Syndromic cholera diagnosis masks diverse causes of diarrhoeal disease in Burundi revealed by portable metagenomics.

BACKGROUND: Cholera outbreaks remain a major public-health challenge in sub-Saharan Africa, where diagnostic capacity is limited and clinical case definitions are non-specific and re ly heavily on syndromic diagnosis. Rapid identification of Vibrio cholerae is critical, yet cholera-suspected diarrhoea can have multiple infectious causes not captured by targeted diagnostics. METHODS: We evaluated a mobile, culture-independent metagenomic sequencing workflow for on-site detection of gastrointestinal pathogens directly from faecal samples in Burundi. The offline workflow combined long-read Oxford Nanopore Technologies (ONT) sequencing with rapid, laptop-based taxonomic and antimicrobial resistance (AMR) screening and was deployed across a health centre, a district hospital, and a refugee transit camp. The frontline and real-time results were verified using both conventional culturing and in-depth bioinformatic analyses. RESULTS: V. cholerae signals were only detected in a subset of suspected cholera cases, while many samples were dominated by alternative bacterial taxa, most frequently Escherichia coli. V. cholerae abundance correlated strongly with detection of the C holera T oxin P hage CTXφ, supporting differentiation between toxigenic signal and background exposure. AMR genes were detected across samples, providing early situational insight into resistance determinants among gastrointestinal bacteria. CONCLUSIONS: Mobile, offline metagenomic sequencing enables rapid frontline characterization of gastrointestinal disease, especially cholera-suspected, in resource-limited settings and complements existing diagnostics by improving etiological resolution and outbreak response.