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.

Characterisation of the forest cobra (Naja melanoleuca) venom using a multifaceted mass spectrometric-based approach.

Snake venoms consist of highly biologically active proteins and peptides that are responsible for the lethal physiological effects of snakebite envenomation. In order to guide the development of targeted antivenom strategies, comprehensive understanding of venom compositions and in-depth characterisation of various proteoforms, often not captured by traditional bottom-up proteomic workflows, is necessary. Here, we employ an integrated 'omics' and intact mass spectrometry (MS)-based approach to profile the heterogeneity within the venom of the forest cobra (Naja melanoleuca), adopting different analytical strategies to accommodate for the dynamic molecular mass range of venom proteins present. The venom proteome of N. melanoleuca was catalogued using a venom gland transcriptome-guided bottom-up proteomics approach, revealing a venom consisting of six toxin superfamilies. The subtle diversity present in the venom components was further explored using reversed phase-ultra performance liquid chromatography (RP-UPLC) coupled to intact MS. This approach showed a significant increase in the number of venom proteoforms within various toxin families that were not captured in previous studies. Furthermore, we probed at the higher-order structures of the larger venom proteins using a combination of native MS and mass photometry and revealed significant structural heterogeneity along with extensive post-translational modifications in the form of glycosylation in these larger toxins. Here, we show the diverse structural heterogeneity of snake venom proteins in the venom of N. melanoleuca using an integrated workflow that incorporates analytical strategies that profile snake venom at the proteoform level, complementing traditional venom characterisation approaches.

Knowledge about Snake Venoms and Toxins from Colombia: A Systematic Review.

Colombia encompasses three mountain ranges that divide the country into five natural regions: Andes, Pacific, Caribbean, Amazon, and Orinoquia. These regions offer an impressive range of climates, altitudes, and landscapes, which lead to a high snake biodiversity. Of the almost 300 snake species reported in Colombia, nearly 50 are categorized as venomous. This high diversity of species contrasts with the small number of studies to characterize their venom compositions and natural history in the different ecoregions. This work reviews the available information about the venom composition, isolated toxins, and potential applications of snake species found in Colombia. Data compilation was conducted according to the PRISMA guidelines, and the systematic literature search was carried out in Pubmed/MEDLINE. Venom proteomes from nine Viperidae and three Elapidae species have been described using quantitative analytical strategies. In addition, venoms of three Colubridae species have been studied. Bioactivities reported for some of the venoms or isolated components-such as antibacterial, cytotoxicity on tumoral cell lines, and antiplasmodial properties-may be of interest to develop potential applications. Overall, this review indicates that, despite recent progress in the characterization of venoms from several Colombian snakes, it is necessary to perform further studies on the many species whose venoms remain essentially unexplored, especially those of the poorly known genus Micrurus.

Proteomic characteristics of six snake venoms from the Viperidae and Elapidae families in China and their relation to local tissue necrosis.

Patients envenomed by snakes from the Viperidae and Elapidae families in China often have varying degrees of local tissue necrosis. Due to the relative clinical characteristics of local tissue necrosis and ulceration following envenoming, this study has analyzed the proteome of six snake venoms from the Viperidae and Elapidae family, and the toxin profiles of each snake were compared and correlated with the clinical manifestations that follow cytotoxic envenoming. Deinagkistrodon acutus and Naja atra envenomation induce severe ulceration, which is absent in Bungarus multicinctus envenomation and mild in the other three vipers. It is interesting to note that the proportion of c-type lectins (CTL) (20.63%) in Deinagkistrodon acutus venom was relatively high, which differs from the venom of other vipers. In addition, three-fingered toxin (3FTx) (2.15%) is present in the venom of Deinagkistrodon acutus, but has not been detected in the remaining three vipers. Snake venom metalloprotease (SVMP) (34.4%-44.7%), phospholipase A2 (PLA2) (9.81%-40.83%), and snake venom serine protease (SVSP) (9.44%-16.2%) represent the most abundant families of toxin in Viperidae venom. The Elapidae venom proteome was mainly composed of neurotoxins and cytotoxins, including 3FTx (39.28%-60.08%) and PLA2 (8.24%-58.95%) toxins, however, the proportion of CRISPS (26.36%) in Naja atra venom was relatively higher compared to Bungarus multicinctus venom. Significant differences in SVMP, SVSP, and 3FTx expression levels exist between the Viperidae and the Elapidae family. The main toxins responsible for the development of tissue necrosis and ulcerations following Viperidae envenoming are hematotoxins (SVSMP, SVSP) and myotoxins (PLA2). Deinagkistrodon acutus venom contains high levels of CTL and traces of 3FTx, leading to more severe local necrosis. However, Naja atra venom can also cause severe local necrosis through the effects of myotoxin (3FTx, CRISP, PLA2). Bungarus multicinctus venom does not contain myotoxins, resulting in pure systemic neurological manifestations no obvious necrosis of local tissue in patients.

