Commensal Escherichia coli has the potential to easily acquire resistance to a broad range of antimicrobials, making it a reservoir for its transfer to other microorganisms, including pathogens. The aim of this study was to determine the prevalence of resistant commensal Escherichia coli isolated from dairy cows’ feces. Phenotypic resistance profiles and categorization were determined by minimum inhibitory concentration (MIC) testing with the broth microdilution method, while the PCR method was used to determine the presence of resistant genes. Out of 159 commensal E. coli isolates, 39 (24.5%) were confirmed to have resistance. According to the MIC values, 37 (97.3%) and 1 (2.7%) isolate were phenotypically categorized as ESBL and ESBL/AmpC, respectively. All isolates showed resistance to ampicillin, while 97.4%, 56.4%, and 36% showed resistance to cefotaxime, ciprofloxacine, and azitromycine, respectively. Not all isolates that showed phenotypic resistance were found to be carrying the corresponding gene. The most prevalent resistant genes were gyrA, tetA, sul2, and tetB, which were present in 61.5%, 64%, 54%, and 49% of the isolates, respectively. The results clearly indicate that, besides their resistance to multiple antimicrobials, the commensal E. coli isolates did not necessarily carry any genes conferring resistance to that particular antimicrobial.
Campylobacter jejuni is one of the most important food borne pathogens. Since the start of the 21 st century C. jejuni is the leading cause for food borne enteritis. Another point of attention is the change in the antimicrobial resistance of this microorganism towards some critical antimicrobials used in the human and veterinary medicine. In this study samples were taken from three points in the broiler meat production (farm, slaughter line and cold storage of the meat before shipping to the market). A total of 283 samples (cloacal swabs, caeca and carcass swabs) were analyzed for the presence of C. jejuni. The isolates of C. jejuni were confirmed with the conventional microbiological method and with the use of multiplex PCR method. Both methods confirmed the overall prevalence of C. jejuni of 39.2%. In the second part of the study 108 confirmed isolates of C. jejuni were analyzed for the presence of resistance genes (CmeB, Bla oxa-61 , tet(O), aph-3-1 and aadE). The analysis in the third part of the study was concentrated on the antimicrobial resistance of the C. jejuni isolates towards three important antimicrobials (ciprofloxacin, erythromycin and tetracycline). The PCR method used revealed highest prevalence for Bla oxa-61 (25%), followed by CmeB and tet(O) genes (19.4%) and aadE with 13.9%. The aph-3-1 gene was not detected in none of the C. jejuni isolates. C. jejuni isolates in this study showed the highest resistance towards ciprofloxacin (63%) and tetracycline (50%) while the resistance towards erythromycin was very low (5.6%).
Staphylococcus aureus is an important foodborne pathogen due to toxin-related virulence, invasiveness and antibiotic resistance. The ability of S. aureus strains to produce one or more staphylococcal enterotoxins (SEs) in food has been associated with the occurrence of staphylococcal food poisoning (SFP), which is the most common foodborne intoxication worldwide. The study aimed to determine the count of S. aureus strains in samples of raw cow’s milk and various cheeses produced in R. North Macedonia and to detect their ability to produce enterotoxins by passive agglutination SET RPLA (OXOID, UK) and by enzyme-linked fluorescence assay (ELFA) VIDAS SET 2 (Biomerieux, France). A total of 130 S. aureus strains were analyzed. The ability to produce SEs was determined in 17 (13.1%) strains using the SET RPLA detection kit and in 20 (15.4%) strains using the VIDAS SET 2. The study detected enterotoxigenic strains in cheese samples, despite the low count of S. aureus which was below the detection limit according to the Book of rules for microbiological criteria (Off. G. of R.M no 100/2013). Based on these and similar findings, S. aureus must be considered as a possible cause of intoxication, despite the undetected and underreported cases of SFP in the scientific literature.
The aim of the study was to identify the isolation rate of thermotolerant campylobacters in a small-scale broiler-meat production farm over a one-year period. The second deliverable of the study was to determine the potential virulence markers. The laboratory investigation was performed on 283 samples (cloacal swabs, caeca, carcass swabs) collected on three sampling points (farm, slaughter line, and cold storage). The isolates obtained with the conventional microbiological method were confirmed with multiplex PCR for identification of campylobacters. The presence of 10 virulence genes was analyzed in the C. jejuni isolates ( flaA, racR, virB11, dnaJ, wlaN, cadF, ciaB, cdtA, cdtB, cdtC). Out of 283 samples, 169 (59.7%) were confirmed as Campylobacter spp., 111 (39.2%) C. jejuni, and 43 (15.2%) C. coli. C. jejuni was the most prevalent in all sampling points. Campylobacter spp. showed a characteristically seasonal prevalence with the highest isolation rate during the warmer period of the year. We detected the cadF and ciaB genes in all C. jejuni isolates. The flaA gene was present in 50% of the examined strains. The cdt genes (cdtA, cdtB, and cdtC) were confirmed in 52.8%, 52.8%, and 47.2% of the C. jejuni strains, respectively. C. jejuni showed 15 profiles of virulence patterns with four predominant profiles.
β-lactamases are a diverse class of enzymes produced by bacteria that present a major cause for resistance to β-lactams. In this study we analysed 159 fecal samples from dairy cows, for the presence of presumptive ESBL, AmpC, and carbapenemase-producing E. coli. Phylotyping was done using Clermont phylo-typing method, targeting arpA, ChuA, and YjaA genes, along with the DNA fragment TspE4.C2. Convetional PCR method was used to confirm the presence of bla genes among 39 phenotypically confirmed ESBL producing E. coli. The results showed presence of CTX-M, SHV, TEM and OXA1 bla genes in 28 (71.79%), 1 (2.56%), 29 (74.35%), 2 (5.12%) of isolates, respectively Twenty (51.28%) isolates showed presence of both blaCTX-M and TEM genes. The strain that carried the blaSHV gene was found to carry blaTEM gene as well, while one of the strains that carried blaOXA1 gene was also carrying blaCTX-M and TEM gene. The ration between isolates and phylo-groups was as follows: 9 (23.07%) strains were assigned to phyllo-group D; 14 (35.89%) to phyllo-group B; 16 (41.02%) to phyllo-group A. Out of the 39 strains where bla genes were identified, 29 (74.35%) were categorized as multi drug resistant.
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