Methicillin-resistant Staphylococcus Aureus Isolates Research Articles

Antibiofilm and synergistic antibacterial effect of Garlic (Allium sativum) against Methicillin resistant Staphylococcus aureus (MRSA)

Background: Methicillin resistant Staphylococcus aureus (MRSA) is one of main bacterial pathogens that is associated with difficult to treat nosocomial and community infections with high mortality. Shiga toxin producing E. Coli (STEC) infections result in bloody diarrhea, vomiting and mild fever. Biofilm formation has been linked with pathogenicity and virulence of these bacterial pathogens. Garlic (Allium sativum) has been traditionally used as an antibacterial agent in India and South East Asia. We have demonstrated that garlic exhibits antibiofilm and antibacterial activity against drug resistant STEC clinical isolates. In addition, organosulphide compounds of garlic have been reported to exhibit antibacterial activity against MRSA. In view of these observations, our laboratory wanted to investigate if crude garlic extract has any antibiofilm activity against MRSA. Further, we wanted to test if for synergistic effect of garlic when used in combination with antibiotics. Methods and materials: The study was performed with three MRSA isolates. Standard disc diffusion, serial dilution and kill curve assays were performed to access the antibacterial and synergistic activity of garlic against MRSA. In vitro microtiter plate assay was performed to quantify the formation of biofilm in the presence and absence of garlic crude extract. In addition, the effect of garlic on biofilm formation was also evaluated using scanning electron microscopy (SEM). Results: The results indicated that crude garlic extract exhibited significant dose dependent antibacterial activity against MRSA and the activity was much better in terms of clarity of zones of inhibition compared to pure organosulphide compounds found in garlic. It was interesting to note that sub-MIC concentrations of garlic were able to sensitize MRSA to methicillin. Garlic at sub-MIC concentrations reduced MRSA biofilm formation by more than 60%. We conclude that crude garlic extract has more antibacterial activity than its active ingredients. This is the first study to report its antibiofilm property against MRSA to best of our knowledge. Conclusion: Garlic organosulphide compounds have the potential to be used in combination with standard antibiotics to treat drug resistant bacterial infections. Garlic organosulphide compounds based novel antibiotic and antibiofilm drugs can be developed to tackle the problem of antibacterial drug resistance.

Prevalence of Methicillin Resistant Staphylococcus aureus (MRSA) in Cattle Milk from dairy Herds in Oyo State, Nigeria

Methicillin-resistant Staphylococcus aureus (MRSA) has received a lot of attention in recent years as a zoonotic organism of global concern. Contaminated milk, especially those from mastitic cows, serve as reservoirs for humans in the epidemiology of antibiotic resistant MRSA. This study was designed to determine the level of contamination of bulk fresh milk from dairy cattle herds with MRSA in Ibarapa, Oyo and Oke-Ogun areas of Oyo State and the antibiotic resistance profile of the isolates. One hundred and sixty-five (165) milk samples were obtained from the study areas and used for the study. Staphylococcus aureus was isolated from the samples using bacterial culture and biochemical tests. Methicillin-resistant Staphylococcus aureus was identified using cefoxitin disk diffusion method. All the S. aureus isolates were subjected to microbial susceptibility test. Ninety (54.5%) milk samples were positive for Staphylococcus spp. out of which 52 (31.5%) were Staphylococcus aureus and 13 (7.9%) yielded MRSA. Antibiogram of S. aureus indicated highest resistance to Cloxacillin (88.5%) followed by (Augmentin 67.3%) and Ceftrazidine (67.3%). Ten out of the 13 MRSA isolates were multidrug resistance while all the isolates were 100% susceptible to ofloxacin. The results of this study showed that milk produced from dairy cattle in Oyo State was contaminated with MRSA. This portends serious food safety and public health risk among the consumers of such milk especially in raw or improperly pasteurized form. Proper dairy herd health management and prudent use of antibiotics and hygienic milking procedures are hereby recommended to prevent contamination of milk and subsequent spread of the organism to humans.

Molecular analysis and the toxin, MSCRAMM, and biofilm genes of methicillin-resistant Staphylococcus aureus strains isolated from pemphigus wounds: A study based on SCCmec and dru typing.