miRNAs derived from cobra venom exosomes contribute to the cobra envenomation.

Currently, there is an increasing amount of evidence indicating that exosomes and the miRNAs they contain are crucial players in various biological processes. However, the role of exosomes and miRNAs in snake venom during the envenomation process remains largely unknown. In this study, fresh venom from Naja atra of different ages (2-month-old, 1-year-old, and 5-year-old) was collected, and exosomes were isolated through ultracentrifugation. The study found that exosomes with inactivated proteins and enzymes can still cause symptoms similar to cobra envenomation, indicating that substances other than proteins and enzymes in exosomes may also play an essential role in cobra envenomation. Furthermore, the expression profiles of isolated exosome miRNAs were analyzed. The study showed that a large number of miRNAs were co-expressed and abundant in cobra venom exosomes (CV-exosomes) of different ages, including miR-2904, which had high expression abundance and specific sequences. The specific miR-2094 derived from CV-exosomes (CV-exo-miR-2904) was overexpressed both in vitro and in vivo. As a result, CV-exo-miR-2904 induced symptoms similar to cobra envenomation in mice and caused liver damage, demonstrating that it plays a crucial role in cobra envenomation. These results reveal that CV-exosomes and the miRNAs they contain play a significant regulatory role in cobra envenomation. Our findings provide new insights for the treatment of cobra bites and the development of snake venom-based medicines.

Metabolome-Based Classification of Snake Venoms by Bioinformatic Tools.

Snakebite is considered a neglected tropical disease, and it is one of the most intricate ones. The variability found in snake venom is what makes it immensely complex to study. These variations are present both in the big and the small molecules found in snake venom. This study focused on examining the variability found in the venom's small molecules (i.e., mass range of 100-1000 Da) between two main families of venomous snakes-Elapidae and Viperidae-managing to create a model able to classify unknown samples by means of specific features, which can be extracted from their LC-MS data and output in a comprehensive list. The developed model also allowed further insight into the composition of snake venom by highlighting the most relevant metabolites of each group by clustering similarly composed venoms. The model was created by means of support vector machines and used 20 features, which were merged into 10 principal components. All samples from the first and second validation data subsets were correctly classified. Biological hypotheses relevant to the variation regarding the metabolites that were identified are also given.

Identification evidence unraveled by strict proteomics rules toward forensic samples.

