Methicillin-susceptible S. Aureus Isolates Research Articles

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.

Cephalotin Versus Dicloxacillin for the Treatment of Methicillin-Susceptible Staphylococcus aureus Bacteraemia: A Retrospective Cohort Study.

First-line treatments for methicillin-susceptible S. aureus (MSSA) bacteraemia are nafcillin, oxacillin, or cefazolin. Regional shortages of these antibiotics force clinicians to use other options like dicloxacillin and cephalotin. This study aims to describe and compare the safety and efficacy of cephalotin and dicloxacillin for the treatment of MSSA bacteraemia. This retrospective study was conducted in a referral centre in Mexico City. We identified MSSA isolates in blood cultures from 1 January 2012 to 31 December 2022. Patients ≥ 18 years of age, with a first episode of MSSA bacteraemia, who received cephalotin or dicloxacillin as the definitive antibiotic treatment, were included. The primary outcome was in-hospital all-cause mortality. We included 202 patients, of which 48% (97/202) received cephalotin as the definitive therapy and 52% (105/202) received dicloxacillin. In-hospital all-cause mortality was 20.7% (42/202). There were no differences in all-cause in-hospital mortality between patients receiving cephalotin or dicloxacillin (20% vs. 21%, p = 0.43), nor in 30-day all-cause mortality (14% vs. 18%, p = 0.57) or 90-day all-cause mortality (24% vs. 22%, p = 0.82). No severe adverse reactions were associated with either antibiotic. Cephalotin and dicloxacillin were equally effective for treating MSSA bacteraemia, and both showed an adequate safety profile.

Genetic characterization of methicillin-resistant / susceptible Staphylococcus aureus (MRSA/MSSA) and Staphylococcus argenteus clinical isolates in Bangladesh: Dominance of ST6-MRSA-IV/t304 and detection of cfr/fexA in ST8-MSSA/t008

ObjectivesCoagulase-positive staphylococcus (CoPS), represented by Staphylococcus aureus, is a major cause of infections in humans. This study aimed to investigate molecular epidemiological characteristics, antimicrobial resistance, and their trends of CoPS in Bangladesh. MethodsClinical isolates of CoPS were collected from two medical institutions in Bangladesh for a 2-year period and analyzed for their species, genotypes, virulence factors, antimicrobial susceptibility, and resistance determinants. Results172 CoPS isolates collected were identified as S. aureus or S. argenteus (170 and two, respectively). Methicillin-resistant S. aureus (MRSA) accounted for 36% (n = 61), having Staphylococcal cassette chromosome mec (SCCmec)-IV (82%) or V (18%). Panton-Valentine leukocidin (PVL) genes were detected at higher rate in methicillin-susceptible S. aureus (MSSA) (62%) than MRSA (26%). MRSA comprised 11 STs, including a dominant type ST6 (46%) associated with mostly SCCmec-IVa/spa-t304, and one isolate had genetic features of the USA300 clone (ST8/SCCmec-IVa/coa-IIIa/spa-t008/ACME-I/ΦSa2USA). STs of CC1, CC88, and CC398 were common in MSSA, with CC88 showing the highest PVL-positive rate. One MSSA isolate (ST8/spa-t008) harbored fexA and cfr showing susceptibility to linezolid. S. argenteus was methicillin-susceptible and belonged to ST2250/coa-XId. ConclusionsGenetic characteristics of current MRSA/MSSA in Bangladesh were revealed, with first identification of S. argenteus at low prevalence.

860. Molecular Characterization of Isolated Staphylococcus aureus Specimens Using Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS)

