CA-MRSA Strains Research Articles

Community-acquired Methicillin-resistant Staphylococcus Aureus Pneumonia: A Diagnostic Challenge

Even though methicillin-resistant Staphylococcus aureus (MRSA) is considered a source of nosocomial pneumonia, a few cases have likewise depicted instances of community-acquired pneumonia (CAP) among healthy individuals that were caused by MRSA. Mostly CA-MRSA causes skin and soft tissue infection but can be responsible for severe invasive infection with higher morbidity. Invasive CA-MRSA requires aggressive management to prevent complications. CA-MRSA strains are now recognized as distinct clonal entities that differ from traditional hospital-acquired MRSA (HA-MRSA) strains. The increasing prevalence of MRSA in the community is an important public health problem, which needs increased vigilance in the diagnosis and management of suspected and confirmed staphylococcal infections. We present an intense case of CAP brought about by MRSA with impending respiratory failure in a 19-year-old male, who presented with acute febrile illness with swelling over lips with no co-morbid and insignificant past medical history. Through this case, we intend to increase awareness among physicians about the growing role of CA-MRSA in the population; they should be meticulous in treating patients with CA-MRSA, and be aware of the complications of this disease and the appropriate empiric and target antibiotics regimen for such infections, whenever it is suspected, to enhance better clinical outcomes.

Phenotypic and genomic analysis of the hypervirulent ST22 methicillin-resistant Staphylococcus aureus in China.

ST22 MRSA (methicillin-resistant Staphylococcus aureus) strains are only sporadically reported in China. Through the phylogenetic reconstruction of 30 ST22 strains from China and 480 ST22 strains from global sources, we found that the global ST22 strains can be divided into three clades (I, II, and III). The China ST22 strains were found primarily in clade II (IIb and IIc) and also in clade III, indicating that the China ST22-MRSA clones have different origins. The China subclade IIb strains (SCCmec Vb-t309) may evolve from the native ST22 MSSA clone, while the China IIc strains may have spread from other countries. Subclade IIc (SCCmecIVa-t309) strains exhibited particularly strong lethality and invasiveness in Galleria mellonella infection and mouse skin abscess models in comparison to USA300 and other dominant China HA-MRSA (ST5 and ST239) or CA-MRSA (ST59) strains. This study described the emergence of a highly virulent ST22 MRSA subclade and improved our insight into the molecular epidemiology of ST22 strains in China. IMPORTANCE ST22 is a successful hospital-associated MRSA lineage which first appeared in the United Kingdom as EMRSA-15. At present, ST22 MRSA clones are spreading rapidly around the world and even replaced other dominant clones in some regions. We placed the Chinese ST22 in the worldwide phylogeny of ST22, demonstrating a distinctive molecular epidemiology and to our knowledge, this is the first time that a novel clade of ST22 has been found in China. Among the 15 ST22 MRSA strains belonging to the novel clade, 14 ST22 SCCmecIVa strains from different regions carried both pvl and tst and displayed significantly higher in vitro and in vivo virulence in comparison to other clade/subclade ST22 strains as well as other common China HA-MRSA or CA-MRSA strains. The further spread of this subclade of strains could pose a serious threat to the health system in China and other regions.

Prevalence and clinical characteristics of methicillin-resistant Staphylococcus aureus infections among dermatology inpatients: A 7-year retrospective study at a tertiary care center in southwest China.