Snake venom is a complex mixture of proteins and peptides secreted by venomous snakes from their poison glands. Although proteomics for snake venom composition, interspecific differences, and developmental evolution has been developed for a decade, current diagnosis or identification techniques of snake venom in clinical intoxication and forensic science applications are mainly dependent on morphological and immunoassay. It could be expected that the proteomics techniques directly offer great help. This work applied a bottom-up proteomics method to identify proteins' types and species attribution in suspected snake venom samples using ultrahigh-performance liquid chromatography-quadrupole-electrostatic field Orbitrap tandem mass spectrometric technique, and cytotoxicity assay was amended to provide a direct evidence of toxicity. Toward the suspicious samples seized in the security control, sample pretreatment (in-sol and in-gel digestion) and data acquisition (nontargeted and targeted screening) modes complemented and validated each other. We have implemented two consequent approaches in identifying the species source of proteins in the samples via the points of venom proteomics and strict forensic identification. First, we completed a workflow consisting of a proteomics database match toward an entire SWISS-PROT (date 2018-11-22) database and a result-directed specific taxonomy database. The latter was a helpful hint to compare master protein kinds and reveal the insufficiency of specific venom proteomics characterization rules. Second, we suggested strict rules for protein identification to meet the requirements of forensic science on improved identification correctness, that is, (1) peptide spectrum matches confidence, peptide confidence, and protein confidence were both high (with the false-discovery ratio less than 1%); (2) the number of unique peptides was more than or equal to two in one protein, and (3) within unique peptides, which at least 75% of the ∆m/z of the matched y and b ions were less than 5 ppm. We identified these samples as cobra venom containing 10 highly abundant proteins (P00597, P82463, P60770, Q9YGI4, P62375, P49123, P80245, P60302, P01442, and P60304) from two snake venom protein families (acid phospholipase A2 and three-finger toxins), and the most abundant proteins were cytotoxins.

Snake Venomics and Antivenomics of Cape Cobra (Naja nivea) from South Africa: Insights into Venom Toxicity and Cross-Neutralization Activity.

Naja nivea (Cape Cobra) is endemic to southern Africa. Envenoming by N. nivea is neurotoxic, resulting in fatal paralysis. Its venom composition, however, has not been studied in depth, and specific antivenoms against it remain limited in supply. Applying a protein decomplexation approach, this study unveiled the venom proteome of N. nivea from South Africa. The major components in the venom are cytotoxins/cardiotoxins (~75.6% of total venom proteins) and alpha-neurotoxins (~7.4%), which belong to the three-finger toxin family. Intriguingly, phospholipase A2 (PLA2) was undetected-this is a unique venom phenotype increasingly recognized in the African cobras of the Uraeus subgenus. The work further showed that VINS African Polyvalent Antivenom (VAPAV) exhibited cross-reactivity toward the venom and immunorecognized its toxin fractions. In mice, VAPAV was moderately efficacious in cross-neutralizing the venom lethality with a potency of 0.51 mg/mL (amount of venom completely neutralized per milliliter of antivenom). In the challenge-rescue model, VAPAV prevented death in 75% of experimentally envenomed mice, with slow recovery from neurotoxicity up to 24 h. The finding suggests the potential para-specific utility of VAPAV for N. nivea envenoming, although a higher dose or repeated administration of the antivenom may be required to fully reverse the neurotoxic effect of the venom.

Venom Variation of Neonate and Adult Chinese Cobras in Captivity Concerning Their Foraging Strategies.

The venom and transcriptome profile of the captive Chinese cobra (Naja atra) is not characterized until now. Here, LC-MS/MS and illumine technology were used to unveil the venom and trascriptome of neonates and adults N. atra specimens. In captive Chinese cobra, 98 co-existing transcripts for venom-related proteins was contained. A total of 127 proteins belong to 21 protein families were found in the profile of venom. The main components of snake venom were three finger toxins (3-FTx), snake venom metalloproteinase (SVMP), cysteine-rich secretory protein (CRISP), cobra venom factor (CVF), and phosphodiesterase (PDE). During the ontogenesis of captive Chinese cobra, the rearrangement of snake venom composition occurred and with obscure gender difference. CVF, 3-FTx, PDE, phospholipase A2 (PLA2) in adults were more abundant than neonates, while SVMP and CRISP in the neonates was richer than the adults. Ontogenetic changes in the proteome of Chinese cobra venom reveals different strategies for handling prey. The levels of different types of toxin families were dramatically altered in the wild and captive specimens. Therefore, we speculate that the captive process could reshape the snake venom composition vigorously. The clear comprehension of the composition of Chinese cobra venom facilitates the understanding of the mechanism of snakebite intoxication and guides the preparation and administration of traditional antivenom and next-generation drugs for snakebite.