Pemphigus is a chronic autoimmune blistering disease. Pemphigus blisters can damage the natural skin barrier and increase the risk of life-threatening conditions. Colonization of pemphigus wounds with methicillin-resistant Staphylococcus aureus (MRSA) prolongs wound healing and increases mortality rate. Assessing MRSA prevalence, types, and toxin and adhesion genes can facilitate the detection of MRSA strains which cause infections, selection of appropriate treatments, and healing of pemphigus wounds. This study aimed to determine the SCCmec, the direct repeat unit (dru) types (dts), and the toxin, MSCRAMM, and biofilm genes of MRSA strains isolated from pemphigus wounds. In this cross-sectional study, 118S. aureus isolates were gathered from 118 patients with pemphigus. MRSA detection was performed using the mecA gene. Using the polymerase chain reaction method, all MRSA isolates were assessed for the presence of the sea, seb, sec, tst, eta, pvl, hla, hlb, MSCRAMM, and ica genes. Typing and subtyping were performed through respectively SCCmec typing and dru typing methods. The Bionumerics software was used for analyzing the data and drawing the minimum spanning tree. From 118S. aureus isolates, 51 were MRSA. SCCmec typing revealed the prevalence of SCCmec II with a prevalence of 64.7% (33 out of 51 isolates) and SCCmec III with a prevalence of 35.3% (18 out of 51 isolates). Dru typing indicated seven dts, namely dts 10a, 10g, 10m, 13i, 8h, 8i, and 9ca in two main clusters. The dt9ca was a new dru type and was registered in the dru-typing database (www.dru-typing.org). The prevalence rates of the hla, sea, and sec genes in MRSA isolates were respectively 54.9%, 27.4%, and 1.9%, while the hlb, seb, eta, and pvl genes were not detected at all. Only one MRSA with SCCmec III and dt10a carried the tst encoding gene. MSCRAMM gene analysis revealed the high prevalence of the eno (31.3%) and the fib (21.5%) genes. The prevalence rates of the icaA and icaD biofilm formation genes were 3.9% and 5.8%, respectively. There were no significant differences between the two detected SCCmec types and between the two detected dts clusters respecting the prevalence of the encoding genes of virulence factors and MSCRAMMs. The toxin genes hla and sea are prevalent among MRSA strains with SCCmec II and III isolated from pemphigus wounds. The most prevalent dts are dt10a and dt10g among MRSA with SCCmec III and dt8h and dt8i among MRSA with SCCmec II.

Development and application of a triplex-PCR assay for rapid detection of methicillin-resistant Staphylococcus aureus from pigs.

A triplex-PCR assay was developed and evaluated for rapid detection of methicillin-resistant Staphylococcus aureus (MRSA) recovered from various biological samples of pig. Three sets of primers were designed to target mecA, 16S rRNA and nuc genes of MRSA. The specific amplification generated three bands on agarose gel, with sizes 280bp for mecA, 654bp for 16S rRNA and 481bp for nuc, respectively. A potential advantage of the PCR assay is its sensitivity with a detection limit of 102 CFU per ml of bacteria. In all, 79 MRSA isolates recovered from various samples of pigs were subjected to the amplification by the triplex-PCR assay and all the isolates yielded three bands corresponding to the three genes under this study. No false-positive amplification was observed, indicating the high specificity of the developed triplex-PCR assay. This assay will be a useful and powerful method for differentiation of MRSA from methicillin-sensitive S. aureus, coagulase-negative methicillin-resistant staphylococci and coagulase-negative methicillin-sensitive staphylococci.

Antibacterial Activity of Prunus Scoparia Root Methanol Extract against Most Common Burn Wound Pathogens.

Background: Burn wound infection and sepsis are serious medical conditions requiring prompt intervention. Plants are a good natural source for the development of novel, safe, and cost-effective antibacterial agents. The objective of the present study was to assess the antibacterial potential of aqueous, chloroform, and methanol extracts of the Prunus scoparia (P. scoparia) root against the most common burn wound pathogens.Methods: The present experimental study was conducted at Shiraz University of Medical Sciences (Shiraz, Iran) during 2018-2019. The antibacterial activity of the total plant extract was assayed using the broth microdilution method. Fractionation was performed using a separation funnel and solvents with different polarities. Broth microdilution and agar well diffusion assays were performed to determine the antibacterial potential of the obtained fractions. Quantitative and qualitative phytochemical analyses were performed to confirm the presence of secondary metabolites in both the total extract and the fractions.Results: Methanolic extract of P. scoparia root exhibited antibacterial activity against all tested bacterial strains, especially against Methicillin-resistant Staphylococcus aureus (MRSA) isolates. This extract, compared to the aqueous and chloroformic extracts, exhibited the presence of active antibacterial compounds. The quantitative and qualitative results of phytochemical screening showed that phenols and flavonoids were the main antibacterial compounds in the methanolic extract of the plant.Conclusion: For the first time, we demonstrated the antibacterial activity of the P. scoparia root against MRSA isolates and other common burn wound pathogens.

