien-Shan could have resulted from repetitive expansion-constriction-isolation events in montane regions during Pleistocene climate fluctuations. The mitochondrial phylogeny pattern conflicts with the current taxonomy., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Past climatic and tectonic events are believed to have strongly influenced species diversity in the Eastern Afromontane Biodiversity Hotspot. We investigated the phylogenetic relationships and historical biogeography of the East African genus Atheris (Serpentes: Viperidae), and explored temporal and spatial relationships between Atheris species across Africa, and the impact of palaeoclimatic fluctuations and tectonic movements on cladogenesis of the genus. Using mitochondrial sequence data, the phylogeny of East African species of Atheris shows congruent temporal patterns that link diversification to major tectonic and aridification events within East Africa over the last 15million years (my). Our results are consistent with a scenario of a delayed direct west-east colonisation of the Eastern Arc Mountains of Atheris by the formation of the western rift. Based on the phylogenetic patterns, this terrestrial, forest-associated genus has dispersed into East Africa across a divided route, on both west-southeasterly and west-northeasterly directions (a C-shaped route). Cladogenesis in the Eastern Arc Mountains and Southern Highlands of Tanzania corresponds to late Miocene and Plio-Pleistocene climatic shifts. Taxonomically, our data confirmed the monophyly of Atheris as currently defined, and reveal four major East African clades, three of which occur in discrete mountain ranges. Possible cryptic taxa are identified in the Atheris rungweensis and A. ceratophora clades., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The facial pits of rattlesnakes, copperheads, lanceheads, bushmasters and other American and Asian pitvipers (Crotalinae) are highly innervated and densely vascularized infrared (IR) receptor organs. For over a century, studies have focused on a small sample of model species from North America and Asia. Based on an expanded survey of Central and South American crotalines, we report a conspicuous accessory structure composed of well-defined papillae that project from the anterior orbital adnexa. The papillae are continuous with the inner chamber of the IR receptor organ and our histological and ultrastructural data suggest that they possess a well-developed nervous network and extensive vascularization; however, they lack the characteristic IR-sensitive terminal nerve masses found in the IR-receptive pit membrane. The function of the IR receptor organ papillae is unknown.

Protobothrops mucrosquamatus is the endangered snake that is distributed in southwestern China and other adjacent countries of Asia. We determined the complete mitochondrial genome of P. mucrosquamatus. The circle genome with the total length of 17,233 bp contained 13 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 2 control regions. Overall base composition of the complete mtDNA was 32.98% A, 25.17% T, 29.19% C and 12.67% G. All the genes in P. mucrosquamatus were distributed on the H-strand, except for the ND6 subunit gene and eight tRNA genes that were encoded on the L-strand.

