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Davinia Pla
Evolutionary and Translational Venomics Laboratory, Instituto de Biomedicina de Valencia, Consejo Superior de Investigaciones Científicas (CSIC), Valencia, Spain

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Research article
Published: 01 February 2021 in PLOS Neglected Tropical Diseases
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Background Bothrops asper represents the clinically most important snake species in Central America and Northern South America, where it is responsible for an estimated 50–80% of snakebites. Compositional variability among the venom proteomes of B. asper lineages across its wide range mirrors clinical differences in their envenomings. Bothropic antivenoms generated in a number of Latin American countries commonly exhibit a certain degree of paraspecific effectiveness in the neutralization of congeneric venoms. Defining the phylogeographic boundaries of an antivenom's effectivity has implications for optimizing its clinical use. However, the molecular bases and impact of venom compositions on the immune recognition and neutralization of the toxic activities of across geographically disparate populations of B. asper lineages has not been comprehensively studied. Methodology/Principal findings Third-generation antivenomics was applied to quantify the cross-immunorecognizing capacity against the individual components of venoms of three B. asper lineages (B. asper (sensu stricto), B. ayerbei and B. rhombeatus) distributed in south-western (SW) Colombia, of six Latin American antivenoms, produced against homologous (Colombia, INS-COL and PROBIOL) and Costa Rica (ICP)), and heterologous (Argentina (BIOL), Perú (INS-PERU) and Venezuela (UCV)) bothropic venoms. In vivo neutralization assays of the lethal, hemorrhagic, coagulant, defibrinogenating, myotoxic, edematogenic, indirect hemolytic, and proteolytic activities of the three SW Colombian B. asper lineage venoms were carried to compare the preclinical efficacy of three (Colombian INS-COL and PROBIOL, and Costa Rican ICP) antivenoms frequently used in Colombia. Antivenomics showed that all the six antivenom affinity matrices efficiently immunoretained most of the B. asper lineages venom proteins and exhibited impaired binding towards the venoms' peptidomes. The neutralization profile of the INS-COL, PROBIOL and ICP antivenoms towards the biological activities of the venoms of SW Colombian B. asper (sensu stricto), B. ayerbei and B. rhombeatus lineages was coherent with the antivenomics outcome. In addition, the combination of in vitro (antivenomics) and in vivo neutralization results allowed us to determine their toxin-specific and venom neutralizing antibody content. Noteworthy, heterologous INS-PERU, BIOL, and UCV bothropic antivenoms had equal or higher binding capacity towards the venoms components of SW Colombian B. asper lineages that the homologous Colombian and Costa Rican antivenoms. Conclusions/Significance The combined in vitro and in vivo preclinical outcome showed that antivenoms manufactured in Colombia and Costa Rica effectively neutralize the major toxic activities of SW Colombian B. asper lineage venoms. The antivenomics profiles of the heterologous antivenoms manufactured in Argentina, Venezuela, and Perú strongly suggests their (pre)clinical adequacy for the treatment of B. asper lineage envenomings in SW Colombia. However, their recommendation in the clinical setting is pending on in vivo neutralization testing and clinical testing in humans. Bothrops asper is a highly adaptable snake species complex, which is considered the most dangerous snake throughout much of its distribution range from the Atlantic lowland of eastern México to northwestern Perú. Antivenoms are the only scientifically validated treatment of snakebite envenomings. Venom variation is particularly common in wide ranging species, such as B. asper, and may result in variable clinical presentations of envenomings, as is the case for the B. asper species complex, potentially undermining the efficacy of snakebite treatments depending on the immunization mixture used in the generation of the antivenom. Conversely, phylogenetic conservation of antigenic determinants confers an unpredictable degree of paraspecificity to homologous antivenoms produced for a geographic area, but also to heterologous congeneric antivenoms, towards the venom components of allopatric conspecific populations. This work aimed at comparing the preclinical profile of a panel of Latin American homologous and heterologous antivenoms against the venoms of B. asper lineages distributed in SW Colombia. The outcome of this study strongly suggests the suitability of considering the heterologous antivenoms BIOL (Argentina), UCV (Venezuela) and INS-PERU (Perú) as alternatives to homologous Colombian INS-COL and PROBIOL and Costa Rican ICP antivenoms for the treatment of envenomings by B. asper (sensu stricto) in W Colombia and Ecuador, B. ayerbei in Cauca and Nariño (Colombia), and B. rhombeatus in Cauca river valley, SW Colombia.

ACS Style

Diana Mora-Obando; Davinia Pla; Bruno Lomonte; Jimmy Alexander Guerrero-Vargas; Santiago Ayerbe; Juan J. Calvete. Antivenomics and in vivo preclinical efficacy of six Latin American antivenoms towards south-western Colombian Bothrops asper lineage venoms. PLOS Neglected Tropical Diseases 2021, 15, e0009073 .

AMA Style

Diana Mora-Obando, Davinia Pla, Bruno Lomonte, Jimmy Alexander Guerrero-Vargas, Santiago Ayerbe, Juan J. Calvete. Antivenomics and in vivo preclinical efficacy of six Latin American antivenoms towards south-western Colombian Bothrops asper lineage venoms. PLOS Neglected Tropical Diseases. 2021; 15 (2):e0009073.

Chicago/Turabian Style

Diana Mora-Obando; Davinia Pla; Bruno Lomonte; Jimmy Alexander Guerrero-Vargas; Santiago Ayerbe; Juan J. Calvete. 2021. "Antivenomics and in vivo preclinical efficacy of six Latin American antivenoms towards south-western Colombian Bothrops asper lineage venoms." PLOS Neglected Tropical Diseases 15, no. 2: e0009073.

Journal article
Published: 20 August 2020 in Journal of Proteomics
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Bothrops asper is a venomous pitviper that is widely distributed and of clinical importance in Mesoamerica and northern South America, where it is responsible for 50–80% of all envenomations by Viperidae species. Previous work suggests that B. asper has a complex phylogeographic structure, with the existence of multiple evolutionarily distinct lineages, particularly in the inter-Andean valleys of north South America. To explore the impact of the evolutionary history of B. asper on venom composition, we have investigated geographic variation in the venom proteome of this species from the populations from the Pacific side of Ecuador and south-western Colombia. Among the 21 classes of venom components identified, proteins from mainly four major toxin families, snake venom metalloproteases (PI- and PII-SVMP), phospholipases A2 (K49- and D49-PLA2s), serine proteinases (SVSP), and C-type lectins-like (CTL) proteins are major contributors to the geographic variability in venom. Principal component analyses demonstrate significant differences in venom composition between B. asper lineages previously identified through combination of molecular, morphological and geographical data, and provide additional insights into the selection pressures modulating venom phenotypes on a geographic scale. In particular, altitudinal zonation within the Andean mountain range stands out as a key ecological factor promoting diversification in venom. In addition, the pattern of distribution of PLA2 molecules among B. asper venoms complements phylogenetic analysis in the reconstruction of the dispersal events that account for the current biogeographic distribution of the present-day species' phylogroups. Ontogenic variation was also evident among venoms from some Ecuadorian lineages, although this age-related variation was less extreme than reported in B. asper venoms from Costa Rica. The results of our study demonstrate a significant impact of phylogenetic history on venom composition in a pitviper and show how analyses of this variation can illuminate the timing of the cladogenesis and ecological events that shaped the current distribution of B. asper lineages. Bothrops asper, called “the ultimate pitviper” due to its defensive behavior, large body size, and medical importance, represents a species complex that is widely distributed from southern México southwards across north‐western South America to north-western Perú. This work reports the characterization of the venom proteomes of B. asper lineages from the Pacific sides of Ecuador and south-western Colombia. Multivariate analyses indicate that variability in venom composition among the B. asper lineages is driven by proteins from four major toxin families, presumably in response to selection pressures created by recent and historical ecological conditions created by geological and climatic events from the Pliocene-Pleistocene to the present along the Central and South American Continental Divide. The emerging biogeographic pattern of venom variation, interpreted in the context of the current phylogenetic hypotheses, support and complement previously proposed evolutionary Plio-Pleistocene dispersal events that shaped the present-day distribution range of B. asper lineages. In addition, our venomics data indicate the occurrence of genetic exchange between Colombian and Pacific Costa Rican populations, which may have occurred during the second wave of B. asper migration into Mesoamerica. Our work represents a foundation for a future broader sampling and more complete “-omics” analyses to deepen our understanding of the patterns and causes of venom variation in this medically important pitviper.

