Hybridizations were performed as suggested by Bruant [15] An amo

Hybridizations were performed as suggested by Bruant [15]. An amount of 500 ng of labelled DNA was dried under vacuum in a rotary desiccator (Savant SpeedVac?, ArrayIt, Holbrook, NY, USA) and resuspended in a hybridization buffer (Dig Ease Buffer, Roche Diagnostics spa, Milan, Italy). Before hybridization, microarrays were pre-hybridized for at least one hour at 42 ��C in a pre-heated pre-hybridization solution containing 5X SSC, 0.1% SDS (Sigma Aldrich spa, Milan, Italy) and 1.0% BSA (Sigma Aldrich spa). After pre-hybridization, the microarrays were hybridized mixing a solution of Dig Easy Hyb buffer (Roche Diagnostics), Bakers Yeast tRNA (10 mg/ml) (Sigma Aldrich spa), Sonicated Salmon Sperm DNA (10 mg/mL) (Sigma Aldrich spa) with previously denatured labelled DNA.

Microarrays were hybridized overnight at 42 ��C in a SlideBooster (Advalytix, ABI, Milan, Italy). After hybridization, the slides were washed with increasing stringency washes (1X SSC, 0.1% SDS preheated to 42 ��C; 1X SSC and 0.1X SSC at room temperature). Microarray slides were scanned using a ScanArray Lite fluorescent microarray analysis system (Perkin Elmer, Milan, Italy) at excitation wavelengths of 532 nm (Cy3) and 635 (Cy5) and then analysed with the ScanArray Gx software (Perkin Elmer). Images were examined using the QuantArray software version 3.1 (Packard Bioscience, Boston, MA, USA).The data were normalized as described previously [17]. For each subarray, after subtraction of local background intensity, the median value for each set of triplicate spotted probes was divided by the empty signal and then logarithmically transformed.

The data file was then elaborated with Cluster software [18,19]. Strains were clustered by hierarchical clustering using the algorithm Centered Pearson Dacomitinib Correlation Distance and Pairwise Maximum Linkage method. For visualization of the elaborated data, Java TreeView, an Open Source program, was utilised [18�C20].3.?Results and DiscussionMultiplex PCR identified 62.75% of the isolates as C. jejuni and 37.24% as C. coli (Table 1). In this study the antimicrobial resistance and two methods (PFGE and microarray) for genome analysis of C. jejuni and C. coli strains were evaluated.The antibiotic resistance profiles of the isolates are shown in Table 2. In particular, 100 (68.97%) isolates were resistant to at least one antibiotic, whereas the remaining strains (31.03%) were susceptible to all antibiotics tested. The highest levels of resistance were found for ciprofloxacin (62.76%), tetracycline (55.86%) and nalidixic acid (55.17%). In contrast, only 19 (13.10%) strains were resistant to erythromycin, 7 (4.83%) strains to streptomycin and only one (0.69%) isolate to chloramphenicol.

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