Fractionation by reversed-phase HPLC of venom from four species of saw-scaled viper: Echis ocellatus, Echis pyramidum leakeyi, Echis carinatus sochureki, and Echis coloratus led to identification in each sample of an abundant protein with cytotoxic activity against human non-small cell lung adenocarcinoma A549 cells. The active component in each case was identified by MALDI-TOF mass fingerprinting of tryptic digests as [Ser⁴⁹]phospholipase A₂ ([Ser⁴⁹]PLA₂). An isoform of [Ser⁴⁹]PLA₂ containing the single Ala¹⁸→ Val substitution and a partially characterized [Asp⁴⁹]PLA₂ were also present in the E. coloratus venom. LC₅₀ values against A549 cells for the purified [Ser⁴⁹]PLA₂ proteins from the four species are in the range 2.9-8.5 μM. This range is not significantly different from the range of LC₅₀ values against human umbilical vein endothelial HUVEC cells (2.5-12.2 μM) indicating that the [Ser⁴⁹]PLA₂ proteins show no differential anti-tumor activity. The LC₅₀ value for [Ser⁴⁹]PLA₂ from E. ocellatus against human erythrocytes is >100 μM and the MIC values against Escherichia coli and Staphylococcus aureus are >100 μM. It is suggested that the [Ser⁴⁹]PLA₂ proteins play a major role in producing local tissue necrosis and hemorrhage at the site of envenomation., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Snake venom constitutes one of the most complex mixtures of naturally-occurring toxic proteins/polypeptides and a large number of these possess very profound biological activities. Disintegrins, that are commonly found in viper venoms, are low molecular weight proteins that usually contain an -Arg-Gly-Asp- (-RGD-) motif that is known to be involved in cell adhesion ligand recognition, binding specifically to cell surface integrin receptors and also exhibiting platelet anti-aggregation activity. Here, we report for the first time, the successful cloning of three cDNAs encoding disintegrin precursors from lyophilised venom-derived libraries of Atheris chlorechis, Atheris nitschei and Atheris squamigera, respectively. All of these disintegrins belong to the short-coding class and all exhibit high degrees of structural identity, both in their amino acid sequences and in the arrangement of their functional domains. Mass spectrometric analyses of the HPLC-separated/in-gel digested venom proteins was performed to characterise the mature disintegrins as expressed in the venom proteome. Studies on both the structures and conserved sites within these disintegrins are of considerable theoretical interest in the field of biological evolution and in the development of new research tools or novel templates for drug design., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Hump-nosed pit vipers of Genus Hypnale are the commonest cause of snake bite in Sri Lanka. Although there are many reports of local effects, coagulopathy and acute kidney injury, it remains unclear how frequent these clinical effects are and therefore the medical importance of this snake genus. The genus has been recently revised to include Hypnale hypnale from Sri Lanka and Western Ghats of Southern India, and the two endemic species to Sri Lanka, Hypnale zara and Hypnale nepa. This was a prospective hospital-based clinical study of definite Hypnale spp. bites from July 2008 to July 2010 in six Sri Lankan hospitals. There were 114 patients included and all snakes were correctly identified by hospital staff as Hypnale spp. Of these, 93 snakes were identified as H. hypnale by an expert, 16 as H. zara and five as H. nepa. Most bites occurred on the lower limbs in the daytime. There was no difference in the clinical effects between the three species. Pain and fang marks were present in all patients, 101 had local swelling and only 16 (14%) developed extensive local swelling that spread proximally and involved more than half of the bitten limb. Systemic symptoms occurred in 18 patients; four patients had an abnormal 20 min whole blood clotting test and one patient developed an acute kidney injury that required haemodialysis. All patients were discharged alive with a median length of stay of 2 days. This study confirms that hump-nosed viper bites cause only minor effects in most cases. Future studies need to undertake formal coagulation studies and identify important early indicators of renal impairment., (Crown Copyright © 2012. Published by Elsevier Ltd. All rights reserved.)

Background: Envenoming by south Asian hump-nosed pit vipers (Genus: Hypnale) is a significant health issue in Sri Lanka and in peninsular India. Bites by these snakes frequently lead to local envenoming, coagulopathy and acute renal failure even resulting in death. Recently the genus was revised and the existence of three species viz H. hypnale, H. nepa and H. zara were recognized. There is, however, a paucity of information on the toxicity of the venoms of these species. Hence, we compared the toxic effects of the three Hypnale venoms using BALB/c mice., Findings: Intraperitoneal median lethal doses (LD50) for H. hypnale, H. zara and H. nepa venoms were 1.6, 6.0 and 9.5 μg protein/g respectively. Minimum haemorrhagic doses for venoms of H. hypnale, H. zara and H. nepa were 3.4, 11.0 and 16.6 μg protein/mouse respectively. The minimum necrotic doses for the same venoms were 15.0, 55.1 and 68.2 μg protein/mouse respectively. Severe congestion and petecheal haemorrhages were observed in lungs, kidneys, liver and the alimentary tract. Histopathogical examination of kidneys revealed proximal tubular cell injury and acute tubular necrosis with intact basement membrane indicating possible direct nephrotoxicity. Hypnale venoms caused pulmonary oedema, hepatocellular degeneration and necrosis, focal neuronal degeneration in brain and extramedullary haemopoiesis in spleen. H. hypnale venom caused all above histopathological alterations at lower doses compared to the other two., Conclusion: Hypnale venoms cause similar pathological changes with marked differences in the severity of the toxic effects in vivo. Therefore, differences in the severity of the clinical manifestations could possibly be seen among bite victims of the three Hypnale species.

