The present study aimed to define the effect of different concentrations of propolis powder or glue extract compared with synthetic antibiotic on ram's semen quality, resistance to bacterial contamination and fertility rate. Semen samples of six mature rams aging 2.2 years old with an average 62.0 kg live body weight were collected twice a week for three weeks using artificial vagina. The semen ejaculates were pooled, extended and divided into seven portions. The final extension rate was 1 semen: 10 extender. The seven portions were (CTL, without any antibiotic type and kept as control group), 400µl pen-step (SA400µl) and 600µl pen-step (SA600µl) as synthetic antibiotic; 400µl and 600µl propolis powder (PP400µl & PP600µl); 400µl and 600µl propolis glue (PG400µl & PG600µl) as natural antibiotic, respectively. Extended semen samples were stored at 5 °C for 6 days. The obtained results showed that, spermatozoa quality was significantly (P<0.05) decreased with the advancement of storage period in all extended samples. In addition, sperm parameters (sperm motility, acrosomal status and normal spermatozoa), enzymatic activities (ALT, AST, ALP and LDH), bacterial count and fertility rate were improved (P<0.05) in extended samples treated with synthetic and natural antibiotics either powder or glue than the control. A higher antimicrobial activity after 24 hrs incubation for PP600µl and PG600µl extenders were significantly (P<0.05) decreased compared with other extended samples. Moreover, the PG600µl antibiotic was insignificantly greater litter size (1.27%) compared with (1.07%) for SA600µl one. Regarding to the results of this study, it could be concluded that antimicrobial effect of natural propolis either powder or glue in extended ram semen was capable to improve ram semen quality and inhibitory bacterial zones, as well as, improvement fertility rate.