ACS Style

Diana Mora-Obando; David Salazar-Valenzuela; Davinia Pla; Bruno Lomonte; Jimmy Alexander Guerrero-Vargas; Santiago Ayerbe; H. Lisle Gibbs; Juan J. Calvete. Venom variation in Bothrops asper lineages from North-Western South America. Journal of Proteomics 2020, 229, 103945 .

AMA Style

Diana Mora-Obando, David Salazar-Valenzuela, Davinia Pla, Bruno Lomonte, Jimmy Alexander Guerrero-Vargas, Santiago Ayerbe, H. Lisle Gibbs, Juan J. Calvete. Venom variation in Bothrops asper lineages from North-Western South America. Journal of Proteomics. 2020; 229 ():103945.

Chicago/Turabian Style

Diana Mora-Obando; David Salazar-Valenzuela; Davinia Pla; Bruno Lomonte; Jimmy Alexander Guerrero-Vargas; Santiago Ayerbe; H. Lisle Gibbs; Juan J. Calvete. 2020. "Venom variation in Bothrops asper lineages from North-Western South America." Journal of Proteomics 229, no. : 103945.

Journal article
Published: 20 April 2020 in Toxicon: X
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We have applied a combination of venomics, in vivo neutralization assays, and in vitro third-generation antivenomics analysis to assess the preclinical efficacy of the monospecific anti-Macrovipera lebetina turanica (anti-Mlt) antivenom manufactured by Uzbiopharm® (Uzbekistan) and the monospecific anti-Vipera berus berus antivenom from Microgen® (Russia) against the venom of Dagestan blunt-nosed viper, Macrovipera lebetina obtusa (Mlo). Despite their low content of homologous (anti-Mlt, 5–10%) or para-specific (anti-Vbb, 4–9%) F(ab')2 antibody fragments against M. l. obtusa venom toxins, both antivenoms efficiently recognized most components of the complex venom proteome's arsenal, which is made up of toxins derived from 11 different gene families and neutralized, albeit at different doses, key toxic effects of M. l. obtusa venom, i.e., in vivo lethal and hemorrhagic effects in a murine model, and in vitro phospholipase A2, proteolytic and coagulant activities. The calculated lethality neutralization potencies for Uzbiopharm® anti-Mlt and anti-Vbb Microgen® antivenoms were 1.46 and 1.77 mg/mL, indicating that 1 mL of Uzbiopharm® and Microgen® antivenoms may protect mice from 41 to 50 LD50s of Mlo venom, respectively. The remarkable degree of conservation of immunogenic determinants between species of the clades of European and Oriental viper, which evolved geographically segregated since the early Miocene, suggests an eventual window of opportunity for the treatment of envenomings by Eurasian snakes. Clearly, the rational use of heterologous antivenoms requires establishing their para-specificity landscapes. This paper illustrates the analytical power of combining in vitro and in vivo preclinical quantitative assays toward this goal.

ACS Style

Davinia Pla; Sarai Quesada-Bernat; Yania Rodríguez; Andrés Sánchez; Mariángela Vargas; Mauren Villalta; Susana Mesén; Álvaro Segura; Denis O. Mustafin; Yulia A. Fomina; Ruslan I. Al-Shekhadat; Juan J. Calvete. Dagestan blunt-nosed viper, Macrovipera lebetina obtusa (Dwigubsky, 1832), venom. Venomics, antivenomics, and neutralization assays of the lethal and toxic venom activities by anti-Macrovipera lebetina turanica and anti-Vipera berus berus antivenoms. Toxicon: X 2020, 6, 100035 .

AMA Style

Davinia Pla, Sarai Quesada-Bernat, Yania Rodríguez, Andrés Sánchez, Mariángela Vargas, Mauren Villalta, Susana Mesén, Álvaro Segura, Denis O. Mustafin, Yulia A. Fomina, Ruslan I. Al-Shekhadat, Juan J. Calvete. Dagestan blunt-nosed viper, Macrovipera lebetina obtusa (Dwigubsky, 1832), venom. Venomics, antivenomics, and neutralization assays of the lethal and toxic venom activities by anti-Macrovipera lebetina turanica and anti-Vipera berus berus antivenoms. Toxicon: X. 2020; 6 ():100035.

Chicago/Turabian Style

Davinia Pla; Sarai Quesada-Bernat; Yania Rodríguez; Andrés Sánchez; Mariángela Vargas; Mauren Villalta; Susana Mesén; Álvaro Segura; Denis O. Mustafin; Yulia A. Fomina; Ruslan I. Al-Shekhadat; Juan J. Calvete. 2020. "Dagestan blunt-nosed viper, Macrovipera lebetina obtusa (Dwigubsky, 1832), venom. Venomics, antivenomics, and neutralization assays of the lethal and toxic venom activities by anti-Macrovipera lebetina turanica and anti-Vipera berus berus antivenoms." Toxicon: X 6, no. : 100035.

Journal article
Published: 01 September 2019 in Journal of Proteomics
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Russell's viper (Daboia russelii) is, together with Naja naja, Bungarus caeruleus and Echis carinatus, a member of the medically important 'Big Four' species responsible for causing a large number of morbidity and mortality cases across the Indian subcontinent. Despite the wide distribution of Russell's viper and the well-documented ubiquity of the phenomenon of geographic variability of intraspecific snake venom composition, Indian polyvalent antivenoms against the "Big Four" venoms are raised against venoms sourced mainly from Chennai in the southeastern Indian state of Tamil Nadu. Biochemical and venomics investigations have consistently revealed notable compositional, functional, and immunological differences among geographic variants of Russell's viper venoms across the Indian subcontinent. However, these studies, carried out by different laboratories using different protocols and involving venoms from a single geographical region, make the comparison of the different venoms difficult. To bridge this gap, we have conducted bioactivities and proteomic analyses of D. russelii venoms from the three corners of the Indian subcontinent, Pakistan, Bangladesh, and Tamil Nandu (India) and Sri Lanka, along with comparative in vivo neutralization and in vitro third-generation antivenomics of antivenoms used in India, Bangladesh and Sri Lanka. These analyses let us to propose two alternative routes of radiation for Russell's viper in the Indian subcontinent. Both radiations, towards the northeast of India and Bangladesh and towards south India and Sri Lanka, have a common origin in Pakistan, and provide a phylovenomics ground for rationalizing the geographic variability in venom composition and their distinct immunoreactivity against available antivenoms. BIOLOGICAL SIGNIFICANCE: Russell's viper (Daboia russelii), the Indian cobra (Naja naja), the common krait (Bungarus caeruleus), and the saw-scaled viper (Echis carinatus) constitute the 'Big Four' snake species responsible for most snakebite envenomings and deaths in the Indian subcontinent. Despite the medical relevance of Daboia russelii, and the well documented variations in the clinical manifestations of envenomings by this wide distributed species, which are doubtless functionally related to differences in venom composition of its geographic variants, antivenoms for the clinical treatment of envenomings by D. russelii across the Indian subcontinent are invariably raised using venom sourced mainly from the southeastern Indian state of Tamil Nadu. We have applied a phylovenomics approach to compare the venom proteomes of Russell's vipers from the three corners of the Indian subcontinent, Pakistan, Bangladesh, and South India/Sri Lanka, and have assessed the in vitro (third-generation antivenomics) and in vivo preclinical efficacy of a panel of homologous antivenoms. The identification of two dispersal routes of ancestral D. russelii into the Indian subcontinent provides the ground for rationalizing the variability in composition and immunoreactivity of the venoms of extant geographic variants of Russell's viper. Such knowledge is relevant for envisioning strategies to improve the clinical coverage of anti- D. russelii antivenoms.

ACS Style

Davinia Pla; Libia Sanz; Sarai Quesada-Bernat; Mauren Villalta; Joshua Baal; Mohammad Abdul Wahed Chowdhury; Guillermo León; José M. Gutiérrez; Ulrich Kuch; Juan J. Calvete. Phylovenomics of Daboia russelii across the Indian subcontinent. Bioactivities and comparative in vivo neutralization and in vitro third-generation antivenomics of antivenoms against venoms from India, Bangladesh and Sri Lanka. Journal of Proteomics 2019, 207, 103443 .

AMA Style

Davinia Pla, Libia Sanz, Sarai Quesada-Bernat, Mauren Villalta, Joshua Baal, Mohammad Abdul Wahed Chowdhury, Guillermo León, José M. Gutiérrez, Ulrich Kuch, Juan J. Calvete. Phylovenomics of Daboia russelii across the Indian subcontinent. Bioactivities and comparative in vivo neutralization and in vitro third-generation antivenomics of antivenoms against venoms from India, Bangladesh and Sri Lanka. Journal of Proteomics. 2019; 207 ():103443.