Cobra Venom Factor Boosts Arteriogenesis in Mice.

Arteriogenesis, the growth of natural bypass blood vessels, can compensate for the loss of arteries caused by vascular occlusive diseases. Accordingly, it is a major goal to identify the drugs promoting this innate immune system-driven process in patients aiming to save their tissues and life. Here, we studied the impact of the Cobra venom factor (CVF), which is a C3-like complement-activating protein that induces depletion of the complement in the circulation in a murine hind limb model of arteriogenesis. Arteriogenesis was induced in C57BL/6J mice by femoral artery ligation (FAL). The administration of a single dose of CVF (12.5 µg) 24 h prior to FAL significantly enhanced the perfusion recovery 7 days after FAL, as shown by Laser Doppler imaging. Immunofluorescence analyses demonstrated an elevated number of proliferating (BrdU+) vascular cells, along with an increased luminal diameter of the grown collateral vessels. Flow cytometric analyses of the blood samples isolated 3 h after FAL revealed an elevated number of neutrophils and platelet-neutrophil aggregates. Giemsa stains displayed augmented mast cell recruitment and activation in the perivascular space of the growing collaterals 8 h after FAL. Seven days after FAL, we found more CD68+/MRC-1+ M2-like polarized pro-arteriogenic macrophages around growing collaterals. These data indicate that a single dose of CVF boosts arteriogenesis by catalyzing the innate immune reactions, relevant for collateral vessel growth.

The Target Selects the Toxin: Specific Amino Acids in Snake-Prey Nicotinic Acetylcholine Receptors That Are Selectively Bound by King Cobra Venoms.

Snake venom is an adaptive ecological trait that has evolved primarily as a form of prey subjugation. Thus, the selection pressure for toxin diversification is exerted by the prey's physiological targets, with this pressure being particularly acute for specialist feeders, such as the King Cobra species, all of which are snake-prey specialists. However, while extensive research has been undertaken to elucidate key amino acids that guide toxin structure-activity relationships, reciprocal investigations into the specific sites guiding prey-lineage selective effects have been lacking. This has largely been due to the lack of assay systems amenable to systematic amino acid replacements of targeted proteins in the prey's physiological pathways. To fill this knowledge gap, we used a recently described approach based upon mimotope peptides corresponding to the orthosteric site of nicotinic acetylcholine receptor alpha-1 subunits, a major binding site for snake venom neurotoxins that cause flaccid paralysis. We investigated the venoms of four different types of King Cobra (Cambodian, Javan, Malaysian, and Thai). This approach allowed for the determination of the key amino acid positions in King Cobra snake prey that are selectively bound by the toxins, whereby replacing these amino acids in the snake-prey orthosteric site with those from lizards or rats resulted in a significantly lower level of binding by the venoms, while conversely replacing the lizard or rat amino acids with those from the snake at that position increased the binding. By doing such, we identified three negatively charged amino acids in the snake orthosteric site that are strongly bound by the positively charged neurotoxic three-finger toxins found in King Cobra venom. This study, thus, sheds light on the selection pressures exerted by a specialist prey item for the evolution of lineage-selective toxins.

The relative efficacy of chemically diverse small-molecule enzyme-inhibitors against anticoagulant activities of Black Snake (Pseudechis spp.) venoms.

