Phylogenetic analysis of Classical swine fever virus from archival formalin fixed clinical tissues reveals vietnamese origin of the isolates

Abstract

Detection of Classical swine fever virus (CSFV) nucleic acid in archival formalin fixed tissue samples and their use for phylogenetic analysis was investigated. Ten samples were examined for the presence of CSFV nucleic acid by reverse transcriptase polymerase chain reaction (RT-PCR) amplification of 5'UTR and E2 gene. RT-PCR was found positive for 5'UTR fragment in eight samples while only one tissue samples showed amplification for E2 gene target fragment. For molecular epidemiology of the disease, 5'UTR PCR product of sample from Darbhanga (Bihar), was cloned and sequenced. The sequence was compared with the sequences available in database. The phylogenetic analysis reveals that the isolate belongs to subgroup 2.2 sharing 98.7% nucleotide identities with Vietnamese isolate (CaTh05-1, AB252170), indicating towards the possible origin of genogroup 2.2 CSFV isolates involved in the outbreak from Vietnam. From the study, it can be concluded that the tissue samples collected and stored in buffer formalin for years can be used to detect CSFV nucleic acid. Results are also suggestive of that the 5'UTR region of genome is more suitable target for RT-PCR based detection of CSFV in archival formalin fixed specimens. The study also indicates the potential of archival formalin fixed tissues for molecular epidemiology and genotyping of the CSF virus.