Molecular characterization and phylogenetic analysis of Cyprinid herpesvirus 3 genotypes in Iran from 2020 to 2023

The aquaculture industry in Iran contributed to about 1% of the world's aquaculture production in 2020 with a volume of 0.7 million tons. A targeted approach was used to identify positive samples by collecting samples from 342 suspected carp farms showing signs of disease. The carp species was considered as the host for the viruses under investigation and Khuzestan, Mazandaran and Gilan provinces were selected for sampling. A total of 251 farms in Gilan, 68 farms in Mazandaran, and 23 farms in Khuzestan provinces were sampled. Cyprinid herpesvirus 3 (CyHV-3) was characterized by a combination of sequence analysis and duplex PCR. Genetic analyzes and phylogenetic tree construction were performed using MEGA7 software. Of the 342 farms sampled, 85 were infected with koi herpes virus (KHV). Asian 1 and Asian 2 genotypes were identified by sequence analysis of the SphI-5 and TK gene regions. One of the positive samples showed a match in all motif positions within the TK gene, specifically genotype A1, except for positions 814 - 813 where they had the sequence AT, which was a rare exception. Duplex PCR analysis of two variable marker regions between ORF29 and ORF30 (marker I) and ORF133 and its upstream region (marker II) revealed viruses of genotype J (I⁺⁺ II ⁺), an intermediate genotype (I⁺⁺ II ^-), and a new genotype, I⁺⁺ II ^-*, identified in viruses from different farms. This new genotype retains the I⁺⁺ allele of marker I and has a 5-bp deletion in the marker II. The global distribution of CyHV-3 genotypes is not yet fully elucidated. Results indicate the high degree of diversification of CyHV-3 in the West Asian regions, where at least three different genotypes (I⁺⁺ II ⁺, I⁺⁺ II ^-, and I⁺⁺ II ^-Δ) currently appear to circulate.