Antibiotic residues and bacteria resistant to antimicrobial have been found in poultry litter, which is popular low cost manure in agriculture. A study was conducted to investigate a pre-treatment method to reduce antibiotic resistant bacteria in poultry litter. Poultry litter was collected from eleven medium-scale broiler farms in Kandy. Total and antibiotic resistant (tetracycline and/or enrofloxacin) culturable bacteria were isolated using Tryptic Soy Agar supplemented with 0 or 100 ppm of each antibiotic. A fresh and a cured broiler litter sample were collected from one farm and treated with different temperatures ( 25o C, 40 o C, 60 o C and 80 o C for 24 hours), neem seed extract (10 and 1000 ppm for 4 day) and neem dry leaves (10% w/w for 4 days) in triplicates. Total and antibiotic resistant bacteria in treated litter were assessed. Broiler litter from all farms contained tetracycline resistant bacteria. Heating at 80 o C reduced tetracycline resistant bacteria by 96-100% compared to 25 o C. Neem seed extract at 10 ppm reduced tetracycline resistant bacteria by 75-96%. Treating broiler litter with high temperature (>60 o C) and neem seed extract (10 ppm) could minimize spread of antibiotic resistant bacteria to the environment.