Transmission dynamics for invasive Non-Typhoidal S almonella serovars (TiNTS): protocol for a household study of transmission and immune response to non-typhoidal Salmonella in Malawi.

Background: Invasive non-typhoidal Salmonella (iNTS) disease is a leading cause of community-onset bloodstream infection in Africa, driving high morbidity in young children. The World Health Organization has published preferred product characteristics for an iNTS vaccine, but lack of transmission data is an impediment to vaccine licensure. Enteric NTS (eNTS) is the asymptomatic carriage of NTS in stool that precedes invasive disease. We do not know how long eNTS shedding lasts, how often infection spreads in endemic settings, or how an eNTS episode shapes immunity against later invasion. These gaps make it difficult to define trial sites, select cohorts, refine target product profiles, and build reliable models of vaccine impact. Here we describe TiNTS, a prospective household study in Blantyre, Malawi, which will measure real-time eNTS incidence, transmission, and antibody responses to close these evidence gaps and accelerate rational vaccine deployment. Methods: We will recruit all members of at least 60 households in Ndirande, Blantyre, Malawi. Stool samples will be collected every other day for at least four weeks and tested for NTS using culture and pan- Salmonella PCR on growth media. Environmental samples collected at enrolment will be tested using the same methods. Symptoms and exposure risks will be recorded throughout.We will collect blood samples at enrolment, after four weeks, and four weeks after the first eNTS episode in each household. We will measure serum IgG responses to Salmonella Typhimurium and Enteritidis LPS antigens. We will extend follow-up if participants continue shedding or if the first household case occurs with fewer than 14 days of follow-up remaining.All culture-positive isolates and PCR-positive broths will undergo Illumina sequencing to enable genome and metagenome reconstruction for transmission inference. Conclusions: TiNTS will define the burden, transmission patterns, and immune response to eNTS. Findings will inform vaccine modelling, trial design, and targeted introduction strategies.