Increased rates of methicillin-resistant Staphylococcus aureus (MRSA) antibiotic resistance and the associated morbidity have increased dermatologists' attention to skin and soft tissue MRSA infections. However, the clinical characterization of MRSA skin and soft tissue infections (SSTIs) in Southwest China is lacking, which precludes optimal prevention and treatment of these infections. This study was conducted to characterize the prevalence, clinical comorbidities and antibiotic susceptibility of MRSA isolates from SSTIs, including community-associated (CA) and healthcare-associated (HA) isolates. In the Dermatology Inpatient Department of the First Affiliated Hospital of Guangxi Medical University, a retrospective study was conducted on data, including patient demographics and clinical information, from culture-confirmed S. aureus isolated from skin and soft tissue between January 1, 2015, and December 31, 2021. Isolate susceptibility to 13 antibiotics was determined using the Vitek 2 system. From among 864 S. aureus strains, we identified 283 MRSA (32.75%) isolates comprising 203 CA-MRSA and 80 HA-MRSA isolates. The average rate of CA-MRSA isolation for MRSA SSTIs was 71.73%. The HA-MRSA isolation rate for MRSA SSTIs increased significantly. HA-MRSA-infected patients were older. The most common dermatological presentation of CA-MRSA infection was staphylococcal scalded skin syndrome, while the comorbidity severe drug eruption was significantly associated with HA-MRSA infection. One CA-MRSA strain was resistant to linezolid, and one HA-MRSA strain had an intermediate phenotype for vancomycin; both strains had low sensitivity to clindamycin and erythromycin (3.70%~19.40%). However, HA-MRSA isolates were more susceptible to trimethoprim/sulfamethoxazole. CA-MRSA is a predominant pathogen causing SSTIs, and HA-MRSA infection incidence is increasing gradually. Both strains showed increasing antibiotic resistance. Our data on MRSA susceptibility may guide dermatologist antibiotic treatment decisions. Dermatologists should consider these identified comorbidities of MRSA SSTIs when patients are admitted and initiate early prevention and treatment of MRSA.

Molecular Mechanisms of Drug Resistance in Staphylococcus aureus.

This paper discusses the mechanisms of S. aureus drug resistance including: (1) introduction. (2) resistance to beta-lactam antibiotics, with particular emphasis on the mec genes found in the Staphylococcaceae family, the structure and occurrence of SCCmec cassettes, as well as differences in the presence of some virulence genes and its expression in major epidemiological types and clones of HA-MRSA, CA-MRSA, and LA-MRSA strains. Other mechanisms of resistance to beta-lactam antibiotics will also be discussed, such as mutations in the gdpP gene, BORSA or MODSA phenotypes, as well as resistance to ceftobiprole and ceftaroline. (3) Resistance to glycopeptides (VRSA, VISA, hVISA strains, vancomycin tolerance). (4) Resistance to oxazolidinones (mutational and enzymatic resistance to linezolid). (5) Resistance to MLS-B (macrolides, lincosamides, ketolides, and streptogramin B). (6) Aminoglycosides and spectinomicin, including resistance genes, their regulation and localization (plasmids, transposons, class I integrons, SCCmec), and types and spectrum of enzymes that inactivate aminoglycosides. (7). Fluoroquinolones (8) Tetracyclines, including the mechanisms of active protection of the drug target site and active efflux of the drug from the bacterial cell. (9) Mupirocin. (10) Fusidic acid. (11) Daptomycin. (12) Resistance to other antibiotics and chemioterapeutics (e.g., streptogramins A, quinupristin/dalfopristin, chloramphenicol, rifampicin, fosfomycin, trimethoprim) (13) Molecular epidemiology of MRSA.

The thematic role of extracellular loop of VraG in activation of the membrane sensor GraS in a cystic fibrosis MRSA strain differs in nuance from the CA-MRSA strain JE2.

Patients with cystic fibrosis (CF) often suffer recurrent bronchial bacterial infections that lead to deterioration of lung function over time. The infections in CF patients are often due to S. aureus and P. aeruginosa that colonize the airways. Significantly, methicillin-resistant S. aureus (MRSA) makes it challenging for treatment in CF patients due to its feature of multiple antibiotic resistance. In bronchial airways, cationic antimicrobial peptides are often present in mucosa cells, neutrophils, and macrophages that interfere with bacterial proliferation. The major mechanism for resistance to the bactericidal activity of cationic peptides in S. aureus is mediated by the GraRS two-component system that activates expression of MprF and DltABCD to increase surface positive charge to repel interactions with cationic peptides. We recently found that VraG, a membrane permease component of the VraFG efflux pumps, harbors a long 200-residue extracellular loop (EL) that utilizes K380 to interact with the negatively charged 9-residue extracellular loop of the membrane sensor GraS to control mprF expression in a community-acquired MRSA strain JE2. In this study, we extended this observation to a CF MRSA strain CF32A1 where we affirmed that the EL loop of VraG controls GraS-mediated signal transduction; however, in contrast to community acquired MRSA strain JE2, the CF MRSA strain CF32A1 requires both K380 and K388 in the EL of VraG to properly modulate signal transduction mediated by GraS. This effect was not attributable to the several single nucleotide polymorphisms that exist between VraG and GraS in the two MRSA strains.

