Isolates Of Methicillin-resistant Research Articles

P-1350. Novel mutations in PBP1 and AcrB associated with methicillin resistance in clinical isolates of S. aureus lacking mec variants

Abstract Background Methicillin-resistant S. aureus (MRSA) is a major global pathogen. In the absence of mecA and mecC, methicillin resistance has been associated with genetic changes, such as PBP1 to PBP4 (Penicillin-Binding Protein) and GdpP (c-di-AMP phosphodiesterase) mutations. However, this unusual genotype has not been reported in Colombia. Here, the aim was to characterize the genetic determinants in two methicillin-resistant clinical isolates of S. aureus lacking mec variants in Colombia. Table 1 Genetic changes associated with methicillin resistance lacking mec variants in SaA and SaB isolates. Methods One isolate (SaA) was recovered in 2015 from a urine culture and the second (SaB) was obtained in 2016 from a peritoneal fluid secretion. OXA susceptibility was assessed via broth microdilution according to CLSI M100 guidelines, and species confirmation and mecA detection were performed using multiplex PCR. Genome characterization by Illumina Whole Genome Sequencing was conducted to evaluate the presence of mec variants (A, B, or C) and determinants associated with OXA resistance lacking mec variants (mutations in PBP1 to PBP4, pbp4 promoter, GdpP, and AcrB). Results Both isolates were identified as MRSA (OXA MIC of 16 ug/mL) but lacked mecA by PCR. In addition, genomic characterization showed that SaA (ST-5/CC-5/agr-III) and SaB (ST-765/CC-30/agr-I) do not harbor any mec variants. Moreover, the bla operon was complete in SaB, whereas was absent in SaA. Notably, we found genetic changes in SaA and SaB previously associated with methicillin resistance lacking mec (Table 1), some of which were common in both isolates (PBP1: N118D, PBP3: T438S and PBP4: T409A). Mutations in PBP2 were only detected in SaB. Remarkably, novel mutations, such as D604G in PBP1 and AcrB mutations (T52S, A282T, P285S, A577T, A677V, A711T, and V747I) were identified (Table 1). Conclusion Novel mutations in PBP1 and AcrB were found in clinical isolates of MRSA lacking mec variants, as well as previously documented genetic changes. This emergent unusual genotype needs further investigation. Disclosures All Authors: No reported disclosures

Asymptomatic carriage and molecular characterization of Staphylococcus aureus in pre-clinical and clinical medical students.

Medical students are exposed to the hospital environment and patients during their studies, increasing the risk of exposure to virulent and antibiotic-resistant isolates of Staphylococcus aureus. The aim of the study is to determine the prevalence of Staphylococcus aureus among medical students who have varying levels of exposure to the hospital environment to provide valuable insights into the risk of colonization and transmission. Nasal swabs and fingerprints were obtained and cultured on a selective medium for staphylococci. The obtained isolates were confirmed as methicillin-sensitive S. aureus (MSSA) or methicillin-resistant (MRSA) using PCR. Antibiotic resistance, the presence of virulence genes including enterotoxin encoding genes, and spa typing were performed. Among pre-clinical students, MSSA was detected on the nose in 45.2% and on the fingerprints in 10.6% of the participants. Among clinical students, MSSA was detected on the nose in 42.0% and on the fingerprints in 25.4%. Only one MRSA isolate was obtained. Genes seg and sei were the most frequently detected in both student groups, with their presence in over 40% of isolates among clinical students. The eta and etb genes were mainly detected from the nose in both student groups. In pre-clinical students, S. aureus carrying eta gene occurred in 6.4% and etb in 8.5%. In clinical students, the occurrence was 5.1% for eta and 8.5% for etb. The tst gene was identified only in the nose and fingerprints of the clinical student group. The most frequently observed resistance was to clindamycin and erythromycin. In total 58 different spa types were identified. High rates of asymptomatic MSSA carriage were observed in both groups of medical students. Detected MSSA strains showed a high degree of genetic variability, with a number of them carrying the virulence and antibiotic resistance genes. Although students do not exhibit increased risk to their patient's, increased hygiene is required in asymptomatic carriage personnel. The overall prevalence of MRSA was low, with a minimal risk of spread.

Emerging Resistance Mechanisms in Gram-Positive Bacteria Isolated From Septicemia Cases in ICUs: A Focus on Genotypic Insights.

