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Potential para-specific and geographical utility of Thai Green Pit Viper (Trimeresurus albolabris) Monovalent Antivenom: Neutralization of procoagulant and hemorrhagic activities of diverse Trimeresurus pit viper venoms

Yong MY, Tan KY, Tan CH

Toxicon, 2021 Nov;203:85-92.
PMID: 34600909 DOI: 10.1016/j.toxicon.2021.09.021

The Trimeresurus complex consists of diverse medically important venomous pit vipers that cause snakebite envenomation. Antivenoms, however, are in limited supply, and are specific to only two out of the many species across Asia. This study thus investigated the immunoreactivities of regional pit viper antivenoms toward selected Trimeresurus pit viper venoms, and examined the neutralization of their hemotoxic activities. Trimeresurus albolabris Monovalent Antivenom (TaMAV, Thailand) exhibited a higher immunoreactivity than Hemato Bivalent Antivenom (HBAV, raised against Trimeresurus stejnegeri and Protobothrops mucrosquamatus, Taiwan) and Gloydius brevicaudus Monovalent Antivenom (GbMAV, China), attributed to its monovalent nature and conserved antigens in the Trimeresurus pit viper venoms. The venoms showed moderate-to-strong in vitro procoagulant and in vivo hemorrhagic effects consistent with hemotoxic envenomation, except for the Sri Lankan Trimeresurus trigonocephalus venom which lacked hemorrhagic activity. TaMAV was able to differentially neutralize both in vitro and in vivo hemotoxic effects of the venoms, with the lowest efficacy shown against the procoagulant effect of T. trigonocephalus venom. The findings suggest that TaMAV is a potentially useful treatment for envenomation caused by hetero-specific Trimeresurus pit vipers, in particular those in Southeast Asia and East Asia. Clinical study is warranted to establish its spectrum of para-specific effectiveness, and dosages need be tailored to the different species in respective regions.

Matched MeSH terms: Trimeresurus*
Characterizing and applying immunoglobulins in snakebite diagnostics: A simple and rapid venom detection assay for four medically important snake species in Southeast Asia

Lee LP, Tan CH, Khomvilai S, Sitprija V, Chaiyabutr N, Tan KY

Int J Biol Macromol, 2023 May 01;236:123727.
PMID: 36863668 DOI: 10.1016/j.ijbiomac.2023.123727

Snakebite envenoming is a medical emergency requiring urgent and specific treatment. Unfortunately, snakebite diagnostics are scarce, time-consuming and lacking specificity. Hence, this study aimed to develop a simple, quick and specific snakebite diagnostic assay using animal antibodies. Anti-venom horse immunoglobulin G (IgG) and chicken immunoglobulin Y (IgY) were produced against the venoms of four major medically important snake species in Southeast Asia, i.e., the Monocled Cobra (Naja kaouthia), Malayan Krait (Bungarus candidus), Malayan Pit Viper (Calloselasma rhodostoma), and White-lipped Green Pit Viper (Trimeresurus albolabris). Different capture:detection configurations of double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) were constructed using both immunoglobulins, and the horse IgG:IgG-HRP configuration was found to be most selective and sensitive in detecting the corresponding venoms. The method was further streamlined to develop a rapid immunodetection assay, which is able to produce a visual color change within 30 min for discrimination between different snake species. The study shows it is feasible to develop a simple, quick and specific immunodiagnostic assay using horse IgG, which can be derived directly from antisera prepared for antivenom production. The proof-of-concept indicates it is a sustainable and affordable approach in keeping with on-going antivenom manufacturing activities for specific species in the region.

Matched MeSH terms: Trimeresurus*
Toxicity and cross-neutralization of snake venoms from two lesser-known arboreal pit vipers in Southeast Asia: Trimeresurus wiroti and Trimeresurus puniceus

Lee LP, Tan KY, Tan CH

Toxicon, 2020 Oct 15;185:91-96.
PMID: 32585219 DOI: 10.1016/j.toxicon.2020.06.012