Staphylococcus aureus Isolated from the Oral Cavity: Phage Susceptibility in Relation to Antibiotic Resistance.

Nowadays, research on bacteriophage therapy and its potential use in combination with antibiotics has been gaining momentum. One hundred and ten oral Staphylococcus aureus isolates were phage-typed and their antibiotic resistance was determined by standard and molecular methods. The prevalence of MSSA and MRSA strains was 89.1% and 10.9%, respectively. Nearly all (91.8%) analyzed isolates, whether MSSA or MRSA, were susceptible to the phages used from the international set. The highest lytic activity showed phages 79 and 52 A from lytic group I. The predominant phage groups were mixed, the I+III group and a mixed group containing phages from at least three various lytic groups. S. aureus strains sensitive to phage group I were usually resistant to penicillin and susceptible to ciprofloxacin, whereas the strains typeable with group V or group V with the 95 phage were susceptible to most antibiotics. Epidemic CA-MRSA strains (SCCmecIV) of phage type 80/81 carried Panton–Valentine leucocidin genes. Considering the high sensitivity of oral S. aureus to the analyzed phages and the promising results of phage therapies reported by other authors, phage cocktails or phage-antibiotic combinations may potentially find applications in both the prevention and eradication of staphylococcal infections.

Community-acquired methicillin-resistant Staphylococcus aureus provoked cytokine storm causing severe infection on BALB/c mice

Methicillin-resistant Staphylococcus aureus (MRSA) has become the most important pathogen of hospital-acquired (HA) or community-acquired (CA) infections. However, it is unclear of the cytokines responsible for pathological hyper-inflammation in sepsis related cytokine storm for MRSA infection. In this study, we selected typical HA-MRSA strain (YNSA163: ST239-t030-SCCmecⅢ) and two CA-MRSA isolates (YNSA7: ST59-t439-SCCmecⅣa and YNSA53: ST59-t437-SCCmecⅤb) from our previous research, infected on BALB/c mice, and analyzed the cytokine storm patterns during infection process. The animal experiments revealed the most serious lethal effect on BALB/c mice caused by YNSA7 strain infection, followed by YNSA53, and no BALB/c mice died for YNSA163 infection. Histopathological analyses revealed that lung was the most seriously damaged organs, followed by spleen and kidney, especially for CA-MRSA infection. The severe inflammatory reactions, tissue destruction, and massive exudation of inflammatory mediators and cells could be identified in CA-MRSA strains infected mice. Interleukin-6 (IL-6) and IL-10 were both highly expressed in spleen and lung of YNSA7 and YNSA53 dead cases compared with YNSA53 survived and YNSA163 cases, which demonstrated cytokine storm pattern for CA-MRSA strains infection. The results of IL-6 intervention experiment verified that the enhanced IL-6 secretion was responsible for the host lethality of YNSA7 infection. RNA-sequencing results among three MRSA isolates indicated most of the differentially expressed genes referred to cellular process, metabolism and genetic information processing of bacteria. Specifically, clpP, chp chemotaxis inhibit, fnbB, pathogencity island protein and virulence associated protein E were highly expressed in YNSA7 strain. In general, CA-MRSA strains provoked cytokine storm on BALB/c mice led to severe infection and lethality, the up-regulated of some virulence genes might play important role in pathogenesis.

The Role of ArlRS and VraSR in Regulating Ceftaroline Hypersusceptibility in Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus infections are a global health problem. New control strategies, including fifth-generation cephalosporins such as ceftaroline, have been developed, however rare sporadic resistance has been reported. Our study aimed to determine whether disruption of two-component environmental signal systems detectably led to enhanced susceptibility to ceftaroline in S. aureus CA-MRSA strain MW2 at sub-MIC concentrations where cells normally continue to grow. A collection of sequential mutants in all fifteen S. aureus non-essential two-component systems (TCS) was first screened for ceftaroline sub-MIC susceptibility, using the spot population analysis profile method. We discovered a role for both ArlRS and VraSR TCS as determinants responsible for MW2 survival in the presence of sub-MIC ceftaroline. Subsequent analysis showed that dual disruption of both arlRS and vraSR resulted in a very strong ceftaroline hypersensitivity phenotype. Genetic complementation analysis confirmed these results and further revealed that arlRS and vraSR likely regulate some common pathway(s) yet to be determined. Our study shows that S. aureus uses particular TCS environmental sensing systems for this type of defense and illustrates the proof of principle that if these TCS were inhibited, the efficacy of certain antibiotics might be considerably enhanced.