Chicago/Turabian Style

Davinia Pla; Libia Sanz; Sarai Quesada-Bernat; Mauren Villalta; Joshua Baal; Mohammad Abdul Wahed Chowdhury; Guillermo León; José M. Gutiérrez; Ulrich Kuch; Juan J. Calvete. 2019. "Phylovenomics of Daboia russelii across the Indian subcontinent. Bioactivities and comparative in vivo neutralization and in vitro third-generation antivenomics of antivenoms against venoms from India, Bangladesh and Sri Lanka." Journal of Proteomics 207, no. : 103443.

Journal article
Published: 01 February 2019 in Toxins
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The common European adder, Vipera berus berus, is a medically relevant species, which is widely distributed in Russia and thus, is responsible for most snakebite accidents in Russia. We have investigated the toxic and enzymatic activities and have determined the proteomic composition of its venom. Phospholipases A2 (PLA2, 25.3% of the venom proteome), serine proteinases (SVSP, 16.2%), metalloproteinases (SVMP, 17.2%), vasoactive peptides (bradykinin-potentiating peptides (BPPs), 9.5% and C-type natriuretic peptides (C-NAP, 7.8%), cysteine-rich secretory protein (CRISP, 8%) and L-amino acid oxidase (LAO, 7.3%) represent the major toxin classes found in V. b. berus (Russia) venom. This study was also designed to assess the in vivo and in vitro preclinical efficacy of the Russian Microgen antivenom in neutralizing the main effects of V. b. berus venom. The results show that this antivenom is capable of neutralizing the lethal, hemorrhagic and PLA2 activities. Third-generation antivenomics was applied to quantify the toxin-recognition landscape and the maximal binding capacity of the antivenom for each component of the venom. The antivenomics analysis revealed that 6.24% of the anti-V. b. berus F(ab’)2 molecules fraction are toxin-binding antibodies, 60% of which represent clinically relevant antivenom molecules.

ACS Style

Ruslan Al-Shekhadat; Ksenia S. Lopushanskaya; Álvaro Segura; José María Gutiérrez; Juan J. Calvete; Davinia Pla. Vipera berus berus Venom from Russia: Venomics, Bioactivities and Preclinical Assessment of Microgen Antivenom. Toxins 2019, 11, 90 .

AMA Style

Ruslan Al-Shekhadat, Ksenia S. Lopushanskaya, Álvaro Segura, José María Gutiérrez, Juan J. Calvete, Davinia Pla. Vipera berus berus Venom from Russia: Venomics, Bioactivities and Preclinical Assessment of Microgen Antivenom. Toxins. 2019; 11 (2):90.

Chicago/Turabian Style

Ruslan Al-Shekhadat; Ksenia S. Lopushanskaya; Álvaro Segura; José María Gutiérrez; Juan J. Calvete; Davinia Pla. 2019. "Vipera berus berus Venom from Russia: Venomics, Bioactivities and Preclinical Assessment of Microgen Antivenom." Toxins 11, no. 2: 90.

Short communication
Published: 29 December 2018 in Toxicon: X
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Bothrops lanceolatus inflicts severe envenomings in the Lesser Caribbean island of Martinique. Bothrofav®, a monospecific antivenom against B. lanceolatus venom, has proven highly effective at the preclinical and clinical levels. Here, we report a detailed third-generation antivenomics quantitative analysis of Bothrofav®. With the exception of poorly-immunogenic peptides, Bothrofav® immunocaptured all the major protein components. These results, along with previous preclinical and clinical observations, underscore the high neutralizing efficacy of the antivenom against B. lanceolatus venom.

ACS Style

Davinia Pla; Yania Rodríguez; Dabor Resiere; Hossein Mehdaoui; José María Gutiérrez; Juan J. Calvete. Third-generation antivenomics analysis of the preclinical efficacy of Bothrofav® antivenom towards Bothrops lanceolatus venom. Toxicon: X 2018, 1, 100004 .

AMA Style

Davinia Pla, Yania Rodríguez, Dabor Resiere, Hossein Mehdaoui, José María Gutiérrez, Juan J. Calvete. Third-generation antivenomics analysis of the preclinical efficacy of Bothrofav® antivenom towards Bothrops lanceolatus venom. Toxicon: X. 2018; 1 ():100004.

Chicago/Turabian Style

Davinia Pla; Yania Rodríguez; Dabor Resiere; Hossein Mehdaoui; José María Gutiérrez; Juan J. Calvete. 2018. "Third-generation antivenomics analysis of the preclinical efficacy of Bothrofav® antivenom towards Bothrops lanceolatus venom." Toxicon: X 1, no. : 100004.

Journal article
Published: 02 October 2018 in Journal of Proteomics
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While envenoming by the southern African shield-nosed or coral snakes (genus Aspidelaps) has caused fatalities, bites are uncommon. Consequently, this venom is not used in the mixture of snake venoms used to immunise horses for the manufacture of regional SAIMR (South African Institute for Medical Research) polyvalent antivenom. Aspidelaps species are even excluded from the manufacturer's list of venomous snakes that can be treated by this highly effective product. This leaves clinicians, albeit rarely, in a therapeutic vacuum when treating envenoming by these snakes. This is a significantly understudied small group of nocturnal snakes and little is known about their venom compositions and toxicities. Using a murine preclinical model, this study determined that the paralysing toxicity of venoms from Aspidelaps scutatus intermedius, A. lubricus cowlesi and A. l. lubricus approached that of venoms from highly neurotoxic African cobras and mambas. This finding was consistent with the cross-genus dominance of venom three-finger toxins, including numerous isoforms which showed extensive interspecific variation. Our comprehensive analysis of venom proteomes showed that the three Aspidelaps species possess highly similar venom proteomic compositions. We also revealed that the SAIMR polyvalent antivenom cross-reacted extensively in vitro with venom proteins of the three Aspidelaps. Importantly, this cross-genus venom-IgG binding translated to preclinical (in a murine model) neutralisation of A. s. intermedius venom-induced lethality by the SAIMR polyvalent antivenom, at doses comparable with those that neutralise venom from the cape cobra (Naja nivea), which the antivenom is directed against. Our results suggest a wider than anticipated clinical utility of the SAIMR polyvalent antivenom, and here we seek to inform southern African clinicians that this readily available antivenom is likely to prove effective for victims of Aspidelaps envenoming. Coral and shield-nosed snakes (genus Aspidelaps) comprise two species and several subspecies of potentially medically important venomous snakes distributed in Namibia, Botswana, Zimbabwe, Mozambique and South Africa. Documented human fatalities, although rare, have occurred from both A. lubricus and A. scutatus. However, their venom proteomes and the pathological effects of envenomings by this understudied group of nocturnal snakes remain uncharacterised. Furthermore, no commercial antivenom is made using venom from species of the genus Aspidelaps. To fill this gap, we have conducted a transcriptomics-guided comparative proteomics analysis of the venoms of the intermediate shield-nose snake (A. s. intermedius), southern coral snake (A. l. lubricus), and Cowle's shield snake (A. l. cowlesi); investigated the mechanism of action underpinning lethality by A. s. intermedius in the murine model; and assessed the in vitro immunoreactivity of the SAIMR polyvalent antivenom towards the venom toxins of A. l. lubricus and A. l. cowlesi, and the in vivo capability of this antivenom at neutralising the lethal effect of A. s. intermedius venom. Our data revealed a high degree of conservation of the global composition of the three Aspidelaps venom proteomes, all characterised by the overwhelming predominance of neurotoxic 3FTxs, which induced classical signs of systemic neurotoxicity in mice. The SAIMR polyvalent antivenom extensively binds to Aspidelaps venom toxins and neutralised, with a potency of 0.235 mg venom/mL antivenom, the lethal effect of A. s. intermedius venom. Our data suggest that the SAIMR antivenom could be a useful therapeutic tool for treating human envenomings by Aspidelaps species.

ACS Style

Gareth Whiteley; Nicholas R. Casewell; Davinia Pla; Sarai Quesada Bernat; Rhiannon A.E. Logan; Fiona M.S. Bolton; Simon C. Wagstaff; José M. Gutiérrez; Juan J. Calvete; Robert A. Harrison. Defining the pathogenic threat of envenoming by South African shield-nosed and coral snakes (genus Aspidelaps), and revealing the likely efficacy of available antivenom. Journal of Proteomics 2018, 198, 186 -198.

AMA Style

Gareth Whiteley, Nicholas R. Casewell, Davinia Pla, Sarai Quesada Bernat, Rhiannon A.E. Logan, Fiona M.S. Bolton, Simon C. Wagstaff, José M. Gutiérrez, Juan J. Calvete, Robert A. Harrison. Defining the pathogenic threat of envenoming by South African shield-nosed and coral snakes (genus Aspidelaps), and revealing the likely efficacy of available antivenom. Journal of Proteomics. 2018; 198 ():186-198.