Clinical characteristics and molecular analysis of USA300 and ST 764 methicillin-resistant Staphylococcus aureus isolates from outpatients in Japan by PCR-Based open reading frame typing

IntroductionUSA300 is the most common community-acquired methicillin-resistant Staphylococcus aureus (MRSA) strain. Sequence type (ST) 764 MRSA is a new local variant of the ST 5 lineage. The objective of this study was to determine the clinical characteristics of USA300 and ST 764 infections among outpatients in Japan. MethodsWe obtained MRSA isolates from 132 outpatients who visited our hospital from January 2016 to December 2017 and compared USA300 infection group to ST 764 infection group. Molecular analysis, including that of various toxins and other virulence factors, of the MRSA isolates were performed. In particular, we investigated the relationships among PCR-based open reading frame typing (POT) scores, MRSA clones, and virulence factors. ResultsTwenty-seven USA300 isolates (20.5%) and 16 ST 764 isolates (12.1%) were identified. Although USA300 and ST 764 had lower rates of risk factors, their infection rates were higher. USA300-infected patients had higher rates of deep skin and soft tissue infections compared with the non-USA300 CA-MRSA-infected patients. Notably, the USA300 and ST 764 isolates had unique POT scores. ConclusionsOur results indicated that USA300 MRSA was spreading in an area 120 km west of Tokyo, Japan. We observed multiple cases of ST 764 MRSA infection, raising concerns about the antimicrobial resistance of ST 764, as it limits the choices of antibiotics to treat infection. The POT score can predict the presence of toxins and virulence factors, as well as the clone identity of MRSA with high accuracy.

The Evolving Reduction of Vancomycin and Daptomycin Susceptibility in MRSA-Salvaging the Gold Standards with Combination Therapy.

Methicillin-resistant Staphylococcus aureus (MRSA) is associated with substantial morbidity and mortality. Vancomycin (VAN) has been used as the gold standard treatment for invasive MRSA infections for decades but, unfortunately, the reliance of VAN as the primary treatment option against these infections has led to a reduction in VAN susceptibility in MRSA isolates. Although daptomycin (DAP) is another common treatment option against invasive MRSA infections, it has been shown that the development of VAN resistance can lead to DAP nonsusceptibility. VAN or DAP backbone regimens in combination with other antibiotics has been advocated as an alternative approach to improve patient outcomes in VAN/DAP-susceptible infections, enhance outcomes in infections caused by isolates with reduced VAN/DAP susceptibility, and/or prevent the emergence of VAN/DAP resistance or further resistance. A peer-reviewed literature search was conducted using Medline, Google Scholar and PubMed databases. The primary purpose of this review is to describe the mechanisms and epidemiology of MRSA isolates with a reduction in VAN and/or DAP susceptibility, evaluate in vitro and in vivo literature describing combination therapy (CT) against MRSA isolates with reduced VAN and/or DAP susceptibility and describe studies involving the clinical outcomes of patients treated with CT against invasive MRSA infections.

Novel Antibiotic Combinations of Diverse Subclasses for Effective Suppression of Extensively Drug-Resistant Methicillin-Resistant Staphylococcus aureus (MRSA).

The emergence of multidrug-resistant pathogens such as methicillin-resistant Staphylococcus aureus (MRSA), the chief etiological agent for a range of refractory infections, has rendered all β-lactams ineffective against it. The treatment process is further complicated with the development of resistance to glycopeptides, primary antibiotics for treatment of MRSA. Antibiotic combination therapy with existing antimicrobial agents may provide an immediate treatment option. Minimum inhibitory concentrations (MICs) of 18 different commercially available antibiotics were determined along with their 90 possible pairwise combinations and 64 triple combinations to filter out 5 best combinations. Time-Kill kinetics of these combinations were then analyzed to find collateral bactericidal combinations which were then tested on other randomly selected MRSA isolates. Among the top 5 combinations including levofloxacin-ceftazidime; amoxicillin/clavulanic acid-tobramycin; amoxicillin/clavulanic acid-cephradine; amoxicillin/clavulanic acid-ofloxacin; and piperacillin/tazobactam-tobramycin, three combinations were found to be collaterally effective. Levofloxacin-ceftazidime acted synergistically in 80% of the tested clinical MRSA isolates. First-line β-lactams of lower generations can be used effectively against MRSA infection when used in combination. Antibiotics other than glycopeptides may still work in combination.

Genotyping of methicillin resistant Staphylococcus aureus from the United Arab Emirates

Reports from Arabian Gulf countries have demonstrated emergence of novel methicillin resistant Staphylococcus aureus (MRSA) strains. To address the lack of data from the United Arab Emirates (UAE), genetic characterisation of MRSA identified between December 2017 and August 2019 was conducted using DNA microarray-based assays. The 625 MRSA isolates studied were grouped into 23 clonal complexes (CCs) and assigned to 103 strains. CC5, CC6, CC22 and CC30 represented 54.2% (n/N = 339/625) of isolates with other common CCs being CC1, CC8, CC772, CC361, CC80, CC88. Emergence of CC398 MRSA, CC5-MRSA-IV Sri Lanka Clone and ST5/ST225-MRSA-II, Rhine-Hesse EMRSA/New York-Japan Clone in our setting was detected. Variants of pandemic CC8-MRSA-[IVa + ACME I] (PVL+) USA300 were detected and majority of CC772 strains were CC772-MRSA-V (PVL+), “Bengal- Bay Clone”. Novel MRSA strains identified include CC5-MRSA-V (edinA+), CC5-MRSA-[VT + fusC], CC5-MRSA-IVa (tst1+), CC5-MRSA-[V/VT + cas + fusC + ccrA/B-1], CC8-MRSA-V/VT, CC22-MRSA-[IV + fusC + ccrAA/(C)], CC45-MRSA-[IV + fusC + tir], CC80-MRSA-IVa, CC121-MRSA-V/VT, CC152-MRSA-[V + fusC] (PVL+). Although several strains harboured SCC-borne fusidic acid resistance (fusC) (n = 181), erythromycin/clindamycin resistance (ermC) (n = 132) and gentamicin resistance (aacA-aphD) (n = 179) genes, none harboured vancomycin resistance genes while mupirocin resistance gene mupR (n = 2) and cfr gene (n = 1) were rare. An extensive MRSA repertoire including CCs previously unreported in the region and novel strains which probably arose locally suggest an evolving MRSA landscape.