The genus Bothriechis comprises a lineage of nine species of Neotropical pitvipers distributed mainly in highlands across Middle America, all adapted to arboreal habitats. Bothriechis supraciliaris is a relatively recently described species that inhabits the Pacific southwest of Costa Rica, whose venom had never been studied. A proteomic and toxicological profiling of its venom is here reported. Proteins or peptides that belong to eleven families were found, with a predominance of bradykinin-potentiating peptides (21.9%), followed by serine proteinases (15.2%) and phospholipases A(2) (13.4%). A group of short polyglycine peptides, resembling the poly-His/poly-Gly metalloproteinase inhibitors described in Atheris and Echis snake venoms, was observed for the first time in a Bothriechis venom. Comparison of the venom proteome of B. supraciliaris with those of Bothriechis schlegelii, Bothriechis lateralis, and Bothriechis nigroviridis, confirms the highly diverse toxicological strategies evolved by these arboreal snakes in each case, as possible alternative solutions to the same trophic purpose. Toxicological profiling of B. supraciliaris venom revealed a potent hemorrhagic action, moderate myotoxicity, and very weak procoagulant activity. Importantly from the medical perspective, the lethal activity of its venom (mouse intraperitoneal LD(50): 7.1 μg/g) was efficiently neutralized by a polyvalent (Viperidae) antivenom of therapeutic use in Central America., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

The last two populations of the Hungarian meadow viper Vipera ursinii rakosiensis were thought to persist in the steppe fragments of Hungary until meadow vipers were discovered in central Romania (Transylvania), suggesting a possible existence of remnant populations elsewhere. We assessed the phylogenetic position of the Transylvanian vipers using 2030 bp of mitochondrial DNA sequence. We showed that they were closely related to the Hungarian vipers, while those from northeastern Romania (Moldavia) and Danube Delta belonged to the subspecies Vipera ursinii moldavica. Montane subspecies from Europe (Vipera ursinii ursinii and Vipera ursinii macrops) formed a sister clade to the two lowland subspecies. Vipera renardi formed a sister clade to V. ursinii, with populations from the Greater Caucasus (Vipera renardi lotievi) and Tien Shan (Vipera renardi tienshanica) as the sister group to Vipera renardi renardi, and Vipera renardi eriwanensis from the Lesser Caucasus as the most basal taxon in the species. Our results illustrate that the divergence between the lowland and montane populations occurred separately in each species and several times in V. renardi. We demonstrated that the recently discovered Transylvanian population is the third surviving population of V. u. rakosiensis and the only known population outside of Hungary., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Introduction: Snakebite is an important and preventable health hazard. Viper bites are more common than other poisonous snakebites in human beings. The present study aimed to collate the victim profile of viper bite cases in the region and to determine the pattern, manifestations, complications and the associated risk factors of these bites., Methods: This was a prospective study of viper bite cases admitted to Kasturba Hospital, Manipal, India between August 2003 and November 2005. The demographic and clinical details of each case were obtained from the patients, their relatives and the patients' hospital records, and analysed., Results: A total of 31 viper bite cases were reported during the study period. The victims were predominantly male and aged 25-55 years. The highest number of cases occurred during daytime in the months of September and October, which coincided with the harvesting season, and involved the lower limbs. 94% of the snakebite victims were farmers, suggesting that this was an occupational hazard. Envenomation was observed in patients with scratch marks, suggesting the importance of keeping the victim under observation in all alleged snakebite cases, even in the absence of clear fang marks. The mortality rate in our study was 6.5%., Conclusion: Immobilising and transporting snakebite victims to the hospital and prompt administration of anti-snake venom remain the best way to reduce morbidity and mortality. It is also important to practise correct first aid measures, as otherwise they may cause more harm than good.

