Organophosphorus pesticides (OP) are used extensively in many arenas including agriculture and industry leading to humans and agroecosystems disorders. Malathion is one of the OP that are used in agriculture to control pest and protect crops. Also, they harm non-target organisms and affect cruelly water sources, air, and soil quality. The present study aimed to isolate and identify a potent bacterial isolate capable of degrading malathion. Bacterial strain that isolated from Al Fayoum governorate, Egypt exhibited high efficiency for malathion biodegradation. Biodegradation process using minimal salt medium (MSM) supplemented with different malathion concentrations indicated that the bacterium was able to degrade and use malathion as a sole carbon source up to 700 mg/l at 37°C.The potent strain that exhibited biodegradation potential was identified as Bacillus sp. FYM31 and deposited into GenBank with the accession number OK325597.  HPLC proved the effectiveness of malathion removal by Bacillus sp. FYM31 after 12 days of incubation to the level of 70.1% malathion (700 mg/l) degradation. Organophosphorus hydrolase (opd) gene was detected in the potent Bacillus sp. FYM31 strain. Due to the widespread usage of malathion in Egypt's agricultural areas, Bacillus sp. FYM31 can help bio-remediate the polluted areas.