Carbapenems drive the collateral resistance to ceftaroline in cystic fibrosis patients with MRSA

Chronic airways infection with methicillin-resistant Staphylococcus aureus (MRSA) is associated with worse respiratory disease cystic fibrosis (CF) patients. Ceftaroline is a cephalosporin that inhibits the penicillin-binding protein (PBP2a) uniquely produced by MRSA. We analyzed 335 S. aureus isolates from CF sputum samples collected at three US centers between 2015–2018. Molecular relationships demonstrated that high-level resistance of preceding isolates to carbapenems were associated with subsequent isolation of ceftaroline resistant CF MRSA. In vitro evolution experiments showed that pre-exposure of CF MRSA to meropenem with further selection with ceftaroline implied mutations in mecA and additional mutations in pbp1 and pbp2, targets of carbapenems; no effects were achieved by other β-lactams. An in vivo pneumonia mouse model showed the potential therapeutic efficacy of ceftaroline/meropenem combination against ceftaroline-resistant CF MRSA infections. Thus, the present findings highlight risk factors and potential therapeutic strategies offering an opportunity to both prevent and address antibiotic resistance in this patient population.

Vancomycin MIC and agr dysfunction in invasive MRSA infections in southern Brazil.

In methicillin-resistant Staphylococcus aureus (MRSA) treatment, the vancomycin minimum inhibitory concentration (MIC) increase, vancomycin heteroresistance (hVISA) presence, and accessory gene regulator (agr) dysfunction are predictors of vancomycin therapy failure. This study evaluated the association between vancomycin MIC (≥ 1.0μg/mL) and agr dysfunction in invasive MRSA isolates. Vancomycin MIC, hVISA phenotype, agr group, and function were determined in 171 MRSA isolates obtained between 2014 and 2019 from hospitals in Porto Alegre, Brazil. All MRSA were susceptible to vancomycin; 16.4% of these had MIC ≥ 1.0μg/mL. Seventeen MRSA isolates expressed the hVISA phenotype; 35.3% of them had MIC of 1.5μg/mL. agr groups I (40.9%) and II (47.1%) were the most found groups for MRSA and hVISA isolates, respectively. The proportion of MRSA with vancomycin MIC ≥ 1.0μg/mL in agr group II was significantly higher than in agr groups I and III (p = 0.002). agr dysfunction was observed in 4.7% (8/171) of MRSA, especially those with vancomycin MIC ≥ 1.0μg/mL (p < 0.001). In addition, six isolates (35.3%; 6/17) with hVISA phenotype presented agr dysfunction, which was significantly higher than that in non-hVISA phenotype (p < 0.001). In conclusion, agr dysfunction in MRSA is associated with vancomycin MIC ≥ 1.0μg/mL and hVISA phenotype, which suggests that agr dysfunction might confer potential advantages on MRSA to survive in invasive infections.

Biofilm formation in Methicillin Resistant Staphylococcus aureus

Methicillin Resistant Staphylococcus aureus (MRSA) is one of the important antibiotic resistance pathogen. Chronic hospital acquired infections by S. aureus have become a major problem in recent years. One of the main mechanisms for chronic hospital acquired infections is defined by biofilm formation. Therefore, the current research aimed to identify the prevalence of biofilm formation among MRSA clinical isolates in Ilam hospitals and to determine the gene responsible for biofilm formation. For this reason, 26 MRSA clinical isolates were identified and subjected to biofilm by micro-plate assay and PCR for identification of icaa and icaD. The results demonstrated that 69.2% (n = 18) of MRSA clinical isolates possess icaD and were interestingly were negative for icaa; while biofilm formation were negative in three isolates that were positive for icaD. The current study strongly recommended icaD as a main factor to produce the biofilm formation in MRSA clinical isolates and suggested this gene as a target for antibiofilm therapy. The reason for negatively in 2 isolates could be explained in the abstract by low accuracy of micro-late assay. Another reason that could be concluded was silencing of icaD in these three isolates.

Genomic Characterization of Methicillin-Resistant Staphylococcus aureus (MRSA) by High-Throughput Sequencing in a Tertiary Care Hospital.

