Bovine blood inoculated and incubated with bacteria was tested to determine if secondary screwworm, Cochliomyia macellaria (F.), would be attracted to the incubated substrate for oviposition. Five species of bacteria, Klebsiella oxytoca (Flugge), Proteus mirabilis Hauser, Proteus vulgaris Hauser, Providencia rettgeri Hadley, Elkins and Caldwell, and Providencia stuartii Ewing, previously isolated from animal wounds infested by primary screwworms, Cochliomyia hominivorax (Coquerel), were used. Incubated substrates were tested in a two-choice cage bioassay to study landing response and oviposition by gravid C. macellaria. Significantly more flies landed on substrates containing P. mirabilis than on substrates with other species of bacteria. Klebsiella oxytoca-treated substrates attracted the least flies. Substrates containing bacteria incubated for 72 h attracted significantly more flies than those incubated for 24-, 48-, or 96-h period. In 3-h duration oviposition tests, substrates with P. rettgeri attracted significantly more flies to oviposit than the other four species. The most eggs were recorded when substrates treated with all five species of bacteria were offered for oviposition. It is likely that multiple active chemicals present in the volatiles from substrates treated with all five species result in greater response than those in a single species. At least 72-h incubation seems to be necessary to obtain the most active volatile chemicals. Results suggest that C. macellaria uses similar chemical cues as C. hominivorax from bacteria volatiles as oviposition attractant/stimulant.