Abstract Background Staphylococcus aureus (S. aureus) is a common cause of skin and soft tissue infections (SSTIs). Both the β-lactam resistant form, methicillin resistant S. aureus (MRSA), and non-resistant form, methicillin susceptible S. aureus (MSSA) are associated with disparate infections across age and race; community-associated infections are often due to S. aureus US300. Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS) is a used in clinical microbiology laboratories to identify bacterial species, but limited data is available on if this tool can identify different strains of S. aureus. We hypothesize that MALDI-TOF MS can be used to identify strain-specific peaks of S. aureus USA300. Methods Children were enrolled from emergency departments and swabbed for S. aureus (anterior nares, axilla, perirectal, oropharyngeal, and wounds) during two time periods (2006-2008 and 2015-2016). MALDI-TOF MS was performed on S. aureus cultured from study participants. Default parameters used for variance stability, smoothing, baseline subtraction, and normalization in R 4.4.3. and MALDIquant v1.22 were applied to raw data. Peak detection and alignment were performed with a signal-to-noise ratio of three and tolerance of 300 ppm. Binning and machine learning algorithms were used to identify intensity differences of each peak identified; significance of peaks was determined using Chi Square statistical analyses, comparing MRSA and MSSA, and then between all S. aureus isolates compared to USA300. Statistical significance was considered with p-value < 0.05 using R and SAS 9.4. Results From 57 MRSA and 53 MSSA isolates, 320 peaks were identified. 31 significant peaks were identified, with 14 unique peaks for MRSA and 17 for MSSA. With S. aureus USA300, eight of the 31 peaks remained statistically significant. Conclusion MALDI-TOF MS can identify unique peak differences between MRSA and MSSA. This tool can be potentially used to distinguish S. aureus at the strain level in a clinical setting. MALDI-TOF MS can be an impactful and rapid diagnostic tool to detect antibiotic resistance in S. aureus in a clinical setting. Disclosures Lilly Immergluck, MD,MS, Melinta: Grant/Research Support|Melinta: Grant/Research Support

Diversity of <em>Spa</em> gene between methicillin-resistant and methicillin-sensitive <em>Staphylococcus aureus</em> bacteria in a tertiary referral hospital, Indonesia

BACKGROUND Spa typing is an effective and fast technique to identify the prevalence and spread of Staphylococcus aureus strains based on their spa gene profiles. The distribution of spa types will contribute to control the spread of S. aureus. Little is known regarding the spa types of S. aureus in Indonesia. This study aimed to investigate the diversity of spa gene among S. aureus carriage isolates in North Sumatra Province, Indonesia.
 METHODS 79 S. aureus isolates consisting of 39 methicillin-resistant S. aureus (MRSA) and 40 methicillin-susceptible S. aureus (MSSA) carriage isolates were identified by VITEK2 Compact (Biomerieux, Indonesia) to detect mecA gene. All samples underwent spa typing and sequencing.
 RESULTS Spa gene was detected among 31/39 (79%) of the MRSA isolates and 24/40 (60%) of the MSSA isolates. Most spa typing genes were identified between 350 and 400 base pair (bp). t258 and t852 were the most prevalence spa types among MRSA and MSSA isolates, respectively.
 CONCLUSIONS Many MRSA and MSSA isolates encoded spa gene. The most genes detected were t258 and t852, identified in Germany and Portugal, respectively; while t18977 was initially identified in Malaysia. This indicated a global spread of MRSA according to spa typing.

Therapeutic potential of natural products and antibiotics against bovine mastitis pathogen of cows and buffaloes.

The present study aims toevaluate the prevalence and antimicrobial sensitivity ofStaphylococcus aureus associated with bovine mastitis toselected antibiotics and plant extracts. Inthe current study, 140milk samples were collected from cows and buffaloes. Among the 140samples, 93samples were positive for sub-clinical mastitis based onthe California Mastitis Test (CMT). Out ofthe total positive samples, 45 were confirmed for S.aureus onaMannitol salt agar media. The antimicrobial susceptibility test revealed that 44.82% ofthe isolates were resistant tocefoxitin (oxacillin) confirming methicillin-resistant S.aureus (MRSA) with ahigher percentage (51.61%) inthe buffalo than inthe cow samples. Furthermore, the PCR assay confirmed the presence ofthemecA gene inall the MRSA isolates. Among the seven tested antibiotics, sulfamethoxazole + trimethoprim showed high efficacy (71.1%) against methicillin-susceptible S.aureus isolates (MSSA). Oxytetracycline and sulfamethoxazole+ trimethoprim showed 20% efficacy against MRSA followed byenrofloxacin (10%). Onthe other hand, the tested samples from Pistacia chinensis revealed that the ethyl acetate extract ofbark showed amaximum zone ofinhibition of21.3mm against MSSA and MRSA isolates at3000μg/disc. Moreover, the methanol extract ofCotoneaster microphyllus formed a12.3mm and 9.1mm zone ofinhibition against the MSSA and MRSA isolates, respectively.

Clonal Complexes Distribution of Staphylococcus aureus Isolates from Clinical Samples from the Caribbean Islands.

