Detection Of mecA Gene Research Articles

Molecular detection of mecA gene in methicillin-resistant Staphylococcus aureus isolated from dairy mastitis in Nineveh governorate, Iraq

Staphylococcus (S.) aureus is universally the leading aetiologic cause of dairy mastitis. Additionally, methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogenic bacterium in veterinary medicine and public health. Sixty-six cattleʼs milk samples were collected randomly from different areas of the Nineveh Province from November 2018 to February 2020. In this study, the classical and molecular biology methods had used to identify the MRSA and detect the target genes. The results revealed that S. aureus was isolated and identified based on classical methods such as catalase, clumping factors, and coagulase test. In addition, the nuc gene was detected in all the positive S. aureus isolates 23 (34.8%), while the mecA gene was detected in 12 (52.2%) MRSA isolates by using polymerase chain reaction (PCR) assay. The present work emerged that the results of classical methods and the PCR technique were similar. MRSA is regarded as a significant causative agent of various types of bovine mastitis in Iraq, and it can to resist all types of beta-lactams. MRSA isolated from different regions in Mosul city. PCR assay is a powerful method for detecting the different genes based on the target sequence of the specific gene.

Detection of mecA gene and methicillin-resistant Staphylococcus aureus (MRSA) isolated from milk and risk factors from farms in Probolinggo, Indonesia.

Background: Staphylococcus aureus is commonly found in dairy cows and is a source of contamination in milk. S. aureus that are resistant to beta-lactam antibiotics (especially cefoxitin) are referred to as methicillin-resistant Staphylococcus aureus (MRSA). The spread of MRSA cannot be separated from sanitation management during milking; it can originate from milk collected from the udder or from the hands of farmers during the milking process. The purpose of this study was to examine the level of MRSA contamination in dairy cow's milk and farmer's hand. Methods: A total of 109 samples of dairy cow's milk and 41 samples of farmer's hand swabs were collected at a dairy farm in Probolinggo, East Java, Indonesia. Samples were cultured and purified using mannitol salt agar (MSA). The profile of S. aureus resistance was established by disk diffusion test using a disk of beta-lactam antibiotics, namely oxacillin and cefoxitin. Results: The S. aureus isolates that were resistant to oxacillin and cefoxitin antibiotics were then tested for oxacillin resistance screening agar base (ORSAB) as a confirmation test for MRSA identity. S. aureus isolates suspected to be MRSA were then tested genotypically by polymerase chain reaction (PCR) method to detect the presence of the mecA gene. The results of the isolation and identification found 80 isolates (53.33%) of S. aureus. The results of the resistance test found that 42 isolates (15%) of S. aureus were resistant to oxacillin and 10 isolates (12.5%) were resistant to cefoxitin. The ORSAB test found as many as 20 isolates (47.62%) were positive for MRSA. In PCR testing to detect the presence of the mecA gene, three isolates (30%) were positive for the mecA gene. Conclusions: This study shows that several S. aureus isolates were MRSA and had the gene encoding mecA in dairy farms.

Molecular epidemiology, virulence and antimicrobial resistance of Bulgarian methicillin resistant Staphylococcus aureus isolates.

Severe infections of virulent methicillin-resistant Staphylococcus aureus (MRSA) are a serious health problem. The present study aimed to investigate clonal spread, virulence and antimicrobial resistance rates of Bulgarian MRSA isolates in 2016-2020. Molecular identification and mecA gene detection were performed with PCR. Clonal relatedness was evaluated by RAPD PCR and MLST. MRSA epidemiology, virulence and resistance patterns were investigated by PCR. All 27 isolates were identified as S. aureus and were mecA positive, and all were susceptible to linezolid, tigecycline and vancomycin. The toxin genes hlg (in 92.6% of isolates), seb (77.8%), sei (77.8%), seh (59.3%), sej (55.6%), and seg (48.1%), were frequently found among the isolates. Epidemiological typing by RAPD identified 4 clones (16 isolates) and 11 were with a unique profile. MLST analysis of the same MRSA isolates showed five MLST clonal complexes and 11 ST types, including CC5 (33.3%) (ST5, ST221, ST4776), CC8 (22.2%) (ST8, ST239, ST72), CC15 (ST582), CC22 (14.8%) (ST217, ST5417), CC30 (ST30) CC398 (ST398), and CC59 (ST59). The isolates from CC5 showed higher virulence potential and almost all were macrolide resistant (ermB or ermC positive). CC8 isolates showed higher level of resistance. To the best of our knowledge, this study is the first describing the clonal spreading of Bulgarian MRSA and the association with their virulence and resistance determinants. Monitoring of MRSA epidemiology, resistance and virulence profile can lead to better prevention and faster therapeutic choice in cases of severe infections.

