Introduction: Industrial livestock farming is a possible source of multi-resistant Gram-negative bacteria, including producers of extended spectrum beta-lactamases (ESBLs) conferring resistance to 3rd generation cephalosporins. Limited information is currently available on the situation of ESBL producers in livestock farming outside of Western Europe. A surveillance study was conducted from January to May in 2014 in four dairy cattle farms in different areas of the Nile delta, Egypt.Materials and Methods: In total, 266 samples were collected from 4 dairy farms including rectal swabs from clinically healthy cattle (n = 210), and environmental samples from the stalls (n = 56). After 24 h pre-enrichment in buffered peptone water, all samples were screened for 3rd generation cephalosporin-resistant Escherichia coli using Brilliance™ ESBL agar. Suspected colonies of putatively ESBL-producing E. coli were sub-cultured and subsequently genotypically and phenotypically characterized. Susceptibility testing using the VITEK-2 system was performed. All suspect isolates were genotypically analyzed using two DNA-microarray based assays: CarbDetect AS-1 and E. coli PanType AS-2 kit (ALERE). These tests allow detection of a multitude of genes and their alleles associated with resistance toward carbapenems, cephalosporins, and other frequently used antibiotics. Serotypes were determined using the E. coli SeroGenotyping AS-1 kit (ALERE).Results: Out of 266 samples tested, 114 (42.8%) ESBL-producing E. coli were geno- and phenotypically identified. 113 of 114 phenotypically 3rd generation cephalosporin-resistant isolates harbored at least one of the ESBL resistance genes covered by the applied assays [blaCTX-M15 (n = 105), blaCTX-M9 (n = 1), blaTEM (n = 90), blaSHV (n = 1)]. Alarmingly, the carbapenemase genes blaOXA-48 (n = 5) and blaOXA-181 (n = 1) were found in isolates that also were phenotypically resistant to imipenem and meropenem. Using the array-based serogenotyping method, 66 of the 118 isolates (55%) could be genotypically assigned to O-types.Conclusion: This study is considered to be a first report of the high prevalence of ESBL-producing E. coli in dairy farms in Egypt. ESBL-producing E. coli isolates with different underlying resistance mechanisms are common in investigated dairy cattle farms in Egypt. The global rise of ESBL- and carbapenemase-producing Gram-negative bacteria is a big concern, and demands intensified surveillance.

Membrane-based spoligotyping has been converted to DNA microarray format to qualify it for high-throughput testing. We have shown the assay's validity and suitability for direct typing from tissue and detecting new spoligotypes. Advantages of the microarray methodology include rapidity, ease of operation, automatic data processing, and affordability.