Chicago/Turabian Style

Gareth Whiteley; Nicholas R. Casewell; Davinia Pla; Sarai Quesada Bernat; Rhiannon A.E. Logan; Fiona M.S. Bolton; Simon C. Wagstaff; José M. Gutiérrez; Juan J. Calvete; Robert A. Harrison. 2018. "Defining the pathogenic threat of envenoming by South African shield-nosed and coral snakes (genus Aspidelaps), and revealing the likely efficacy of available antivenom." Journal of Proteomics 198, no. : 186-198.

Review
Published: 01 June 2018 in Toxicon
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Snakebite envenoming represents a major issue in rural areas of tropical and subtropical regions across sub-Saharan Africa, South to Southeast Asia, Latin America and Oceania. Antivenoms constitute the only scientifically validated therapy for snakebite envenomings, provided they are safe, effective, affordable, accessible and administered appropriately. However, the lack of financial incentives in a technology that has remained relatively unchanged for more than a century, has contributed to some manufacturers leaving the market and others downscaling production or increasing the prices, leading to a decline in the availability and accessibility for these life-saving antidotes to millions of rural poor most at risk from snakebites in low income countries. The shortage of antivenoms can be significantly alleviated by optimizing the use of current antivenoms (through the assessment of their specific and paraspecific efficacy against the different medically relevant homologous and heterologous snake venoms) and by generating novel polyspecific antivenoms exhibiting broad clinical spectrum and wide geographic distribution range. Research on venoms has been continuously enhanced by advances in technology. Particularly, the last decade has witnessed the development of omics strategies for unravelling the toxin composition of venoms (“venomics”) and to assess the immunorecognition profile of antivenoms (“antivenomics”). Here, we review recent developments and reflect on near future innovations that promise to revolutionize the mutually enlightening relationship between evolutionary and translational venomics.

ACS Style

Juan J. Calvete; Yania Rodríguez; Sarai Quesada-Bernat; Davinia Pla. Toxin-resolved antivenomics-guided assessment of the immunorecognition landscape of antivenoms. Toxicon 2018, 148, 107 -122.

AMA Style

Juan J. Calvete, Yania Rodríguez, Sarai Quesada-Bernat, Davinia Pla. Toxin-resolved antivenomics-guided assessment of the immunorecognition landscape of antivenoms. Toxicon. 2018; 148 ():107-122.

Chicago/Turabian Style

Juan J. Calvete; Yania Rodríguez; Sarai Quesada-Bernat; Davinia Pla. 2018. "Toxin-resolved antivenomics-guided assessment of the immunorecognition landscape of antivenoms." Toxicon 148, no. : 107-122.

Journal article
Published: 19 April 2018 in Communications Biology
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Snake envenoming causes several potentially lethal pathologies. The specific pathology is dictated by the toxin composition of venom, which varies by species, geography and ontogeny. This variation severely restricts the paraspecific efficacy of antivenoms used to treat snakebite victims. With a view to devising pathology-specific snakebite treatments, we assessed the procoagulant activity of 57 snake venoms and investigated the efficacy of various antivenoms. We find that procoagulant venoms act differentially on key steps of the coagulation cascade, and that certain monospecific antivenoms work in a previously unrecognised paraspecific manner to neutralise this activity, despite conventional assumptions of congener-restricted efficacy. Moreover, we demonstrate that the metal chelator EDTA is also capable of neutralising venom-induced lethality in vivo. This study illustrates the exciting potential of developing new, broad-spectrum, toxin-targeting antivenoms capable of treating key snakebite pathologies, and advocates a thorough re-examination of enzyme inhibiting compounds as alternative therapies for treating snakebite victims. Stuart Ainsworth and colleagues characterized the coagulopathic activity of snake venom and demonstrated that certain monospecific antivenoms can neutralize procoagulant venoms from more than one species. This study suggests a possibility of developing broad-spectrum, toxin-targeting antivenoms to treat snakebite victims.

ACS Style

Stuart Ainsworth; Julien Slagboom; Nessrin Alomran; Davinia Pla; Yasir Alhamdi; Sarah I. King; Fiona M. S. Bolton; José María Gutiérrez; Freek J. Vonk; Cheng-Hock Toh; Juan J. Calvete; Jeroen Kool; Robert A. Harrison; Nicholas R. Casewell. The paraspecific neutralisation of snake venom induced coagulopathy by antivenoms. Communications Biology 2018, 1, 1 -14.

AMA Style

Stuart Ainsworth, Julien Slagboom, Nessrin Alomran, Davinia Pla, Yasir Alhamdi, Sarah I. King, Fiona M. S. Bolton, José María Gutiérrez, Freek J. Vonk, Cheng-Hock Toh, Juan J. Calvete, Jeroen Kool, Robert A. Harrison, Nicholas R. Casewell. The paraspecific neutralisation of snake venom induced coagulopathy by antivenoms. Communications Biology. 2018; 1 (1):1-14.

Chicago/Turabian Style

Stuart Ainsworth; Julien Slagboom; Nessrin Alomran; Davinia Pla; Yasir Alhamdi; Sarah I. King; Fiona M. S. Bolton; José María Gutiérrez; Freek J. Vonk; Cheng-Hock Toh; Juan J. Calvete; Jeroen Kool; Robert A. Harrison; Nicholas R. Casewell. 2018. "The paraspecific neutralisation of snake venom induced coagulopathy by antivenoms." Communications Biology 1, no. 1: 1-14.

Journal article
Published: 01 March 2018 in Journal of Proteomics
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The Brown Treesnake (Boiga irregularis) is an arboreal, nocturnal, rear-fanged venomous snake native to northern and eastern regions of Australia, Papua New Guinea and the Solomon Islands. It was inadvertently introduced onto the island of Guam during the late 1940's to early 1950's, and it has caused massive declines and extirpations of the native bird, lizard, and mammal populations. In the current study, we report the characterization of the venom proteome of an adult and a neonate B. irregularis specimens from Guam by a combination of venom gland transcriptomic and venomic analyses. Venom gland transcriptomic analysis of an adult individual identified toxins belonging to 18 protein families, with three-finger toxin isoforms being the most abundantly expressed transcripts, comprising 94% of all venom protein transcript reads. Transcripts for PIII-metalloproteinases, C-type lectins, cysteine-rich secretory proteins, acetylcholinesterases, natriuretic peptides, ficolins, phospholipase A2 (PLA2) inhibitors, PLA2s, vascular endothelial growth factors, Kunitz-type protease inhibitors, cystatins, phospholipase Bs, cobra venom factors, waprins, SVMP inhibitors, matrix metalloproteinases, and hyaluronidases were also identified, albeit, at very low abundances ranging from 0.05% to 1.7% of the transcriptome. The venom proteomes of neonate and adult B. irregularis were also both overwhelmingly (78 and 84%, respectively) dominated by monomeric and dimeric 3FTxs, followed by moderately abundant (21% (N) and 13% (A)) CRISPs, low abundance (1% (N), 3% (A)) PIII-SVMPs, and very low abundance (< 0.01%) PLA2 and SVMP inhibitors. The differences in relative toxin abundances identified between neonate and adult snakes likely correlates to shifts in prey preference between the two age classes, from nearly-exclusively lizards to lizards, birds and small mammals. Immunoaffinity antivenomics with experimentally designed rabbit anti-Brown Treesnake (anti-BTS) venom IgGs against homologous venom from adult snakes demonstrated that CRISPs, PIII-SVMPs, and 60–70% of 3FTxs were effectively immunocaptured. Western blot analysis showed that all venom proteins were recognized by anti-BTS IgGs, and cross-reactivity with other rear-fanged snake venoms was also observed. Incubation of anti-BTS venom IgGs with crude B. irregularis venom resulted in a significant decrease in proteolytic (SVMP) activity against azocasein. These results provide the first comparative venomic and anti-venomic analysis of neonate and adult B. irregularis from Guam, further highlighting evolutionary trends in venom composition among rear-fanged venomous snakes. The Brown Treesnake (Boiga irregularis) has caused extensive ecological and economic damage to the island of Guam where it has become a classic example of the negative impacts of invasive species. In the current study, we report the first combined transcriptomic and proteomic analysis of B. irregularis venom of Guam origin. The transcriptome of an adult snake contained toxin sequences belonging to 18 protein families, with three-finger toxin (3FTx) isoforms being the most abundant and representing 94% of all venom protein transcript reads. Our bottom-up and top-down venomic analyses confirmed that 3FTxs are the major components of B. irregularis venom, and a comparative analysis of neonate and adult venoms demonstrate a clear ontogenetic shift in toxin abundance, likely driven by dietary variation between the two age classes. Second-generation antivenomics and Western blot analysis using purified anti-Brown Treesnake rabbit serum IgGs (anti-BTS IgGs) showed strong immunoreactivity toward B. irregularis venom. Interestingly, our anti-BTS IgGs did not cross-react with 3FTxs found in several other rear-fanged snake venoms, or against 3FTxs in the venom of the elapid Ophiophagus hannah, indicating that epitopes in these 3FTx molecules are quite distinct.

