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Sequence Divergence in Venom Genes Within and Between Montane Pitviper (Viperidae: Crotalinae: Cerrophidion) Species is Driven by Mutation–Drift Equilibrium

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Abstract

Snake venom can vary both among and within species. While some groups of New World pitvipers—such as rattlesnakes—have been well studied, very little is known about the venom of montane pitvipers (Cerrophidion) found across the Mesoamerican highlands. Compared to most well-studied rattlesnakes, which are widely distributed, the isolated montane populations of Cerrophidion may facilitate unique evolutionary trajectories and venom differentiation. Here, we describe the venom gland transcriptomes for populations of C. petlalcalensis, C. tzotzilorum, and C. godmani from Mexico, and a single individual of C. sasai from Costa Rica. We explore gene expression variation in Cerrophidion and sequence evolution of toxins within C. godmani specifically. Cerrophidion venom gland transcriptomes are composed primarily of snake venom metalloproteinases, phospholipase A\(_2\)s (PLA\(_2\)s), and snake venom serine proteases. Cerrophidion petlalcalensis shows little intraspecific variation; however, C. godmani and C. tzotzilorum differ significantly between geographically isolated populations. Interestingly, intraspecific variation was mostly attributed to expression variation as we did not detect signals of selection within C. godmani toxins. Additionally, we found PLA\(_2\)-like myotoxins in all species except C. petlalcalensis, and crotoxin-like PLA\(_2\)s in the southern population of C. godmani. Our results demonstrate significant intraspecific venom variation within C. godmani and C. tzotzilorum. The toxins of C. godmani show little evidence of directional selection where variation in toxin sequence is consistent with evolution under a model of mutation–drift equilibrium. Cerrophidion godmani individuals from the southern population may exhibit neurotoxic venom activity given the presence of crotoxin-like PLA\(_2\)s; however, further research is required to confirm this hypothesis.

Resumen

El veneno de las serpientes puede variar entre y dentro de las especies. Mientras algunos grupos de viperidos del Nuevo Mundo—como las cascabeles—han sido bien estudiadas, muy poco se sabe acerca del veneno de las nauyacas de frío (Cerrophidion) que se encuentran en las zonas altas de Mesoamérica. Comparadas con las extensamente estudiadas cascabeles, que estan ampliamente distribuidas, las poblaciones de Cerrophidion, aisladas en montañas, pueden poseer trayectorias evolutivas y diferenciación en su veneno unicos. En el presente trabajo, describimos el transcriptoma de las glándulas de veneno de poblaciones de C. petlalcalensis, C. tzotzilorum, y C. godmani de México, y un individuo de C. sasai de Costa Rica. Exploramos la variación en la expresión de toxinas en Cerrophidion y la evolución en las secuencias geneticas en C. godmani específicamente. El transcriptoma de la glándula de veneno de Cerrophidion esta compuesto principalmente de Metaloproteinasas de Veneno de Serpiente, Fosfolipasas A\(_2\) (PLA\(_2\)s), y Serin Proteasas de Veneno de Serpiente. Cerrophidion petlalcalensis presenta poca variación intraespecífica; sin embargo, los transcriptomas de la glandula de veneno de C. godmani y C. tzotzilorum difieren significativamente entre poblaciones geográficamente aisladas. Curiosamente, la variación intraespecífica estuvo atribuida principalmente a la expresión de las toxinas ya que no encontramos señales de selección en las toxinas de C. godmani. Adicionalmente, encontramos miotoxinas similares a PLA\(_2\) en todas las especies excepto C. petlalcalensis, y PLA\(_2\)s similares a crotoxina en la población sureña de C. godmani. Nuestros resultados demuestran la presencia de variacion intraespecífica presente en el veneno de C. godmani y C. tzotzilorum. Las toxinas de Cerrophidion godmani muestran poca evidencia de selección direccional, y la variación en la secuencias de las toxinas es consistente con evolucion bajo un modelo de equilibrio de mutación-deriva. Algunos individuos de C. godmani de la población del sur potencialmente tienen un veneno neurotóxico dada la presencia de PLA\(_2\)s similares a la crotoxina, sin embargo, se necesita más evidencia para corroborar esta hipótesis.

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Data Availability

The raw data-sets generated and analyzed during the current study are available in the National Center for Biotechnology Information (NCBI) repository under the accession numbers SRR19227107, SRR19227106, SRR19227101, SRR19227100, SRR19227099, SRR19227098, SRR19227097, SRR19227096, SRR19227095, SRR19227094, SRR19227105, SRR19227104, SRR19227103, and SRR19227102. The consensus transcriptomes generated and analyzed during the current study are available at the github repository https://github.com/RamsesRosales/Cerrophidion_Selection, a similar version is available at the DDBJ/EMBL/GenBank under the accession numbers: C. sasai GJZX00000000, the version described in this paper is the first version, GJZX01000000; C. petlalcalensis GKAV00000000, the version described in this paper is the first version, GKAV01000000; C. godmani GKCA00000000, the version described in this paper is the first version, GKCA01000000. C. tzotzilorum GKIB00000000, the version described in this paper is the first version, GKIB01000000. Scripts utilized in this work can be found in the online resources 3 and 4, and in the github repositories: https://github.com/RamsesRosales/Cerrophidion_Selection, https://github.com/RamsesRosales/ModelCrotA.

