Tryptose agar

A total of 110 consecutive and non-duplicated A. baumannii clinical isolates were collected from different departments [intensive care unit (ICU), gastroenterology, respiratory, neurosurgery and other wards] at the First Affiliated Hospital of Chengdu Medical College, Chengdu, Sichuan, China from 2012 to 2013. Isolates were identified by standard laboratory methods and ATB New (bioMérieux, France). A. baumannii was further verified when two PCR products were yielded as reported: a 425-bp internal control amplicon corresponding to the recA gene of Acinetobacter spp. and the 208-bp fragment of the 16S rRNA (Chiang et al., 2011 (link)) intergenic spacer region of A. baumannii (Table 1). All strains were stored at –80°C, and bacteria were grown on tryptose agar or Mueller–Hinton broth or agar (Oxoid, England).

Ten drops of a stool suspension prepared in sterile saline were deposited on a 0.6 µm pore-size membrane filter which was applied on the center of tryptose agar (Oxoid, Hampshire, UK) supplemented with 5% unlysed horse blood. Following a 15–20-min filtration process, the filter was removed under aseptic conditions and the medium was incubated at 37 °C under a microaerobic environment enriched with H₂ for six days [9 (link)].