The aim of this study was to comprehensively characterise S. aureus from the Caribbean Islands of Trinidad and Tobago, and Jamaica. A total of 101 S. aureus/argenteus isolates were collected in 2020, mainly from patients with skin and soft tissue infections. They were characterised by DNA microarray allowing the detection of ca. 170 target genes and assignment to clonal complexes (CC)s and strains. In addition, the in vitro production of Panton-Valentine leukocidin (PVL) was examined by an experimental lateral flow assay. Two isolates were identified as S. argenteus, CC2596. The remaining S. aureus isolates were assigned to 21 CCs. The PVL rate among methicillin-susceptible S. aureus (MSSA) isolates was high (38/101), and 37 of the 38 genotypically positive isolates also yielded positive lateral flow results. The isolate that did not produce PVL was genome-sequenced, and it was shown to have a frameshift mutation in agrC. The high rate of PVL genes can be attributed to the presence of a known local CC8-MSSA clone in Trinidad and Tobago (n = 12) and to CC152-MSSA (n = 15). In contrast to earlier surveys, the USA300 clone was not found, although one MSSA isolate carried the ACME element, probably being a mecA-deficient derivative of this strain. Ten isolates, all from Trinidad and Tobago, were identified as MRSA. The pandemic ST239-MRSA-III strain was still common (n = 7), but five isolates showed a composite SCCmec element not observed elsewhere. Three isolates were sequenced. That showed a group of genes (among others, speG, crzC, and ccrA/B-4) to be linked to its SCC element, as previously found in some CC5- and CC8-MRSA, as well as in S. epidermidis. The other three MRSA belonged to CC22, CC72, and CC88, indicating epidemiological connections to Africa and the Middle East.

Clonal diversity of Staphylococcus aureus isolates in clinical specimens from selected health facilities in Ethiopia

Staphylococcus aureus is among the top three causative agents of nosocomial infection in Ethiopia. The majority of studies in Ethiopia have focused on the epidemiology of S. aureus in hospital settings, with limited molecular genotyping results. Molecular characterization of S. aureus is essential for identification of strains, and contributes to the control and prevention of S. aureus infection. The aim of the current study was to determine the molecular epidemiology of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) isolates recovered from clinical specimens in Ethiopia. A total of 161 MSSA and 9 MRSA isolates were characterized using pulsed-field gel electrophoresis (PFGE) and staphylococcal protein A (spa) typing. Based on the PFGE analysis, MSSA isolates were grouped into eight pulso-types groups (from A to I), while MRSA isolates clustered into three (A, B and C) pulso-types with more than 80% similarity. The spa typing analysis showed diversity of S. aureus with 56 distinct spa types. Spa type t355 was most prevalent (56/170, 32.9%), while eleven new spa types were detected including t20038, t20039, and t20042. The identified spa types were clustered into 15 spa-clonal complexes (spa-CCs) using BURP analysis; novel/unknown spa types were further subjected to MLST analysis. The majority of isolates belonged to spa-CC 152 (62/170, 36.4%), followed by spa-CC 121 (19/170, 11.2%), and spa-CC 005 (18 /170, 10.6%). Of the nine MRSA isolates, 2 (22.2%) were spa-CC 239 with staphylococcal cassette chromosome (SCC)mec III. These findings highlight the diversity of S. aureus strains in Ethiopia, as well as the presence of potentially epidemic strains circulating in the country necessitating further characterization of S. aureus for antimicrobial resistance detection and infection prevention purposes.

Decrease in vancomycin MICs and prevalence of hGISA in MRSA and MSSA isolates from a German pediatric tertiary care center