ACS Style

Davinia Pla; Daniel Petras; Anthony J. Saviola; Cassandra Modahl; Libia Sanz; Alicia Pérez; Elena Juárez; Seth Frietze; Pieter C. Dorrestein; Stephen P. Mackessy; Juan J. Calvete. Transcriptomics-guided bottom-up and top-down venomics of neonate and adult specimens of the arboreal rear-fanged Brown Treesnake, Boiga irregularis, from Guam. Journal of Proteomics 2018, 174, 71 -84.

AMA Style

Davinia Pla, Daniel Petras, Anthony J. Saviola, Cassandra Modahl, Libia Sanz, Alicia Pérez, Elena Juárez, Seth Frietze, Pieter C. Dorrestein, Stephen P. Mackessy, Juan J. Calvete. Transcriptomics-guided bottom-up and top-down venomics of neonate and adult specimens of the arboreal rear-fanged Brown Treesnake, Boiga irregularis, from Guam. Journal of Proteomics. 2018; 174 ():71-84.

Chicago/Turabian Style

Davinia Pla; Daniel Petras; Anthony J. Saviola; Cassandra Modahl; Libia Sanz; Alicia Pérez; Elena Juárez; Seth Frietze; Pieter C. Dorrestein; Stephen P. Mackessy; Juan J. Calvete. 2018. "Transcriptomics-guided bottom-up and top-down venomics of neonate and adult specimens of the arboreal rear-fanged Brown Treesnake, Boiga irregularis, from Guam." Journal of Proteomics 174, no. : 71-84.

Journal article
Published: 22 September 2017 in Plant Biotechnology Journal
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Antivenoms developed from the plasma of hyperimmunized animals are the only effective treatment available against snakebite envenomation but shortage of supply contributes to the high morbidity and mortality toll of this tropical disease. We describe a synthetic biology approach to affordable and cost-effective antivenom production based on plant-made recombinant polyclonal antibodies (termed pluribodies). The strategy takes advantage of virus superinfection exclusion to induce the formation of somatic expression mosaics in agroinfiltrated plants, which enables the expression of complex antibody repertoires in a highly reproducible manner. Pluribodies developed using toxin-binding genetic information captured from peripheral blood lymphocytes of hyperimmunized camels recapitulated the overall binding activity of the immune response. Furthermore, an improved plant-made antivenom (plantivenom) was formulated using an in vitro selected pluribody against Bothrops asper snake venom toxins and has been shown to neutralize a wide range of toxin activities and provide protection against lethal venom doses in mice.

ACS Style

Jose Manuel Julve Parreño; Estefanía Huet; Asun Fernández‐Del‐Carmen; Alvaro Segura; Micol Venturi; Antoni Gandia; Wei‐Song Pan; Irene Albaladejo; Javier Forment; Davinia Pla; Andrés Wigdorovitz; Juan J. Calvete; Carlos Gutiérrez; José María Gutiérrez; Antonio Granell; Diego Orzáez. A synthetic biology approach for consistent production of plant-made recombinant polyclonal antibodies against snake venom toxins. Plant Biotechnology Journal 2017, 16, 727 -736.

AMA Style

Jose Manuel Julve Parreño, Estefanía Huet, Asun Fernández‐Del‐Carmen, Alvaro Segura, Micol Venturi, Antoni Gandia, Wei‐Song Pan, Irene Albaladejo, Javier Forment, Davinia Pla, Andrés Wigdorovitz, Juan J. Calvete, Carlos Gutiérrez, José María Gutiérrez, Antonio Granell, Diego Orzáez. A synthetic biology approach for consistent production of plant-made recombinant polyclonal antibodies against snake venom toxins. Plant Biotechnology Journal. 2017; 16 (3):727-736.

Chicago/Turabian Style

Jose Manuel Julve Parreño; Estefanía Huet; Asun Fernández‐Del‐Carmen; Alvaro Segura; Micol Venturi; Antoni Gandia; Wei‐Song Pan; Irene Albaladejo; Javier Forment; Davinia Pla; Andrés Wigdorovitz; Juan J. Calvete; Carlos Gutiérrez; José María Gutiérrez; Antonio Granell; Diego Orzáez. 2017. "A synthetic biology approach for consistent production of plant-made recombinant polyclonal antibodies against snake venom toxins." Plant Biotechnology Journal 16, no. 3: 727-736.

Research article
Published: 07 August 2017 in PLOS Neglected Tropical Diseases
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Bothrops, Crotalus and Lachesis represent the most medically relevant genera of pitvipers in Central and South America. Similarity in venom phenotype and physiopathological profile of envenomings caused by the four nominal Lachesis species led us to hypothesize that an antivenom prepared against venom from any of them may exhibit paraspecificity against all the other congeneric taxa. To assess this hypothesis, in this work we have applied antivenomics and immunochemical methods to investigate the immunoreactivity of three monovalent antivenoms and two polyvalent antivenoms towards the venoms from different geographic populations of three different Lachesis species. The ability of the antivenoms to neutralize the proteolytic, hemorrhagic, coagulant, and lethal activities of the seven Lachesis venoms was also investigated. A conspicuous pattern of immunorecognition and cross-neutralization for all effects was evident by the polyspecific antivenoms, indicating large immunoreactive epitope conservation across the genus during more than 10 million years since the Central and South American bushmasters diverged. Despite the broad geographic distribution of Lachesis, antivenoms against venoms of different species are effective in the neutralization of congeneric venoms not used in the immunization mixture, indicating that they can be used equivalently for the clinical treatment of any lachesic envenoming. This study demonstrates that antivenoms raised against venom of different Lachesis species are indistinctly effective in the neutralization of congeneric venoms not used in the immunization mixture, indicating that antivenoms against conspecific venoms may be used equivalently for the clinical treatment of envenomings caused by any bushmaster species. Snakebite envenoming is a neglected public health problem in many developing countries and antivenom administration constitutes the mainstay in the treatment of such envenomings. Therapeutic antivenoms contain animal-derived antibodies against venom toxins and are produced by immunizing animals with the venom from one or several snake species from a defined geographical area. Defining the geographic boundaries of the efficiency of an antivenom therefore has implications for its rational and efficient use. In Central and South America most accidents are caused by pitvipers of the genus Bothrops, Crotalus and Lachesis. There are four Lachesis species distributed in a variety of habitats ranging from the Caribbean coast of Central America to the Atlantic rainforest of Brazil. Lachesis species cause severe envenomings in humans due to the toxicity of their venoms and also to the large amount of venom they inject into their victims. In this work we investigate the capability of several antivenoms to neutralize the toxic activities of a panel of Lachesis venoms. The results demonstrate that antivenoms raised by immunizing horses with the venoms of different Lachesis species are effective at neutralizing congeneric venoms not used in the immunization, indicating that they could be used equivalently for the clinical treatment of any lachesic envenoming.

ACS Style

Marvin Madrigal; Davinia Pla; Libia Sanz; Elexandra Barboza; Cynthia Arroyo-Portilla; Carlos Corrêa-Netto; José María Gutiérrez; Alberto Alape-Girón; Marietta Flores-Díaz; Juan J. Calvete. Cross-reactivity, antivenomics, and neutralization of toxic activities of Lachesis venoms by polyspecific and monospecific antivenoms. PLOS Neglected Tropical Diseases 2017, 11, e0005793 .

AMA Style

Marvin Madrigal, Davinia Pla, Libia Sanz, Elexandra Barboza, Cynthia Arroyo-Portilla, Carlos Corrêa-Netto, José María Gutiérrez, Alberto Alape-Girón, Marietta Flores-Díaz, Juan J. Calvete. Cross-reactivity, antivenomics, and neutralization of toxic activities of Lachesis venoms by polyspecific and monospecific antivenoms. PLOS Neglected Tropical Diseases. 2017; 11 (8):e0005793.