The lesser-known Sundaic lance-headed pit vipers Trimeresurus wiroti (Malaysia) and Trimeresurus puniceus (Indonesia) contribute to the disease burden of snakebite envenomation in Southeast Asia, but their venom toxicity and neutralization remain insufficiently investigated. This study demonstrated that both venoms were procoagulant (involving thrombin-like activity), hemorrhagic, and lethal to mice, with T. wiroti venom being more lethal (LD50 = 0.78 μg/g c.f. 1.21 μg/g). The hetero-specific antivenom from Thailand, Green Pit Viper Antivenom (GPVAV, raised against Trimeresurus albolabris) cross-reacted with T. wiroti and T. puniceus venoms with a higher efficacy of immunological binding activity for the latter. The antivenom was also effective in cross-neutralizing the procoagulant, hemorrhagic and lethal effects of the venoms. In lethality neutralization, GPVAV showed a potency of 0.79-1.05 mg venom per mL antivenom, corresponding to the complete neutralization of approximately 8-10 mg venom per unit vial of antivenom for T. wiroti and T. puniceus venoms. Taken together, it was inferred that T. wiroti, T. puniceus, and T. albolabris venoms share common toxin epitopes, thus enabling the cross-neutralization observed. These findings suggest that GPVAV may be potentially useful in the management of envenomation by T. wiroti and T. puniceus while awaiting clinical trial and validation.

Matched MeSH terms: Trimeresurus*
Cross-reactivity of monovalent and polyvalent Trimeresurus antivenoms with venoms from various species of Trimeresurus (lance-headed pit viper) snake

Tan NH, Choy SK, Chin KM, Ponnudurai G

Toxicon, 1994 Jul;32(7):849-53.
PMID: 7940592

Trimeresurus bite is a serious medical problem in Asia. However, at present only a few monospecific Trimeresurus antivenoms are available. Investigation of the cross-neutralization capacity of three Trimeresurus antivenoms indicates that the antivenoms exhibit broad cross-reactivity. A polyvalent Trimeresurus antivenom was also found to be effective in neutralization of the haemorrhagic, necrotizing and thrombin-like activities of heterologous Trimeresurus venoms.

Matched MeSH terms: Trimeresurus/immunology*
Evaluation of the cross-neutralization capacity of Thai green pit viper antivenom against venom of Myanmar green pit viper

Yee KT, Maw LZ, Kyaw AM, Khow O, Oo AW, Oo TKK, et al.

Toxicon, 2020 Apr 15;177:41-45.
PMID: 32056833 DOI: 10.1016/j.toxicon.2020.02.003

Green pit viper (Trimeresurus sp.) bite occurred throughout Myanmar, but there is no specific antivenom produced in the country for related envenomation. Instead, Myanmar Russell's viper antivenom (Anti-MRV) was often misused because of prolonged clotting time was observed from both species. Thai green pit viper antivenom (Anti-TGPV) raised against Trimeresurus albolabris was found to be effective against venoms of more than ten Trimeresurus sp. from Thailand, Malaysia and Indonesia. The present study compared the neutralization capacities of Anti-TGPV and Anti-MRV towards the venom from T. erythrurus from Myanmar. Anti-TGPV was more efficacious than Anti-MRV in cross-neutralizing the lethal and haemorrhagic activities of the venom by a potency of a least 1.4 times higher. Although Anti-TGPV effectively cross-neutralized the coagulation activity of the venom, Anti-MRV failed to do so. Immunodiffusion and immunoblot experiments showed that Anti-TGPV cross-reacted with more protein components of the venom than Anti-MRV. In conclusion, Anti-TGPV is a better choice for patients bitten by Myanmar green pit viper, but further clinical investigation is required. The current findings highlight the development of a specific antivenom against Myanmar green pit viper venom.

Matched MeSH terms: Trimeresurus*
Fulltext Frequency, geographical distribution and outcomes of pit viper bites in Malaysia consulted to Remote Envenomation Consultancy Services (RECS) from 2017 to 2020

Qamruddin RM, Safferi RS, Mohamed Ismail Z, Salleh MS, Abd Hamid MNH, Frederic Ng VER, et al.

PLoS Negl Trop Dis, 2023 Aug;17(8):e0011569.
PMID: 37585486 DOI: 10.1371/journal.pntd.0011569

Not all pit viper species are present in every state of Malaysia and their distribution varies according to altitude. There is limited information on pit viper bite incidence and its geographical distribution. This was a cross-sectional study of confirmed pit viper bite cases referred to Remote Envenomation Consultancy Services (RECS) from January 2017 to December 2020. Data was collected following the approval of institutional research ethics committee. Universal sampling methods were used. Confirmed pit viper bite cases in each state, geographical location and the antivenom used were reported. A total of 523 confirmed pit viper bite injuries occurred over the 4-year study period. The majority were Malaysians, male and young adults. Most were non-occupational related (83.9%) and involved the upper limbs (46.8%). The commonest pit viper species involved was Trimeresurus purpureomaculatus (23.7%). Green pit viper antivenom (GPAV) was the most frequent antivenom used (n = 51) with the majority of patients requiring only one dose (3 vials). This study provides a better appreciation of indigenous pit viper species distribution for each state and reflects the requirement of appropriate antivenom to be stocked in each state or district hospital.

