Background and Aims: Hospital linen is widely acknowledged to be a potential reservoir for microorganisms and a vector of disease transmission. There are many types of microorganisms and they can be found in different types of hospital linen. This study attempted to investigate the types of hospital linen microorganisms and their distribution in hospital linen. Methods: This descriptive cross-sectional study was conducted in a public hospital namely Sher-E-Bangla Medical College Hospital (SBMCH), and in a private hospital namely Rahat Anower Hospital in Barishal district, Bangladesh. 64 (34 from public hospital and 30 from private hospital) swab samples were selected purposively from various wards and departments for microbiological test. Data processing and analysis were done using SPSS (Statistical Package for Social Science) version 23. Results: Public hospital’s sample illustrated the higher microbiological growth 19 (56%) than private hospital’s sample 10 (33%). Of these, public hospital showed 42% and 36.84% cocci and bacilli group, respectively. On the other hand, 50% and 40% cocci and bacilli group microorganism was found in the private hospital linen. The highest 4 (80%) positive microbiological growth was found in the Gynae operation theater, male surgery ward and female orthopedic ward of public hospital. In contrast, about 6 (40%) positive microbiological growth from the linen store of linen department of private hospital was noticed. Conclusions: It is concluded that different types of microorganisms such as cocci and bacilli can be found in every department and ward of both private and public hospital.