Characterisation, whole-genome sequencing and phylogenetic analysis of three H3N2 avian influenza viruses isolated from domestic ducks at live poultry markets of Iran, 2017: First report.

BACKGROUND: Avian influenza type A viruses (AIV) can infect a broad range of hosts including human and birds, making them an important viral pathogen with zoonotic potential. Ducks are a known reservoir for many avian viruses including the AIV. OBJECTIVES: To sequence the entire genome of duck-derived H3N2 and ran comprehensive phylogenetic analysis on them to study their origin. METHODS: In this study, 962 cloacal swabs were collected from domestic ducks at several live poultry markets (LPMs) of Gilan, Mazandaran and Golestan provinces of Iran in the year 2017. RESULTS: Preliminary assays such as haemagglutination inhibition assay (HI), Neuraminidase Inhibition assay(NI) and RT-qPCR suggested that 0.5% of the birds were infected by H3 low pathogenic influenza viruses (LPAI). Three isolates were selected for whole genome sequencing. The cleavage site of the HA genes showed a PEKQTR/GLF motif, an indicator of LPAI. Furthermore, BLAST and phylogenetic analyses of the HA gene showed high homology to the Eurasian lineage of H3N8 AIV (95.5%-97.1% to several European and East Asian isolates). However, the NA genes showed high homology (at most 96.5-96.9%) to those belonging to AIV N2 subtype. Furthermore, internal genes showed high homology (96%-98%) to a variety of duck-origin subtypes and glycoprotein combinations, which were different for each segment. This showed a complex reassortment between different subtypes. DISCUSSION: This report is the first whole genome sequencing and complete characterisation of H3N2 AIV from Iran. CONCLUSION: Such surveillance should continue to study the evolution and possible emergence of viruses with pandemic potential.

Full sequence analysis of hemagglutinin and neuraminidase genes and proteins of highly pathogenic avian influenza H5N1 virus detected in Iran, 2015.

Over the last two decades, the highly pathogenic avian influenza H5N1 virus has gained a lot of attention due to its zoonotic and mutative nature. Iran is among the countries significantly affected by the virus as it hosts migratory birds during seasonal migration. In this study, the molecular characterizations of hemagglutinin (HA) and neuraminidase (NA) genes and proteins of H5N1 strain A/chicken/Iran/8/2015 detected in backyard poultry, Mazandaran province, were investigated. Phylogenetic analysis classified this virus as a member of subclade 2.3.2.1c, with the cleavage site motif of "PQRERRRK-R/GLF". HA carried a few mutations altering affinity to mammalian cells; however, the virus was categorized as avian. NA protein had the 20-amino acid deletion at aa position 49-69 similar to those isolated since 2000. Mutations of H253Y and H274Y contributing to antiviral resistance were present in NA. From this analysis, it can be concluded that the wild migratory birds flying from Western Asia to Eastern Africa are probably the main carriers of seasonal H5N1 in the country.

Full-length infectious clone of an Iranian isolate of chicken anemia virus.

An Iranian field strain of chicken anemia virus (CAV), designated IR CAV, was isolated in the Marek's disease virus-transformed lymphoblastoid cell line MDCC-MSB1 (MSB1) culture for the first time. The full-length CAV DNA of this strain was cloned in the bacterial plasmid pTZ57R/T to create the molecular clone pTZ-CAV. The nucleotide and deduced amino acid sequences of viral proteins of IR CAV were compared with those of representative CAV sequences including reference and commercial vaccine strains. IR CAV was not related to vaccine strains and also found to have glutamine at positions 139 and 144 confirming previous studies in which such mutations were associated with a slow rate of virus spread in cell culture. pTZ-CAV was digested with PstI to release IR CAV DNA and then transfected into MSB1 cell by electroporation. The transfected cells showed cytopathic effect similar to virion-initiated infection. One-day old specific pathogen-free chicks were inoculated with the regenerated virus, which had been obtained from transfected MSB1 cells, and compared with the chicks inoculated with IR CAV. Gross lesions in the birds inoculated with the regenerated virus illustrated the infectious nature of the regenerated virus from the cloned IR CAV DNA.

Molecular characterization of the surface glycoprotein genes of highly pathogenic H5N1 avian influenza viruses detected in Iran in 2011.

Highly pathogenic avian influenza (HPAI) H5N1 virus is causing the death of a large number of wild birds and poultry. HPAI H5N1 was reported in the north of Iran in 2011. In this study, two A/Chicken/Iran/271/2011 and A/Duck/Iran/178/2011 viruses were genetically characterized by sequence analysis of Hemagglutinin (HA) and Neuraminidase (NA) genes. Phylogenetic analysis revealed that these viruses were different from previous Iranian isolates (Clade 2.2) and belonged to the subclade 2.3.2.1. The results showed that the detected viruses are almost identical to each other and closely related to HPAI H5N1 strains isolated in Mongolia in 2010. Based on the amino acid sequence analysis, these viruses at their HA cleavage sites contained the multibasic amino acid motif PQRERRRK-R/GLF lacking a lysine residue compared with the previous reports of the same motif. There is also a 20-amino acid deletion (resides 49-69) in the NA stalk similar to other viruses isolated after 2000. It seems that introduction of HPAI H5N1 to Iran might have happened by wild birds from Mongolian origin virus.