Chicago/Turabian Style

Marvin Madrigal; Davinia Pla; Libia Sanz; Elexandra Barboza; Cynthia Arroyo-Portilla; Carlos Corrêa-Netto; José María Gutiérrez; Alberto Alape-Girón; Marietta Flores-Díaz; Juan J. Calvete. 2017. "Cross-reactivity, antivenomics, and neutralization of toxic activities of Lachesis venoms by polyspecific and monospecific antivenoms." PLOS Neglected Tropical Diseases 11, no. 8: e0005793.

Original research article
Published: 24 July 2017 in Frontiers in Plant Science
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Drought is one of the main constraints determining forest species growth, survival and productivity, and therefore one of the main limitations for reforestation or afforestation. The aim of this study is to characterize the drought response at the physiological and molecular level of different Pinus halepensis (common name Aleppo pine) seed sources, previously characterized in field trials as drought-sensitive or drought-tolerant. This approach aims to identify different traits capable of predicting the ability of formerly uncharacterized seedlings to cope with drought stress. Gas-exchange, water potential, photosynthetic pigments, soluble sugars, free amino acids, glutathione and proteomic analyses were carried out on control and drought-stressed seedlings in greenhouse conditions. Gas-exchange determinations were also assessed in field-planted seedlings in order to validate the greenhouse experimental conditions. Drought-tolerant seed sources presented higher values of photosynthetic rates, water use efficiency, photosynthetic pigments and soluble carbohydrates concentrations. We observed the same pattern of variation of photosynthesis rate and maximal efficiency of PSII in field. Interestingly drought-tolerant seed sources exhibited increased levels of glutathione, methionine and cysteine. The proteomic profile of drought tolerant seedlings identified two heat shock proteins and an enzyme related to methionine biosynthesis that were not present in drought sensitive seedlings, pointing to the synthesis of sulphur amino acids as a limiting factor for drought tolerance in Pinus halepensis. Our results established physiological and molecular traits useful as distinctive markers to predict drought tolerance in Pinus halepensis provenances that could be reliably used in reforestation programs in drought prone areas.

ACS Style

Khaled Taïbi; Antonio del Campo; Alberto Vilagrosa; José M. Bellés; María Pilar López-Gresa; Davinia Pla; Juan J. Calvete; José M. López-Nicolás; José M. Mulet. Drought Tolerance in Pinus halepensis Seed Sources As Identified by Distinctive Physiological and Molecular Markers. Frontiers in Plant Science 2017, 8, 1 .

AMA Style

Khaled Taïbi, Antonio del Campo, Alberto Vilagrosa, José M. Bellés, María Pilar López-Gresa, Davinia Pla, Juan J. Calvete, José M. López-Nicolás, José M. Mulet. Drought Tolerance in Pinus halepensis Seed Sources As Identified by Distinctive Physiological and Molecular Markers. Frontiers in Plant Science. 2017; 8 ():1.

Chicago/Turabian Style

Khaled Taïbi; Antonio del Campo; Alberto Vilagrosa; José M. Bellés; María Pilar López-Gresa; Davinia Pla; Juan J. Calvete; José M. López-Nicolás; José M. Mulet. 2017. "Drought Tolerance in Pinus halepensis Seed Sources As Identified by Distinctive Physiological and Molecular Markers." Frontiers in Plant Science 8, no. : 1.

Review
Published: 13 May 2017 in Toxins
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Animal-derived antivenoms constitute the mainstay in the therapy of snakebite envenoming. The efficacy of antivenoms to neutralize toxicity of medically-relevant snake venoms has to be demonstrated through meticulous preclinical testing before their introduction into the clinical setting. The gold standard in the preclinical assessment and quality control of antivenoms is the neutralization of venom-induced lethality. In addition, depending on the pathophysiological profile of snake venoms, the neutralization of other toxic activities has to be evaluated, such as hemorrhagic, myotoxic, edema-forming, dermonecrotic, in vitro coagulant, and defibrinogenating effects. There is a need to develop laboratory assays to evaluate neutralization of other relevant venom activities. The concept of the 3Rs (Replacement, Reduction, and Refinement) in Toxinology is of utmost importance, and some advances have been performed in their implementation. A significant leap forward in the study of the immunological reactivity of antivenoms against venoms has been the development of “antivenomics”, which brings the analytical power of mass spectrometry to the evaluation of antivenoms. International partnerships are required to assess the preclinical efficacy of antivenoms against snake venoms in different regions of the world in order to have a detailed knowledge on the neutralizing profile of these immunotherapeutics.

ACS Style

José María Gutiérrez; Gabriela Solano; Davinia Pla; María Herrera; Álvaro Segura; Mariángela Vargas; Mauren Villalta; Andrés Sánchez; Libia Sanz; Bruno Lomonte; Guillermo León; Juan J. Calvete. Preclinical Evaluation of the Efficacy of Antivenoms for Snakebite Envenoming: State-of-the-Art and Challenges Ahead. Toxins 2017, 9, 163 .

AMA Style

José María Gutiérrez, Gabriela Solano, Davinia Pla, María Herrera, Álvaro Segura, Mariángela Vargas, Mauren Villalta, Andrés Sánchez, Libia Sanz, Bruno Lomonte, Guillermo León, Juan J. Calvete. Preclinical Evaluation of the Efficacy of Antivenoms for Snakebite Envenoming: State-of-the-Art and Challenges Ahead. Toxins. 2017; 9 (5):163.

Chicago/Turabian Style

José María Gutiérrez; Gabriela Solano; Davinia Pla; María Herrera; Álvaro Segura; Mariángela Vargas; Mauren Villalta; Andrés Sánchez; Libia Sanz; Bruno Lomonte; Guillermo León; Juan J. Calvete. 2017. "Preclinical Evaluation of the Efficacy of Antivenoms for Snakebite Envenoming: State-of-the-Art and Challenges Ahead." Toxins 9, no. 5: 163.

Article
Published: 10 May 2017 in Toxins
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Second generation antivenomics is a translational venomics approach designed to complement in vivo preclinical neutralization assays. It provides qualitative and quantitative information on the set of homologous and heterologous venom proteins presenting antivenom-recognized epitopes and those exhibiting impaired immunoreactivity. In a situation of worrying antivenom shortage in many tropical and sub-tropical regions with high snakebite mortality and morbidity rates, such knowledge has the potential to facilitate the optimal deployment of currently existing antivenoms and to aid in the rational design of novel broad specificity antidotes. The aim of the present work was to expand the analytical capability of the immunoaffinity second-generation antivenomics platform, endowing it with the ability to determine the maximal binding capacity of an antivenom toward the different toxins present in a venom, and to quantify the fraction of venom-specific antibodies present in a given antivenom. The application of this new platform, termed third generation (3G) antivenomics, in the preclinical evaluation of antivenoms is illustrated in this paper for the case of antivenom EchiTAb-Plus-ICP® reactivity towards the toxins of homologous (B. arietans) and heterologous (N. melanoleuca) venoms.

ACS Style

Davinia Pla; Yania Rodríguez; Juan J. Calvete. Third Generation Antivenomics: Pushing the Limits of the In Vitro Preclinical Assessment of Antivenoms. Toxins 2017, 9, 158 .

AMA Style

Davinia Pla, Yania Rodríguez, Juan J. Calvete. Third Generation Antivenomics: Pushing the Limits of the In Vitro Preclinical Assessment of Antivenoms. Toxins. 2017; 9 (5):158.

Chicago/Turabian Style

Davinia Pla; Yania Rodríguez; Juan J. Calvete. 2017. "Third Generation Antivenomics: Pushing the Limits of the In Vitro Preclinical Assessment of Antivenoms." Toxins 9, no. 5: 158.