Snakebite remains a worldwide public health burden and a severely neglected tropical disease. Recent research has begun to focus on the potential use of repurposed small-molecule enzyme-inhibitors as early treatments to neutralise the effects of snake venoms. Black snakes (Pseudechis spp.) are a widespread and dangerously venomous group found throughout Australia and New Guinea. Utilising validated coagulation assays, our study assessed the efficacy of two chemically different small molecule inhibitors, a phospholipase A2 inhibitor (varespladib) and a metalloproteinase inhibitor (prinomastat), in vitro neutralisation of the anticoagulant prothrombinase-inhibiting activity of venom from seven species within the Pseudechis genus (P. australis, P. butleri, P. coletti, P. guttatus, P. papuanus, P.rossignolii, P. sp (NT).). Varespladib was shown to be highly effective at neutralising this anticoagulant activity for all seven species, but with P. coletti notably less so than the others. In contrast, prinomastat showed strong neutralisation for five out of the seven species, but was ineffective at neutralising the activity of P. coletti or P. rossignolii venoms. This suggests that varespladib binds to a highly conserved site but that prinomastat binds to a more variable site. These results build upon recent literature indicating that metalloproteinase inhibitors have cross-neutralising potential towards snake venom phospholipase A2 toxins, but with higher degrees of variability that PLA2-specific inhibitors. An important caveat is that these are in vitro tests and while suggestive of potential clinical utility, in vivo animal testing and clinical trials are required as future work.

The structural and functional divergence of a neglected three-finger toxin subfamily in lethal elapids.

Bungarus multicinctus is a widely distributed and medically important elapid snake that produces lethal neurotoxic venom. To study and enhance existing antivenom, we explore the complete repertoire of its toxin genes based on de novo chromosome-level assembly and multi-tissue transcriptome data. Comparative genomic analyses suggest that the three-finger toxin family (3FTX) may evolve through the neofunctionalization of flanking LY6E. A long-neglected 3FTX subfamily (i.e., MKA-3FTX) is also investigated. Only one MKA-3FTX gene, which evolves a different protein conformation, is under positive selection and actively transcribed in the venom gland, functioning as a major toxin effector together with MKT-3FTX subfamily homologs. Furthermore, this lethal snake may acquire self-resistance to its ß-bungarotoxin via amino acid replacements on fast-evolving KCNA2. This study provides valuable resources for further evolutionary and structure-function studies of snake toxins, which are fundamental for the development of effective antivenoms and drug candidates.

Generation of Multivalent Nanobody-Based Proteins with Improved Neutralization of Long α-Neurotoxins from Elapid Snakes.

Recombinantly produced biotherapeutics hold promise for improving the current standard of care for snakebite envenoming over conventional serotherapy. Nanobodies have performed well in the clinic, and in the context of antivenom, they have shown the ability to neutralize long α-neurotoxins in vivo. Here, we showcase a protein engineering approach to increase the valence and hydrodynamic size of neutralizing nanobodies raised against a long α-neurotoxin (α-cobratoxin) from the venom of the monocled cobraNaja kaouthia. Based on the p53 tetramerization domain, a panel of anti-α-cobratoxin nanobody-p53 fusion proteins, termed Quads, were produced with different valences, inclusion or exclusion of Fc regions for endosomal recycling purposes, hydrodynamic sizes, and spatial arrangements, comprising up to 16 binding sites. Measurements of binding affinity and stoichiometry showed that the nanobody binding affinity was retained when incorporated into the Quad scaffold, and all nanobody domains were accessible for toxin binding, subsequently displaying increased blocking potency in vitro compared to the monomeric format. Moreover, functional assessment using automated patch-clamp assays demonstrated that the nanobody and Quads displayed neutralizing effects against long α-neurotoxins from both N. kaouthia and the forest cobra N. melanoleuca. This engineering approach offers a means of altering the valence, endosomal recyclability, and hydrodynamic size of existing nanobody-based therapeutics in a simple plug-and-play fashion and can thus serve as a technology for researchers tailoring therapeutic properties for improved neutralization of soluble targets such as snake toxins.

MT9, a natural peptide from black mamba venom antagonizes the muscarinic type 2 receptor and reverses the M2R-agonist-induced relaxation in rat and human arteries.

