Api e

The material to microbiological tests included nasal and pharyngeal swabs collected from patients before a planned vaccination against S. pneumoniae (0m-1st test) and 1-2 months after vaccination (Im-2nd test). The swabs were collected using a kit with a transport medium, and next a culture of microorganisms was performed on the following media: Columbia agar with 5% sheep blood, Mannitol salt agar, McConkey, Chocolate agar, and Sabouraud (bioMerieux, France), and the phenotypic identification with kits: the APINH, the APIStrep, the APIE, the APINE APIStaph, and the API AUX (bioMerieux, France). The numeric code of the identified strain was read out with the apiweb™ program (bioMerieux, France). The tests were conducted according to the routinely applied microbiological diagnostics.

The patient samples used in the study were bone, tissue, or aspiration fluid, which had been taken intraoperatively from the patients with total hip or total knee arthroplasty (more than one sample had been collected from each patient), as routine diagnostic procedure. Bacteriological examination included conventional culture on growth media for aerobic and anaerobic bacteria and direct sample microscopy of Gram stained smears. The identification of the isolated bacteria was performed by conventional bacteriological methods: API-E and API-NE (Biomerieux, Marcy-I’ Etoile/France) for gram-negative bacteria and coagulase and DNAase for Staphylococcus spp.

Between January and March 2017, 46 samples were analyzed at the Experimental Zooprophylactic Institute of Sicily “A. Mirri.” All the samples, in individually sealed packages, were purchased from different supermarkets in Palermo. They consisted of 23 poultry, 13 beef and 10 pork samples. According to the labels, all the samples came from intensive farms based in Sicily.
The samples were immediately sent to the laboratory on ice and subsequently processed in asepsis.
10 g of each sample was added to 90 ml of saline peptone solution (SSP). After homogenization by Stomacher and incubation for one hour at room temperature, samples were plated into Tryptone Bile X-Glucuronide (TBX) Agar. Colonies developed 18 to 24 hours after incubation at 44°C. They were tested by disk diffusion for resistance to fluoroquinolones, in particular to ciprofloxacin (CIP, 5 μg), norfloxacin (NOR, 10 μg), and levofloxacin (LVX, 5 μg) (Oxoid). A resistant colony was selected from each sample and then identified by the API E (BioMérieux) system.
A single colony was suspended in 200 μl sterile bidistilled water. DNA extraction was then performed by High Pure PCR Template Preparation Kit (Roche), according to the manufacturer's instructions.