Background Bloodstream infections (BSIs) are associated with high morbidity and mortality, especially in intensive care unit (ICU) settings. The most common Gram-positive pathogens, methicillin-resistantStaphylococcus aureus (MRSA) and coagulase-negative Staphylococci (CoNS), are likely to cause BSIs. The phenotypic and genotypic characteristics of multidrug-resistant (MDR)Staphylococcal species isolated from patients admitted to the ICUwith septicemia were evaluated for better treatment outcomes. Materials and methods A cross-sectional study was conducted at Sree Balaji Medical College and Hospital over two years (July 2022 to June 2024). Blood samplesin blood culture bottles received in the laboratory from ICU patients with suspected sepsis were included in this study. The BacT/ALERT® 3D system (bioMérieux, France) was used to assess the bacterial growth. TheVITEK®2 system (bioMérieux, France) and conventional methods were used to identify Gram-positive isolates. Antibiotic susceptibility was determined by the Kirby-Bauer disc diffusion method, and polymerase chain reaction (PCR) was used to detect genes like mecA, icaA, and icaD genes. Results Out of 274 blood samples, eight (2.9%) were contaminants, and 121 (44.2%) were culture-positive as true pathogens. Eighty-seven (71.9%) Gram-positive isolates were identified from the positive blood cultures, of whichCoNS was predominant (51, 58.6%), followed by Staphylococcus aureus(25, 28.7%). Methicillin resistance was observed in 10 (13.1%) Staphylococcusaureus and 14 (18.4%) CoNS isolates. PCR detected the mecA gene in 20 (83.3%) methicillin-resistant isolates and biofilm-related genes icaD in 64 (84.2%) and icaA in 58 (76.3%). Vancomycin and teicoplanin showed high effectiveness. Conclusions This study has emphasized the importance of molecular screening for the mecA, icaA, and icaD genes in framing antibiotic regimens. The findings emphasize the efficacy of vancomycin, teicoplanin, and linezolid in combating MDR Staphylococcus infections in ICUs of hospitals and demonstrate the importance of antibiotic stewardship.

Clonal shift and impact of azithromycin use on antimicrobial resistance of Staphylococcus aureus isolated from bloodstream infection during the COVID-19 pandemic

Staphylococcus aureus is a relevant pathogen in bloodstream infections (BSI), and the emergency of the COVID-19 pandemic increased its antimicrobial resistance. S. aureus isolates from BSI (September/2019 - March/2021) were analyzed phenotypically and molecularly, in addition to the clinical features of the patients. Of 88 S. aureus isolates recovered from 85 patients, 25 were isolated before the pandemic and 63 during it, and 16 were from patients with COVID-19. A rate of 45.5% of methicillin-resistant isolates (MRSA) were found, and 5% of them were ceftaroline susceptible dose-dependent. Daptomycin non-susceptibility was observed in 9.1% of isolates. The USA800/ST5/SCCmecIV lineage was prevalent among MRSA isolates (41.8%). Besides, 30.2% of the isolates were associated with community-associated MRSA (CA-MRSA) genotypes. There was a significant impact on the resistance rates for cefoxitin, clindamycin and erythromycin among S. aureus isolates from BSI in COVID-19 patients and association with the previous use of azithromycin by them (p < 0.05). A clonal alternation and an increase in the emergence of CA-MRSA lineages were also found, highlighting the importance of constant microbiological surveillance.

Methicillin-Resistant S. aureus Carrying the PVL and Toxic Shock Syndrome Toxin in Healthy Dogs in Algeria.

Background/Objectives: Staphylococcus aureus and Staphylococcus pseudintermedius are major opportunistic pathogens in both humans and dogs. In pets, the dissemination of methicillin-resistant isolates (MRSA or MRSP) is problematic for the treatment of animals and is a public health issue due to their zoonotic potential. MRSA and MRSP may also harbor virulent genes that increase their dangerousness. This study aimed to assess the prevalence of (MR)SA and (MR)SP in healthy dogs and their owners in Algeria. Methods: Swabs were collected from various body sites of healthy dogs (n = 88) and from the nose of their owners (n = 38). Antimicrobial susceptibility testing was performed by antibiograms according to the disc diffusion method, and clonality was assessed using Pulsed-Field Gel Electrophoresis (PFGE). All methicillin-resistant isolates were short-read whole-genome sequenced using the Illumina technology. Results: 26 S. aureus and 17 S. pseudintermedius isolates were respectively collected from 13 dogs (13/88, 14.8%). No MRSP isolate was detected, while MRSA was found in six dogs (6.8%). Isolates belonged to ST1 (n = 3), ST 80 (n = 1), and ST 22 (n = 2, including the single-locus variant ST7118). All MRSA displayed the immune evasion cluster (IEC) type E. The ST80 isolate presented the Panton-Valentine toxin, and the ST22/ST7118 isolates carried the tst gene coding for the toxic shock syndrome toxin. Conclusions: The epidemiology of MRSA in healthy Algerian dogs mirrors the one in Algerian people. This poses a zoonotic and public health concern due to the virulence and resistance genes displayed by these isolates. Our results indicate the need for developing One Health strategies to avoid a large-scale dissemination of MRSA in Algerian dogs.

