Unbiased metagenomic next generation sequencing to investigate possible infective etiologies of 2021 SHAPU outbreak in Nepal

Background Seasonal Hyperacute Panuveitis (SHAPU) is a rare, rapidly progressive ocular disease that leads to severe vision loss. Its etiology remains uncertain and observed seasonal outbreaks have been temporally linked to white moths. Several bacterial and viral pathogens have been reported, but confirmative studies are lacking. This study utilized unbiased mNGS on ocular samples from the 2021 SHAPU outbreak to investigate potential infectious etiologies. Methods A hospital-based cross-sectional study was conducted from August to December 2021 in four different tertiary-care hospitals in Nepal. The clinical data and ocular samples (aqueous, vitreous, and conjunctival swabs from affected and unaffected eyes) were collected from 135 SHAPU patients. Twenty-one samples were selected for mNGS. Their DNA and RNA libraries were sequenced on Illumina iSeq100 and analyzed using the Chan Zuckerberg ID with healthy conjunctival swabs as background controls. Results The median patient age was 12 years with 41.5% presenting severe SHAPU. Direct contact with white moths was only reported by 34.8% of patients. It was observed that visual outcomes improved after treatment, with severe vision impairment decreasing from 31.9% to 14.1%. mNGS revealed a diverse ocular microbiome containing Streptococcus pneumoniae and Acinetobacter bereziniae , with commensals including Cutibacterium acnes and Escherichia coli . S. pneumoniae was consistently detected in vitreous samples and conjunctival swabs, with higher abundance in affected eyes. Multiple S. pneumoniae strains (serotype-04, 11A, Swiss_NT, 19F) with varied antimicrobial resistance profiles were identified. Opportunistic pathogens such as Acinetobacter bereziniae and Enterococcus spp. were also observed in vitreous samples, though their role remains uncertain. Conclusion In this study, mNGS characterized the ocular microbiome during SHAPU, depicting bacterial etiology including S. pneumoniae, alongside opportunistic pathogens. The external exposures such as moth contact may act as triggers. Larger, multi-sample metagenomic and immunological studies are recommended to understand pathogenic mechanisms and inform interventions.