PurposeResistance of Staphylococcus aureus to vancomycin includes a general increase of minimal inhibitory concentrations (MIC) within the susceptible range over time (Vancomycin MIC Creep) and the presence of a subset of the bacterial population that expresses resistance (heterogeneous glycopeptide-intermediate S. aureus; hGISA). Increased MICs have been associated with adverse clinical outcomes. However, the vancomycin MIC creep is not a uniform trend suggesting the importance of regional surveys.MethodsWe performed a retrospective analysis at a German pediatric tertiary care hospital. Isolates from 2002 to 2017 were selected which were newly identified methicillin-resistant S. aureus (MRSA) or samples from invasive methicillin-susceptible S. aureus (MSSA) or MRSA infections. Vancomycin and oxacillin MICs as well as GISA/hGISA were measured using MIC test strips and resistance was evaluated over time.ResultsA total of 540 samples were tested, 200 from the early (2002–2009) and 340 from the later period (2010–2017). All samples were vancomycin susceptible, but the MIC was higher for the earlier samples as compared to the later ones (1.11 vs 0.99; p < 0.001). 14% of the samples were hGISA, GISA strains were not detected. Again, vancomycin resistance decreased over time with 28 vs. 6% hGISA (p < 0.001). There was no significant difference between MRSA and MSSA samples with respect to vancomycin MIC and hGISA prevalence.ConclusionThis study shows a decreasing trend for both MIC values and presence of hGISA strains highlighting the importance of monitoring local susceptibilities. Vancomycin remains a first-line treatment option for suspected severe infection with Gram-positive cocci and proven infection with MRSA.

Phenotypic and Molecular Detection of Biofilm Formation in Methicillin-Resistant Staphylococcus Aureus Isolated from Different Clinical Sources in Erbil City.

Staphylococcus aureus is an important causative pathogen. The production of biofilms is an important factor and makes these bacteria resistant to antimicrobial therapy. the current study aimed to assess the prevalence of resistance to antibacterial agents and to evaluate the phenotypic and genotypic characterization of biofilm formation among S. aureus strains. This study included 50 isolates of Methicillin-resistant S. aureus (MRSA) and Methicillin-Susceptible S. aureus (MSSA). S. aureus was identified by molecular and conventional methods, and antimicrobial resistance was tested with a disc diffusion method. The biofilm formation was performed through the Microtiter plate method. Strains were subjected to PCR to determine the presence of nuc, mecA, icaA, icaB, icaC, and icaD genes. Of the 50 S. aureus isolates, 32(64%) and 18(36%) were MRSA and MSSA, respectively. A large number of MRSA and MSSA isolates showed resistance to Penicillin and Azithromycin, and a lower number of MRSA and MSSA isolates showed resistance to Amikacin Gentamicin. None of the isolates was resistant to Vancomycin. The MRSA strains had significantly higher resistance against antibiotics than MSSA strains (P = 0.0154). All isolates (MRSA and MSSA) were able to produce biofilm with levels ranging from strong (31.25 %), (16.6%) to moderate (53.12%), (50%) to weak (15.6%), (33.3%) respectively. The MRSA strains had a significantly higher biofilm formation ability than the MSSA strains (P = 0.0079). The biofilm-encoding genes were detected among isolates with different frequencies. The majority of S. aureus isolates, 42 (84%), were positive for the icaA. The prevalence rates of the icaB, icaC and icaD genes were found to be 37 (74%), 40 (80%) and 41 (82%), respectively. The prevalence of biofilm encoding genes associated with multidrug resistance in S. aureus strains is high. Therefore, identifying epidemiology, molecular characteristics, and biofilm management of S. aureus infection would be helpful.

Genomic Epidemiology and Phenotypic Characterization of Staphylococcus aureus from a Tertiary Hospital in Tianjin Municipality, Northern China.

Staphylococcus aureus remains a dangerous pathogen and poses a great threat to public health worldwide. The prevalence of the S. aureus clonotype is temporally and geographically variable. The genomic and phenotypic characteristics of S. aureus isolates in Tianjin, which is among the four big municipalities in China, are unclear. In the present study, 201 nonduplicate S. aureus isolates, including 70 methicillin-resistant S. aureus (MRSA) and 131 methicillin-susceptible S. aureus (MSSA), were collected from 2015 to 2021 in a tertiary hospital in Tianjin. Whole-genome sequencing of S. aureus isolates was carried out to investigate bacterial molecular characteristics, genomic phylogeny, antimicrobial resistance (AMR) gene carriage, and virulence factor gene distribution. The antibiotic resistance profiles, hemolytic activities, and biofilm formation abilities of the S. aureus isolates were also determined. In total, 31 distinct sequence types (STs) and 68 spa types were identified. ST59 (15.9%, 32/201) was the predominant clonotype, followed by ST398 (14.9%, 30/201) and several other major STs (ST1, ST5, ST6, ST22, ST25, ST188, and the newly emerging ST5527). ST59 and ST5527 mainly included MRSA isolates, while ST398 and the other major STs mainly included MSSA isolates. The unique characteristics of the S. aureus isolates belonging to the major STs were determined. ST59 isolates exhibited strong hemolytic activity, and ST398 strains had high biofilm formation capacity, while ST5527 isolates presented the greatest AMR. The genomic epidemiology and phenotypic characteristics of S. aureus isolates determined in this study will help in disease control in nosocomial environments. IMPORTANCE Staphylococcus aureus is an important bacterium pathogen in tertiary hospitals, which provide rich medical resources. Tianjin is one of the four municipalities in China with a population of more than 13 million. However, the epidemiology and molecular characteristics of S. aureus isolates in Tianjin are unknown. In this study, the genomic and phenotypic analyses were performed to investigate 201 S. aureus isolates collected from a tertiary hospital in Tianjin over a time span of 6 years. The refined analysis of predominant clones ST59, ST398, the newly emerging clone ST5527, as well as other major clones, will undoubtedly aid in the control and prevention of infections caused by S. aureus in tertiary hospitals.