Within the Neotropical pit vipers, a lineage of primarily Middle American snake species referred to as the "Porthidium group" includes the genera Atropoides, Cerrophidion, and Porthidium. In this study, the venom proteomes of Porthidium nasutum, P. ophryomegas, and Cerrophidion godmani from Costa Rica were analyzed, and correlated to their toxic and enzymatic activities. Their HPLC profiles revealed a higher similarity between the two Porthidium species than between these and C. godmani. Proteins belonging to nine (P. nasutum), eight (P. ophryomegas), and nine (C. godmani) families were identified by mass spectrometry or N-terminal sequencing. Final cataloging of proteins and their relative abundances confirmed the close relationship between venoms of P. nasutum and P. ophryomegas, departing from that of C. godmani. Since the latter species had been taxonomically classified as Porthidium godmani previously, our venomic analyses agree with its current generic status. Venoms of P. nasutum and P. ophryomegas, despite containing abundant metalloproteinases and serine proteinases, lack procoagulant activity on human plasma, in contrast to venom of C. godmani. The latter induced strong myotoxicity in mice, which correlates with its high proportion of phospholipases A(2), whereas venoms from the two Porthidium species, containing lower amounts of these enzymes, induced only mild muscle damage., (Copyright © 2011 Elsevier B.V. All rights reserved.)

The saw-scaled viper (SSV) (Echis carinatus) is considered to be a highly venomous snake in Sri Lanka despite any published clinical justification. Being a rarity, the clinical profile of SSV bites is not well established in Sri Lanka. We report a series of 48 (n-48) SSV bites from the Northern Province of Sri Lanka. The majority (65%) of victims had evidence of local envenoming at the site of the bite; however, 29% showed spontaneous bleeding and 71% had coagulopathy. There were no deaths in the series. The envenoming was mild in contrast to the mortality and significant morbidity associated with SSV bites in West Africa and some parts of India. These observations need to be further explored with laboratory studies to identify the venom components, study of morphological characteristics, and genetic profiling of the Sri Lankan SSV to see if it is different from the subspecies found elsewhere.

The rapid and accurate recognition of dangerously venomous snakes following bites is crucial to making appropriate decisions regarding first aid, evacuation, and treatment. Past recommendations for identification of dangerous North American pit vipers have often required subjective determinations of head shape or relied on traits shared with some nondangerous species (elliptical pupils and undivided subcaudal scales). Heat-sensitive facial pits are diagnostic but require close examination of the dangerous head, and cephalic traits are useless when working with a decapitated carcass. Exclusive of cephalic traits, pit vipers north of Mexico can be recognized by the combination of keeled middorsal scales and undivided subcaudal scales. The order of colored rings is usually suggested to identify coral snakes in the United States, yet extension of the colored rings across the ventral scales must be added as an essential identifying factor to ensure elimination of all harmless look-alikes. A novel 3-step flow chart is presented that allows dangerous snakes in the United States and Canada to be recognized quickly and dependably without relying on cephalic traits. This process cannot be used in other countries, however, due to greater variability of these characteristics in snakes from other parts of the world. Finally, close examination of potentially venomous snakes is extraordinarily dangerous and steps to safeguard those making such observations are discussed., (Copyright © 2011 Wilderness Medical Society. Published by Elsevier Inc. All rights reserved.)

We conducted an extensive study of neutralization of lethality of 11 species and one subspecies of snakes of the genus Vipera, and of five species of Macrovipera, by two experimental equine antisera. One antiserum was a trivalent preparation raised against the venoms of Vipera aspis aspis, Vipera berus berus and Vipera ammodytes ammodytes; the other was a pentavalent preparation that also included venoms of Vipera (now Montivipera) xanthina and Macrovipera lebetina obtusa. We measured specific neutralization of lethality against all venoms included in the immunization schemes, and paraspecific neutralization against the venoms of Vipera ammodytes montandoni, Vipera (Montivipera) bornmuelleri, Vipera latastei, Vipera (Mo.) latifii, Vipera (Mo.) lotievi, Vipera (Daboia) palaestinae, Vipera (Mo.) raddei and Vipera seoanei, as well as against Macrovipera (D.) deserti, Macrovipera lebetina cernovi, Macrovipera lebetina turanica and Macrovipera schweitzeri. We found an important degree of paraspecific protection within each genera (omitting recent reclassification) that was quite independent of both the lethal potency of the venoms and their geographic origin. This information may be of use to clinicians charged with the treatment of Vipera or Macrovipera envenomations with non-specific antivenoms., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

