Microorganisms associated with commercial motorcycle helmets were investigated in the commercial city of Lagos, Nigeria. 300 motorcycle helmets were randomly collected from different commercial motor cyclists in two densely populated areas of Lagos: Yaba College of Technology (YABATECH) and Lagos University Teaching Hospital (LUTH) main gates respectively. Two sterile swabs moistened with sterile water were rotated over the inner surface of each helmet and cultured on MacConkey Agar and Nutrient Agar for bacterial growth and Sabouraud Dextrose Agar for fungi growth. The plates for bacteria growth were incubated aerobically at 37 ºC for 48 h, while plates for fungi at 28 ºC for 2 weeks. Biochemical tests were used to identify bacteria; while, cultural characteristics were used for fungi identification. The microorganisms consistently common to the samples investigated in the two locations were similar and included (with respective frequency of occurrence for both location): Staphylococcus aureus (80%; 7%), Pseudomonas aeruginosa (75%; 12%), Staphylococcus epidermis (60%; 8%), Enterobacter aerogenes (52%; 27%), Escherichia coli (40%; 13%), Bacillus spp (37%; 10%), Aspergillus spp (82%; 7%), Candida spp (55%; 22%), Rhizopus spp (40%; 27%), and Penicilium spp (35%; 12%). The motorcycle helmets collected at YABATECH had higher microbial colonization than LUTH irrespective of the isolates. This trend was similar for bacterial and fungi. Results showed that helmets could serve as vehicles for transmission of pathogens. Good hygiene practice (GHP) and regular cleaning of motor cycle helmets with sterilants is strongly advocated in order to reduce the incidence of microbial transmission and its associated infection.