All five muscarinic receptors have important physiological roles. The endothelial M2 and M3 subtypes regulate arterial tone through direct coupling to Gq or Gi/o proteins. Yet, we lack selective pharmacological drugs to assess the respective contribution of muscarinic receptors to a given function. We used mamba snake venoms to identify a selective M2R ligand to investigate its contribution to arterial contractions. Using a bio-guided screening binding assay, we isolated MT9 from the black mamba venom, a three-finger toxin active on the M2R subtype. After sequencing and chemical synthesis of MT9, we characterized its structure by X-ray diffraction and determined its pharmacological characteristics by binding assays, functional tests, and ex vivo experiments on rat and human arteries. Although MT9 belongs to the three-finger fold toxins family, it is phylogenetically apart from the previously discovered muscarinic toxins, suggesting that two groups of peptides evolved independently and in a convergent way to target muscarinic receptors. The affinity of MT9 for the M2R is 100 times stronger than that for the four other muscarinic receptors. It also antagonizes the M2R/Gi pathways in cell-based assays. MT9 acts as a non-competitive antagonist against acetylcholine or arecaine, with low nM potency, for the activation of isolated rat mesenteric arteries. These results were confirmed on human internal mammary arteries. In conclusion, MT9 is the first fully characterized M2R-specific natural toxin. It should provide a tool for further understanding of the effect of M2R in various arteries and may position itself as a new drug candidate in cardio-vascular diseases.

Black-necked spitting cobra (Naja nigricollis) phospholipases A2 may cause Trypanosoma brucei death by blocking endocytosis through the flagellar pocket.

African trypanosomes, such as Trypanosoma brucei, are flagellated protozoa which proliferate in mammals and cause a variety of diseases in people and animals. In a mammalian host, the external face of the African trypanosome plasma membrane is covered by a densely packed coat formed of variant surface glycoprotein (VSG), which counteracts the host's adaptive immune response by antigenic variation. The VSG is attached to the external face of the plasma membrane by covalent attachment of the C-terminus to glycosylphosphatidylinositol. As the trypanosome grows, newly synthesised VSG is added to the plasma membrane by vesicle fusion to the flagellar pocket, the sole location of exo- and endocytosis. Snake venoms contain dozens of components, including proteases and phospholipases A2. Here, we investigated the effect of Naja nigricollis venom on T. brucei with the aim of describing the response of the trypanosome to hydrolytic attack on the VSG. We found no evidence for VSG hydrolysis, however, N. nigricollis venom caused: (i) an enlargement of the flagellar pocket, (ii) the Rab11 positive endosomal compartments to adopt an abnormal dispersed localisation, and (iii) cell cycle arrest prior to cytokinesis. Our results indicate that a single protein family, the phospholipases A2 present in N. nigricollis venom, may be necessary and sufficient for the effects. This study provides new molecular insight into T. brucei biology and possibly describes mechanisms that could be exploited for T. brucei targeting.

Serum metabolomics of Bama miniature pigs bitten by Bungarus multicinctus.

Bungarus multicinctus is one of the top ten venomous snakes in China, and its bite causes acute and severe diseases, but its pathophysiology remains poorly elucidated. Thus, an animal model of Bungarus multicinctus bite was established by intramuscular injection of 30µg/kg of Bungarus multicinctus venom, and then the serum metabolites were subsequently screened, identified and validated by ultra-performance liquid chromatography-quadrupole-time of flight-mass spectrometry (UPLC-Q-TOF-MS) methods to explore the potential biomakers and possible metabolic pathways. Untargeted metabolomics analysis showed that 36 and 38 endogenous metabolites levels changed in ESI+ and ESI-, respectively, KEGG pathway analysis showed that 5 metabolic pathways, including mineral absorption, central carbon metabolism in cancer, protein digestion and absorption, aminoacyl-tRNA biosynthesis and ABC transporters might be closely related to Bungarus multicinctus bite. Targeted metabolomics analysis showed that there were significant differences in serum D-proline, L-leucine and L-glutamine after Bungarus multicinctus bite (P < 0.05). In addition, receiver operating characteristic (ROC) analysis showed that the diagnostic efficiency of L-Glutamine was superior to other potential biomarkers and the AUC value was 0.944. Moreover, we found evidence for differences in the pathophysiology of glutamine between Bungarus multicinctus bite group and normal group, specifically with the content of glutamine synthetase (GS) and glutaminase (GLS). Taken together, the current study has successfully established an animal model of Bungarus multicinctus bite, and further identified the links between the metabolic perturbations and the pathophysiology and the potential diagnostic biomakers of Bungarus multicinctus bite, which provided valuable insights for studying the mechanism of Bungarus multicinctus bite.