Detection of mecA gene and methicillin-resistant Staphylococcus aureus (MRSA) isolated from milk and risk factors from farms in Probolinggo, Indonesia.

Background: Staphylococcus aureus is commonly found in dairy cows and is a source of contamination in milk. S. aureus that are resistant to beta-lactam antibiotics (especially cefoxitin) are referred to as methicillin-resistant Staphylococcus aureus (MRSA). The spread of MRSA cannot be separated from sanitation management during milking; it can originate from milk collected from the udder or from the hands of farmers during the milking process. The purpose of this study was to examine the level of MRSA contamination in dairy cow's milk and farmer's hand. Methods: A total of 109 samples of dairy cow's milk and 41 samples of farmer's hand swabs were collected at a dairy farm in Probolinggo, East Java, Indonesia. Samples were cultured and purified using mannitol salt agar (MSA). The profile of S. aureus resistance was established by disk diffusion test using a disk of beta-lactam antibiotics, namely oxacillin and cefoxitin. Results: The S. aureus isolates that were resistant to oxacillin and cefoxitin antibiotics were then tested for oxacillin resistance screening agar base (ORSAB) as a confirmation test for MRSA identity. S. aureus isolates suspected to be MRSA were then tested genotypically by polymerase chain reaction (PCR) method to detect the presence of the mecA gene. The results of the isolation and identification found 80 isolates (53.33%) of S. aureus. The results of the resistance test found that 42 isolates (15%) of S. aureus were resistant to oxacillin and 10 isolates (12.5%) were resistant to cefoxitin. The ORSAB test found as many as 20 isolates (47.62%) were positive for MRSA. In PCR testing to detect the presence of the mecA gene, three isolates (30%) were positive for the mecA gene. Conclusions: This study shows that several S. aureus isolates were MRSA and had the gene encoding mecA in dairy farms.

Characterization of the virulence, agr typing and antimicrobial resistance profile of Staphylococcus aureus strains isolated from food handlers in Brazil.

Staphylococcus aureus is one of the main pathogens associated with foodborne outbreaks in Brazil and food handlers can carry toxigenic and resistant S. aureus strains. The aims of this study were to verify the frequency of virulence genes, to identify the agr groups and to determine the antimicrobial resistance profile of S. aureus strains isolated from food handlers of pilot kitchens located in São Paulo, Brazil. A total of 74 strains of the Staphylococcus genus were detected and 50% were identified as of the species S. aureus. The enterotoxin genes detection, tst and luk-PV detection, agr typing, mecA detection, ccr complex detection and SCCmec typing were performed using PCR. The antimicrobial resistance testing was performed by the disk diffusion method. The enterotoxin genes were identified in 36 S. aureus, including sea (83.8%). The tst gene was detected in 18.92% of the strains and the luk-PV was detected in only one isolate. Agr typing classified 58.3% of the strains as type I. Seven (18.92%) strains were classified as MRSA and the ccr2 complex was detected in six of these isolates. The SCCmec typing characterized strains as type II, III, IV and V. Moreover, there were also a greater number of resistant strains to penicillin (83.78%) and clarithromycin (67.57%). In conclusion, the study revealed a significant prevalence of S. aureus, and the presence of different virulence genes and a worrying resistance profile in S. aureus strains isolated from food handlers in this country.

