Complete genome characterization of Genogroup II norovirus strains from India: Evidence of recombination in ORF2/3 overlap

Noroviruses (NoVs) are considered as important causative agents of non-bacterial acute gastroenteritis, worldwide. The data on NoV genomes, their diversity and evolution from Indian subcontinent are not available to date. The present study describes the characterization of full-length genomes of Indian NoV strains for the first time to establish their phylogenetic and evolutionary relationship with those circulating worldwide. Amplification of full-length genomes of three NoV strains (PC15, PC51 and PC52) was carried out using nine overlapping sets of forward and reverse primers. Full-length genomes of all of the three strains were characterized by phylogenetic, SimPlot, selection pressure and hydrophilicity analyses. The strain, PC15 was placed in the GII.4-Hunter subcluster. An intragenotype recombination event between ORFs 2 (new GII.4 variant) and 3 (Den Haag subcluster) of the strain, PC51 was detected for the first time in this study. The strain, PC52 showed the presence of commonly detected intergenotype recombination, GII.b/GII.3. A 16 amino-acid signature code (TDVVYYAGASQPRDDI) was identified in the ORF2 of recombinant GII.3 specificity strains, which may serve as a genetic marker for differentiation of these strains from non-recombinant GII.3 strains. The amino-acid substitutions in the ORF2 of PC51 and PC52 strains in comparison to the reference strains (Toyama1 and TV24) resulted in an increase in the hydrophilicity suggested alterations in the antigenic regions of Indian NoV strains. A unique pattern of amino-acid substitutions was observed within seven subclusters of GII.4 at 19 sites (including 13 sites under positive selection pressure) spanning entire ORF2. The study indicates adaptation of NoVs in the environment to escape the host immune response and to persist in the population. It also provides in-depth analyses of NoV genomes from India and determines the extent of conserved and variable features of the Indian NoV strains.