Journal article
Published: 01 March 2017 in Comparative Biochemistry and Physiology Part D: Genomics and Proteomics
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The high egg-laying capacity of the modern domestic chicken (i.e. White Leghorn, WL) has arisen from the low egg-laying ancestor Red Junglefowl (RJF) via continuous trait selection and breeding. To investigate whether this long-term selection impacted the seminal fluid (SF)-proteome, 2DE electrophoresis-based proteomic analyses and immunoassays were conducted to map SF-proteins/cytokines in RJF, WL and a 9th generation Advanced Intercross Line (AIL) of RJF/WL-L13, including individual SF (n = 4, from each RJF, WL and AIL groups) and pools of the SF from 15 males of each group, analyzed by 2DE to determine their degree of intra-group (AIL, WL, and RJF) variability using Principal Component Analysis (PCA); respectively an inter-breed comparative analysis of intergroup fold change of specific SF protein spots intensity between breeds. The PCA clearly highlighted a clear intra-group similarity among individual roosters as well as a clear inter-group variability (e.g. between RJF, WL and AIL) validating the use of pools to minimize confounding individual variation. Protein expression varied considerably for processes related to sperm motility, nutrition, transport and survival in the female, including signaling towards immunomodulation. The major conserved SF-proteins were serum albumin and ovotransferrin. Aspartate aminotransferase, annexin A5, arginosuccinate synthase, glutathione S-transferase 2 and l-lactate dehydrogenase-A were RJF-specific. Glyceraldehyde-3-phosphate dehydrogenase appeared specific to the WL-SF while angiotensin-converting enzyme, γ-enolase, coagulation factor IX, fibrinogen α-chain, hemoglobin subunit α-D, lysozyme C, phosphoglycerate kinase, Src-substrate protein p85, tubulins and thioredoxin were AIL-specific. The RJF-SF contained fewer immune system process proteins and lower amounts of the anti-inflammatory/immunomodulatory TGF-β2 compared to WL and AIL, which had low levels- or lacked pro-inflammatory CXCL10 compared to RJF. The seminal fluid proteome differs between ancestor and modern chicken, with a clear enrichment of proteins and peptides related to immune-modulation for sperm survival in the female and fertility.Funding agencies: Research Council FORMAS, Stockholm, Sweden [221-2011-512]; Ministerio de Ciencia e Innovacion (Madrid, Spain) [BFU2013-42833-P]

ACS Style

Mohammad Atikuzzaman; Libia Sanz; Davinia Pla; Manuel Alvarez-Rodriguez; Marie Rubér; Dominic Wright; Juan J. Calvete; Heriberto Rodriguez-Martinez. Selection for higher fertility reflects in the seminal fluid proteome of modern domestic chicken. Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 2017, 21, 27 -40.

AMA Style

Mohammad Atikuzzaman, Libia Sanz, Davinia Pla, Manuel Alvarez-Rodriguez, Marie Rubér, Dominic Wright, Juan J. Calvete, Heriberto Rodriguez-Martinez. Selection for higher fertility reflects in the seminal fluid proteome of modern domestic chicken. Comparative Biochemistry and Physiology Part D: Genomics and Proteomics. 2017; 21 ():27-40.

Chicago/Turabian Style

Mohammad Atikuzzaman; Libia Sanz; Davinia Pla; Manuel Alvarez-Rodriguez; Marie Rubér; Dominic Wright; Juan J. Calvete; Heriberto Rodriguez-Martinez. 2017. "Selection for higher fertility reflects in the seminal fluid proteome of modern domestic chicken." Comparative Biochemistry and Physiology Part D: Genomics and Proteomics 21, no. : 27-40.

Journal article
Published: 24 January 2017 in Biochimica et Biophysica Acta (BBA) - General Subjects
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Non-front-fanged colubroid snakes comprise about two-thirds of extant ophidian species. The medical significance of the majority of these snakes is unknown, but at least five species have caused life-threatening or fatal human envenomings. However, the venoms of only a small number of species have been explored. A combined venomic and venom gland transcriptomic approach was employed to characterise of venom of Dispholidus typus (boomslang), the snake that caused the tragic death of Professor Karl Patterson Schmidt. The ability of CroFab™ antivenom to immunocapture boomslang venom proteins was investigated using antivenomics. Transcriptomic-assisted proteomic analysis identified venom proteins belonging to seven protein families: three-finger toxin (3FTx); phospholipase A2 (PLA2); cysteine-rich secretory proteins (CRISP); snake venom (SV) serine proteinase (SP); C-type lectin-like (CTL); SV metalloproteinases (SVMPs); and disintegrin-like/cysteine-rich (DC) proteolytic fragments. CroFab™ antivenom efficiently immunodepleted some boomslang SVMPs. The present work is the first to address the overall proteomic profile of D. typus venom. This study allowed us to correlate the toxin composition with the toxic activities of the venom. The antivenomic analysis suggested that the antivenom available at the time of the unfortunate accident could have exhibited at least some immunoreactivity against the boomslang SVMPs responsible for the disseminated intravascular coagulation syndrome that caused K.P. Schmidt's fatal outcome. This study may stimulate further research on other non-front-fanged colubroid snake venoms capable of causing life-threatening envenomings to humans, which in turn should contribute to prevent fatal human accidents, such as that unfortunately suffered by K.P. Schmidt.

ACS Style

Davinia Pla; Libia Sanz; Gareth Whiteley; Simon Wagstaff; Robert Harrison; Nicholas R. Casewell; Juan J. Calvete. What killed Karl Patterson Schmidt? Combined venom gland transcriptomic, venomic and antivenomic analysis of the South African green tree snake (the boomslang), Dispholidus typus. Biochimica et Biophysica Acta (BBA) - General Subjects 2017, 1861, 814 -823.

AMA Style

Davinia Pla, Libia Sanz, Gareth Whiteley, Simon Wagstaff, Robert Harrison, Nicholas R. Casewell, Juan J. Calvete. What killed Karl Patterson Schmidt? Combined venom gland transcriptomic, venomic and antivenomic analysis of the South African green tree snake (the boomslang), Dispholidus typus. Biochimica et Biophysica Acta (BBA) - General Subjects. 2017; 1861 (4):814-823.

Chicago/Turabian Style

Davinia Pla; Libia Sanz; Gareth Whiteley; Simon Wagstaff; Robert Harrison; Nicholas R. Casewell; Juan J. Calvete. 2017. "What killed Karl Patterson Schmidt? Combined venom gland transcriptomic, venomic and antivenomic analysis of the South African green tree snake (the boomslang), Dispholidus typus." Biochimica et Biophysica Acta (BBA) - General Subjects 1861, no. 4: 814-823.

Journal article
Published: 01 January 2017 in Journal of Proteomics
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Bothriechis is a genus of eleven currently recognized slender and arboreal venomous snakes, commonly called palm-pitvipers that range from southern Mexico to northern South America. Despite dietary studies suggesting that palm-pitvipers are generalists with an ontogenetic shift toward endothermic prey, venom proteomic analyses have revealed remarkable divergence between the venoms of the Costa Rican species, B. lateralis, B. schlegelii, B. supraciliaris, and B. nigroviridis. To achieve a more complete picture of the venomic landscape across Bothriechis, the venom proteomes of biodiversity of the northern Middle American highland palm-pitvipers, B. thalassinus, B. aurifer, and B. bicolor from Guatemala, B. marchi from Honduras, and neonate Costa Rican B. lateralis and B. schlegelii, were investigated. B. thalassinus and B. aurifer venoms are comprised by similar toxin arsenals dominated by SVMPs (33-39% of the venom proteome), CTLs (11-16%), BPP-like molecules (10-13%), and CRISPs (5-10%), and are characterized by the absence of PLA2 proteins. Conversely, the predominant (35%) components of B. bicolor are D49-PLA2 molecules. The venom proteome of B. marchi is similar to B. aurifer and B. thalassinus in that it is rich in SVMPs and BPPs, but also contains appreciable amounts (14.3%) of PLA2s. The major toxin family found in the venoms of both neonate B. lateralis and B. schlegelii, is serine proteinase (SVSP), comprising about 20% of their toxin arsenals. The venom of neonate B. schlegelii is the only palm-pitviper venom where relative high amounts of Kunitz-type (6.3%) and γPLA2 (5.2%) inhibitors have been identified. Despite notable differences between their proteomes, neonate venoms are more similar to each other than to adults of their respective species. However, the ontogenetic changes taking place in the venom of B. lateralis strongly differ from those that occur in the venom of B. schlegelii. Thus, the ontogenetic change in B. lateralis produces a SVMP-rich venom, whereas in B. schlegelii the age-dependent compositional shift generates a PLA2-rich venom. Overall, genus-wide venomics illustrate the high evolvability of palm-pitviper venoms. The integration of the pattern of venom variation across Bothriechis into a phylogenetic and biogeographic framework may lay the foundation for assessing, in future studies, the evolutionary path that led to the present-day variability of the venoms of palm-pitvipers.Bothriechis represents a monophyletic basal genus of eleven arboreal palm-pitvipers that range from southern Mexico to northern South America. Despite palm-pitvipers' putative status as diet generalists, previous proteomic analyses have revealed remarkable divergence between the venoms of Costa Rican species, B. lateralis, B. schlegelii, B. supraciliaris, and B. nigroviridis. Our current proteomic study of Guatemalan species, B. thalassinus, B. aurifer, and B. bicolor, Honduran B. marchi, and neonate B. lateralis and B. schlegelii from Costa Rica was undertaken to deepen our understanding of the evolutionary pattern of venom proteome diversity across Bothriechis. Ancestral characters are often, but not always, preserved in an organism's development. Venoms of neonate B. lateralis and B. schlegelii are more similar to each other than to adults of their respective species, suggesting that the high evolvability of palm-pitviper venoms may represent an inherent feature of Bothriechis common ancestor. Our genus-wide data identified four nodes of venom phenotype differentiation across the phylogeny of Bothriechis. Integrated into a phylogenetic and biogeographic framework, the pattern of venom variation across Bothriechis may lay the groundwork to establish whether divergence was driven by selection for efficient resource exploitation in arboreal 'islands', thereby contributing to the ecological speciation of the genus.

