Abstract: Mixed microorganisms in a biofilter medium were acclimatized with xylene vapor continuously for 8 h day-1 for 30 and 60 days in laboratory. Biofilter medium was coconut husk: manure compost: wastewater sludge at 75: 20: 5 (v/v). The initial moisture content of the biofilter medium was adjusted to 50% of dry weight. Two bacteria and 4 fungi from biofilter were identified at the end of experiment in enriched medium. All dominant species were fungi which were subsequently isolated and identified as Aspergillus flavus (EF592170), Aspergillus terreus (EF592171), Penicillium glabrum (EF592172) and Aspergillus niger (EF592173), respectively. The result indicates that the fungi last longer in biofilter with xylene environment and thus, may be suitable to be cultured for use in biofilter for xylene removal in industrial applications.