Detection of mecA gene-associated methicillin-resistance coagulase-negative staphylococci (MRCoNS) from bovine mastitis in Gujarat

Exploration of pathogenic potential of Coagulase-Negative Staphylococci (CoNS), in human as well as animals,gained the importance during the past two decades. Emergence of Methicillin-Resistant Coagulase-NegativeStaphylococci (MRCoNS) in bovine milk poses to be a major public health concern. Presence of the mecA gene isone of the most common reasons reported for the methicillin resistant bacteria. Since the scientific data concerningthe presence of mecA gene and MRCoNS in bovine milk is very limited in India, particularly from Gujarat state,the present study was planned to detect mecA gene in 25 MRCoNS and study the antibiogram of 33 CoNS isolatedfrom 185 bovine mastitis cases between January 2018 to December 2019. Staphylococci were isolated from themilk samples of bovine mastitis with a prevalence of 38.9% (72/185), out of which 39 (54.16%) and 33 (45.83%)isolates were identified as coagulase-positive Staphylococci (CoPS) and coagulase-negative Staphylococci (CoNS),respectively. Among the CoNS, S. xylosus was the most predominantly isolated species (9/33, 27.27%), followedby S. epidermis (6/33, 18.18%) and S. haemolyticus (4/33, 12.12%). Out of these 33 isolated CoNS, 21 (63.63%)and 25 (75.75%) were identified as MRCoNS by disc diffusion method and CHROM agar, respectively. Contraryto the likelihood, only 3 MRCoNS showed the presence of mecA gene using PCR method. Antibiogram revealedthat most of the CoNS isolates (84.85%) were multi-drug resistant emphasizing the urgent need of restricting theindiscriminate use of antimicrobial drugs in the area of study.

It's Not You, It's SOSA: A Case Study on Breaking Up With an FDA-Cleared Susceptibility Testing System's Oxacillin Results for Staphylococcus spp. Other Than S. aureus and S. lugdunensis.

BackgroundIn 2021, the Clinical and Laboratory Standards Institute revised its susceptible oxacillin minimum inhibitory concentration (MIC) breakpoint for Staphylococcus spp. other than S. aureus and S. lugdunensis (SOSA) from ≤0.25 to ≤0.5 µg/mL. Here, we describe the response to this breakpoint change, which at the time of this study was not yet recognized by the US Food and Drug Administration (FDA), in our laboratory, where the primary method for antimicrobial susceptibility testing (AST) of SOSA is VITEK 2. VITEK 2 uses the Automated Expert System (AES) to integrate the results of oxacillin MIC and cefoxitin screen tests into a final interpretation; our laboratory also adjudicates discordant oxacillin and cefoxitin results using a PBP2a test.MethodsWe retrospectively reviewed and assessed the yield of PBP2a testing for 189 SOSA isolates with discordant (when applying the FDA susceptible oxacillin breakpoint of ≤0.25 µg/mL) VITEK 2 oxacillin and cefoxitin results, and then prospectively incorporated PBP2a testing for isolates with oxacillin MICs of 0.5 µg/mL and positive cefoxitin screens into our algorithm.ResultsCompared with accepting the VITEK 2 AES interpretation, PBP2a testing substantially improved the accuracy of mecA-mediated resistance classification in both scenarios, especially for the ∼4.7% of isolates with oxacillin MICs ≤0.5 µg/mL and positive cefoxitin screens.ConclusionsAlthough detection of mecA or PBP2a is the gold standard for assessment of β-lactam resistance in staphylococci, targeting a subset of isolates for mecA or PBP2a testing based on phenotypic AST results that predict an increased risk of misclassification may be a pragmatic, labor- and cost-saving approach.

Molecular Characterization of Staphylococci Recovered from Hospital Personnel and Frequently Touched Surfaces in Tianjin, China.

Staphylococci are major hospital-associated pathogens, and the dissemination of methicillin-resistant staphylococci in hospitals poses a great challenge for managing hospital-acquired infections. Little is known about the dissemination of staphylococci recovered from the hospital environment and personnel in China. In this study, antimicrobial susceptibility tests, mecA gene detection, SCCmec typing, and multilocus sequence typing (MLST) were performed to clarify the molecular epidemiology of staphylococci in a large hospital in Tianjin, China. One hundred and ninety-five staphylococci were recovered, and 94% of isolates were resistant to at least one antibiotic. Eighty-five staphylococci were mecA gene-positive, and 40% of them harbored SCCmec IV and V. The genotype of Staphylococcus aureus (S. aureus) was ST25, and the dominant genotype of methicillin-resistant Staphylococcus epidermidis (MRSE) was ST59. Three new sequence types were assigned as ST840, ST841, and ST842. One (2%) frequently touched surface was contaminated by S. aureus, which suggested that environmental contamination occurred in the hospital in China. Nineteen (39%) frequently touched surfaces were contaminated by methicillin-resistant coagulase-negative staphylococci (MRCoNS), and 46% of HP carried MRCoNS. Varied staphylococcal species and antimicrobial-resistance rates were observed between staphylococci that were recovered from hospital personnel and frequently touched surfaces. The transmission of MRSE and S. aureus between hospital personnel and frequently touched surfaces was detected. Hospital items and personnel may act as reservoirs of antimicrobial-resistant staphylococci, and cleaning strategies should be carried out to decrease the dissemination of antimicrobial-resistant staphylococci in hospitals in China.