Ten-Year Prevalence Trends of Phenotypically Identified Community-Associated Methicillin-Resistant Staphylococcus aureus Strains in Clinical Specimens.

BackgroundCommunity-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strains were first detected in hospitals in Korea between the late 2000s and early 2010s. However, there is limited information regarding the prevalence of CA-MRSA strains among hospital isolates and their phenotypic changes over the last decade. We investigated the prevalence trend of CA-MRSA strains isolated from different clinical specimens and their phenotypic changes between September 2009 and September 2019.MethodsCA-MRSA strains were phenotypically identified by confirming their resistance to penicillin (PCN) and oxacillin (OXA) and evaluating their susceptibility to trimethoprim-sulfamethoxazole, rifampin, fusidic acid, tetracycline, and at least one of the following four antimicrobials: clindamycin (CLI), erythromycin (ERY), ciprofloxacin (CIP), and gentamicin (GEN). A CA-MRSA strain that exhibited resistance to ERY, CLI, CIP, or GEN was classified as having resistance pattern I, II, III, or IV, respectively, regardless of its resistance to other antimicrobial agents.ResultsOf the 8,278 MRSA isolates identified in specimens obtained two days after admission, 1,385 (16.73%) were CA-MRSA strains. The prevalence of CA-MRSA strains increased from 12.2% to 26.6% (3.21% per period, P=0.05). Resistance type analysis revealed an increasing trend in the prevalence of PCN/OXA-resistant (1.84%; P=0.049) and PCN/OXA/ERY/CLI/CIP-resistant (0.98%; P=0.04) CA-MRSA strains and in resistance pattern III strains (2.08%; P=0.004).ConclusionsThe prevalence of CA-MRSA strains in Korea has increased significantly over the last decade, and CA-MRSA strains have gained phenotypic diversity beyond PCN/OXA-resistance, including antimicrobial resistance to non-β-lactams, especially CIP.

Clonal diversity and genomic characterization of Panton-valentine Leukocidin (PVL)-positive Staphylococcus aureus in Tehran, Iran

BackgroundSome Staphylococcus aureus strains produce Panton-Valentine leukocidin (PVL), a bi-component pore-forming toxin, which causes leukocyte lysis and tissue necrosis. Currently, there is very limited information on the molecular epidemiology of PVL-encoding S. aureus strains in Iran. This study aimed to determine the molecular epidemiology and genetic background of PVL-positive S. aureus clinical strains isolated from Iranian patients.MethodsA total of 28 PVL-positive S. aureus strains were detected from 600 S. aureus isolates between February 2015 and March 2018 from different hospitals in Tehran, Iran. Antimicrobial susceptibility testing was performed according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. Molecular genotyping was performed using SCCmec and accessory gene regulator (agr) typing, PVL haplotyping, multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE).ResultsThe highest antibiotic resistance rate was found to be against erythromycin (57.1%), followed by ciprofloxacin (42.8%) and clindamycin (35.7%). Moreover, 19 (67.9%) out of 28 S. aureus isolates were identified as MRSA, including CA-MRSA (14/19, 73.7%) and HA-MRSA (5/19, 26.3%). SCCmec type IVa was detected as the predominant type (10/19, 52.6%), followed by type III (5/19, 26.3%) and type V (4/19, 21.1%). The agr type I was identified as the most common type (14/28, 50%), and H and R haplotype groups were observed at frequencies of 67.9 and 32.1%, respectively. Among H variants, the predominant variant was H2 (78/9%). The isolates encompassed 21 different sequence types (STs), including 16 new STs (ST5147 to ST5162). Based on eBURST analysis, the isolates were clustered into five CCs, including CC30, CC22, CC1, CC8, and CC5 (ST5160), and nine singletons. PFGE typing showed that 24 isolates were clustered into A (4 pulsotypes), B (9 pulsotypes), and C (11 pulsotypes) clusters.ConclusionsA high prevalence of PVL-positive CA-MRSA strains was detected in Iran. The majority of PVL-positive isolates were of H (mostly H2) variant, while R variant was harbored by 100% of PVL-positive MRSA strains. Also, CC8, CC22, and CC30 were identified as the dominant clones among PVL-encoding S. aureus strains. This study promotes a better understanding of the molecular epidemiology and evolution of PVL-positive S. aureus strains in Iran.

