Genetic analysis of NS5B gene from bovine viral diarrhea virus-infected cattle
Abstract Background and Aim: A previous study divided Indonesian bovine viral diarrhea virus (BVDV)-1 into subgenotypes BVDV-1a to BVDV-1d based on the partial NS5B gene using strain Bega as reference for BVDV-1a. In fact, it is clustered into BVDV-1c with strain Bega-like Australia. BVDV genotyping has been done on isolates from Jakarta, West and Central Java, but East Java isolates have not been genotyped. This study aimed to analyze genetic variability and amino acid residues in the nucleotide-binding pocket of the NS5B gene from infected cattle. Materials and Methods: Samples were obtained from the Sera Bank originating from active and passive surveillance of cattle that had been tested for BVDV antigen from 2013 to 2017. Detection of the p80 antibody and BVDV genotyping was carried out using ELISA and nested-multiplex-polymerase chain reaction (PCR), respectively. We defined 15 nested PCR products for partial sequencing of NS5B. Those field samples were selected from each location and year using proportional calculation as a representative sample. Homological and phylogenetic analyses of the partial NS5B gene were performed using BLAST and MEGA version 6. Results: Based on the phylogenetic tree analysis using 360 nucleotides as the partial NS5B gene, Indonesian BVDV-1 isolates from Central and East Java were subdivided to BVDV-1a (n=9), BVDV-1b (n=1), and BVDV-1c (n=5). In the present study, the homology of BVDV subgenotype -1a, -1b, and -1c was compared to the BVDV GenBank data and found 90-93%, 93%, and 92-95% respectively with the average pairwise distance of 0.207. A point mutation was shown at R283K of all BVDV isolates based on the sequence of three amino acid residues R283, R285, and I287 in the nucleotide-binding pocket as a part of the encoded RNA-dependent RNA polymerase. Conclusion: This study revealed the genetic variability of BVDV infecting cattle in Central Java and East Java, Indonesia, the subtypes BVDV-1a, BVDV-1b, BVDV-1c, and a point mutation at the R283K residue. Keywords: bovine viral diarrhea virus, NS5B gene, phylogenetic analysis, point mutation, subgenotype.