We investigated phylogenetic structure and morphological variation in Asian mountain pitvipers of the genus Ovophis (comprising 3-4 species some of which are considered polytypic) by sequencing four mitochondrial markers (cytochrome b, NADH dehydrogenase subunit 4, 12S and 16S rRNA) from 72 specimens, and analysed them in a Bayesian framework together with another 26 sequences from closely related genera. We reconstructed the region of origin and direction of dispersal of the major clades, and of Ovophis as a whole, using likelihood framework analysis. We also defined morphogroups from 280 specimens from across the range of Ovophis to allow the geographic extent of the major clades to be determined, as well as to allow inclusion of specimens lacking sequence data. Phylogenetic analyses confirmed the monophyly of Ovophis as currently defined, and revealed that it contains two major lineages, eastern (mainly Chinese) and western, with both occurring in southwestern China, central and northern Viet Nam. The most likely origin of the genus, and of individual lineages, coincides with the northeastern boundary of the Indomalayan hotspot. Major diversification in this species group likely corresponded to major climatic changes arising from the uplift of the Tibetan Plateau in the early to mid Miocene. With reference to the defined morphogroups, we suggest that at least five species are present and provide appropriate names. With a few exceptions, the newly defined species boundaries do not correspond to the existing taxonomy., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Despite much effort towards resolving the molecular phylogenetic tree for pitvipers, some aspects remain unresolved. In particular, the sister group of the diverse New World radiation has remained impossible to identify with any certainty. In this study, which for the first time includes nuclear intron data from all major groups of Asian pitvipers as well as representatives of the New World radiation, Bayesian inference allows Gloydius to be identified as the most likely sister group to the New World radiation and sheds light on other ambiguous relationships among the Old World pitvipers. The sister group relationship of "Ovophis"okinavensis and "Trimeresurus"gracilis is confirmed by the addition of nuclear genes, and we hypothesise that they form a sister group to the Gloydius+New World clade, best supported when the phylogenetic signal from gaps is included in the form of a simple-coded matrix.

We analyze the phylogeny of the medically important and taxonomically unresolved viper genus Echis using four mitochondrial gene fragments. The results show that the populations of the genus fall into four main clades: the Echis carinatus, E. coloratus, E. ocellatus and E. pyramidum groups. The E. pyramidum and E. coloratus groups are sister taxa but the interrelationships of this clade and the E. ocellatus and E. carinatus groups are unresolved. The initial divergence of the genus appears to coincide with the collision between Afro-Arabia and Eurasia, and that between the E. coloratus and E. pyramidum clades appears to be associated with the opening of the Red Sea. Later land connections between Africa and Arabia may have contributed to shaping the distribution of the E. pyramidum complex. The present distribution of E. carinatus may be the result of range expansion from southern India. Taxonomically, our results provide molecular evidence for the validity of Echis omanensis, E. khosatzkii, E. borkini and E. jogeri, for the presence of unsuspected genetic diversity within the E. pyramidum complex in eastern Africa, and for the conspecificity of E. carinatus and E. multisquamatus. The status of E. leucogaster remains to be confirmed.

The identification of adaptations and key innovations has long interested biologists because they confer on organisms the ability to exploit previously unavailable ecological resources and respond to novel selective pressures. Although it can be extremely difficult to test for the effects of a character on the rate of lineage diversification, the convergent evolution of a character in multiple lineages provides an excellent opportunity to test for the effect of that character on lineage diversification. Here, I examine the effect of parity mode on the diversification of vipers, which have independently evolved viviparity in at least 13 lineages. I find strong statistical evidence that viviparous species diversify at a greater rate than oviparous species and correlate major decreases in the diversification rate of oviparous species with periods of global cooling, such as the Oligocene. These results suggest that the evolution of viviparity buffered live-bearing species against the negative effects of global climate change during the Cenozoic, and was a key innovation in the evolution and diversification of live-bearing vipers.