Detection and genetic characterization of multidrug-resistant staphylococci isolated from public areas in an international airport

The environmental realm has been acknowledged as a pivotal arena for the emergence and propagation of antimicrobial resistance. To further explore insight into antimicrobial resistance dynamics beyond clinical and veterinary settings, we embarked on an environmental surveillance initiative targeting the prevalence of antibiotic-resistant bacteria within the bustling confines of an international airport in Japan. Our findings illuminate a high prevalence of methicillin-resistant staphylococci (46.3%) on frequently contacted surfaces in the public domain. Notably, Staphylococcus haemolyticus and S. epidermidis emerged as the preeminent carriers of the mecA gene. Intriguingly, we encountered a virulent strain of livestock-associated MRSA harboring a PVL-positive ST1232 clone, CC398 lineage. Further scrutiny unveiled a repertoire of resistance mechanisms, the methicillin-resistant isolates exhibited two or more resistance genes conferring resistance against different types of antibiotics, including beta-lactams, macrolides, lincosamides, aminoglycosides, chloramphenicol, and fosfomycin. Revealing multidrug-resistant CoNS and a LA-MRSA across various surfaces in urban public areas unearths a looming public health hazard. Thus, implementation of molecular surveillance is imperative, augmenting our capacity for early detection and mitigation of the insidious spread and potential transfer of antibiotic resistance genes and virulence factors amidst urban settings, notably within pivotal nodes such as airports.

MIC trends of vancomycin and teicoplanin among methicillin resistant CoNS isolates from new born blood cultures in a tertiary care centre in Southern India.

Coagulase-negative-Staphylococci (CoNS) are important etiological agent of bacteraemia in newborn babies. Methicillin resistant CoNS infections have only limited treatment options. The antimicrobial susceptibility pattern of Methicillin Resistant CoNS isolates from newborn blood cultures was studied with special reference to MICs of Vancomycin and Teicoplanin. The study population included Methicillin Resistant Coagulase Negative Staphylococcal isolates (MRCoNS) from newborn blood cultures, during a one-year period. Minimum inhibitory concentration (MIC) of Vancomycin and Teicoplanin in Methicillin resistant CoNS isolates was determined by macrobroth dilution method as per CLSI guidelines and by automated methods. Coagulase Negative Staphylococci were the etiological agent in 73.7% (n=56) cases of neonatal bacteremia. Methicillin resistance in newborn CoNS was found to be 58.9%. All the MRCoNS isolates had vancomycin and teicoplanin MICs in the susceptible range. There were MRCoNS isolates with MICs in the upper limit of susceptible range for both vancomycin and teicoplanin, which can result in poor clinical response. Continuous large scale multi-centre surveillance studies with special attention to study the MIC pattern of the high-end anti-MRSA agents like vancomycin, teicoplanin, linezolid are to be carried out. This will help the clinicians to judiciously prescribe the antibiotics, which is very essential for antimicrobial stewardship.

Bidens sulphurea and Tanacetum vulgare L. extracts against Staphylococcus spp. mecA positive in pregnant women

The present study aimed to evaluate the antibacterial activity of aqueous extracts of Bidens sulphurea and Tanacetum vulgare L. against Staphylococcus spp. methicillin-resistant (mecA) isolates from the vaginal microbiota of pregnant women. Fifteen isolates of Staphylococcus spp. with the presence of the mecA gene, from the vaginal swabs of pregnant women, aged between 16 and 38 years, who underwent microbiological examination during the first trimester of pregnancy. The aqueous extracts of B. sulphurea and Tanacetum vulgare L. were obtained by infusion, as recommended by popular use, and then, the chemical identification of the extracts was performed by gas chromatography. The antibacterial activity of plant extracts was performed by the method of microdilution in broth. B. sulphurea extract had as major compounds: custonolide (8.06%), Isohumulene (6.19%), artemetin (21,13%), β-sitosterol (28.68%), phytol (7.36%) e 7,8-epoxylanostan-11-ol, 3-acetoxy- (7.09%). T. vulgare extract presented as the majority artemetin (13.38%), verrucarol (13.27%) and phytol (11.93%), ergosterol (5.43%), ethyl iso-allocholate (6.95%), 7,8-epoxylanostan-11-ol, 3-acetoxy- (14,46%), lycopene, 1,1',2,2'-tetrahydro-1,1'-dimethoxy-, all-trans (7.09%) e 9,19-cyclochloestene-3,7-diol, 4,14-dimethyl-, 3-acetate (8.58%). The isolates tested against aqueous extracts of B. sulphurea and T. vulgare L., obtained minimum inhibitory concentrations (MIC) between 1.17mg/mL and 37.5mg/mL, minimum bactericidal concentrations (MBC) of B. sulphurea varied between 75mg/mL and 150mg/mL, on the other hand, T. vulgare L. MBC had lower values ranging from 9.37mg/mL to 150mg/mL. It was confirmed the bactericidal and bacteriostatic activity of the tested extracts against isolates that have the mecA resistance gene, and it is possible to attest that these extracts are a therapeutic alternative.

