Introduction:The most common oral hygiene aid used to improve the oral health of an individual is the toothbrush. Hence, it could play a role in initiation and transmission of infections and diseases. This study aimed at investigating the presence of haemolytic bacteria on used toothbrushes.Methods: A close-ended questionnaire was designed and distributed to a hundred Nigerian students. This was employed to ascertain the attitudes of students towards their toothbrush. Twenty samples of already-used toothbrush (1-3 months) were randomly collected from students. The heads containing bristles were detached soaked in peptone water and vortexed. Serial dilution was done and appropriate dilution factor was plated out on Nutrient agar medium. The isolates were tentatively identified with morphological and biochemical properties. They were also assayed for haemolytic activity and antibiotics susceptibility. Results: Results indicated that 80% of the students responded to the questionnaire, out of which > 60% agreed to have sometime injured themselves while brushing their teeth. A little above half of the participants do not care about the purity of the water they use in brushing. Majority (70%) saw no use submitting their toothbrush for microbiological assessment. However, 81.25% replace their toothbrush within 5 months of use. Seventeen out of the twenty samples examined had aerobic bacterial count of > 10 4 cfu/toothbrush. The 24 isolates obtained were found to belong to Lactobacillus, Staphylococcus, Streptococcus, and Bacillus genera with 55%, 30%, 20%, and 15% occurrence, respectively. Moreover, 40% of the isolates were found to be either alpha or beta haemolytic indicating their potential to cause disease. Conclusions: Generally, all isolates were susceptible to most of the commonly used antibiotics. It is envisaged that the results of this study would chaperone oral hygiene practice among individuals such as the immunocompromised.