Methicillin-resistant Staphylococcus aureus (MRSA) strains are associated with serious complications and poor clinical outcome. In Egypt, they contribute to more than 70% of S. aureus healthcare-associated infections. This study combined whole-genome sequencing, bioinformatics, and statistical analyses to identify the phylogeny, resistome, virulome and potential genotype–phenotype–clinical correlation among 18 clinical isolates of MRSA in a tertiary hospital in Cairo, Egypt. The ST1535-V MRSA clone was the most frequently isolated (16.6%), followed by ST5-VI, ST1-V and ST239-III (11.1% each). SCCmec V, VI, IV and III types were detected at frequencies of 50%, 16.6%, 11.1% and 11.1%, respectively. None of the tested virulence genes were detected in all isolates, but they ranged in distribution from 1/18 to 17/18. The Panton–Valentine leukocidin (PVL)-encoding genes were detected in only four isolates and were enriched in isolates causing non-severe cases. Phylogenetic analysis revealed relatedness between three ST1535-Vs, two ST5-VIs, two ST239-IIIs and two ST1-Vs; however, only the two genetically related ST1-V isolates were epidemiologically linked. While disease outcome and source of infection had no correlation with a particular genotypic pattern, the sequence type was the most correlated factor with phylogeny and genotypic patterns, and a few genes were associated with non-severe cases.

In vitro antibacterial activity of Acorus calamus extract on methicillin-resistant Staphylococcus aureus wound isolates and reduced invasion into mucosal fibroblasts

Acorus calamus is widely used as an Indian medicinal plant and extensively reported for treatments of skin, digestive and nervous disorders. A methanol extract of A. calamus (rhizome) showed maximum extraction yield of 4% (w/w) followed by 3% (w/w) in water. Total 8 Staphylococcus aureus isolates were obtained from nose, groin, wounds, axilla, throat of various patients exhibited strong β-lactamase activity and antibiotic resistance for ampicillin, oxacillin, and methicillin coded for mec A gene. Thus, all isolates belong to methicillin-resistant Staphylococcus aureus (MRSA) class. Methanol extract is the most potent MRSA antibacterial extract followed by water, n-butanol and chloroform. Minimum inhibitory concentration (MIC) of methanol extract varied from 1.25 to 2.50 mg/ml for all MRSA isolates which also exhibited highest MIC values of 512 µg/ml and 128 µg/ml for ampicillin and oxacillin antibiotics, respectively. Interestingly, the synergistic interaction of 128 µg/ml of A. calamus methanol extract with antibiotics (ampicillin, 64 µg/ml and oxacillin, 32 µg/ml) showed lowered MICs. All extracts of A. calamus and β-asarone showed no significant cytotoxic activity against non-cancerous 3T3 fibroblasts cells which was confirmed by MTT assay on MRSA-08 isolate. Antibacterial effect and preventive invasion properties of MRSA of methanol extract of A. calamus were found to be potent when compared with β-asarone, an active component of A. calamus. The present investigation showed a correlation of polyphenolics and flavonoid based major phytochemical constituents with bioactivities on MRSA revealed A. calamus as a potential non-toxic drug candidate for various skin diseases.

Differentiation of qacA and qacB using high-resolution melt curve analysis, and both qacA and qacB but not qacC or norA types increase chlorhexidine minimal inhibitory concentrations in Staphylococcus aureus isolates.

Chlorhexidine is one of the most essential ingredients in infection control applications. Except qacA, the effects of other various efflux-medicated biocide genes (including qacB, qacC, qacEΔ1, qacH or norA) on biguanides resistance are still controversial. In addition, most of the studies have discussed the effect of qacA/B on clinical S. aureus isolates but not that qacA or qacB individually. In total, 254 methicillin-resistant Staphylococcus aureus (MRSA), selected 30 methicillin-susceptible S. aureus (MSSA) clinical isolates from different patients during 2014-2015 and 15 S. aureus quality control strains (including Mu3 and Mu50) were included in the study. Various biocide genes, including qacA/B, qacC, qacH, qacEΔ1, and different types of norA, were determined through conventional PCR. S. aureus isolates with qacA/B (+) were analyzed using high-resolution melting curve (HRM) to differentiate qacA from qacB. The chlorhexidine MIC was determined using the agar dilution method. Univariate and multivariate statistics were analyzed to see which biocide resistant genes had effects on chlorhexidine MIC. Results of all HRM analyses (n = 22) were consistent with those of Sanger sequencing for differentiation of qacA from qacB. None of the isolates harbored qacH and only one MRSA harbored qacEΔ1. The harboring rate of qacA, qacB, and qacC among MRSA/MSSA isolates was 7.1% (n = 18)/0%, 38.2% (n = 97)/0%, and 7.5% (n = 19)/3.3% (n = 1), respectively. The most type of norA was norAI (n = 158), followed by norAIII (n = 87) and norAII (n = 9) among MRSA isolates. Based on the results of multivariate logistic regression analyses, only qacA and qacB would increase chlorhexidine MIC from ≤ 1 ug/ml to ≥ 2 ug/ml in MRSA isolates (P < 0.001) but not qacC or norA types (P=0.976 and 0.633 or 0.933, respectively). In addition, only qacA but not qacB was contributed to elevate chlorhexidine from ≤ 1 ug/ml to 4 ug/ml in MRSA isolates (P < 0.001 and 0.017, respectively). HRM analysis can be a great method to differentiate qacA from qacB. The biocide gene with the most effect on chlorhexidine MIC in S. aureus isolates was qacA, followed by qacB, but qacC and different types of norA did not have any effect on chlorhexidine susceptibility. Further investigation on the influence of qacB, qacC and types of norA on chlorhexidine susceptibility is necessary.

