Tsa agar

From 2006 to 2011,105 isolates were collected from 11 provinces in China, and their sources included infant food, potable water, and rectal swabs of healthy humans. Three rectal swabs from healthy humans were collected by Jinan municipal CDC, and the participants were informed and had provided their written consent to participate in this study, which was approved by the Jinan municipal CDC Ethics committee. As controls, the isolates ATCC 51329 and ATCC 29544 were provided by the Chinese Academy of Inspection and Quarantine (CAIQ). All isolates were confirmed by biochemical test using kit API 20E (bioMérieux, Marcy l′Etoile, France) and real-time PCR using a primer set and probe targeting the dnaG gene on the macromolecular synthesis operon [19] (link). Isolates were cultured on TSA agar (Oxoid, Basingstoke, United Kingdom), and they were cryopreservation at −80°C for long-term storage.

The initial screening was performed using SEY agar [18 (link)], chocolate agar (Oxoid, Basingstoke, UK) [7 (link)] and TSA agar (Oxoid, Basingstoke, UK) [9 (link)]. All isolates were incubated in Brain and Heart Infusion Broth (Oxoid, Basingstoke, UK) at 37 °C/24 h. After this period, an aliquot of each strain was seeded onto the different agars and incubated for 48 h at 37 °C. After this period, strains showing white colonies or without pigmentation (gray) were suspected of S. argenteus and selected for molecular analysis. The operon crtOPQMN genes (crtO, crtP, crtQ, crtM and crtN) were screened by PCR, according to Zhang et al. [9 (link)] in all nonpigmented isolates. The nrps gene [9 (link)] was tested only in isolates that did not have at least one of the crtOPQMN operon genes. S. aureus MW2, S. aureus WB49, S. aureus N315 and S. aureus COL were used as positive controls for the reactions.

Seven Gram-stain positive, catalase-positive cocci isolated from human clinical material in different routine clinical laboratories (Table 1) were sent to the Reference Laboratory for Staphylococci (Prague, Czech Republic) for identification. Cells grown on TSA agar (Oxoid) were used for biotyping and genotyping analyses. All isolates were maintained on glass beads at −70°C in BHI broth supplemented with 15% glycerol (v/v). Type strains representing known Macrococcus species and reference M. caseolyticus strains CCM 8657, CCM 8658, P262, and P885 were obtained from the Czech Collection of Microorganisms (Masaryk University, Brno). Macrococcus canis DSM 101690^T (= KM45013^T) was obtained from the Leibniz-Institut DSMZ-Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ) and deposited in the Czech Collection of Microorganisms under strain number CCM 8748^T in this work.