Molecular Epidemiological Characterization of Staphylococcus aureus and Staphylococcus argenteus Clinical Isolates from a National Tertiary Care Hospital in Myanmar: Co-Isolation of Multiple Clones and Identification of Novel Staphylocoagulase Genotype.

Spread of antimicrobial resistance and virulence factors among Staphylococcus aureus/Staphylococcus argenteus poses a potential public health concern in Myanmar. In this study, a total of 226 clinical isolates of S. aureus (n = 211) and S. argenteus (n = 15) collected in Yangon General Hospital during a two-year period were analyzed for their antimicrobial susceptibility and genetic features. Methicillin-resistant S. aureus (MRSA) accounted for 19% of S. aureus isolates, associated with mostly staphylococcal cassette chromosome mec (SCCmec) type IV, or V. Panton-Valentine leukocidin (PVL) genes were detected in methicillin-susceptible S. aureus (MSSA) at significantly higher rate (39%) than in MRSA (22%). Among MRSA, ST361 (clonal complex [CC] 361), ST772 (CC1), and ST239 (CC8) were frequently identified, while the most common clone in MSSA was ST2990 (CC1), followed by ST121 and CC8 comprising five STs. Novel coagulase gene genotype XVI was identified in four MSSA isolates. All the S. argenteus isolates were assigned to ST2250 and mecA negative, including only one PVL-positive isolate. MSSA and S. argenteus were co-isolated from two patients, while two different MSSA clones were simultaneously identified in eight patients. This study revealed clonal diversity and genetic characteristics of current MRSA/MSSA/S. argenteus clinical isolates in the national tertiary care hospital in Myanmar.

Going Back in Time: Increasing Penicillin Susceptibility among Methicillin-Susceptible Staphylococcus aureus Osteoarticular Infections in Children.

In the late 1940s to 1950s, Staphylococcus aureus isolates first-gained resistance to penicillin. Recently, some centers have described an increase in the proportion of methicillin susceptible S. aureus (MSSA) which are also susceptible to penicillin (PSSA). There are little data on the frequency of PSSA infections in children. We investigated the prevalence of penicillin susceptibility among pediatric MSSA acute hematogenous osteoarticular infection (OAI) isolates. MSSA OAI isolates were obtained through surveillance studies at Texas Children's and St. Louis Children's Hospitals from January 2011 to December 2019. All isolates underwent PCR for blaZ β-lactamase, PVL genes and agr group. All blaZ negative isolates then underwent penicillin MIC determination. blaZ negative isolates with penicillin MIC ≤ 0.125 μg/mL were considered PSSA. Multilocus sequence typing (MLST) was conducted on a subset of isolates. A total of 329 unique isolates were included in the study. The median patient age was 9.2 years (IQR:5.1 to 12.2). Overall, 6.7% of isolates were penicillin susceptible. No PSSA were detected prior to 2015 but increased yearly thereafter. By the final study year, 20.4% of isolates were PSSA (P = 0.001). PSSA were similar to penicillin-resistant MSSA (PR-MSSA) isolates in terms agr group and PVL carriage as well as clinical presentation and outcomes. PSSA were of distinct sequence types compared to PR-MSSA. PSSA appears to be increasing among OAI in U.S. children. Overall, PSSA isolates are associated with a similar clinical presentation as penicillin-resistant isolates. The potential for use of penicillin treatment in PSSA OAI warrants further study.