Matched MeSH terms: Trimeresurus*
Isolation, characterization and antigenic cross-reactivities of the major hemorrhagin from Cryptelytrops purpureomaculatus venom

Fung SY, Tan NH

Indian J Exp Biol, 2013 Dec;51(12):1063-9.
PMID: 24579371

The major hemorrhagin from C. purpureomaculatus (mangrove pit viper) venom was purified to homogeneity and termed Maculatoxin. Maculatoxin has a molecular weight of 38 kDa as determined by SDS-PAGE. It is an acidic protein (pI= 4.2) and exhibited proteolytic and hemorrhagic activities (MHD10 = 0.84 microg in mice) but was not lethal to mice at a dose of 1 microg/g. The hemorrhagic activity of Maculatoxin was completely inactivated by EDTA and partially inhibited by ATP and citrate. The N-terminal sequence of Maculatoxin (TPEQQRFPPTYIDLGIFVDHGMYAT) shares a significant degree of homology with the metalloprotease domain of other venom hemorrhagins. Indirect ELISA showed anti-Maculatoxin cross reacted with protein components of many snake venoms. In the double-sandwich ELISA, however, anti-Maculatoxin cross-reacted only with venoms of certain species of the Trimeresurus (Asia lance-head viper) complex, and the results support the recent proposed taxonomy changes concerning the Trimeresurus complex.

Matched MeSH terms: Trimeresurus/immunology*; Trimeresurus/physiology
Ecological diversification in a group of Indomalayan pitvipers (Trimeresurus): convergence in taxonomically important traits has implications for species identification

Sanders KL, Malhotra A, Thorpe RS

J Evol Biol, 2004 Jul;17(4):721-31.
PMID: 15271071

We analyse molecular and phenotypic evolution in a group of taxonomically problematic Indomalayan pitvipers, the Trimeresurus sumatranus group. Mitochondrial DNA sequencing provides a well-resolved phylogeny, with each species representing a distinct lineage. Multivariate morphological analysis reveals a high level of phenotypic differentiation, which is congruent between the sexes but does not reflect phylogenetic history. An adaptive explanation for the observed pattern of differentiation is supported by independent contrasts analysis, which shows significant correlations between current ecology and the characters that most account for the variation between taxa, including those that are presently used to identify the species. Reduced precipitation and altitude, and increased temperature, are correlated with higher numbers of scales on the head, body and tail. It is hypothesized that scale number plays an important role in heat and water exchange by influencing the area of exposed of interstitial skin, and that colour pattern variation reflects selection pressures involving camouflage and thermoregulation. Ecological convergence in traits used for classification is found to have important implications for species identification where taxa are distributed over varying environments.

Matched MeSH terms: Trimeresurus/genetics*; Trimeresurus/physiology
Fulltext Description of cranial elements and ontogenetic change within Tropidolaemus wagleri (Serpentes: Crotalinae)

Hill N

PLoS One, 2019;14(2):e0206023.
PMID: 30785876 DOI: 10.1371/journal.pone.0206023

Tropidolaemus wagleri is a species of Asian pitviper with a geographic range including Thailand, Vietnam, Malaysia, Singapore, Bruniei, parts of Indonesia, and the Philippines. Tropidolaemus is a member of the Crotalinae subfamily, within Viperidae. The genus Tropidolaemus includes five species, and was once included within the genus Trimeresurus. While some osteologic characteristics have been noted a comprehensive description of cranial elements has not been produced for T. wagleri. An in-depth description of the cranial skeleton of Tropidolaemus wagleri lays the foundation for future projects to compare and contrast other taxa within Crotalinae and Viperidae. The chosen reference specimen was compared to the presumed younger specimens to note any variation in ontogeny. The study here provides a comprehensive description of isolated cranial elements as well as a description of ontogenetic change within the specimens observed. This study contributes to the knowledge of osteological characters in T. wagleri and provides a foundation for a long term project to identify isolated elements in the fossil record.

Matched MeSH terms: Trimeresurus
Cross reactivity and lethality neutralization of venoms of Indonesian Trimeresurus complex species by Thai Green Pit Viper Antivenom

Tan CH, Liew JL, Tan NH, Ismail AK, Maharani T, Khomvilai S, et al.

