Multidrug‐resistant tuberculosis (MDR‐TB) is one of the major public health concerns worldwide particularly in developing countries, including Bangladesh. Thus far, there are no well‐validated clinical guidelines for the prevention of MDR‐TB. This study aims to evaluate the improvement in nurses' practice using the newly developed Nursing Practice Guidelines for the Prevention of MDR‐TB (NPG: MDR‐TB) among hospitalized adult patients in Bangladesh. The guidelines were developed, disseminated and evaluated among 64 nurses by assessing nursing practice for the prevention of MDR‐TB during pre‐ and postimplementation of the guidelines. Significant differences between pretest and post‐test mean scores of nursing practice for the prevention of MDR‐TB in case finding and case holding were found in three levels of wards, including Level 0 (non‐TB), Level 1 (TB) and Level 2 (MDR‐TB) (P < 0.001). This indicated that the guidelines might be applicable to reduce the development of MDR‐TB in hospitals. However, this was a preliminary study with a limited time frame. Further evaluation is, therefore, needed.