All sequences obtained for VP4(P), VP7(G), VP6(I) and NSP4(E) genes were aligned with the corresponding gene sequences of RVA strains available in the GenBank Linsitinib in vitro by using Clustal W [21]. The phylogenetic analysis was carried out in MEGA 5 by using Kimura –2 parameter and neighbour-joining method [22]. The reliability of different phylogenetic groupings was confirmed by using the bootstrap test (1000 bootstrap replications). The RV NSP4, VP4, VP6 and VP7 gene sequences from this study have been deposited in GenBank under the accession numbers KF951361-KF951404. Group-A RV antigen was detected in 9.4% (35/371) of the specimens collected from adolescent

and adult cases of acute gastroenteritis. The distribution showed a decline in the RV positivity over time (Fig. 1). Genotyping of VP7 and VP4 genes was conducted for all 35 strains detected in adolescent and adult cases of acute gastroenteritis. The VP7 and VP4 genes were both successfully genotyped in 6 cases and one additional VP7 was typed. For the remaining 28 samples, VP7 and VP4 genes could not be amplified despite the use of specific primers. The number of strains non-typeable for both genes (n = 28) was significantly high as compared with the typeable strains

(p < 0.01). Among the strains (n = 6) typeable for both VP7 and VP4 genes, G2P[4] (n = 3;

2 in 2009 and 1 in 2012), G9P[4] (n = 2; 1 each in 2010 and 2011) and G1P[8] (n = 1 in 2009) genotypes were detected. TSA HDAC price All 6 and 1 additional typed VP7 sequences clustered with their respective genotypes (Fig. 2). G2 strains were placed in lineage II sublineages C and D. G9 and G1 strains were classified in lineages L3 and L1, respectively. Analysis of VP4 gene sequences showed clustering of all of the P[4] strains (n = 5) not in the P[4]- 5 lineage and that of the P[8] strain (n = 1) in the P[8]-3 lineage. Two of the P[4] strains did not amplify sufficiently in the first round of PCR and hence were not included in the phylogeny (Fig. 3). Twenty seven of the 35 strains which typed or did not type for VP7 and VP4 genes were amplified in the VP6 PCR and sequenced. Analysis of VP6 gene sequences showed clustering of the majority (24/27; 89%) in the I2 genotype, in two clusters with the remaining 3 strains (3/27, 11%) clustering in the I1 genotype (Fig. 4). Six of the 35 strains were amplified by NSP4 PCR and sequenced, 4 of 6 amplified genes clustered in the two different groups of E2 genotype and the remaining two clustered with the E6 genotype (Fig. 5). The VP6 and NSP4 genes amplified from 20 and 2 strains, respectively, which were non-typeable for VP7 and VP4 genes were most homologous to human RV strains.