Antibiogram and detection of mecA gene among MRSA at Specialist Hospital Sokoto

Antibiogram and detection of mecA gene among MRSA at Specialist Hospital Sokoto

“Evaluation Of Several Phenotypic Methods Of Antibiotic Susceptibility Pattern For The Detection Of MRSA With Molecular Profiling RT-PCR With The Detection Of MecA Gene”

INTRODUCTION:- Methicillin-resistant Staphylococcus aureus (MRSA) is a well-recognized public health problem throughout the world and community MRSA strains are now epidemic in India and others countries. The resistance rates of S. aureus infection and multidrug resistant strains are increasing, making the clinical anti-infective treatment and diagnosis more difficult.
 AIM:- To evaluate several phenotypic methods of the antibiotic susceptibility pattern for the detection of MRSA with Molecular Profiling Rt-PCR with the detection of MecA gene.
 MATERIALS AND METHODS:- The present study was a prospective observational study carried out at the Department of Microbiology, Santosh Medical College, Ghaziabad in a tertiary care hospital over a 12-month period from August 1, 2020 to July 31, 2021. Both IPD and OPD clinical sample comprised 385 clinical isolates as Staphylococcus aureus from various clinical specimens were included. Antibiotic susceptibility by the Kirby-Baure disc diffusion methods was performed according to the Clinical and Laboratory Standard Institute (CLSI) guidelines. Detection of MRSA by various methods like, Cefoxitin Disk diffusion Method, Oxacillin Disk diffusion method, E-Test method and detection of MecA gene by RT-PCR was carried out.
 RESULT:- In the present study out of 384 clinical samples, the majority of isolates from MRSA were found to be resistant to (E-Test) strip Oxacillin 114 (29.7%), followed by cefoxitin 113 (29.4%) and disc diffusion oxacillin 99 (25.8%). However, we observed a high incidence of resistance to other antibiotics such as Erythromycin 265 (69.0%), followed by Clotrimozole 228 (59.4%), Tetracyclin 144 (37.5), Vancomycin 102 (26.6%) and Refampicin 102 (26.6%). We also observed that Linezolid 354 (92.2%) followed by Teicoplanin 325 (84.6%), Gentamycin 272 (70.8%), Clindamycin 249 (64.8%) and Amoxyclave 281 (73.2%). In this study molecular RT-PCR test for the detection of mecA gene observed more in pus with 36 samples out of 113 followed by blood 17, urine 33, Sputum 05, pleural fluid 04, wound swab 16, Vaginal swab 06, CSF 01, Throat swab 2.
 CONCLUSION:- The gold standard assay for determining methicillin resistance is the PCR technique for detecting the mecA gene. But Because PCR is still time intensive and expensive, and it is not yet available in the 95% of routine clinical laboratories the phenotypic methods is still the choice where E-test is more reliable method than the disc diffusion method in detecting the drug resistance and can be utilised on a regular basis for better results.

Analytical performance and potential clinical utility of the GenMark Dx ePlex® blood culture identification gram-positive panel

The test performance and potential clinical utility of the ePlex® BCID Gram-Positive (GP) Panel was evaluated relative to MALDI-TOF mass spectrometry on bacterial isolates and traditional antimicrobial susceptibility testing. All GP bacteria (n=100) in the study were represented on the panel including 50 common skin contaminants, and 7/7 coinfections. The positive percent agreement (PPA) was 97/97 with 2 false positives. Detection of vanA yielded a PPA of 4/4 and NPA of 9/9. mecA gene detection exhibited a PPA of 14/14 and NPA of 14/14 for S. aureus and a PPA of 31/32(97%) and NPA of 16/16 for CNS with 1 false negative. Chart reviews (n=80) identified a mean 24.4h faster time to organism identification, 53.4h earlier optimization in 15(18.8%) patients based on AMR gene detection, 29.2h earlier optimization for 8(10%) patients infected with organisms, such as streptococci, with very low resistance rates, and 42.9h earlier discontinuation of antimicrobials for 14(17.5%) patients with contaminant cultures.

