Detection _ differentiation of Coxsackie A 24 variant isolated from an epidemic of XXXX in north India by RT-PCR using a novel primer pair.

@entity1 @entity19 @entity5518 @entity204 @entity912
BACKGROUND _ OBJECTIVES: An epidemic of @entity5518 ( @entity5518 ) occurred in north India during July to September 1994. We report a reverse transcription-polymerase chain reaction (RT-PCR) using known and novel primers to differentiate and identify the CA 24 virus isolated from the epidemic of @entity5518 . METHODS: Conjunctival swabs were collected from 46 @entity1 (in 12 @entity1 from both the eyes) yielding 58 swabs. The swabs were inoculated in RD 19S and HeLa-199 cell monolayers and observed for cytopathic effect. Serum neutralizing antibodies were tested in 17 acute and 10 convalescent phase serum samples. RT-PCR was done on 9 isolates (7 Coxsackie A 24 and 2 ECHO-1 as identified by neutralization test) using known and a novel primer. Fourteen virus isolates (9 CA 24, 3 ECHO-1 and 2 untyped) were inoculated in suckling @entity19 and these @entity19 were observed daily for 10 days for flaccid @entity912 of hind limb or @entity204 . RESULTS: Cytopathic virus was isolated from conjunctival swabs in 21 of 46 (45.6%) @entity1 subjected to virus isolation. Sixteen of 21 (76.2%) isolates were neutralized by CA 24 specific antisera, 3 isolates were identified as ECHO-1 with Schmidt enteroviruses antiserum pools while 2 remained untypable. Of these 21 isolates, 9 representative isolates (7 CA 24 and 2 ECHO-1) tested by RT-PCR had enterovirus common region DNA but did not show any amplification in RT-PCR with EV-70 specific primers (VP-1 and VP-3). Using CA 24 specific novel (VP 3-1) primers amplification was seen in 6 of 7 CA 24 isolates while 2 ECHO-1 remained unamplified. In contrast with 3C-proteinase region primers, only 2 of 7 CA 24 were amplified along with false amplification of both ECHO-1. Serum neutralizing antibodies were seen in 2 of 17 (11.7%) acute phase sera and 6 of the 10 (60%) convalescent phase sera while in paired sera (available in two @entity1 ) a four-fold rise in titres were observed. Hind-limb @entity912 and/or @entity204 occurred in all suckling @entity19 inoculated with CA 24 isolates while @entity19 remained healthy after inoculation with 3 isolates of ECHO-1 and 2 untypable isolates. INTERPRETATION _ CONCLUSION: The epidemic of @entity5518 was caused by a variant of CA 24. Molecular typing can detect and differentiate between CA 24 and EV-70 viruses. Novel primer pair was found useful in the identification and confirmation of CA 24 isolates.
