Using nanopore metagenomics to characterize pathogens in febrile patients from a highland of Western Kenya

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Introduction: infection, leaving their causes unresolved. As a result of the diagnostic gap, patients receive presumptive antimalarial treatment, unnecessary antibiotics and experience delayed therapy. Methods: We conducted a nanopore metagenomic next-generation sequencing (mNGS) investigation of 168 archived blood samples collected from febrile patients in Kipsamoite and Kapsisywa sites in Nandi County, a highland region in western Kenya experiencing declining malaria transmission, between 2012 and 2020. The study used exploratory taxonomic profiling to identify microbial DNA/RNA signatures in archived plasma samples and described organisms with known or potential clinical relevance, while classifying detected organisms into common commensal, skin-associated, and environmental taxa. Demographic and clinical data linked to samples were used to fit multivariable logistic regression models to assess associations. Results: Sequencing revealed substantial microbial heterogeneity, including frequent detection of common skin/environmental taxa, opportunistic organisms, ubiquitous viruses, and a smaller number of organisms with established fever-causing potential. The detections are interpreted as molecular evidence of microbial nucleic acid, not as proof of active infection or fever causality. Malaria-positive subjects had a higher mean number of identified pathogens compared to malaria-negative subjects (3.78 vs. 2.14; p = 0.002), despite a baseline microbial landscape dominated by commensal flora, environmental organisms, and ubiquitous viruses. Discussion: Nanopore mNGS is a viable tool for identifying non-malarial febrile pathogens in western Kenya. Future efforts must combine systematic sampling with field-deployable contamination controls and causal confirmation frameworks to optimize regional antimicrobial stewardship and surveillance.

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