Promising Antibiofilm Agents: Recent Breakthrough against Biofilm Producing Methicillin-Resistant Staphylococcus aureus.

Multidrug resistant (MDR) methicillin-resistant Staphylococcus aureus (MRSA) is a superbug pathogen that causes serious diseases. One of the main reasons for the lack of the effectiveness of antibiotic therapy against infections caused by this resistant pathogen is the recalcitrant nature of MRSA biofilms, which results in an increasingly serious situation worldwide. Consequently, the development of innovative biofilm inhibitors is urgently needed to control the biofilm formation by this pathogen. In this work, we thus sought to evaluate the biofilm inhibiting ability of some promising antibiofilm agents such as zinc oxide nanoparticles (Zno NPs), proteinase K, and hamamelitannin (HAM) in managing the MRSA biofilms. Different phenotypic and genotypic methods were used to identify the biofilm producing MDR MRSA isolates and the antibiofilm/antimicrobial activities of the used promising agents. Our study demonstrated strong antibiofilm activities of ZnO NPs, proteinase K, and HAM against MRSA biofilms along with their transcriptional modulation of biofilm (intercellular adhesion A, icaA) and quorum sensing (QS) (agr) genes. Interestingly, only ZnO NPs showed a powerful antimicrobial activity against this pathogen. Collectively, we observed overall positive correlations between the biofilm production and the antimicrobial resistance/agr genotypes II and IV. Meanwhile, there was no significant correlation between the toxin genes and the biofilm production. The ZnO NPs were recommended to be used alone as potent antimicrobial and antibiofilm agents against MDR MRSA and their biofilm-associated diseases. On the other hand, proteinase-K and HAM can be co-administrated with other antimicrobial agents to manage such types of infections.

Trimethoprim-Sulfamethoxazole Resistance Patterns Among Methicillin-Resistant Staphylococcus aureus, 2012–2018

Background: Trimethoprim-sulfamethoxazole is commonly used for the treatment of noninvasive methicillin-resistant Staphylococcus aureus (MRSA) infections. Following a report from 2 facilities of increased trimethoprim-sulfamethoxazole resistance among MRSA infections, we assessed changes in resistance nationally and by state. Methods: We reviewed antibiotic susceptibility testing (AST) data for trimethoprim-sulfamethoxazole among S. aureus isolates associated with surgical site infections (SSIs), central-line–associated bloodstream infections (CLABSIs), and catheter-associated urinary tract infections (CAUTIs) from acute-care hospitals reported to the NHSN Device and Procedure Module from 2012 to 2018. We compared the pooled mean percentage of isolates nonsusceptible to trimethoprim-sulfamethoxazole in 2012 and 2018, stratified by MRSA and methicillin-sensitive Staphylococcus aureus (MSSA). Among MRSA isolates, we compared the percentage nonsusceptible to trimethoprim-sulfamethoxazole by healthcare-associated infection (HAI) type and state in 2012 and 2018. States with ≥20 MRSA isolates with AST reported each year were included in the state-level analysis. Results: Overall, 36,587 MRSA isolates and 46,824 MSSA isolates were reported from 2012 to 2018. Moreover, &gt;80% of MRSA and MSSA isolates had trimethoprim-sulfamethoxazole AST reported each year. Nationally, the percentage of trimethoprim-sulfamethoxazole nonsusceptible among MRSA isolates was 3.9% in 2012 compared to 6.5% in 2018 (P &lt; .001), but it was unchanged among MSSA isolates during the same period (1.1% in 2012 vs 1.4% in 2018; P = .08). Among MRSA surgical site infections (SSIs), the proportion of trimethoprim-sulfamethoxazole nonsusceptible isolates was 3.1% in 2012 versus 6.1% in 2018 (P &lt; .001) but did not change significantly for CLABSIs or CAUTIs (Fig. 1). Among the 32 states that met the inclusion criteria, there were no significant decreases, whereas 4 (12.5%) showed significant increases in the percentage of MRSA that were trimethoprim-sulfamethoxazole nonsusceptible in 2018 compared to 2012: New Jersey (2.4% in 2012 vs 19.3% in 2018; P &lt;.001); Florida (9.1% in 2012 vs 22.4% in 2018; P &lt; .001); Maryland (0.0% in 2012 vs 10.9% in 2018; P &lt; .01); and Pennsylvania (1.7% in 2012 vs 6.5% in 2018; P &lt; .001). Conclusions: Nationally, there was a modest but significant increase in the percentage of MRSA HAI isolates nonsusceptible to trimethoprim-sulfamethoxazole in 2018 compared to 2012; however, 3 of 4 states with significant increases in nonsusceptibility had substantial, potentially clinically relevant increases (&gt;10%). Ongoing characterization of MRSA isolates from Florida and New Jersey may provide insight into the underlying cause of these shifting patterns in trimethoprim-sulfamethoxazole resistance among MRSA. Healthcare personnel should select appropriate antibiotic regimens based on local resistance patterns, should monitor patients for treatment failure, and should report changes in resistance to the appropriate public health department.Funding: NoneDisclosures: None