Phenotypic and Genotypic Characterization, and Detection of PVL Encoding Gene in Methicillin Resistant Staphylococcus Aureus Strains Isolated From Patients Admitted to a Tertiary Hospital In Kuantan, Malaysia

Introduction: Methicillin-resistant Staphylococcus aureus is globally a major public health threat. Resistance to methicillin originates from a modified protein (PBP2a) encoded by the mecA gene. The PVL gene as an important virulence factor increases the pathogenicity of MRSA. Epidemiology and characteristics of MRSA differ in different geographical regions. This study was conducted to characterize and determine the antibiotic susceptibility profile of MRSA strains isolated from patients in Hospital Tengku Ampuan Afzan, Pahang, Malaysia and to detect the presence of the mecA and PVL genes in the isolates. Materials and methods: In this study a total of 36 isolates of MRSA have been collected during a period of three months (1stFebruary –30thApril 2018). The susceptibility pattern of the isolates to ten different commonly used antibiotics were determined and the target genes were addressed by real-time PCR experiment. Results: Based on the identifying criteria, 44.4% of the isolates were CA-MRSA, and 55.5% were HA-MRSA. Resistance to oxacillin, cefoxitin and penicillin was 100%, gentamicin 88.8%, erythromycin 33.3%, tetracycline 77.7%, trimethoprim-sulfamethoxazole 61.1%, clindamycin 13.8%, chloramphenicol 11.1%, but no resistant strain of vancomycin was detected. Most of the isolates were resistant to more than three groups of antibiotics. Realtime PCR revealed that all the isolates were mecA positive and 4 isolates were PVL-positive. PVL-positive strains were CA-MRSA and susceptible to clindamycin. Conclusion: The study confirms multi-drug resistant MRSA in the study area, and shows that resistance to methicillin is mecA mediated. PVL carrier strains were present and related to CA-MRSA strains of the isolates.

4165 Mechanisms of Prophage-Mediated Virulence Driving Community-Acquired MRSA Contagion

OBJECTIVES/GOALS: We recently identified a CA-MRSA strain in Brooklyn, New York (USA300-BKV) causing an outbreak of severe skin infections in predominantly healthy children. The evolution of USA300-BKV included acquisition of a novel prophage, and our objective is to identify the prophage-encoded gene(s) and mechanism responsible for increased bacterial virulence. METHODS/STUDY POPULATION: We deleted candidate genes from a novel mosaic block of phage-encoded genes in USA300-BKV that have been shown to enhance virulence in a murine skin infection model. Deletion mutants and complemented clones will be evaluated in vivo to identify culprit genes and determine the effect of lineage-specific genetic variation on the phenotype. Complementary studies include a comprehensive characterization of phage and bacterial genes expressed during lysogeny in vitro using RNA sequencing (RNA-Seq), and in vivo using a targeted approach focusing on known bacterial virulence and phage lytic pathways as well as candidate genes identified by in vitro studies. RESULTS/ANTICIPATED RESULTS: Comparison of otherwise isogenic lab strains showed that the mosaic block of phage genes present in USA300-BKV enhance skin abscess size in mice, confirming previous results. As this region of the phage, named mΦ11, does not contain known toxin genes, we hypothesize that mΦ11 modulates expression of bacterial host genes to enhance virulence. Thus, transcriptional profiles of CA-MRSA containing mΦ11 and selected deletion mutants are expected to reveal changes in known or novel virulence factors compared to controls. Candidate regulators specific to the mosaic block include an adenine methyltransferase linked to changes in global gene expression of other bacterial species. DISCUSSION/SIGNIFICANCE OF IMPACT: Our results will broaden scientific understanding of phage-bacterial interactions and determine the mechanisms by which phage impact virulence independent from toxin gene carriage. Identification of phage-encoded gene(s) enhancing CA-MRSA contagion will inform surveillance efforts and identify novel therapeutic targets.

Do trabecular metal and cancellous titanium implants reduce the risk of late haematogenous infection? An experimental study in rabbits.