Evaluation of a nucleic acid amplification system, GENECUBE, for rapid detection of staphylococcal nuc and mecA in blood culture samples

ObjectiveThis study determined whether the GENECUBE rapid nucleic acid amplification test could directly detect nuc and mecA genes in clinical blood culture samples of Staphylococcus and various other pathogens. MethodsBetween September 2020 and December 2021, 537 blood culture samples from 192 patients with suspected bacteremia were tested using conventional assays (MicroScan WalkAway96 or VITEK 2 systems) and GENECUBE nuc and mecA assays. Isolates from samples with discrepant results between the conventional and GENECUBE assays were further evaluated using MALDI-TOF mass spectrometry, disk diffusion testing using cefoxitin, broth microdilution testing using oxacillin, and sequencing for mecA. Bacterial solutions containing a mixture of methicillin-susceptible Staphylococcus aureus (MSSA) and methicillin-resistant Staphylococcus epidermidis (MRSE) were prepared to evaluate the limit of detection (LOD) of mecA. ResultsUsing conventional assays as the reference, the sensitivity, specificity, and positive and negative predictive values (95 % confidence interval) of GENECUBE were 100 % (96.8–100 %), 100 % (99.1–100 %), 100 % (96.8–100 %), and 100 % (99.1–100 %), respectively, for nuc detection and 100 % (96.1–100 %), 98.9 % (97.4–99.6 %), 94.9 % (88.5–98.3 %), and 100 % (99.2–100 %), respectively, for mecA detection. Sequencing analysis of five samples identified as methicillin-sensitive staphylococci using conventional assays and methicillin-resistant staphylococci using GENECUBE revealed the presence of methicillin-resistant isolates in all samples. The estimated LOD of mecA was 104 colony-forming units (CFU)/mL of MRSE with GENECUBE, compared with 105 CFU/mL with conventional assays. ConclusionThe GENECUBE assay accurately detected mecA in positive blood culture samples and had higher sensitivity than conventional assays.

Livestock associated Staphylococcus aureus in cystic fibrosis patients in Spain: Detection of MRSA and MSSA CC398

PurposeStaphylococcus aureus is one of the most prevalent pathogens in cystic fibrosis (CF), being of special relevance those methicillin-resistant (MRSA). The livestock-associated (LA)-MRSA lineage CC398 is an emerging problem, specially related to pig-farming (PF) environments. The objective was to characterize the S. aureus isolates recovered from CF-patients in a Spanish hospital located in a region with high-PF activity. MethodsForty-two isolates were obtained (January–October/2022) and characterised (one/patient). The antimicrobial resistance phenotype/genotype was evaluated by Microscan/PCR. The presence of virulence and Immune Evasion Cluster (IEC) genes as well as the agr type was determined by PCR. MLST and spa-typing were studied by PCR-sequencing. ResultsNine of the 42 isolates were MRSA (21.4 %), and 8 of them multidrug resistant (MDR). Among MRSA, 6 spa-types were detected, assigned to CC1, CC5, CC8, CC30, and CC398. Four MRSA isolates belonged to the lineage CC398-t011-IEC negative (animal adapted-clade, LA-MRSA). The remaining 33 isolates were methicillin-susceptible (MSSA), of 26 spa-types and associated with 11 CCs (predominant: CC5, CC30, and CC398). Seven MSSA isolates were of the lineage CC398 (spa-types t034, t108, t571, t20352); four of them were IEC-positive and erm(T)-positive (t571, and t20352, human-adapted CC398 clade), being IEC-negative the remaining three. The tst and eta/etb genes were identified in 12 and 2 isolates, respectively (none CC398). Small-colony-variants were demonstrated in 9 isolates (two CC398, both MDR). ConclusionThe lineage CC398 was very frequent among CF-patients (26.2 %), both among MSSA and MRSA. The emergence of LA-MRSA-CC398 in CF-patients requires monitorization, especially in hospitals of high-PF-regions.