Use of Varying Single-Nucleotide Polymorphism Thresholds to Identify Strong Epidemiologic Links Among Patients with Methicillin-Resistant Staphylococcus aureus (MRSA)

Background: Whole-genome sequencing (WGS) has a high discriminatory power in confirming outbreaks. Outbreak investigation models that categorize the possibility of an outbreak based on the degree of genetic relatedness of isolates are highly dependent on the single-nucleotide polymorphism (SNP) threshold used. Methods: NYU Langone Medical center is a 725-bed academic center that has implemented WGS of methicillin-resistant Staphylococcus aureus (MRSA) isolates since 2016. Patients admitted to a medical or intensive care unit were screened on admission and transfer. The first surveillance and clinical MRSA isolate during each hospitalization was sequenced. We conducted a retrospective analysis to identify strong epidemiologic links among patients involved in genetically related clusters. We used different SNP thresholds to define genetic relatedness to identify the optimal threshold that should prompt an outbreak investigation. We considered strong hospital epidemiologic links sharing the same room or unit or having resided in the same room or unit within 7 days. A pairwise analysis was conducted to compare the epidemiologic links among patients involved in genetically related clusters. Results: Among 1,070 isolates, our analysis focused on 777 belonging to USA100 and USA300 clones. For USA100 isolates, we identified 8, 14, and 20 clusters comprising of 16, 29, and 42 patients when the threshold for genetic relatedness was set at 20, 40, and 60 SNP differences, respectively. Patients identified in a cluster yielded a strong hospital epidemiologic link in 62.5%, 87.5%, and 91.7% of cases (Fig. 1). For USA300 isolates, SNP differences of 10, 20, and 30 were used, identifying 20, 34, and 40 clusters of 43, 79, and 127 patients. The expansion of the threshold from 10 to 30 resulted in a decrease of the percentage of pairwise analyses with a strong hospital epidemiologic link from 57.7% to 13.6% by increasing 13-fold the number of analyses that were conducted to identify only 3 times more cases with strong epidemiologic links (Fig. 2). Conclusions: The results of our study indicate that SNPs thresholds determined by intrapatient variability of MRSA isolates might need to be tailored to the individual setting to guide infection control interventions because optimal thresholds might vary depending on characteristics of the population, MRSA isolates, and screening practices. Establishing conservative thresholds might allow the identification and quantification over time of the locations (eg, rooms or units) where transmission is occurring as well as the investigation of the clusters without strong epidemiologic links that might be valuable in elucidating unrecognized routes of transmission.Funding: NoneDisclosures: None

Antimicrobial Nonsusceptibility Among Invasive MRSA USA300 Strains by Healthcare Exposure, Three Sites, 2005–2016