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Acknowledgements

We want to thank Jason Jones, Leo Badillo, and Cristobal Moreno for help in the field. We greatly appreciate Tristan Schramer’s assistance with illustrations, and digital photography. Additionally, he and N. Jade Mellor are thanked for the many great discussions and comments on the manuscript. We greatly appreciate Carl Whittington for his comments on the PLA\(_2\) model analysis. Parallel computing resources were provided by the Clemson Palmetto High-Performance Computing Cluster and in part, with support from the Clemson University Genomics and Bioinformatics Facility, which receives support from two Institutional Development Awards (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant numbers P20GM109094 and P20GM139767.

Funding

This work was supported by the National Science Foundation Grants DUE 1161228, DEB 1638879, and DEB 1822417 to C.L.P., DEB 1638902 to D.R.R, and a Fulbright García Robles graduate fellowship to R.A.R.G.

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Authors and Affiliations

  1. Department of Biological Sciences, Clemson University, 190 Collings St., Clemson, SC, 29634, USA

    Ramses Alejandro Rosales-García, Rhett M. Rautsaw, Erich P. Hofmann, Jason L. Strickland, Andrew J. Mason, Matthew L. Holding, Edward A. Myers & Christopher L. Parkinson

  2. Science Department, Cape Fear Community College, Wilmington, NC, 28401, USA

    Erich P. Hofmann

  3. Herp.mx A.C., Colima, Mexico

    Christoph I. Grünwald, Hector Franz-Chavez, Ivan T. Ahumada-Carrillo & Ricardo Ramirez-Chaparro

  4. Biodiversa A. C., Chapala, Jalisco, 45900, Mexico

    Christoph I. Grünwald, Hector Franz-Chavez, Ivan T. Ahumada-Carrillo & Ricardo Ramirez-Chaparro

  5. Instituto Lorancai, Ocotepec, Veracruz, 24105, Mexico

    Miguel Angel de la Torre-Loranca

  6. Department of Biology, University of South Alabama, Mobile, AL, 36688, USA

    Jason L. Strickland

  7. Department of Evolution, Ecology, and Organismal Biology, The Ohio State University, Columbus, OH, 43210, USA

    Andrew J. Mason

  8. Life Sciences Institute, University of Michigan, Ann Arbor, MI, 48109, USA

    Matthew L. Holding

  9. Facultad de Ciencias Biológicas, Universdad Juárez del Estado de Durango, Gómez Palacio, Durango, 35010, Mexico

    Miguel Borja & Gamaliel Castañeda-Gaytan

  10. Centro Investigaciones en Biodiversidad y Ecología Tropical and Instituto Clodomiro Picado, Universidad de Costa Rica, San José, Costa Rica

    Mahmood Sasa

  11. Department of Biological Science, Florida State University, Tallahassee, FL, 32306, USA

    Darin R. Rokyta

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  1. Ramses Alejandro Rosales-García
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Contributions

RARG, EPH, and CLP conceived the study. CIG, HFC, ITAC, RRC, MADL, MB, GCG, MS, MLH, and RMR collected samples used in this study. RMR, EPH, MLH, JLS, AJM, and RARG generated data, RARG, RMR, and EPH processed data and performed analyses. RARG drafted the manuscript. JLS, AJM, DRR, EAM, MLH, and CLP provided analytical and conceptual input. RMR, EAM, EPH, AJM, JLS, MLH, and CLP reviewed and edited the manuscript. All authors reviewed and approved the final manuscript.

Corresponding authors

Correspondence to Ramses Alejandro Rosales-García or Christopher L. Parkinson.

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Ethics Approval

All protocols involving live snakes followed the ASIH guidelines and were approved by the University of Central Florida and Clemson University Animal Care and Use Committees (16-17W: UCF and 2017-067: CU). The sample (LIAP244) was collected in and exported from Costa Rica under Investigation Permit SINAC-ACC-OSJ-re-428-2021. Samples (CHFCB-0271, CHFCB-0272, CHFCB-0274, CHFCB-0290, CHFCB-0291, CHFCB-0300, CHFCB-0236, CHFCB-0238, CHFCB-0239, CHFCB-0276, CHFCB-027, CHFCB-0296, CHFCB-0471) were collected in and exported from Mexico under SEMARNAT:SGPA/DGVS/01090/17;SGPA/DGVS/002288/18;SGPA/DGVS/08831/20.

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Rosales-García, R.A., Rautsaw, R.M., Hofmann, E.P. et al. Sequence Divergence in Venom Genes Within and Between Montane Pitviper (Viperidae: Crotalinae: Cerrophidion) Species is Driven by Mutation–Drift Equilibrium. J Mol Evol 91, 514–535 (2023). https://doi.org/10.1007/s00239-023-10115-2

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