ACS Style

Davinia Pla; Libia Sanz; Mahmood Sasa; Manuel E. Acevedo; Quetzal Dwyer; Jordi Durban; Alicia Pérez; Yania Rodriguez; Bruno Lomonte; Juan J. Calvete. Proteomic analysis of venom variability and ontogeny across the arboreal palm-pitvipers (genus Bothriechis ). Journal of Proteomics 2017, 152, 1 -12.

AMA Style

Davinia Pla, Libia Sanz, Mahmood Sasa, Manuel E. Acevedo, Quetzal Dwyer, Jordi Durban, Alicia Pérez, Yania Rodriguez, Bruno Lomonte, Juan J. Calvete. Proteomic analysis of venom variability and ontogeny across the arboreal palm-pitvipers (genus Bothriechis ). Journal of Proteomics. 2017; 152 ():1-12.

Chicago/Turabian Style

Davinia Pla; Libia Sanz; Mahmood Sasa; Manuel E. Acevedo; Quetzal Dwyer; Jordi Durban; Alicia Pérez; Yania Rodriguez; Bruno Lomonte; Juan J. Calvete. 2017. "Proteomic analysis of venom variability and ontogeny across the arboreal palm-pitvipers (genus Bothriechis )." Journal of Proteomics 152, no. : 1-12.

Journal article
Published: 01 January 2017 in Journal of Proteomics
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The Papuan black snake (Pseudechis papuanus Serpentes: Elapidae) is endemic to Papua New Guinea, Indonesian Papua and Australia's Torres Strait Islands. We have investigated the biological activity and proteomic composition of its venom. The P. papuanus venom proteome is dominated by a variety (n≥18) of PLA2s, which together account for ~90% of the venom proteins, and a set of low relative abundance proteins, including a short-neurotoxic 3FTx (3.1%), 3-4 PIII-SVMPs (2.8%), 3 cysteine-rich secretory proteins (CRISP; 2.3%) 1-3 l-amino acid oxidase (LAAO) molecules (1.6%). Probing of a P. papuanus cDNA library with specific primers resulted in the elucidation of the full-length nucleotide sequences of six new toxins, including vespryn and NGF not found in the venom proteome, and a calglandulin protein involved in toxin expression with the venom glands. Intravenous injection of P. papuanus venom in mice induced lethality, intravascular haemolysis, pulmonary congestion and oedema, and anticoagulation after intravenous injection, and these effects are mainly due to the action of PLA2s. This study also evaluated the in vivo preclinical efficacy of Australian black snake and polyvalent Seqirus antivenoms. These antivenoms were effective in neutralising the lethal, PLA2 and anticoagulant activities of P. papuanus venom in mice. On the other hand, all of the Seqirus antivenoms tested using an antivenomic approach exhibited strong immunorecognition of all the venom components. These preclinical results suggest that Australian Seqirus1 antivenoms may provide paraspecific protection against P. papuanus venom in humans. The toxicological profile and proteomic composition of the venom of the Papuan black snake, Pseudechis papuanus, a large diurnal snake endemic to the southern coast of New Guinea and a handful of close offshore islands, were investigated. Intravenous injection of P. papuanus venom in mice induced intravascular hemolysis, pulmonary congestion and edema, anticoagulation, and death. These activities could be assigned to the set of PLA2 molecules, which dominate the P. papuanus venom proteome. This study also showed that Australian Seqirus black snake or polyvalent antivenoms were effective in neutralising the lethal, PLA2 and anticoagulant activities of the venom. These preclinical results support the continued recommendation of these Seqirus antivenoms in the clinical management of P. papuanus envenoming in Australia, Papua New Guinea or Indonesian Papua Province.

ACS Style

Davinia Pla; Benjamin W. Bande; Ronelle E. Welton; Owen K. Paiva; Libia Sanz; Álvaro Segura; Christine Wright; Juan J. Calvete; José María Gutiérrez; David J. Williams. Proteomics and antivenomics of Papuan black snake ( Pseudechis papuanus ) venom with analysis of its toxicological profile and the preclinical efficacy of Australian antivenoms. Journal of Proteomics 2017, 150, 201 -215.

AMA Style

Davinia Pla, Benjamin W. Bande, Ronelle E. Welton, Owen K. Paiva, Libia Sanz, Álvaro Segura, Christine Wright, Juan J. Calvete, José María Gutiérrez, David J. Williams. Proteomics and antivenomics of Papuan black snake ( Pseudechis papuanus ) venom with analysis of its toxicological profile and the preclinical efficacy of Australian antivenoms. Journal of Proteomics. 2017; 150 ():201-215.

Chicago/Turabian Style

Davinia Pla; Benjamin W. Bande; Ronelle E. Welton; Owen K. Paiva; Libia Sanz; Álvaro Segura; Christine Wright; Juan J. Calvete; José María Gutiérrez; David J. Williams. 2017. "Proteomics and antivenomics of Papuan black snake ( Pseudechis papuanus ) venom with analysis of its toxicological profile and the preclinical efficacy of Australian antivenoms." Journal of Proteomics 150, no. : 201-215.

Review
Published: 01 November 2016 in Toxicon
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The application of proteomic tools to the study of snake venoms has led to an impressive growth in the knowledge about their composition (venomics), immunogenicity (antivenomics), and toxicity (toxicovenomics). About one-third of all venomic studies have focused on elapid species, especially those of the Old World. The New World elapids, represented by coral snakes, have been less studied. In recent years, however, a number of venomic studies on Micrurus species from North, Central, and South America have been conducted. An overview of these studies is presented, highlighting the emergence of some patterns and trends concerning their compositional, functional, and immunological characteristics. Results gathered to date, encompassing 18 out of the approximately 85 species of Micrurus, reveal a dichotomy of venom phenotypes regarding the relative abundance of the omnipresent phospholipases A2 (PLA2) and 'three-finger' toxins (3FTx): a group of species express a PLA2-predominant venom composition, while others display a 3FTx-predominant compositional pattern. These two divergent toxin expression phenotypes appear to be related to phylogenetic positions and geographical distributions along a North-South axis in the Americas, but further studies encompassing a higher number of species are needed to assess these hypotheses. The two contrasting phenotypes also show correlations with some toxic functionalities, complexity in the diversity of proteoforms, and immunological cross-recognition patterns. The biological significance for the emergence of a dichotomy of venom compositions within Micrurus, in some cases observed even among sympatric species that inhabit relatively small geographic areas, represents a puzzling and challenging area of research which warrants further studies.

ACS Style

Bruno Lomonte; Paola Rey Suarez; Julián Fernández; Mahmood Sasa; Davinia Pla; Nancy Vargas; Melisa Benard Valle; Libia Sanz; Carlos Corrêa-Netto; Vitelbina Núñez; Alberto Alape-Girón; Alejandro Alagón; José María Gutiérrez; Juan J. Calvete. Venoms of Micrurus coral snakes: Evolutionary trends in compositional patterns emerging from proteomic analyses. Toxicon 2016, 122, 7 -25.

AMA Style

Bruno Lomonte, Paola Rey Suarez, Julián Fernández, Mahmood Sasa, Davinia Pla, Nancy Vargas, Melisa Benard Valle, Libia Sanz, Carlos Corrêa-Netto, Vitelbina Núñez, Alberto Alape-Girón, Alejandro Alagón, José María Gutiérrez, Juan J. Calvete. Venoms of Micrurus coral snakes: Evolutionary trends in compositional patterns emerging from proteomic analyses. Toxicon. 2016; 122 ():7-25.

Chicago/Turabian Style

Bruno Lomonte; Paola Rey Suarez; Julián Fernández; Mahmood Sasa; Davinia Pla; Nancy Vargas; Melisa Benard Valle; Libia Sanz; Carlos Corrêa-Netto; Vitelbina Núñez; Alberto Alape-Girón; Alejandro Alagón; José María Gutiérrez; Juan J. Calvete. 2016. "Venoms of Micrurus coral snakes: Evolutionary trends in compositional patterns emerging from proteomic analyses." Toxicon 122, no. : 7-25.