Virulence Genes of Staphylococcus aureus Associated With Keratitis, Conjunctivitis, and Contact Lens-Associated Inflammation.

Purpose Staphylococcus aureus, cause a range of ocular diseases in humans, including noninfectious corneal infiltrative events (niCIE), infectious conjunctivitis and sight threatening microbial keratitis (MK). This study aimed to determine the possession of known virulence genes of S. aureus associated with MK and conjunctivitis, in strains isolated from these conditions and niCIE.MethodsSixty-three S. aureus strains—23 from MK, 26 from conjunctivitis, and 14 from niCIE—were evaluated for possession of genes. Polymerase chain reaction was used for the detection of mecA and 10 known virulence genes involved in MK (clfA, fnbpA, eap, coa, scpA, sspB, sspA, hla, hld, and hlg), 2 associated with conjunctivitis (pvl and seb).Results mecA was present in 35% of infections and 7% of niCIE strains (P = 0.05). It was not seen in infection strains from Australia. Adhesion genes were found in all strains except clfA, which was found in 75% of infection and 93% of niCIE strains. Invasion genes were found in higher frequency in infections strains—hlg (100% vs. 85%; P = 0.04) and hld (94% vs. 50%; P = 0.005)—compared with niCIE strains. Evasion genes were common in infection strains except scpA, which was found at a significantly higher frequency in niCIE strains (86%) compared with infection strains (45%; P = 0.001).ConclusionsThe higher rates of hlg and hld in strains isolated from infections than niCIE may have a role in pathogenesis, whereas scpA may be an important virulence factor during niCIEs.Translational RelevanceThis study has identified virulence factors involved in the ocular pathogenesis of S. aureus infections and niCIE.

Detection of mecA gene and methicillin-resistant Staphylococcus aureus (MRSA) isolated from milk and risk factors from farms in Probolinggo, Indonesia.

Background: Staphylococcus aureus is commonly found in dairy cows and is a source of contamination in milk. S. aureus that are resistant to beta-lactam antibiotics are referred to as methicillin-resistant Staphylococcus aureus (MRSA). The spread of MRSA cannot be separated from sanitation management during milking; it can originate from milk collected from the udder or from the hands of farmers during the milking process. The purpose of this study was to examine the level of MRSA contamination in dairy cow's milk and farmer's hand swabs. Methods: A total of 109 samples of dairy cow's milk and 41 samples of farmers' hand swabs were collected at a dairy farm in Probolinggo, East Java, Indonesia. Samples were cultured and purified using mannitol salt agar (MSA). The profile of S. aureus resistance was established by disk diffusion test using a disk of beta-lactam antibiotics, namely oxacillin and cefoxitin. Results: The S. aureus isolates that were resistant to oxacillin and cefoxitin antibiotics were then tested for oxacillin resistance screening agar base (ORSAB) as a confirmation test for MRSA identity. S. aureus isolates suspected to be MRSA were then tested genotypically by polymerase chain reaction (PCR) method to detect the presence of the mecA gene. The results of the isolation and identification found 80 isolates (53.33%) of S. aureus. The results of the resistance test found that 42 isolates (15%) of S. aureus were resistant to oxacillin and 10 isolates (12.5%) were resistant to cefoxitin. The ORSAB test found as many as 20 isolates (47.62%) were positive for MRSA. In PCR testing to detect the presence of the mecA gene, three isolates (30%) were positive for the mecA gene. Conclusions: This study shows that several S. aureus isolates were MRSA and had the gene encoding mecA in dairy farms.