Please cite this article as: Monecke S, Engelmann I, Archambault M, Coleman DC, Coombs GW, Cortez de Jäckel S, Pelletier-Jacques G, Schwarz S, Shore AC, Slickers P, Ehricht R, Distribution of SCCmecassociated phenol-soluble modulin in staphylococci, Molecular and Cellular Probes (2012), doi: 10.1016/ j.mcp.2012 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. AB547236.1) and S. vitulinus (AB546780.1).The aim of this study was to screen staphylococcal isolates for the presence of PSM-mec in order to obtain more data on its distribution. 102 S. aureus isolates and 38 coagulase-negative staphylococci were genotyped using DNA microarrays (StaphyType, Alere Technologies GmbH, Jena, Germany, [7,9]). This allowed characterising their SCCmec elements [10]. In addition, SCCmec elements of some isolates have previously been studied comprehensively [11,12]. PCR primers pr_psmMEC_02 (5'-CGAAAGCCTGAATGCAAGTCT-3') and pr_psmMEC_03(5'-GGATTTCACTGGTGTTATTACAAGC-3') were used to detect PSM-mec. Reaction conditions included an initial denaturation (2 min at 96°C) followed by 35 cycles (20 sec at 96°C, 20 sec at 70°C and 20 sec at 72°C). A second PCR covered a region spanning from xylR to PSMmec (pr_psmMEC_02 and pr_xylR_04, 5'-AAGCGTCATCTTCTCATTTAGTTGA-3') and a third PCR covered the region from PSM-mec to mecR1 (pr_mecR_01, 5'-CCAGAAAGTAAACAACGATATTCACC-3' and pr_psmMEC_03). Both reactions comprised denaturation (2 min at 96°C) followed by 35 cycles (20 sec at 96°C, 20 sec at 55°C and 70 sec at The results are summarised in Table 1 vitulinus. The presence of PSM-mec did not depend on the host species, and was found in isolates from humans, dogs, cats, goats, cattle, pigs and turkeys. PSM-mec was absent in SCCmec types I, IIC, IIE, IV, V or XI and in SCC elements lacking the mec complex. Some isolates, including CC12-MRSA, WA-MRSA-59, harboured SCCmec elements comprising mecR1 but lacking xylR.These strains were PSM-mec-negative.There was no evidence for alternative locations of PSM-mec. Isolates lacking SCCmec elements, or harbouring SCCmec types I, IIC, IIE, IV and V were PSM-mec-negative regardless of clonal complex or species affiliation. A PCR covering the region from xylR to PSM-mec using primers pr_psmMEC_02/pr_xylR_04 yielded results which were in all cases but one in accordance with the PSM-mec PCR. In a PSM-mec-positive CC5-MRSA, xylR was absent, and primer pair pr_psmMEC_02/pr_xylR_04 did not yield a result.Results of a third PCR (pr_psmMEC/pr_mecR_01) differed from those of the other two PCRs by yielding negative results for ST8-MRSA-IIA or -IID, and for deletion variants of this strain. This can be attrib...

Background Antimicrobial resistance (AMR) is an increasing global health concern reducing options for therapy of infections and also for perioperative prophylaxis. Many Enterobacteriaceae cannot be treated anymore with third generation cephalosporins (3GC) due to the production of certain 3GC hydrolysing enzymes (extended spectrum beta-lactamases, ESBLs). The role of animals as carriers and vectors of multi-resistant bacteria in different geographical regions is poorly understood. Therefore, we investigated the occurrence and molecular characteristics of ESBL-producing Escherichia coli (E. coli) in wild birds and slaughtered cattle in Ibadan, Nigeria. Cattle faecal samples (n = 250) and wild bird pooled faecal samples (cattle egrets, Bubulcus ibis, n = 28; white-faced whistling duck, Dendrocygna viduata, n = 24) were collected and cultured on cefotaxime-eosin methylene blue agar. Antimicrobial susceptibility was determined by agar diffusion assays and all 3GC resistant isolates were genotypically characterised for AMR genes, virulence associated genes (VAGs) and serotypes using DNA microarray-based assays. Results All 3GC resistant isolates were E. coli: cattle (n = 53), egrets (n = 87) and whistling duck (n = 4); cultured from 32/250 (12.8%), 26/28 (92.9%), 2/24(8.3%), cattle, egrets and whistling duck faecal samples, respectively. blaCTX-M gene family was prevalent; blaCTX-M15 (83.3%) predominated over blaCTX-M9 (11.8%). All were susceptible to carbapenems. The majority of isolates were resistant to at least one of the other tested antimicrobials; multidrug resistance was highest in the isolates recovered from egrets. The isolates harboured diverse repositories of other AMR genes (including strB and sul2), integrons (predominantly class 1) and VAGs. The isolates recovered from egrets harboured more AMR genes; eight were unique to these isolates including tetG, gepA, and floR. The prevalent VAGs included hemL and iss; while 14 (including sepA) were unique to certain animal isolates. E. coli serotypes O9:H9, O9:H30 and O9:H4 predominated. An identical phenotypic microarray profile was detected in three isolates from egrets and cattle, indicative of a clonal relationship amongst these isolates. Conclusion Wild birds and cattle harbour diverse ESBL-producing E. coli populations with potential of inter-species dissemination and virulence. Recommended guidelines to balance public health and habitat conservation should be implemented with continuous surveillance.