Genetic Characterization of Methicillin-Resistant Staphylococcus aureus Isolated From Diabetic Foot Ulcers in a Tertiary Care Hospital in Mysuru, South India.

Background Diabetic foot ulcers (DFUs) are common complications in diabetes patients, often leading to sepsis and leg amputation. Methicillin-resistant Staphylococcus aureus (MRSA) infections in DFUs pose challenges due to methicillin resistance with mecA and mecC genes. This study aims to assess the prevalence of MRSA in clinical isolates from DFUs,analyze the antibiogram of MRSA isolates, and detect the presence of the mecA and mecC genes among MRSA isolates. Methodology The isolated S. aureus colonies were identified and antimicrobial susceptibility was performed using the Vitek-2 Compact system. Methicillin resistance was also confirmed through the disc diffusion method. Confirmed methicillin-resistant isolates were subjected to real-time polymerase chain reaction (RT-PCR) to detect mecA and mecC genes. Results A total of 474 purulent samples from DFUs yielded 541 distinct isolates, comprising 201 gram-positive and 340 gram-negative organisms. Among the gram-positive organisms, Staphylococcus species predominated, with 79 S. aureus isolates, 34 of which were methicillin-resistant. All MRSA isolates (100%) were sensitive to tetracycline, linezolid, teicoplanin, and vancomycin, and 94% were sensitive to cotrimoxazole but least susceptible to ciprofloxacin and levofloxacin. RT-PCR confirmed the presence of mecA genes in all 34 isolates and mecC genes in three isolates. Conclusions The presence of mecA in all 34 MRSA isolates underscores consistent methicillin resistance. The co-occurrence of mecA and mecC in three isolates hints at genetic diversity. Two MRSA isolates positive for mecC were isolated from rural patients involved in farming and animal husbandry, suggesting an occupational risk. The third patient was from a non-rural area, indicating potential alternative transmission pathways warranting further investigation.

Detection Staphylococcus aureus contaminated some door handles and table surfaces and their sensitive to antibiotics

The current study's objective was to identify and isolate Staphylococcus spp. germs from a few of Omar Al-Mukhtar University's faculty laboratories. It is regarded as the first investigation carried out at Omar Al-Mukhtar University to assess the level of Staphylococcal bacterial contamination of door handles and table surfaces inside the laboratories. 33 isolates from the College of Science's Botany Department were used in the study, which took place in the department's laboratories during the fall semester of the academic year (2022). After evaluating the expanding colonies and using morphological inspections and biochemical tests to diagnose the isolates, it was showed that 90.9% of the samples from the Botany Department were Staphylococcus and that the percentage of S. aureus reached 36.7% of isolates of Staphylococcus. A biofilm-forming capacity was demonstrated by 80% of the isolated bacteria, of which 29% were S. aureus. In order to ascertain the effectiveness of the antibiotics and which ones the bacteria are resistant to, the sensitivity of thirty S. aureus isolates to hospital-grade antibiotics against infection was tested. The findings revealed a high level of resistance to the majority of antibiotics and a definite resistance to gentamicin. Methicillin-resistant MRSA isolates made up 73.3% of the isolates, whereas vancomycin-resistant VRSA isolates made up 70%.

Genomic portraits of methicillin-resistant staphylococci (MRS) from food fish unveiled the genes associated with staphylococcal food poisoning (SFP), virulence and antimicrobial resistance

BackgroundCharacteristics of non-clinical strains of methicillin-resistant Staphylococcus aureus (MRSA) especially from fishery environment are poorly understood. This research, in addition to comprehensive characterisation, sought to delineate the genetic relatedness between the MRSA strains originating from clinical as well as non-clinical settings. Out of 39 methicillin-resistant staphylococcal isolates from 197 fish samples, 6 (Three each of methicillin-resistant S. haemolyticus (MRSH) and MRSA) with distinct resistance profiles were selected for whole-genome sequencing. Using respective bioinformatics tools, MRSA genomes were comprehensively characterized for resistome, virulomes, molecular epidemiology and phylogenetic analysis. Simultaneously, MRSH genomes were specifically examined to characterize antimicrobial resistance genes (ARGs), owing to the fact that MRSH is often recognized as a reservoir for resistance determinants.ResultsThree MRSA clones identified in this study include ST672-IVd/t13599 (sequence type-SCCmec type/spa type), ST88-V/t2526, and ST672-IVa/t1309. Though, the isolates were phenotypically vancomycin-sensitive, five of the six genomes carried vancomycin resistance genes including the VanT (VanG cluster) or VanY (VanM cluster). Among the three MRSA, only one harbored the gene encoding Panton-Valentine Leukocidin (PVL) toxin, while staphylococcal enterotoxin (SEs) genes such as sea and seb, associated with staphylococcal food poisoning were identified in two other MRSA. Genomes of MRSH carried a composite of type V staphylococcal cassette chromosome mec (SCCmec) elements (5C2 & 5). This finding may be explained by the inversion and recombination events that may facilitate the integration of type V elements to the SCC elements of S. aureus with a methicillin-susceptible phenotype. Phylogenetically, MRSA from a non-clinical setting displayed a considerable relatedness to that from clinical settings.ConclusionThis study highlights the genetic diversity and resistance profiles of MRSA and MRSH, with non-clinical MRSA showing notable relatedness to clinical strains. Future research should explore resistance gene transfer mechanisms and environmental reservoirs to better manage MRSA spread.