Venom-Derived Neurotoxins Targeting Nicotinic Acetylcholine Receptors.

Acetylcholine was the first neurotransmitter described. The receptors targeted by acetylcholine are found within organisms spanning different phyla and position themselves as very attractive targets for predation, as well as for defense. Venoms of snakes within the Elapidae family, as well as those of marine snails within the Conus genus, are particularly rich in proteins and peptides that target nicotinic acetylcholine receptors (nAChRs). Such compounds are invaluable tools for research seeking to understand the structure and function of the cholinergic system. Proteins and peptides of venomous origin targeting nAChR demonstrate high affinity and good selectivity. This review aims at providing an overview of the toxins targeting nAChRs found within venoms of different animals, as well as their activities and the structural determinants important for receptor binding.

The clinical course and treatment of black mamba (Dendroaspis polylepis) envenomations: a narrative review.

CONTEXT: The black mamba (Dendroaspis polylepis) is, due to its extremely toxic venom, one of the most dangerous snake species in Sub-Saharan Africa. A D. polylepis bite is a medical emergency and requires adequate action to prevent severe complications. However, there are no comprehensive reviews available based on clinical cases, and no readily accessible guidelines for standardized treatment. Therefore, we aim to provide an overview regarding the currently available clinical literature on D. polylepis envenomations; in order to promote knowledge on symptomatology and treatment options. METHODS: We searched for cases reporting humans bitten by D. polylepis in PubMed, Embase, Scopus, and Sabinet. We searched the reference lists of all eligible articles for additional articles. After quality assessment, 29 cases were included in this review. We used descriptive analysis to create an overview of the collected parameters. DISCUSSION: Among the included case reports and case series, D. polylepis envenomations most frequently resulted in decreased respiratory function, sweating and paralysis. The onset of symptoms usually occurred within 60 minutes. Neurological symptoms occurred more often than symptoms of autonomic dysfunction. In the reported cases most patients (26/29) received antivenom and most survived (25/29). We recommend the reporting of additional structured case reports to improve future analyses on the clinical course of envenomations, in order to improve public health response to D. polylepis envenomations.

ptFVa (Pseudonaja Textilis Venom-Derived Factor Va) Retains Structural Integrity Following Proteolysis by Activated Protein C.

OBJECTIVE: The Australian snake venom ptFV (Pseudonaja textilis venom-derived factor V) variant retains cofactor function despite APC (activated protein C)-dependent proteolysis. Here, we aimed to unravel the mechanistic principles by determining the role of the absent Arg306 cleavage site that is required for the inactivation of FVa (mammalian factor Va). APPROACH AND RESULTS: Our findings show that in contrast to human FVa, APC-catalyzed proteolysis of ptFVa at Arg306 and Lys507 does not abrogate ptFVa cofactor function. Remarkably, the structural integrity of APC-proteolyzed ptFVa is maintained indicating that stable noncovalent interactions prevent A2-domain dissociation. Using Molecular Dynamics simulations, we uncovered key regions located in the A1 and A2 domain that may be at the basis of this remarkable characteristic. CONCLUSIONS: Taken together, we report a completely novel role for uniquely adapted regions in ptFVa that prevent A2 domain dissociation. As such, these results challenge our current understanding by which strict regulatory mechanisms control FVa activity.