Exploring the Role of Microbiome in Susceptibility, Treatment Response and Outcome among Tuberculosis Patients from Pakistan; Study Protocol for a Prospective Cohort Study (Micro-STOP)

Introduction Tuberculosis (TB) caused by Mycobacterium tuberculosis is a common infectious disease associated with significant morbidity and mortality, especially in low and middle-income countries. Successful treatment of the disease requires prolonged intake (6 – 8 months) of multiple antibiotics with potentially detrimental consequences on the composition and functional potential of the human microbiome. The protocol described in the current study aims to identify microbiome (oral and gut) signatures associated with TB pathogenesis, treatment response and, outcome in humans. Methods and analysis Four hundred and fifty, newly diagnosed TB patients from three district levels (Peshawar, Mardan, Swat) TB diagnosis and treatment centers will be recruited in this non-interventional, prospective cohort study and will be followed and monitored until treatment completion. Demographic and dietary intake data, anthropometric measurement and blood, stool and salivary rinse samples will be collected at baseline, day 15, month-2 and end of the treatment. Additionally, we will recruit age (± 3 years) and sex-matched healthy controls (n=30). Blood sampling will allow monitoring of the immune response during the treatment, while salivary rinse and fecal samples will allow monitoring of dynamic changes in oral and gut microbiome diversity. Within this prospective cohort study, a nested case-control study design will be conducted to assess perturbations in oral and gut microbiome diversity (microbial dysbiosis) and immune response and compare between the patients groups (treatment success vs failure). Ethics and dissemination The study has received ethics approval from the Ethic Board of Khyber Medical University Peshawar, and administrative approval from Provincial TB Control Program of Khyber Pakhtunkhwa, Pakistan. The study results will be presented in national and international conferences and published in peer-reviewed journals. Trial registration number NCT04985994; pre-results Strengths and limitations of the study Prospective study design. Large sample size, several sampling time points, and samples taken before, during and after anti-tuberculosis treatment will ensure capture of an accurate picture of any emerging microbial dysbiosis and immune modulation; this will provide new understanding of how these two factors are mutually influenced and how they are impacted by treatment and outcome in TB patients. In addition to microbial signatures, collection of socio-demographic, anthropometric, nutrition and co-morbidities data will permit further subgroup analysis and meaningful results that will be shared with the scientific community. A potential weakness is that not all patients may be able to provide samples for all of the specified time points. Another weakness is the raised likelihood that some participants will withdraw since the study is carried out in resource poor communities