Antibiotic resistance and molecular typing of clinical Staphylococcus aureus isolates from Malaysian military hospital

Objective: To determine the antibiotic resistance profile (ARP) of Staphylococcus (S.) aureus isolates and molecular typing of the methicillin-resistant S. aureus (MRSA) isolates from Tuanku Mizan Armed Forces Hospital (TMAFH), Kuala Lumpur. Methods: The ARP and presence of the pvl gene were determined for 209 S. aureus isolates from clinical specimens. Of these, 123 were methicillin-susceptible S. aureus (MSSA) isolates and 86 were MRSA isolates. All MRSA isolates were characterized using SCCmec typing and spa typing. Descriptive analysis was performed to compare the demographic data with the phenotypic and genotypic variables of the S. aureus isolates. Results: No vancomycin-intermediate and -resistant S. aureus (VISA and VRSA, respectively) were detected among the study isolates. The MSSA isolates showed low resistance rates to all tested antibiotics, were commonly invasive (28/42, 66.7%), and mostly harboured pvl (35/42, 83.3%). Meanwhile, MRSA isolates showed high resistance to penicillin (86/86, 100%), ampicillin (86/86, 100%), sulbactam/ampicillin (86/86, 100%), cefuroxime (81/86, 94.19%), cefoperazone (76/86, 88.37%), azithromycin (56/86, 65.12%), and erythromycin (54/86, 62.79%). The majority of MRSA isolates were of SCCmec type IVh (65/86, 75.58%), spa type t032 (55/85, 63.95%), and grouped into spaCC-t022 (66/85, 77.65%). The t032 type was found to be associated with resistance traits to azithromycin and erythromycin (P<0.05). We also found several spa types that are typically associated with hospital-, community-, and livestock-associated MRSA co-existing in our MRSA population. Conclusions: This study reflected the consistent absence of VISA and VRSA and corroborated the clonal shifting of MRSA isolates in the Malaysian MRSA isolates.

Distribution of chlorhexidine resistance genes among Staphylococcus aureus clinical isolates: the challenge of antiseptic resistance.

Chlorhexidine is an antiseptic agent which is extensively used to prevent nosocomial infections; however, this could result in reduction of its susceptibility. The aim of this work was to determine chlorhexidine susceptibility among Staphylococcus aureus isolates and to detect qacA/B and smr antiseptic resistance genes among these isolates. Furthermore, we aimed to identify possible risk factors for the reduction of chlorhexidine susceptibility among S. aureus isolates. Various clinical samples were collected from patients with evidence of S. aureus infection. Antimicrobial susceptibilities of identified S. aureus isolates were determined by disk diffusion method. Resistance to methicillin was identified by cefoxitin disk diffusion test besides mecA gene detection by PCR. Chlorhexidine minimum inhibitory concentration (MIC) values were measured by broth microdilution method while qacA/B and smr resistance genes were detected by multiplex PCR. A total percentage of 25.9% of S. aureus isolates showed reduced susceptibility to chlorhexidine. Methicillin resistant S. aureus (MRSA) had a reported percentage of 39.5%, which was significantly higher than the 11.3% reported for methicillin susceptible S. aureus (MSSA), p<0.001. S. aureus isolates were found to harbor qacA/B and smr genes at 23.2% and 7.7% respectively. Risk factors for reduced susceptibility to chlorhexidine included; ICU setting (OR=2.02, 95%CI: 0.3-1.6), prolonged ICU stay (OR=1.7, 95%CI: 0.4-1.1), presence of central vascular catheter (OR=2.3, 95%CI: 0.2-1.9), mechanical ventilation (OR=1.88, 95%CI: 0.4-1.7) and acquisition of qacA/B (OR=15.7, 95%CI: 3.4-12.1) or smr gene (OR=15.7, 95%CI: 3.4-12.1). Our work highlighted the current challenge of antiseptic resistance in our locality. The frequencies of qacA/B and smr genes were significantly higher among MRSA than MSSA isolates. About two thirds of chlorhexidine tolerant isolates displayed an MDR profile. To maintain chlorhexidine efficiency, biocidal stewardship program and ongoing surveillance are essential.

