Microbial biofilm is one of the most serious problems facing poultry farms all over the world and especially in Egypt. Salmonella, E. coli and S.aureus were the highest implicated bacteria in biofilm formation in poultry farms. Consequently, 440 samples were collected from 8-broiler and 8-layer farms at El- Sharkia Province, Egypt, during the period from (July 2021 till August 2022). The objective of the study was to evaluate biofilm development capacity of the tested bacterial species by the microtiter plate (MTP) assay. Also, the efficacy of five disinfectants commonly used in poultry farms (Sodium hypochlorite, hydrogen peroxide, Virkon S, glutaraldehyde and copper sulphate) with different concentrations (1, 2 and 5%) and different contact times (10, 60 and 120 m) on reducing the biofilms produced by S. Enteritidis, E. coli O78 and S. aureus was estimated. Results showed that out of 440 collected samples, 17 (3.8%), 200(45.5%) and 66 (15%) strains were identified as Salmonella, E. coli and S. aureus, respectively. 88.2%, 92% and 87.8% of the isolates of Salmonella, E.coli and S. aureus were biofilm producers. The most effective disinfectant was sodium hypochlorite which eliminated the biofilms of S. Enteritidis and E. coli O78 when used at concentration 5% for 120 m while 5% for 60 m against S. aureus biofilm. Additionally, hydrogen peroxide showed great efficiency and complete removal of biofilm of S. Enteritidis when used at concentration 2% for 120m and 5%f or 120 m against S.aureus biofilm, meanwhile removed 91% of E. coli O78 biofilm when used at concentration of 5% for 120 m. However, Copper Sulphate was insufficient disinfectant to be used against the biofilms. It can be concluded that the anti-biofilm efficiency of the disinfectants increases with the increase concentration and contact time with biofilms especially when using oxidizing disinfectants (hypochlorite and peroxides).