Toxicon, 2017 Dec 15;140:32-37.
PMID: 29051104 DOI: 10.1016/j.toxicon.2017.10.014

Arboreal pit vipers of the Trimeresurus complex group are medically important species in Indonesia (west of Wallace's line), but there is no specific antivenom produced in the country for treating related envenomation. Instead, the exiting trivalent Indonesian antivenom, Biosave® Serum Anti Bisa Ular (SABU, indicated for envenoming by Malayan pit viper, Javan spitting cobra and banded krait) is often misused to treat Trimeresus envenoming resulting in poor therapeutic outcome. Here, we investigated the cross-reactivity and neutralization capability of Thai Green Pit Viper Antivenom (GPVAV) against the venoms of four Indonesian Trimeresurus species. Consistently, the venoms of Trimeresurus (Trimeresurus) insularis, Trimeresurus (Trimeresurus) purpureomaculatus, Trimeresurus (Parias) hageni and Trimeresurus (Craspedocephalus) puniceus of Indonesia showed stronger immunoreactivity on ELISA to GPVAV than to Biosave®. The findings correlated with in vivo neutralization results, whereby GPVAV was far more effective than Biosave® in cross-neutralizing the lethality of the venoms by a potency of at least 13 to 80 times higher. The efficacy of GPVAV is partly attributable to its cross-neutralization of the procoagulant effect of the venoms, thereby mitigating the progression of venom-induced consumptive coagulopathy. The paraspecific effectiveness of GPVAV against Trimeresurus species envenoming in Indonesia await further clinical investigation.

Matched MeSH terms: Trimeresurus*
A confirmed bite by a Beautiful Pit Viper (Trimeresurus venustus) resulting in local symptoms

Fuchs J, Bessire K, Weiler S

Toxicon, 2019 Mar 20;163:44-47.
PMID: 30902684 DOI: 10.1016/j.toxicon.2019.03.019

This case report describes the effect of an envenomation by the Beautiful Pit Viper Trimeresurus venustus (or Cryptelytrops venustus), which is a green pit viper native to Thailand and Malaysia. A previously healthy 60-year-old snake breeder with no relevant medical history was bitten by his adult T. venustus in the third finger of his right hand while taking it out of the feeding box. The bite was painful and swelling progressed to include his whole hand within an hour after the bite. He was treated symptomatically with pre-emptive antibiotics and analgesics, never developed any hematological disorders such as coagulopathy and bleeding or disseminated intravascular coagulation and was discharged 26 hours after the bite. The clinical course in our patient matched two other well-documented cases reported to our Poisons Centre, and one further case presented as a conference-poster. All patients recovered with symptomatic therapy and never required antivenom. Therefore, bites by T. venustus seem to present with less severe symptoms compared to other Trimeresurus species.

Matched MeSH terms: Trimeresurus
Fulltext Cytotoxic and anticancer properties of the Malaysian mangrove pit viper (Trimeresurus purpureomaculatus) venom and its disintegrin (purpureomaculin)

Tan CH, Liew JL, Navanesan S, Sim KS, Tan NH, Tan KY

J Venom Anim Toxins Incl Trop Dis, 2020;26:e20200013.
PMID: 32742279 DOI: 10.1590/1678-9199-JVATITD-2020-0013

Background: The Asiatic pit vipers from the Trimeresurus complex are medically important venomous snakes. These pit vipers are often associated with snakebite that leads to fatal coagulopathy and tissue necrosis. The cytotoxic venoms of Trimeresurus spp.; however, hold great potential for the development of peptide-based anticancer drugs.
Methods: This study investigated the cytotoxic effect of the venom from Trimeresurus purpureomaculatus, the mangrove pit viper (also known as shore pit viper) which is native in Malaysia, across a panel of human cancer cell lines from breast, lung, colon and prostate as well as the corresponding normal cell lines of each tissue.
Results: The venom exhibited dose-dependent cytotoxic activities on all cell lines tested, with median inhibition concentrations (IC50) ranging from 0.42 to 6.98 µg/mL. The venom has a high selectivity index (SI = 14.54) on breast cancer cell line (MCF7), indicating that it is significantly more cytotoxic toward the cancer than to normal cell lines. Furthermore, the venom was fractionated using C18 reversed-phase high-performance liquid chromatography and the anticancer effect of each protein fraction was examined. Fraction 1 that contains a hydrophilic low molecular weight (approximately 7.5 kDa) protein was found to be the most cytotoxic and selective toward the breast cancer cell line (MCF7). The protein was identified using liquid chromatography-tandem mass spectrometry as a venom disintegrin, termed purpureomaculin in this study.
Conclusion: Taken together, the findings revealed the potent and selective cytotoxicity of a disintegrin protein isolated from the Malaysian T. purpureomaculatus venom and suggested its anticancer potential in drug discovery.

Matched MeSH terms: Trimeresurus

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