Identification of previously untypable RD cell line isolates and detection of EV-A71 genotype C1 in a child with AFP in Nigeria

Abstract

We previously attempted to identify 96 nonpolio enteroviruses (EVs) recovered on RD cell culture from children <15 years with acute flaccid paralysis (AFP) in Nigeria. We succeeded in identifying 69 of the isolates. Here, we describe an attempt to identify the remaining 27 isolates.Twenty-six (the 27 th isolate was exhausted) isolates that could not be typed previously were further analyzed. All were subjected to RNA extraction, cDNA synthesis, enterovirus 5"-UTR -VP2 PCR assay and a modified VP1 snPCR assay. Both the 5"-UTR -VP2 and VP1amplicons were sequenced, isolates identified and subjected to phylogenetic analysis.Twenty of the 26 isolates analyzed were successfully identified. Altogether, 23 EV strains were recovered. Thesebelong to 11 EV (one EVA, nine EVB and one EVC) types which were EVA71 genotype C1 (1 strain), CVB3 (7 strains), CVB5 (1 strain), E5 (2 strain), E11 (3 strains), E13 (2 strain), E19 (1 strain), E20 (1 strain), E24 (2 strains), EVB75 (1 strain) and EVC99 (2 strains). Of the 11 EV types, the 5"-UTR-VP2 assay identified seven while the VP1 assay identified 10. Both assays simultaneously detected 7 of the 11 EV types identified in this study with 100% congruence.In this study we identified 20 of 26 samples that were previously untypable. In addition, we provided evidence that suggests that a clade of EVA71 genotype C1 might have been circulating in sub-Saharan Africa since 2008. Finally, we showed that the 5"-UTR -VP2 assay might be as valuable as the VP1 assay in EV identification.