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Coverage of HIV testing services (HTS) is generally low among men who have sex with men (MSM) and transgender women ( hijra ) in Bangladesh, thus impeding the national goal of attaining the 90-90-90 target. In this context, this article delineates HTS uptake barriers among these populations. This qualitative study entailed 30 in-depth interviews, six focus groups and seven key-informant interviews with purposively selected MSM and hijra , alongside service providers. Participants cited individual and interpersonal barriers such as low risk perception and misconceptions about HIV testing, programmatic barriers such as knowledge gaps among peer service providers, as well as community and structural barriers such as the criminalization and stigmatization of male-to-male sex. Considering these contexts, it is essential for stakeholders to improve the HTS modality using multipronged approaches to address the multifaceted barriers of HTS uptake.