Genotypes of Staphylococcus aureus Clinical Isolates Are Associated with Phenol-Soluble Modulin (PSM) Production

Phenol-soluble modulins (PSMs) are important S. aureus virulence factors that cause cytolysis, mast cell degranulation, and stimulate inflammatory responses. In this study, PSM production by S. aureus clinical isolates was measured by liquid chromatography/mass spectrometry (LC-MS) and correlated with staphylococcal protein A (spa) type and staphylococcal cassette chromosome mec (SCCmec) type. Of 106 S. aureus clinical isolates, 50 (47.2%) corresponded to methicillin-susceptible S. aureus (MSSA) and 56 (52.8%) to methicillin-resistant S. aureus (MRSA). LC-MS analysis revealed no significant difference in average PSMα3, PSMα4, PSMβ2, and δ-toxin production between MSSA and MRSA isolates, but PSMα1, PSMα2, and PSMβ1 production were higher in MSSA than MRSA. This study demonstrated that average PSMα1–α4, PSMβ1–β2, and δ-toxin production by SCCmec type II strains was significantly lower than the IV, IVA, and V strains. Most of the SCCmec type II strains (n = 17/25; 68.0%) did not produce δ-toxin, suggesting a dysfunctional Agr system. The spa type t111 (except one strain) and t2460 (except one strain producing PSM α1–α4) did not produce PSMα1–α4 and δ-toxin, while average PSM production was higher among the t126 and t1784 strains. This study showed that the genotype of S. aureus, specifically the spa and SCCmec types, is important in characterizing the production of PSMs.

Virulence genes distributed among Staphylococcus aureus causing wound infections and their correlation to antibiotic resistance

BackgroundStaphylococcus aureus causes many human infections, including wound infections, and its pathogenicity is mainly influenced by several virulence factors.AimThis study aimed to detect virulence genes (hla, sea, icaA, and fnbA) in S. aureus isolated from different wound infections among Egyptian patients admitted to Minia University Hospital. This study also aimed to investigate the prevalence of these genes in methicillin-resistant S. aureus (MRSA), methicillin-susceptible S. aureus (MSSA), and vancomycin-resistant S. aureus isolates and the resistance and sensitivity to different antibiotic classes.MethodsA cross-sectional study was carried out from November 2019 to September 2021. Standard biochemical and microbiological tests revealed 59 S. aureus isolates. The Kirby-Bauer disc diffusion method was used to determine antibiotic susceptibility. DNA was extracted using a DNA extraction kit, and polymerase chain reaction was used to amplify all genes.ResultsA total of 59 S. aureus isolates were detected from 51 wound samples. MRSA isolates accounted for 91.5%, whereas MSSA isolates accounted for 8.5%. The multidrug resistance (MDR) percentage in S. aureus isolates was 54.2%. S. aureus showed high sensitivity pattern against vancomycin, linezolid, and chloramphenicol. However, a high resistance pattern was observed against oxacillin and piperacillin. sea was the most predominant gene (72.9%), followed by icaA (49.2%), hla (37.3%), and fnbA (13.6%). sea was the commonest virulence gene among MRSA isolates (72.2%), and a significant difference in the distribution of icaA was found. However, sea and icaA were the commonest genes among MSSA isolates (79.9%). The highest distribution of sea was found among ciprofloxacin-resistant isolates (95.2%).ConclusionThe incidence of infections caused by MDR S. aureus significantly increased with MRSA prevalence. sea is the most predominant virulence factor among antibiotic-resistant strains with a significant correlation to piperacillin, gentamicin, and levofloxacin.

Antibacterial Activities and Synergistic Interaction of Citrus Essential Oils and Limonene with Gentamicin against Clinically Isolated Methicillin-Resistant Staphylococcus aureus.

Citrus reticulata Blanco and Citrus aurantifolia are the edible plants which contain several biological properties including antibacterial activity. The aims of the present study were to determine the chemical compositions and evaluate antibacterial activities of citrus essential oils extracted from the fruit peels of C. reticulata (CREO) and C. aurantifolia (CAEO), alone and in combination with gentamicin, against a panel of clinically isolated methicillin-resistant S. aureus (MRSA) (n = 40) and methicillin-susceptible S. aureus (MSSA) (n = 45). Gas chromatography-mass spectrometry analysis revealed that 12 and 25 compounds were identified in CREO and CAEO with the most predominant compound of limonene (62.9–72.5%). The antibacterial activities were determined by agar disk diffusion and resazurin-based microdilution methods. The results found that almost all MRSA isolates were resistant to ciprofloxacin, erythromycin, and clindamycin, and some isolates were resistant to gentamicin. CREO and CAEO exhibited inhibitory effects toward clinical isolates (MIC: 1.0–32.0 and 8.0–32.0 mg/mL, respectively), with a similar trend to limonene (MIC: 1.0–32.0 mg/mL). However, the higher antibacterial effects were found in CREO and limonene when compared to CAEO (p < 0.01). In combination effect, the results showed the synergistic interaction of gentamicin with CREO and limonene on the MRSA and MSSA isolates (FIC indexes: 0.012–0.258 and 0.012–0.375), but that interaction of gentamicin with CAEO was observed only on MRSA (FIC index: 0.012–0.016). These findings demonstrated the potential of these citrus essential oils as natural antibacterial agents that may contribute to reduce the emerging of antimicrobial-resistant bacteria.