Prevalence and Phage-Based Biocontrol of Methicillin-Resistant Staphylococcus aureus Isolated from Raw Milk of Cows with Subclinical Mastitis in Vietnam.

S. aureus, particularly methicillin-resistant S. aureus, has been recognized as a main cause of bovine mastitis and food poisoning. This study investigated the prevalence, antibiotic resistance, and phage-based biocontrol of S. aureus and methicillin-resistant S. aureus isolated from raw milk of cows with subclinical mastitis. The results showed that the prevalence of S. aureus and methicillin-resistant S. aureus was 12% (48/400) and 1.5% (6/400), respectively. The S. aureus isolates were highly resistant to penicillin (72.92%), erythromycin (43.75%), and tetracycline (39.58%). Out of 48 S. aureus isolates, 6 were identified as methicillin-resistant strains. Among them, one isolate was found to harbor the sea gene. A total of 5 phages were recovered from 50 pork and 50 chicken meat samples, 1 from pork and 4 from chicken meat samples. Phage PSA2 capable of lysing all 6 methicillin-resistant isolates was selected for characterization. The use of phage PSA2 completely inactivated methicillin-resistant S. aureus SA33 in raw milk at both 24 °C and 4 °C, indicating its potential as a promising antibacterial agent in controlling methicillin-resistant S. aureus in raw milk and treating bovine mastitis.

Unveilling genetic profiles and correlations of biofilm-associated genes, quorum sensing, and antibiotic resistance in Staphylococcus aureus isolated from a Malaysian Teaching Hospital

BackgroundStaphylococcus aureus is a notorious multidrug resistant pathogen prevalent in healthcare facilities worldwide. Unveiling the mechanisms underlying biofilm formation, quorum sensing and antibiotic resistance can help in developing more effective therapy for S. aureus infection. There is a scarcity of literature addressing the genetic profiles and correlations of biofilm-associated genes, quorum sensing, and antibiotic resistance among S. aureus isolates from Malaysia.MethodsBiofilm and slime production of 68 methicillin-susceptible S. aureus (MSSA) and 54 methicillin-resistant (MRSA) isolates were determined using a a plate-based crystal violet assay and Congo Red agar method, respectively. The minimum inhibitory concentration values against 14 antibiotics were determined using VITEK® AST-GP67 cards and interpreted according to CLSI-M100 guidelines. Genetic profiling of 11 S. aureus biofilm-associated genes and agr/sar quorum sensing genes was performed using single or multiplex polymerase chain reaction (PCR) assays.ResultsIn this study, 75.9% (n = 41) of MRSA and 83.8% (n = 57) of MSSA isolates showed strong biofilm-forming capabilities. Intermediate slime production was detected in approximately 70% of the isolates. Compared to MSSA, significantly higher resistance of clindamycin, erythromycin, and fluoroquinolones was noted among the MRSA isolates. The presence of intracellular adhesion A (icaA) gene was detected in all S. aureus isolates. All MSSA isolates harbored the laminin-binding protein (eno) gene, while all MRSA isolates harbored intracellular adhesion D (icaD), clumping factors A and B (clfA and clfB) genes. The presence of agrI and elastin-binding protein (ebpS) genes was significantly associated with biofilm production in MSSA and MRSA isolates, respectively. In addition, agrI gene was also significantly correlated with oxacillin, cefoxitin, and fluoroquinolone resistance.ConclusionsThe high prevalence of biofilm and slime production among MSSA and MRSA isolates correlates well with the detection of a high prevalence of biofilm-associated genes and agr quorum sensing system. A significant association of agrI gene was found with cefoxitin, oxacillin, and fluoroquinolone resistance. A more focused approach targeting biofilm-associated and quorum sensing genes is important in developing new surveillance and treatment strategies against S. aureus biofilm infection.

Antibiotic Susceptibility of Staphylococcus Aureus with VanA and MecA Genes.

