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Deep tubewells are important sources of arsenic mitigation in rural Bangladesh. Compared to commonly available shallow tubewells, deep tubewells tap into deeper low-arsenic aquifers and greatly reduce exposure to arsenic in drinking-water. However, benefits from these more distant and expensive sources may be compromised by higher levels of microbial contamination at point-of-use (POU). This paper examines differences in microbial contamination levels at source and POU among households using deep tubewells and shallow tubewells, and investigates factors associated with POU microbial contamination among deep tubewell users. We assessed a prospective longitudinal cohort of 500 rural households in Matlab, Bangladesh, across 135 villages. Concentration of Escherichia coli (E. coli) in water samples at source and POU using Compartment Bag Tests (CBTs) was measured across rainy and dry seasons. We employed linear mixed-effect regression models to measure the effect of different factors on log E. coli concentrations among deep tubewell users. CBT results show that log E. coli concentrations are similar at source and at POU during the first dry and rainy season, but are significantly higher at POU among deep tubewell users during the second dry season. Log E. coli at POU among deep tubewell users is positively associated with both presence (exponentiated beta exp(b) = 2.52, 95% Confidence Interval (CI) = 1.70, 3.73) and concentration of E. coli (exp(b) = 1.36, 95% CI = 1.19, 1.54) at source, and walking time to the tubewell source (exp(b) = 1.39, 95% CI = 1.15, 1.69). Drinking-water during the second dry season is associated with reduced log E. coli (exp(b) = 0.33, 95% CI = 0.23, 0.57) compared to the rainy season. These results suggest that while households that use deep tubewells have lower arsenic exposure, they may be at higher risk of consuming microbially contaminated water compared to households that use shallow tubewells., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Mark Sobsey, a co-investigator and author on this publication is an inventor on the technology being used in this research. Sobsey is also part owner of the company, Aquagenx LLC, which has licensed this technology. UNC-Chapel Hill has the patent on the technology and has a financial interest in the company. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results., (Copyright © 2023 Elsevier Ltd. All rights reserved.)