Virulence, agr groups, antimicrobial resistance and epidemiology of Staphylococcus aureus isolated from bovine subclinical mastitis

Fifty-two Staphylococcus aureus recovered from papillary ostium and milk samples collected from cows with subclinical mastitis and milking environments in three small dairy herds located in southeastern Brazil were subjected to PCR identification based on the thermonuclease (nuc) gene. All the strains were submitted to in vitro antimicrobial susceptibility testing, and we investigated the sequence types (STs), agr groups (I-IV), virulence genes encoding for Microbial Surface Components Recognizing Adhesive Matrix Molecules (MSCRAMMs), biofilm-associated proteins, bi-component toxins, pyrogenic toxin superantigens, and enterotoxins. Screening for oxacillin resistance (2-6 μg/ml oxacillin), beta-lactamase activity assays, and PCR for the mecA/mecC genes detected 26 methicillin-susceptible S. aureus (MSSA) and 26 mec-independent oxacillin-nonsusceptible S. aureus (MIONSA). While MSSA isolates were found to be susceptible to all antimicrobial agents tested, or only resistant to penicillin and ampicillin, MIONSA isolates were multidrug-resistant. ST126-agr group II MSSA isolates were prevalent in milk (n=14) and carried a broad set of virulence genes (clfA, clfB, eno, fnbA, fiB, icaA, icaD, lukED, hla, and hlb), as well as the ST126-agr group II MIONSA isolated from milking liners (n=1), which also carried the eta gene. ST1-agr group III MIONSA isolates (n=4) were found in papillary ostium and milk, but most MIONSA isolates (n=21), which were identified in both papillary ostium and milking liners, were agr-negative and assigned to ST126. The agr-negative and agr group III lineages showed a low potential for virulence. Studies on the characterization of bovine-associated MSSA/MIONSA are essential to reduce S. aureus mastitis to prevent economic losses in dairy production and also to monitor the zoonotic potential of these pathogens associated with invasive infections and treatment failures in healthcare.

Whole-genome sequence analysis of Staphylococcus aureus from retail fish acknowledged the incidence of highly virulent ST672-MRSA-IVa/t1309, an emerging Indian clone, in Assam, India.

The epidemiology and toxigenicity of MRSA in the fishery environment are poorly understood. In this study, methicillin-resistant Staphylococcus aureus (MRSA) (n= 1) and methicillin-susceptible S. aureus (MSSA) (n= 2) from retail fish were subjected to comprehensive genome analysis. Here, we report the occurrence of ST672-MRSA-IV/t1309 and ST5-MSSA/t105 for the first time from India in the fishery environment. The resistome of the isolates was in concordance with their phenotypic resistance pattern. Phenotypically, the resistance profile of MSSA isolates (n= 2) was AMP-CLI-ERY-NOR-PEN. For MRSA (n= 1), it was AMP-CFZ-CLI-ERY-NOR-OXA-PEN. The antibiotic efflux genes and mutations in the antibiotic target accounted for fluoroquinolone resistance whereas methicillin resistance was conferred through possession of a mecA gene. Similarly, all three isolates carried a similar array of virulence factors. The conjugative plasmid inc18 and rep family 10 plasmids were found in two of the three isolates. This study documents the MRSA carrying SCCmec IVa elements which are the markers of community-associated MRSA (CA-MRSA). Through the possession of SCCmec IV elements, which are smaller than other types of SCCmec, MRSA can contribute to the rapid dissemination of antimicrobial resistance and virulence factors. In short, our findings highlighted that the presence of ST672-MRSA in fishery environments may pose a risk to human health.