This study evaluated the late resistance to haematogenous contamination by microbial pathogens of implants and bone-implant interface and the development of late clinical infection when cementless components with different surface or structural properties are implanted. 50 adult male New Zealand white rabbits were divided into 5 groups of 10 animals each. In Group A smooth titanium, in Group B grit blasted titanium, in Group C HA-coated titanium, in Group D trabecular metal and in group E cancellous titanium rods were implanted in the right proximal tibia. Four weeks later, 1 ml of inoculum of a standardised CA-MRSA strain (3 × 108 cfu/ml) was injected through a femoral artery catheter (groups B, C, D, E) while in group A, 1 ml of sterile saline was injected in a similar way (control group). Subjects were killed 8 weeks after the initial procedure and 3 samples of each tibial specimen were subjected to conventional cultures and PCR studies. The number of the specimens (conventional cultures and PCR studies) contaminated by the standardized pathogen was as follows: Group A: 0/10, Group B: 7/10, Group C: 6/10, Group D; 5/10 and Group E: 5/10. Comparing the number of colony form units isolated from the implant samples, Group B (GB titanium) showed statistically significantly higher values (Mann-Whitney test) compared to Group C (p = 0.044), Group D (p = 0.040) and Group E (p = 0.038). Local active infection was observed in 6 animals: 3 in Group B; 1 in Group C, 1 in Group D, and 1 in Group E. Modern cementless implants (trabecular metal and cancellous titanium) showed a lower risk of implant contamination and late clinical haematogenous infection.

Studying Staphylococcal Leukocidins: A Challenging Endeavor.

Staphylococcus aureus is a well-known colonizer of the human skin and nose, but also a human pathogen that causes a wide spectrum of diseases. It is well established that S. aureus secretes an arsenal of virulence factors that have evolved to circumvent the human immune system. A major group of S. aureus virulence factors is the bi-component β-barrel pore-forming toxins, also known as leukocidins. These pore-forming toxins target specific cells of the innate and adaptive immune system by interacting with specific receptors expressed on the cell membrane. Even though still heavily debated, clinical and epidemiological studies suggest the involvement of one of the bi-component toxin, Panton-Valentine Leukocidin (PVL), as an important factor contributing to the epidemic spread and increased virulence of CA-MRSA strains. However, the host- and cell-specificity of PVL and other leukocidins, and the lack of adequate in vivo models, fuels the controversy and impairs the appropriate assessment of their role in S. aureus pathophysiology. Currently, the mechanisms of pore-formation and the contribution of PVL and other leukocidins to S. aureus pathophysiology are incompletely understood. This review summarizes our current understanding of leukocidin pore-formation, knowledge gaps, and highlights recent findings identifying novel host-factors involved in the toxin-host interface. As a result, this review furthers emphasizes the complexity behind S. aureus leukocidin cytotoxicity and the challenges associated in the quest to study and understand these major virulence factors.

Skin and Soft Tissue Infections Caused by Different Genotypes of PVL-Positive Community-Acquired Methicillin-Resistant Staphylococcus aureus Strains.

Panton-Valentine leukocidin (PVL) is a causative agent of lethal necrotizing pneumonia and is associated with epidemic strains of community-acquired (CA) methicillin-resistant Staphylococcus aureus (MRSA). PVL-producing strains have rarely been isolated in Japan. However, PVL-positive CA-MRSA has been isolated much more frequently in recent years. To investigate the relevance of pvl genes (lukS/F-PV) and clinical traits in epidemic S. aureus strains, we genotyped four PVL-positive CA-MRSA strains isolated from patients with skin and soft tissue infections and measured their susceptibility to antibiotics. Three of the isolates matched the genotype of the USA300 clone, which has predominantly been isolated in the USA. The remaining strain matched the ST217 genotype, and its spa type was identical to that of PVL-positive strains previously reported in India and China. Abscess drainage was necessary in all cases, and deep cutaneous ulcers were formed in three out of four cases regardless of the genotype. The ST217 genotype strain was resistant to clindamycin, in addition to quinolones, macrolides, and aminoglycosides. Thus, diagnostic determination of lukS/F-PV should be used as a guide for selecting the treatment regimen.