Distribution of Leukocidins, Exfoliative Toxins, and Selected Resistance Genes Among Methicillin-resistant and Methicillin-sensitive Staphylococcus aureus Clinical Strains in Egypt

Background: Infection with Staphylococcus aureus (S.aureus) is an increasing health problem worldwide. This pathogen has multiple virulence factors that contribute to its pathogenesis in a wide range of diseases. The present study aimed to investigate the prevalence of leukocidins, exfoliative toxins, and common antimicrobial resistance genes among Methicillin-Resistant Staphylococcus aureus (MRSA) and Methicillin-Sensitive Staphylococcus aureus (MSSA) strains collected from various clinical sources in Egypt. Methods: Isolates were identified as S.aureus by the standard microbiological methods. Methicillin resistance was detected phenotypically by cefoxitin disc diffusion method and genotypically by PCR for detection of mecA gene. PCR was also used to detect the presence of leukocidin genes (LukD, LukE, LukF-PV, and LukS-PV), exfoliative toxin genes (eta and etb), and antibiotic resistance genes (tetK, tetM, ermA, ermC, msrA, and aacA-aphD). Results: About 50.5% of tested isolates were methicillin resistant by cefoxitin disc assay, while mecA gene was amplified in 64.6% of isolates. The highest prevalent toxin gene was lukE (93%) and the least prevalent one was eta (1%). The resistance genes tetK and tetM were detected in nearly 50% of the tested strains but lower prevalence rates were recorded for aacA-aphD, msrA, ermA, and ermC genes. Conclusion: Methicillin resistance was highly prevalent among tested S.aureus strains. Regarding the studied virulence and resistance genes, no significant difference was detected between MRSA and MSSA strains, except for ermA gene p<0.05 which was highly prevalent in MRSA strains. So, the variation between MRSA and MSSA strains in the response to treatment may be attributed to the resistance of MRSA strains to all β-lactams in addition to other possible acquired resistance mechanisms. Accordingly, fewer options of antimicrobial medications are available to treat MRSA infections.

Diversity of staphylococcal cassette chromosome mec (SCCmec) types among methicillin-resistant coagulase-negative staphylococci (MR-CoNS)

Methicillin-resistant coagulase-negative staphylococci (MR-CoNS) have become one of the important causes of nosocomial infections yet their clinical data in Malaysia is scarce compared to methicillin-resistant Staphylococcus aureus (MRSA). Staphylococcal Cassette Chromosome mec (SCCmec) genes play roles in their pathogenicity. This study thus aimed to determine species distribution, antimicrobial susceptibility pattern and SCCmec types among MR-CoNS isolated from blood cultures. A laboratory-based descriptive study was involved with non-probability sampling method. One hundred CoNS isolated from blood cultures were collected from Microbiology laboratory, Hospital Serdang and proceeded with phenotypic identification, antimicrobial susceptibility testing, mecA gene detection and SCCmec types classification. Staphylococcus epidermidis was the most common isolated MR-CoNS species. All 100 isolates were resistant to penicillin while being sensitive to vancomycin. The predominant SCCmec Type IV was observed in S. epidermidis which exhibited 100% resistant to penicillin and erythromycin besides dominating multiple antibiotic resistance. Meanwhile, the combination type was observed in type I & IVa (n=9, 9%) whereas 31 strains (31%) were non-typeable. Besides demonstrating MRCoNS susceptibility pattern variations to commonly used antimicrobials for treatment of staphylococcal infections, this study could also preliminarily contribute in providing more local epidemiological data regarding MR-CoNS.

Occurrence and characterization of methicillin-resistant Staphylococcus spp. in diseased dogs in Brazil.

Staphylococcus pseudintermedius is a major commensal bacterium of the skin and mucosae of dogs and an opportunistic agent responsible for several clinical infections, such as pyoderma, otitis, and surgical wound infections. The emergence of methicillin-resistant S. pseudintermedius (MRSP) has become a problem of great concern in veterinary and human medicine because it is multidrug resistant (MDR) and can also infect humans. This study aimed to identify the occurrence of Staphylococcus spp. in infected patients and investigate the antimicrobial resistance profiles and molecular structure of MRSP isolates. Samples were obtained from two different veterinary clinics; suggestive colonies were submitted to matrix-assisted laser desorption ionization-time of flight (MALDI-ToF) mass spectrometry and confirmed at the species level by polymerase chain reaction (PCR). Sequencing of the 16S rRNA and rpoB genes were used in selected samples that were not identified by MALDI-ToF and by the species-specific PCR. Antimicrobial susceptibility and PCR detection of mecA were performed. MRSP isolates were subjected to multilocus sequence typing. Of all the clinical staphylococci (n = 131), 98 (74.8%) were identified as S. pseudintermedius. Multidrug resistance (resistance to ≥3 classes of antimicrobials) was observed in 63.2% of S. pseudintermedius isolates, and 24.5% of S. pseudintermedius isolates were methicillin-resistant. Half of the MRSP isolates were isolated from surgical site infections. Among the ten sequence types (ST) identified, nine were novel. ST71 was the most prevalent and associated with resistance to fluoroquinolones. Prior antimicrobial therapy, hospitalization, and surgical site infections seemed to be risk factors for MRSP acquisition. The present study showed a high rate of MDR staphylococci in infected dogs. MRSP was isolated from different clinical conditions, mainly surgical site infections. Additionally, this is the first study to extensively investigate the population structure of MRSP in Brazil, which revealed the dispersion of CC71 and nine novel ST. These findings raise concerns for both animal and human health due to the zoonotic potential of this species and limited therapeutic options available for MRSP infections.

