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Novel genomic targets for proper subtyping of bovine viral diarrhea virus 1 (BVDV-1) and BVDV-2

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Abstract

Whole-genome phylogenetic analysis, the most suitable strategy for subtyping bovine viral diarrhea virus 1 (BVDV-1) and BVDV-2, is not feasible for many laboratories. Consequently, BVDV isolates/strains have been frequently subtyped based on analysis of single genomic regions, mainly the 5’ untranslated region (UTR). This approach, however, may lead to inaccurate and/or poorly statistically supported viral classification. Herein, we describe novel primer sets whose amplicons may be easily sequenced and used for BVDV subtyping. Initially, genomic regions previously described as the most suitable targets for BVDV subtyping were analyzed for design of high-coverage primers. The putative amplicons were analyzed in silico for their suitability to reproduce the phylogenetic classification of 118 BVDV-1 and 88 BVDV-2 complete/near-complete genomes (CNCGs) (GenBank). This analysis was also performed considering the region amplifiable by primers HCV90-368, 324-326 and BP189-389 (5’UTR), which have been used for BVDV diagnosis and/or classification. After confirming the agreement between the analyses of our primers’ amplicon versus the CNCGs, we optimized the RT-PCRs and evaluated their performance for amplification of BVDV isolates/strains (n = 35 for BVDV-1; n = 33 for BVDV-2). Among the potential targets for BVDV subtyping, we designed high-coverage primers for NS3-NS4A (BVDV-1) (526 bp amplicon) and NS5B (BVDV-2) (728 bp). The classification based on these regions fully reproduced the subtyping of all CNCGs. On the other hand, subtyping based on the putative amplicons from primers HCV90-368, 324-326 and BP189-389 showed disagreements in relation the CNCG analysis. The NS3-NS4A and NS5B primers also allowed the amplification of all BVDV isolates/strains tested. Finally, we suggest the use of these primers in future phylogenetic and epidemiological studies of BVDVs.

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Data availability

Datasets generated and/or analyzed during the current study will be available on GenBank and may also be made available by the corresponding author upon reasonable request.

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Acknowledgements

CIM and PSBO thank the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) (Brazil) and Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), respectively, for the scholarship. JVJSJr was supported by Financiadora de Estudos e Projetos (FINEP) DTI-A-1. EFF (process 301414/2010-6) and RW (process 305867/2018-0) were supported by CNPq research fellowships. The research was also supported partially by Coordenação de Aperfeiçocamento de Pessoal de Nível Superior (CAPES) (Brazil), finance code 001.

Funding

This study was partially funded by Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) (Brazil), finance code 001.

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Authors and Affiliations

  1. Setor de Virologia, Departamento de Medicina Veterinária Preventiva, Universidade Federal de Santa Maria, Santa Maria, Av. Roraima, 1000, Prédio 63A, Camobi, Santa Maria, Rio Grande do Sul, 97105-900, Brazil

    Carolina Isabela Mucellini, José Valter Joaquim Silva Júnior, Pablo Sebastian Britto de Oliveira, Rudi Weiblen & Eduardo Furtado Flores

  2. Programa de Pós-graduação em Medicina Veterinária, Universidade Federal de Santa Maria, Santa Maria, Rio Grande do Sul, Brazil

    Carolina Isabela Mucellini & Pablo Sebastian Britto de Oliveira

  3. Setor de Virologia, Laboratório de Imunopatologia Keizo Asami, Universidade Federal de Pernambuco, Recife, Pernambuco, Brazil

    José Valter Joaquim Silva Júnior

  4. Departamento de Análises Clínicas, Centro de Ciências da Saúde, Universidade Federal de Santa Maria, Santa Maria, Rio Grande do Sul, Brazil

    José Valter Joaquim Silva Júnior

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  1. Carolina Isabela Mucellini
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  2. José Valter Joaquim Silva Júnior
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  3. Pablo Sebastian Britto de Oliveira
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  4. Rudi Weiblen
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  5. Eduardo Furtado Flores
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Contributions

CIM, JVJSJr, PSBO, EFF and RW conceived the study design; CIM, JVJSJr and PSBO designed the primer sets; CIM and JVJSJr performed data collection; CIM, JVJSJr and PSBO analyzed the data; CIM and JVJSJr prepared the draft manuscript; EFF and RW critically reviewed the manuscript. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Eduardo Furtado Flores.

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Mucellini, C.I., Silva Júnior, J.V.J., de Oliveira, P.S.B. et al. Novel genomic targets for proper subtyping of bovine viral diarrhea virus 1 (BVDV-1) and BVDV-2. Virus Genes 59, 836–844 (2023). https://doi.org/10.1007/s11262-023-02022-x

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