204. Antagonistic Effect of Colistin on Vancomycin Activity Against Methicillin-Resistant Staphylococcus aureus in in vitro and in vivo Studies

BackgroundThere is a concern that the vancomycin MIC of methicillin-resistant Staphylococcus aureus (MRSA) could be increased by concomitant colistin administered against multidrug-resistant gram-negative pathogen.MethodsWe confirmed the molecular genotypes of MRSA blood isolates collected in a tertiary hospital in Seoul, South Korea, and selected representative strains from the community-associated MRSA strains (CA-MRSA, ST72-SCCmec IV) and hospital-acquired MRSA strains (HA-MRSA, ST5-SCCmec II). USA CA-MRSA (USA300, ST8-SCCmec IV) and MRSA standard strain (ATCC 43300, ST39-SCCmec II) were also used for comparison with representative. We identified changes of the vancomycin MIC in MRSA by colistin exposure in a checkerboard assay and performed a time-kill assay to evaluate the combined effect of vancomycin and colistin on MRSA. In addition, we administered vancomycin, colistin, and combination of two antibiotics, respectively, to a neutropenic murine thigh infection model to evaluate the in vivo antagonistic effect of colistin on vancomycin treatment.ResultsIn the checkerboard assay, all 4 MRSA strains showed a tendency for the vancomycin MIC to increase along with increasing concentrations of colistin. However, the time-kill assay showed the antagonism of vancomycin and colistin only against ST5-MRSA, when vancomycin concentration was 2 times the vancomycin MIC (Figure 1). No antagonism was observed in other strains. In the murine thigh infection model of ST5-MRSA, vancomycin monotherapy showed a significant log CFU reduction compared with a combination of vancomycin and colistin at 24 hours, demonstrating the antagonistic effect of vancomycin and colistin combination (Figure 2).ConclusionThis study showed that exposure of colistin to certain MRSA strains may reduce the susceptibility to vancomycin. Combination therapy with vancomycin and colistin for MDR pathogens infections might result in treatment failure for concurrent MRSA infection. Disclosures All authors: No reported disclosures.

Evolution of a 72-Kilobase Cointegrant, Conjugative Multiresistance Plasmid in Community-Associated Methicillin-Resistant Staphylococcus aureus Isolates from the Early 1990s.

Horizontal transfer of plasmids encoding antimicrobial resistance and virulence determinants has been instrumental in Staphylococcus aureus evolution, including the emergence of community-associated methicillin-resistant S. aureus (CA-MRSA). In the early 1990s, the first CA-MRSA strain isolated in Western Australia (WA), WA-5, encoded cadmium, tetracycline, and penicillin resistance genes on plasmid pWBG753 (∼30 kb). WA-5 and pWBG753 appeared only briefly in WA; however, fusidic acid resistance plasmids related to pWBG753 were also present in the first European CA-MRSA isolates at the time. Here, we characterize a 72-kb conjugative plasmid, pWBG731, present in multiresistant WA-5-like clones from the same period. pWBG731 was a cointegrant formed from pWBG753 and a pWBG749 family conjugative plasmid. pWBG731 carried mupirocin, trimethoprim, cadmium, and penicillin resistance genes. The stepwise evolution of pWBG731 likely occurred through the combined actions of IS257, IS257-dependent miniature inverted-repeat transposable elements (MITEs), and the BinL resolution system of the β-lactamase transposon Tn552 An evolutionarily intermediate ∼42-kb nonconjugative plasmid, pWBG715, possessed the same resistance genes as pWBG731 but retained an integrated copy of the small tetracycline resistance plasmid pT181. IS257 likely facilitated the replacement of pT181 with conjugation genes on pWBG731, thus enabling autonomous transfer. Like conjugative plasmid pWBG749, pWBG731 also mobilized nonconjugative plasmids carrying oriT mimics. It seems likely that pWBG731 represents the product of multiple recombination events between the WA-5 pWBG753 plasmid and other mobile genetic elements present in indigenous community-associated methicillin-sensitive S. aureus (CA-MSSA) isolates. The molecular evolution of pWBG731 saliently illustrates how diverse mobile genetic elements can together facilitate rapid accrual and horizontal dissemination of multiresistance in S. aureus CA-MRSA.

Evolution of a major bovine mastitic genotype (rpoB sequence type 10-2) of Staphylococcus aureus in cows.