High Levels of Methicillin-Resistant Staphylococcus aureus Carriage Among Healthcare Workers at a Teaching Hospital in Addis Ababa Ethiopia: First Evidence Using mecA Detection.

BackgroundStaphylococcus aureus is a major human pathogen and causes healthcare and community-acquired infection. Data on the extent of MRSA colonization among health-care workers (HCWs) in sub-Saharan Africa are limited. Hence, we determined the burden of MRSA colonisation among HCWs and administrative staff in Tikur Anbessa Specialised Hospital (TASH), College of Health Sciences (CHS), Addis Ababa University, Ethiopia.MethodsUsing a cross-sectional study design, participants were screened for MRSA colonisation between June 2018 and August 2019 using nasal swabs. The swabs were analysed using standard laboratory methods including antibiotic resistance gene, mecA. Anonymised sociodemographic data were collected by pretested questionnaires to evaluate HCWs factors associated with MRSA carriage.ResultsA total of 588 HCWs and 468 administrative staff were screened for MRSA. Women were over-represented. Overall, 49.1% (289/588) of HCWs were nurses and 25% (117/468) of the administrative staff were cleaners or laundry workers. Overall, 138 S. aureus isolates were retrieved from the nasal swabs of both groups (16.3%, 96/588 from HCWs). The burden of MRSA colonisation was 4.8% (28/580, 95% CI: 3.1–6.5%) among HCWs compared to 0.2% (1/468, 95% CI: 0.18–0.6%) of administrative staff (p value <0.05). The majority of S. aureus and all MRSA isolates were resistant to penicillin. Isolates from HCWs were more resistant to tested antibiotics than administrative staff (P-value <0.05).ConclusionThis is the first report in Ethiopia on MRSA colonization using mecA and revealed that; (i) overall carriage rates of MRSA in HCWs are comparable with observations reported in some other countries and (ii) HCWs exhibit a higher burden of MRSA carriage than administrative staff. Our data support strategic screening of MRSA and antimicrobial stewardship for better intervention measures.

O03 ST97 Staphylococcus aureus and oxacillin resistance: an emerging challenge for microbiologists?

BackgroundThe presence of cefoxitin and oxacillin resistance in Staphylococcus aureus isolates is suggestive of methicillin resistance (MRSA) with molecular detection of the mecA or mecC genes providing confirmation. The UK SMI and EUCAST guidelines for MRSA detection describe a subset of strains that exhibit reduced susceptibility to oxacillin and cefoxitin, though negative for mecA and mecC genes—frequently named borderline oxacillin-resistant S. aureus (BORSA). Local data noted a number of S. aureus isolates that matched the above phenotype leading to laboratory, clinical and epidemiological questions.MethodsRoutine samples were collected between August 2020 and April 2021 at West Midlands Health Security Agency Laboratory, Birmingham. If provisional results suggested an MRSA isolate, the following confirmatory tests were undertaken. Locally, clinical samples underwent susceptibility testing with cefoxitin disc diffusion and oxacillin gradient diffusion (MRSA screens) or Vitek 2 (non-MRSA screens). Isolates with discrepant results [susceptible to cefoxitin; resistant to oxacillin (MIC >2 mg/L) or vice versa] were sent to the reference laboratory at Colindale, UK for further testing, which included mecA or mecC gene detection by PCR and serotyping with Illumina sequencing for all isolates.ResultsIn total, 53 isolates were sent to the reference lab of which 22 were confirmed to have an ST97 serotype. All 53 were negative by PCR for mecA and mecC genes. For the 22 isolates of ST97, local oxacillin MIC values ranged from 2 to 6 mg/L. Cefoxitin susceptibility was confirmed locally in all isolates. The 22 isolates were from 15 patients; wound swabs (n = 18); blood culture (n = 1), sputum (n = 1) and MRSA screen (n = 2). Median patient age was 48 years (IQR 38–59) with 5 being female and 10 male. A history of injecting drug use was documented in 53% (8/15).ConclusionsBORSA remains a problem from a laboratory, clinical and infection control perspective. This work raises two important questions: what is the most appropriate local laboratory testing pathway and what is the clinical relevance of these isolates (i.e. can flucloxacillin be relied upon in treatment)? The ST97 serotype appears to be associated with skin and soft tissue infection and may be linked to people who inject drugs.