Background: Incidence of community-associated (CA) and healthcare-associated, community-onset (HACO) USA300 methicillin-resistant Staphylococcus aureus (MRSA) bloodstream infections has remained unchanged in recent years. Traditionally considered a CA strain, USA300 is increasingly associated with healthcare settings. We examined whether antimicrobial nonsusceptibility among USA300 strains could distinguish epidemiologic class (community vs hospital), and whether divergences in susceptibility were occurring over time. Methods: We used data on invasive MRSA infections from active, population, and laboratory-based surveillance during 2005–2016 from 11 counties in 3 states. Invasive cases were defined as MRSA isolation from a normally sterile site in a surveillance area resident. Cases were considered hospital-onset (HO) if the culture was obtained &gt;3 days after hospitalization and HACO if ≥1 of the following risk factors was present: hospitalization, surgery, dialysis, or residence in a long-term care facility in the past year; or central vascular catheter ≤2 days before culture. Otherwise, cases were considered CA. Sites submitted a convenience sample of clinical MRSA isolates for molecular typing and antimicrobial susceptibility testing. Molecular typing was performed by pulsed-field gel electrophoresis until 2008, when typing was inferred using a validated algorithm based on molecular characteristics. Reference broth microdilution was performed for 8 antimicrobials and interpreted based on CLSI interpretive criteria. We compared USA300 nonsusceptibility for HO and CA isolates. For antimicrobials with &gt;5% nonsusceptibility and for which HO isolates had greater nonsusceptibility than CA isolates, we compared nonsusceptibility for HACO and CA and analyzed annual trends in nonsusceptibility within each epidemiologic class (ie, CA, HACO, and HO) using linear regression. Results: Of 17,947 MRSA cases during 2005–2016, isolates were available for 6,685 (37%), and 2,120 were USA300 (34% CA, 52% HACO, 14% HO). HO isolates had more nonsusceptibility than CA isolates to gentamicin (2.2% vs 0.6%; P = .03), levofloxacin (47.8% vs 39.7%; P = .02), rifampin (3.7 vs 1.1%; P = .01), and trimethoprim-sulfamethoxazole (3.4% vs 0.6%; P = .04). HACO isolates also had more nonsusceptibility than CA isolates to levofloxacin (50.9% vs 39.7%; P &lt; .01). Levofloxacin nonsusceptibility increased during 2005–2016 for HACO and CA isolates (P &lt; .01), but not among HO isolates (P = .36) (Fig. 1). Conclusions: Overall, nonsusceptibility across drugs cannot distinguish USA300 isolates causing HO versus CA disease. Although HO isolates had higher levofloxacin nonsusceptibility than CA and HACO isolates early on, USA300 MRSA HACO isolates now have levofloxacin nonsusceptibility most similar to that of HO isolates. Further study could help to explore whether increases in fluoroquinolone nonsusceptibility among CA and HACO cases may be contributing to the persistence of USA300 strains.Disclosures: NoneFunding: None

Methicillin-Resistant Staphylococcus aureus (MRSA) Risk Factors: Comparison Between Acute-Care, and Subacute- and Long-Term Care Facilities in a Healthcare Network

Background: The risk factors for methicillin-resistant Staphylococcus aureus (MRSA) colonization can differ between acute-care, subacute-care, and long-term care facilities, but comparative information is lacking. We compared risk factors for MRSA colonization contemporaneously between an acute-care hospital (ACH) and its affiliated intermediate- and long-term care facilities (ILTCFs). Methods: Serial cross-sectional studies were conducted in a 1,600-bed tertiary-care ACH and its 6 affiliated ILTCFs in Singapore, in June–July 2014–2016. Separate nasal, axillary, and groin swabs were taken and cultured for MRSA. MRSA isolates were subject to whole-genome sequencing. Clinical and epidemiological data were obtained from medical records. To account for clustering, multivariable 2-level multinomial logistic regression models were constructed to assess factors associated with colonization of specific MRSA clones, in the ACH and ILTCFs, respectively. Results: In total, 8,873 samples from 2,985 patients in the ACH and 7,172 samples from 2,409 patients and residents in ILTCFs were included in the study. Patients and residents in the ILTCFs (29.7%) were more likely to be colonized with MRSA than patients in the ACH (12.6%) (P &lt; .0001). The predominant MRSA clones were clonal complexes (CC)22 (n = 692, 46.7%) and CC45 (n = 494, 33.4%), contributing to 80% of MRSA isolates. For ACH patients, after adjusting for age, gender, comorbidities, prior exposures to antibiotics and percutaneous devices, presence of wounds, and screening year, prior MRSA carriage in the preceding 12 months was the strongest predictor of colonization with all MRSA clones: CC22 (aOR, 14.71; 95% CI, 6.17–34.48); CC45 (aOR, 7.75; 95% CI, 2.70–22.22); and others (aOR, 22.22; 95% CI, 3.83–125.00). Hospital stay &gt;14 days was also positively associated with colonization with MRSA CC22 (aOR, 2.67; 95% CI, 1.22–5.88), but not the other clones. For ILTCF patients and residents, after adjusting for age, comorbidities, prior exposure to antibiotics, presence of wounds, and screening year, prior MRSA carriage was a significant predictor of colonization with MRSA CC22 (aOR, 2.72; 95% CI, 1.35–5.46), and CC45 (aOR, 2.36; 95 % CI, 1.06–5.24), but not with other clones. Additionally, prior exposure to a percutaneous device and being male were respectively positively associated with colonization by MRSA CC22 (aOR, 2.70; 95% CI, 1.19–6.17) and CC45 (aOR, 2.17; 95% CI, 1.11–4.26). Conclusions: Prior MRSA carriage was a common risk factor for colonization with the predominant MRSA clones in both the ACH and ILTCFs. Hospital stay &gt;14 days and exposure to percutaneous devices were additional risk factors for CC22 colonization in the ACH and ILTCFs, respectively. Pre-emptive contact precautions for prior MRSA-carriers on admission and active screening for long-stayers in the ACH could prevent intra- and interinstitutional MRSA transmission.Funding: NoneDisclosures: None