AIM: To detect the quantitative expression levels of the pro-inflammatory interleukin-8 (IL8), antimicrobial peptides human beta defense-2 (HBD2), and human beta defense-3 (HBD3) genes in bacterial conjunctivitis. METHODS: The human conjunctival epithelial cells were obtained using the impression cytology technique from healthy controls and patients. The genes expression levels were determined utilizing a reverse transcription quantitative polymerase chain reaction (RT-qPCR). The contribution of causative agent type, the number of isolates and severity of clinical features, in the increase of genes expression was also determined. RESULTS: The RT-qPCR showed that IL8, HBD2, and HBD3 expression increased in bacterial conjunctivitis as compared to healthy control (P<0.001). In gram-negative bacterial conjunctivitis, HBD2 was highly up-regulated (P<0.001) compared to other types of bacterial conjunctivitis. In mixed bacterial conjunctivitis, a direct correlation between HBD2 up-regulation and HBD3 up-regulation was observed (P<0.05). The severity of clinical features was related to the up-regulation of IL8 and HBD2 (P<0.05). CONCLUSION: IL8, HBD2, and HBD3 are immune-effectors in infectious conjunctivitis. HBD2 is active during different bacterial conjunctivitis but is more released with gram-negative bacteria compared to gram-positive bacteria. HBD3 is an obvious defender in different bacterial conjunctivitis.