Microbiological profile of diabetic foot infections and the detection of mecA gene in predominant Staphylococcus aureus

BACKGROUNDDiabetes mellitus (DM) is a serious health problem that is rapidly expanding worldwide. Staphylococcus aureus is a pathogenic bacterium which has a number of drug resistant strains. Different variants of this pathogen have been isolated from patients with diabetic foot ulcers - in persons having uncontrolled blood sugar level - all over the world, resulting in high rates of morbidity and mortality. The objective of this study was to determine the prevalence of drug resistant Staphylococcus aureus in diabetic foot infections (DFIs). METHODS An epidemiological survey was conducted and 300 pus samples were collected from wounds, abscesses, skin and soft tissue lesions of patients having type II diabetes with foot ulcer infections at a tertiary care hospital. Further, the antibacterial susceptibility patterns of all the isolated Staphylococcus aureus were determined against methicillin, oxacillin, vancomycin and novobiocin. RESULTS Pathogenic bacterial species including coagulase positive and coagulase negative Staphylococcus aureus, Escherichia coli, Klebsiella sp., Proteus sp., Pseudomonas sp., and Citrobacter sp. were identified, among which Staphylococcus was the main genus identified. A total of 13 (4.3%) isolates of coagulase positive Staphylococcus aureus were resistant to methicillin. Using PCR, 7 (53.8%) staphylococcal isolates were detected with the mecA gene. CONCLUSIONStaphylococcus aureus is the most common cause of DFIs. This study demonstrates that about 53.8% of all methicillin resistant Staphylococcus aureus isolates have mecA genes. Such a finding is the primary step in understanding and tackling the resistance mechanism.

Detection of Resistant and Enterotoxigenic Strains of Staphylococcus warneri Isolated from Food of Animal Origin.

The topic of this work is the detection of antimicrobial resistance to Staphylococcus warneri strains and the genes encoding staphylococcal enterotoxins. It is considered a potential pathogen that can cause various—mostly inflammatory—diseases in immunosuppressed patients. The experimental part of the paper deals with the isolation of individual isolates from meat samples of Oryctolagus cuniculus, Oncorhynchus mykiss, Scomber scombrus, chicken thigh, beef thigh muscle, pork thigh muscle, and bryndza cheese. In total, 45 isolates were obtained and subjected to phenotypic (plasma coagulase activity, nuclease, pigment, hemolysis, lecithinase, and lipase production) and genotypic analyses to confirm the presence of the S. warneri species. The presence of genes encoding staphylococcal enterotoxins A (three isolates) and D (six isolates) was determined by PCR. Using the Miditech system, the minimum inhibitory concentration for various antibiotics or antibiotics combinations was determined, namely for ampicillin; ampicillin + sulbactam; oxacillin; cefoxitin; piperacillin + tazobactam; erythromycin; clindamycin; linezolid; rifampicin; gentamicin; teicoplanin; vancomycin; trimethoprim; chloramphenicol; tigecycline; moxifloxacin; ciprofloxacin; tetracycline; trimethoprim + sulfonamide; and nitrofurantoin. Resistance to ciprofloxacin and tetracycline was most common (73%). At the same time, out of a total of 45 isolates, 22% of the isolates were confirmed as multi-resistant. Isolates that showed phenotypic resistance to β-lactam antibiotics were subjected to mecA gene detection by PCR.