The occurrence of increasing hospital-acquired infections and the reports of the occurrence of ants in the hospital had become a research focus owing to the exposure of patients and health professionals to the risk associated with insects. The study aimed to evaluate the potential of ants as mechanical vectors of pathogenic bacteria and assess the antibiotic resistance pro ile of gram negative bacilli isolated in selected level one hospitals in Davao City. The study utilized a descriptive design. The researcher collected the ant samples in female, male and pediatrics ward in the selected level one hospitals. Afterwards, the captured ant species were immersed in double strength TSB for bacterial cultivation. After distinct pure colonies of bacteria were isolated, they were subjected to tests for bacterial identi ication and antibacterial susceptibility tests using VITEK 2 Systems. The indings of the study suggest that Tapinoma melacephalum, Monomorium pharaonis and Anoplolepis grecilepis are mechanical vectors of opportunistic pathogen which commonly cause hospital acquired infections. Moreover, two (2) isolates of E. cloacae from T. melanocephalum and one (1) isolate of E. cloacae from M. pharaonis demonstrated multi-drug resistance. Further, the results also suggest possible poor implementation of sanitation and pest control protocols. The researcher recommends for further identi ication and pro iling of gram positive bacteria and to explore the different hospital areas including critical care units as sampling points.