Staphylococcus aureus (S.aureus) is an emerging antibiotic resistant bacterium responsible for various infections in human. Resistance to methicillin and vancomycin are of prime concern in S. aureus. The study aims to determine the minimum inhibitory concentration (MIC) of Vancomycin and evaluate the existence of mecA and vanA genes, associated with antibiotic resistance. Clinical specimens from three Kathmandu hospitals were processed and S. aureus was identified using conventional microbiological procedures. MRSA was phenotypically identified with cefoxitin (30µg) disc diffusion, while vancomycin susceptibility was assessed using the Ezy MICTM stripes. The mecA and vanA genes were detected by polymerase chain reaction (PCR). Out of 266 S. aureus samples from various clinical specimen subjected for analysis, 77 (28.9%) were found methicillin-resistant (MRSA) and 10 (3.8%) were observed vancomycin-resistant (VRSA). Vancomycin resistant isolates showed a significant correlation between resistance to ampicillin, chloramphenicol, and cefoxitin. The mecA gene was found in 39 of the MRSA isolates, having 50.64% of MRSA cases, while the vanA gene was detected in 4 of the VRSA cases, constituting 40% of VRSA occurrences. The strains with higher vancomycin minimum inhibitory concentration values (≥ 1.5 μg/ml) displayed increased resistance rates to various antibiotics compared to strains with lower minimum inhibitory concentration values (< 1.5 μg/ml). The presence of vanA genes was strongly associated (100%) with vancomycin resistance, while the 10.3% mecA gene was identified from MRSA having resistance towards vancomycin also.

Molecular Prevalence of MecA and MecC Genеs in Coagulasе-Positive Staphylococci Isolated From Dogs with Dermatitis and Otitis in Belgrade, Serbia: A One Year Study

Abstract The escalating global concern of antimicrobial resistance in human and veterinary medicine is exacerbated by the inappropriate prescription of antibiotics for bacterial infections in companion animals. This study aimed to determine the distribution of coagulase-positive staphylococci causing clinical skin and ear infections in dogs and to determine methicillin-resistant isolates. A total of 78 staphylococcal strains were isolated from clinical samples taken from patients at the Dermatology Clinic at the Faculty of Veterinary Medicine in Belgrade, Serbia. Multiplex PCR was used for species-specific identification, and mecA and mecC genes were used to determine methicillin resistance, in addition to phenotypic determination, MIC values and detection of PBP2a. Out of the 78 samples analyzed, 65.8% were identified as Staphylococcus pseudintermedius, 22.4% as S. aureus, 7.9% as S. coagulans, and 3.9% as S. intermedius. Four S. aureus isolates exhibited methicillin resistance confirmed by cefoxitin disk diffusion, while five were confirmed with MIC testing and latex agglutination. MecA gene was detected in 29.4% of S. aureus and 30% of S. pseudintermedius isolates. These isolates were classified as methicillin-resistant S. aureus (MRSA) and methicillin-resistant S. pseudintermedius (MRSP), respectively. No isolates carried the mecC gene. This study provides insights into the prevalence of CoPS species and methicillin resistance in isolates from dogs. Continued surveillance is essential to monitor and understand the emergence and dissemination of antimicrobial resistance in veterinary medicine and the results of this study accent the need for establishment of a continuous antimicrobial resistance surveillance program in the Republic of Serbia.

Atopic dermatitis pediatric patients show high rates of nasal and intestinal colonization by methicillin-resistant Staphylococcus aureus and coagulase-negative staphylococci

BackgroundAtopic dermatitis (AD) patients have high rates of colonization by Staphylococcus aureus, which has been associated with worsening of the disease. This study characterized Staphylococcus spp isolates recovered from nares and feces of pediatric patients with AD in relation to antimicrobial susceptibility, staphylococcal cassette chromosome mec (SCCmec) type, presence of pvl genes and clonality. Besides, gut bacterial community profiles were compared with those of children without AD.ResultsAll 55 AD patients evaluated had colonization by Staphylococcus spp. Fifty-three (96.4%) patients had colonization in both clinical sites, whereas one patient each was not colonize in the nares or gut. Staphylococcus aureus was identified in the nostrils and feces of 45 (81.8%) and 39 (70.9%) patients, respectively. Methicillin-resistant Staphylococcus spp. isolates were found in 70.9% of the patients, and 24 (43.6%) had methicillin-resistant S. aureus (MRSA). S. aureus (55.6%) and S. epidermidis (26.5%) were the major species found. The prevalent lineages of S. aureus were USA800/SCCmecIV (47.6%) and USA1100/SCCmecIV (21.4%), and 61.9% of the evaluated patients had the same genotype in both sites. Additionally, gut bacterial profile of AD patients exhibits greater dissimilarity from the control group than it does among varying severities of AD.ConclusionsHigh rates of nasal and intestinal colonization by S. aureus and methicillin-resistant staphylococci isolates were found in AD patients. Besides, gut bacterial profiles of AD patients were distinctly different from those of the control group, emphasizing the importance of monitoring S. aureus colonization and gut microbiome composition in AD patients.

