While it was reported that there was no statistically significant difference in vaccine efficacy against G1 and non-G1 genotypes Anti-cancer Compound Library in vitro in the clinical trial [8], we considered it important to

examine whether the strain variation observed for the two surface protein genes extended to the other genome segments. Of note, there is a considerable lack of overall genomic RNA homology between human rotavirus strains with long RNA patterns (as represented by the Wa strain; hence called the Wa genogroup to which RIX4414 belongs), and human rotavirus strains with short RNA patterns (as represented by the DS-1 strain; hence called the DS-1 genogroup to which strains including genotype G2P[4] belong) [18], [19] and [20]. The aim of this study was to compare by RNA–RNA hybridization the whole genomic RNA constellation of circulating wild-type rotaviruses detected during the clinical trial in Malawi with RIX4414 (the strain contained in Rotarix™). This study also aimed to determine the nucleotide sequence similarities between RIX4414

and circulating wild-type rotaviruses in Malawi, as compared with RIX4414 and other globally circulating strains, in the genome segments coding for the neutralisation proteins Mdm2 inhibitor VP7 (G genotype) and VP4 (P genotype), the middle capsid protein (VP6: I genotype), and the viral enterotoxin (NSP4: E genotype). Rotavirus-positive specimens (N = 147) collected from vaccine and placebo recipients in the clinical trial in Blantyre, Malawi, were previously examined for G and P types at DDL Diagnostic Laboratory (Voorburg, too the Netherlands) by a testing algorithm using RT-PCR followed by a reverse hybridization assay [21]. Of those, only specimens containing a minimum volume of 500 μl as 10% suspension in Earl’s Balanced Salt Solution (N = 88) were utilized in this study. Rotavirus specimens examined comprised G12P[6] (N = 25),

G8P[4] (N = 28), G1P[8] (N = 11), G9P[8] (N = 9), G12P[8] (N = 5), G2P[4] (N = 3), G8P[8] (N = 2), G12P[4] (N = 1), G1P[6] (N = 1), G8P[6] (N = 1), G12P[6]/P[8] (N = 1) and G8P[4]/P[6] (N = 1). The vaccine strain (RIX4414) used in this study was recovered following inoculation into MA104 cell culture of a portion of the Rotarix™ commercial vaccine. Rotavirus RNA was extracted using an RNAeasy kit (Qiagen Ltd., Sussex, UK) according to the manufacturer’s instructions, and separated by electrophoresis on a 10% polyacrylamide gel followed by staining with silver nitrate as described previously [22]. Electropherotypes were assigned for those strains that showed 11 segments of double-stranded (ds) RNA, and were determined by comparison of the individual RNA migration patterns of genome segments on the gel.