Isolation and cultivation of Haemophilus ducreyi

Abstract

A useful method for isolating and recognizing H. ducreyi from chancres and buboes of male patients is presented. A total of 41 clinical isolates of H. ducreyi were recovered from 33 patients over an 8 yr period; the experience with the 15 most recent isolates is presented in detail. Chocolate agar supplemented with 1% IsoVitaleX and 5% sheep blood agar were prepared, using Trypticase soy and Mueller-Hinton agar bases; incubation conditions included ambient, capneic and anaerobic environments. Mueller-Hinton agar was clearly superior over Trypticase soy agar for isolation of H. ducreyi, although there was little difference between 5% sheep blood and supplemented chocolate agar. Growth in ambient air and under anaerobiasis was poor or lacking; growth in 5-7% CO2 was good to luxuriant. Heat-inactivated and fresh (unheated) human blood clot tubes also were used for selective isolation. Although the rates of isolation from the 2 types of clot tubes were not significantly different, unheated clot tubes were superior to heated clot tubes because of reduced level of contaminants. Gram stain characteristics taken from blood clot tubes and solid media, cellular and colonial morphology of the bacilli, and lack of oxidase, catalase and biochemical activity except nitrate reductase were determinant factors. Successful isolation of H. ducreyi evidently can be achieved with a minimal amount of resources and expertise.