Antibiotic resistance and the mode of transmission were investigated in bacteria isolated from poultry litter. Total aerobic heterotrophic bacteria were screened and identified for their resistance to different antibiotics such as ampicillin, streptomycin, erythromycin, tetracycline, chloramphenicol, kanamycin, tobramycin, and rifampicin. The distribution of bacteria found in the litter was Staphylococcus (29.1%), which was the predominant group, followed by Streptococcus (25%), Micrococcus (20.8%), Escherichia coli (12.5%), Salmonella (8.3%), and Aeromonas (4.1%). Fifty percent of these isolates were susceptible to ampicillin, 57% to erythromycin, 25% to tetracycline, 4% to chloramphenicol, 40% to kanamycin, 75% to streptomycin, 54% to tobramycin, and 4% to rifampicin. Three randomly selected isolates representing Staphylococcus, Streptococcus, and Micrococcus were examined for plasmids, and plasmid-curing and plasmid-induced transformation studies were conducted. Streptococcus and Micrococcus harbored a plasmid of 4.2 and 5.1 kb, respectively, whereas Staphylococcus did not harbor any plasmids. Plasmids were cured in Streptococcus and Micrococcus at a concentration of 75 and 100 microg/ mL of acridine orange, respectively, and transformation of 4.2- and 5.1-kb plasmids isolated from the Streptococcus and Micrococcus to plasmid-free E. coli DH5alpha strain was possible. In conjugation experiments, the antibiotic resistance profiles of transconjugant cells were found to be the same as the donors with the exception of Staphylococcus. The results of this study suggest that transformation and conjugation could be an important mechanism for horizontal gene transfer between bacteria in poultry litter. An understanding of the mechanism and magnitude of resistance gene transfer may provide a strategy to reduce the potential for dissemination of these genes.