Repurposing antibiotic resistance surveillance data to support treatment of recurrent infections in a remote setting

In northern Australia, a region with limited access to healthcare and a substantial population living remotely, antibiotic resistance adds to the complexity of treating infections. Focussing on Escherichia coli urinary tract infections (UTIs) and Staphylococcus aureus skin & soft tissue infections (SSTIs) captured by a northern Australian antibiotic resistance surveillance system, we used logistic regression to investigate predictors of a subsequent resistant isolate during the same infection episode. We also investigated predictors of recurrent infection. Our analysis included 98,651 E. coli isolates and 121,755 S. aureus isolates from 70,851 patients between January 2007 and June 2020. Following an initially susceptible E. coli UTI, subsequent recovery of a cefazolin (8%) or ampicillin (13%) -resistant isolate during the same infection episode was more common than a ceftriaxone-resistant isolate (2%). For an initially susceptible S. aureus SSTI, subsequent recovery of a methicillin-resistant isolate (8%) was more common than a trimethoprim-sulfamethoxazole-resistant isolate (2%). For UTIs and SSTIs, prior infection with a resistant pathogen was a strong predictor of both recurrent infection and resistance in future infection episodes. This multi-centre study demonstrates an association between antibiotic resistance and an increased likelihood of recurrent infection. Particularly in remote areas, a patient’s past antibiograms should guide current treatment choices since recurrent infection will most likely be at least as resistant as previous infection episodes. Using population-level surveillance data in this way can also help clinicians decide if they should switch antibiotics for patients with ongoing symptoms, while waiting for diagnostic results.

Deciphering the genomic character of the multidrug-resistant &lt;i&gt;Staphylococcus aureus&lt;/i&gt; from Dhaka, Bangladesh

&lt;p&gt;&lt;italic&gt;Staphylococcus aureus&lt;/italic&gt; is one of the leading agents of nosocomial and community-acquired infections. In this study, we explored the genomic characterization of eight methicillin-resistant clinical isolates of &lt;italic&gt;S. aureus&lt;/italic&gt; from Dhaka, Bangladesh. Notably, all strains were resistant to penicillin, cephalosporins, and monobactams, with partial susceptibility to meropenem and complete susceptibility to amikacin, vancomycin, and tigecycline antibiotics. The strains were found to have an average genome size of 2.73 Mbp and an average of 32.64% GC content. Multi-locus sequence typing analysis characterized the most predominant sequence type as ST361, which belongs to the clonal complex CC361. All isolates harbored the &lt;italic&gt;mecA&lt;/italic&gt; gene, often linked to SCC&lt;italic&gt;mec&lt;/italic&gt;_type IV variants. Multidrug resistance was attributed to efflux pumps NorA, NorC, SdrM, and LmrS alongside genes encoding beta-lactamase BlaZ and factors like ErmC and MepA. Additionally, virulence factors including &lt;italic&gt;adsA&lt;/italic&gt;, &lt;italic&gt;sdrC&lt;/italic&gt;, &lt;italic&gt;cap8D&lt;/italic&gt;, &lt;italic&gt;harA&lt;/italic&gt;, &lt;italic&gt;esaA&lt;/italic&gt;, essC, &lt;italic&gt;isdB&lt;/italic&gt;, &lt;italic&gt;geh&lt;/italic&gt;, and &lt;italic&gt;lip&lt;/italic&gt; were commonly identified. Furthermore, genes associated with heme uptake and clumping were present, highlighting their roles in &lt;italic&gt;S. aureus&lt;/italic&gt; colonization and pathogenesis. Nine secondary metabolite biosynthetic gene clusters were found, of which six were common in all the strains. Numerous toxin-antitoxin systems were predicted, with ParE and ParB-like nuclease domains found to be the most prevalent toxin and antitoxin, respectively. Pan-genome analysis revealed 2007 core genes and 229 unique genes in the studied strains. Finally, the phylogenomic analysis showed that most Bangladeshi strains were grouped into two unique clades. This study provides a genomic and comparative insight into the multidrug resistance and pathogenicity of &lt;italic&gt;S. aureus&lt;/italic&gt; strains, which will play a crucial role in the future antibiotic stewardship of Bangladesh.&lt;/p&gt;