Staphylococcus aureus is the major pathogen leading to bovine mastitis globally while livestock-associated methicillin resistant S. aureus (LA-MRSA) has become a potential threat to public health. MRSA from bovine mastitis is not common but a methicillin susceptible S. aureus (MSSA) genotype, rpoB sequence type (RST)10-2 (RST10-2), is prevalent in Korea. To date, many genomic sequences from S. aureus have been elucidated, but the complete genome sequences of RST10-2 MSSA from bovine mastitis has never been reported. In this study, we determined the complete genome sequence of two RST10-2 MSSA that differ from each other in staphylococcal protein A and molecular prophage types [PMB64-1 (t2489/ mPPT0) and PMB81-4 (t127/mPPT1-2-3)] and conducted a comparative genomics study. The genomic sequences of PMB64-1 and PMB81-4 were more homologous to the representative human RST10-2 strains (MSSA476, MW2 etc.) compared to other RSTs. Most of them shared five common pseudogenes, along with high amino acid identity of four variable virulence genes that were identified in this study. However, PMB64-1 and PMB81-4 acquired different strainspecific pseudogenes and mobile genetic elements than the human strains. The unique pseudogene profile and high identity of the virulence genes were verified in RST10-2 field strains from bovine mastitis. Thus, bovine mastitic RST10-2 MSSA may have an evolutionary relationship with the human RST10-2 community-associated (CA) MSSA and CA-MRSA strains but may have adapted to cows.

Methicillin-sensitive and methicillin-resistant Staphylococcus aureus strains and their toxin genes in the nostrils of dogs and workers at an animal shelter.

Methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MSSA and MRSA, respectively) in the nostrils of dogs and workers at an animal shelter were cultured. Staphylococcal toxin genes were analysed to identify potential health concerns. Samples were obtained from 441 dogs and 9 workers. The respective isolation rates of S. aureus and MRSA were 49·0% (216/441) and 1·6% (7/441) for shelter dogs and 44·4% (4/9) and 33·3% (3/9) for workers, respectively. Isolation of S. aureus in summer (61·9%) and in adult dogs (59·2%) were significantly higher than those in winter (35·8%) and in juvenile dogs (33·3%) (P<0·001), respectively. The predominant enterotoxin genotypes and combination profiles of S. aureus were (sea, seb, seg, sei, sem, sen, seo, seu) and (sea, sea-seb, and seg-sei-sem-sen-seo-seu), respectively, and 20% of isolates carried food poisoning-associated enterotoxins. The se profiles in shelter dogs were different from those in general pet dogs and their owners. MRSA isolates were identified as SCCmec IV and VII, and they shared se combination profiles of (sec-seg-sei-sel-sem-sen-seo-seu) and (seb-sek-seq). MRSA in this shelter had similar microbiological characteristics as those reported in CA-MRSA ST59 in humans. Human health-associated bacteria and food poisoning-related toxin genes were identified. Further evaluations of health concerns in animal shelters are necessary. This is the first study to focus on se prevalence and MRSA characteristics in an animal shelter in Taiwan. The MRSA characteristics determined in this study were similar to those of CA-MRSA strains isolated from communities in the past, indicating potential health risks in cities.

What is behind phylogenetic analysis of hospital-, community- and livestock-associated methicillin-resistant Staphylococcus aureus?

Methicillin-resistant Staphylococcus aureus (MRSA) has been shown to be the predominant life-threatening pathogen in Egypt. MRSA is a major cause of severe healthcare-associated (HA) infections. During the last decades, the incidence of community-associated (CA) MRSA infections has a complex epidemiology arising from the circulation of different strains in the general population. Moreover, livestock-associated (LA) MRSA emerged recently becomes an emerging threat to public health. Therefore, it is important to illuminate the differences between CA-, HA- and LA-MRSA to shed light on their genetic diversity and evolution. This study presents the first data on analysing the correlation between CA-, LA- and HA-MRSA using antibiogram typing, molecular characteristics and antimicrobial resistance and virulence genes' profiles. Overall, HA-MRSA strains tended to be multidrug resistant and less virulent than both LA- and CA-MRSA strains. Importantly, CA-MRSA strains had a high homology with each of HA- and LA-MRSA. However, no similarity was observed between HA- and LA-MRSA. Our findings suggest that the epidemiological changes in genetic behaviour between HA- and LA-MRSA are due to the presence of CA-MRSA confirming that CA-MRSA has created a public health crisis worldwide.