MOLECULAR DETECTION OF JAPANESE ENCEPHALITIS VIRUS 5′ UNTRANSLATED REGION IN SEROLOGICALLY REACTIVE PAEDIATRIC CASES FROM SOUTHERN CHHATTISGARH, INDIA

Abstract

Background: Japanese encephalitis virus (JEV), a mosquito-borne flavivirus, remains an important cause of acute encephalitis syndrome (AES) in India. Molecular detection of viral RNA provides direct evidence of infection and also constitutes the first step toward genomic sequencing and genotype characterization. However, the short and low-level viraemic phase of JEV frequently limits the sensitivity of reverse-transcription polymerase chain reaction (RT-PCR), particularly when specimens are collected after the appearance of virus-specific IgM antibodies. Objectives: To evaluate molecular detection of the conserved 5′ untranslated region (5′UTR) of JEV in serum specimens from pediatric patients reactive for JEV-specific IgM and to describe their demographic, clinical and serological characteristics. Methods: A descriptive laboratory-based investigation was done on ten pediatric serum specimens obtained between March 6 and July 13, 2026 from patients in the Bastar, Dantewada, and Kondagaon districts of Chhattisgarh, India. All specimens tested positive for JEV-specific IgM using the NIV JEV IgM ELISA kit. Molecular testing for JEV RNA was then performed at Pt. J.N.M. Medical College, Raipur, using the DiAGSure® JEV Detection Kit, which targeted the JEV 5′UTR region. Demographic variables, clinical presentation, AES status, epidemiological history, ELISA optical density values, and RT-PCR results were all analyzed descriptively. Results: The mean age of the patients was 5.45 years, with a median age of 4.75 years and a range of 3–11 years. Five patients were male and five were female. Six cases originated from Bastar, two from Dantewada and two from Kondagaon. Four patients (40%) fulfilled the recorded AES classification, while six (60%) did not. Convulsions or abnormal body movements were documented in four patients. The mean JEV IgM ELISA optical-density value was 1.50 ± 0.69, with values ranging from 0.63 to 2.88. Despite IgM reactivity, JEV RNA targeting the 5′UTR was not detected in any of the 10 serum specimens. Thus, the observed molecular detection rate among the IgM-reactive samples was 0%. Conclusion: JEV-specific IgM reactivity in these pediatric cases was not associated with detectable viral RNA in serum, according to a 5′UTR-targeted RT-PCR assay. The results demonstrate the diagnostic distinction between serological evidence of recent infection and direct molecular detection of circulating virus RNA. Negative serum RT-PCR results should not be used to rule out JEV infection, especially if the samples were acquired after the short viraemic phase. Early acute-phase serum, whole blood (validated), and CSF sample combined with serology may boost molecular yield. To assess the circulating JEV genotypes and genetic diversity in Chhattisgarh, prospective genomic surveillance using sequencing of RT-PCR-positive specimens will be necessary.