Community-associated Strains Research Articles

Clonal Diversity, Antibiotic Resistance, and Virulence Factor Prevalence of Community Associated Staphylococcus aureus in Southeastern Virginia.

Staphylococcus aureus is a significant human pathogen with a formidable propensity for antibiotic resistance. Worldwide, it is the leading cause of skin and soft tissue infections (SSTI), septic arthritis, osteomyelitis, and infective endocarditis originating from both community- and healthcare-associated settings. Although often grouped by methicillin resistance, both methicillin-resistant (MRSA) and methicillin-sensitive (MSSA) strains are known to cause significant pathologies and injuries. Virulence factors and growing resistance to antibiotics play major roles in the pathogenicity of community-associated strains. In our study, we examined the genetic variability and acquired antibiograms of 122 S. aureus clinical isolates from SSTI, blood, and urinary tract infections originating from pediatric patients within the southeast region of Virginia, USA. We identified a suite of clinically relevant virulence factors and evaluated their prevalence within these isolates. Five genes (clfA, spA, sbi, scpA, and vwb) with immune-evasive functions were identified in all isolates. MRSA isolates had a greater propensity to be resistant to more antibiotics as well as significantly more likely to carry several virulence factors compared to MSSA strains. Further, the carriage of various genes was found to vary significantly based on the infection type (SSTI, blood, urine).

Population structure of food-borne Staphylococcus aureus in China

Objective: To understand the population structure of food-borne Staphylococcus (S.) aureus in China. Methods: Whole genome sequencing was used to analyze 763 food-borne S. aureus strains from 16 provinces in China from 2006 to 2020. Multilocus sequence typing (MLST), staphylococcal protein A gene (spa) typing, and staphylococcal chromosome cassettemec (SCCmec) typing were conducted, and minimum spanning tree based on ST types (STs) was constructed by BioNumerics 7.5 software. Thirty-one S. aureus strains isolated from imported food products were also included in constructing the genome phylogenetic tree. Results: A total of 90 STs (20 novel types) and 160 spa types were detected in the 763 S. aureus isolates. The 72 STs (72/90, 80.0%) were related to 22 clone complexes. The predominant clone complexes were CC7, CC1, CC5, CC398, CC188, CC59, CC6, CC88, CC15, and CC25, accounting for 82.44% (629/763) of the total. The STs and spa types in the predominant clone complexes changed over the years. The methicillin-resistant S. aureus (MRSA) detection rate was 7.60%, and 7 SCCmec types were identified. The ST59-t437-Ⅳa (17.24%, 10/58), ST239-t030-Ⅲ (12.07%, 7/58), ST59-t437-Ⅴb (8.62%, 5/58), ST338-t437-Ⅴb (6.90%, 4/58) and ST338-t441-Ⅴb (6.90%, 4/58) were the main types in MRSA strains. The genome phylogenetic tree had two clades, and the strains with the same CC, ST, and spa types clustered together. All CC7 methicillin sensitive S. aureus strains were included in Clade1, while 21 clone complexes and all MRSA strains were in Clade2. The MRSA strains clustered according to the SCCmec and STs. The strains from imported food products in CC398, CC7, CC30, CC12, and CC188 had far distances from Chinese strains in the tree. Conclusions: In this study, the predominant clone complexes of food-borne strains were CC7, CC1, CC5, CC398, CC188, CC59, CC6, CC88, CC15, and CC25, which overlapped with the previously reported clone complexes of hospital and community-associated strains in China, suggesting that close attention needs to be paid to food, a vehicle of pathogen transmission in community and food poisoning.

Prevalence of Methicillin-Resistant and Methicillin Sensitive Staphylococcus aureus Nasal Carriage and their Antibiotic Resistant Patterns in Kirkuk City, Iraq

The carriage of community-acquired methicillin-resistant S. aureus (CA-MRSA) has become a source of community-associated infections, particularly in the anterior nares of people worldwide. This study was aimed at determining the prevalence of community-acquired S. aureus (CA-SA) and CA-MRSA nasal carriage among healthy individuals of various ages in Kirkuk City and evaluating the susceptibility of isolates to various antibiotics. A total of 597 healthy individuals were recruited in the community between December 1, 2021, and December 30, 2022. Nasal swabs obtained from participants were taken to the laboratory, where bacteria were isolated and identified using phenotypic characteristics. The MRSA isolates were identified by applying the modified Kirby Bauer disc diffusion technique. The results showed that the prevalence of CA-SA and CA-MRSA nasal carriers was 16.6 and 4.5%, respectively. The average age of the participants was 33.2 years, with a male-to-female ratio of 1.1: 1. The highest resistance of the isolates was observed against oxacillin (27.3%), followed by penicillin G (24.3%), amoxicillin (15.2%), erythromycin (12.1%), and tetracycline (6.1%). There was a 9.1% resistance rate to clindamycin, rifampin, gentamycin, and ciprofloxacin. However, all CA-MRSA isolates were multi-drug resistant. However, all the isolates were sensitive (100 %) to vancomycin, linezolid, and mupirocin. The findings of the present study highlight the potential for CA-SA and CA-MRSA acquisition in this population, which may be related to antibiotic abuse or overuse as well as poor hygiene. To lessen the impact of community-associated strains of MRSA nasal carriage, this necessitates the probable need for infection prevention measures and adequate antibiotic therapy.

Antimicrobial Effect of Syzygium cumini Extract Against Methicillin Non-Susceptible Staphylococcus aureus

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is a leading cause of infections in Community- Associated (CA) as well as Hospital-Associated (HA) settings. Identification of new antibacterial agents from natural sources takes the forefront in research. Objectives: The aim of the present study was to identify the resistance pattern of S. aureus in the clinical samples causing disease in Dhaka city, molecular typing of the methicillin non-susceptible S. aureus isolates and identifying new herbal components with anti-microbial effect against S. aureus. Methods: We screened total of 78 clinical specimen of various nature (pus, urine, tracheal aspirate, conjunctiva and wound swab) with confirm S. aureus infection between March 2018 to October 2018. The specimen were cultured on mannitol salt agar to isolate S. aurues, which were later tested for antibiotic resistance according to disc diffusion method. The MRSA isolates were confirmed with PCR and typed for SCCmec element to know the distribution of hospital-associated and community-associated strains. Finally, the MRSA isolates were cultured in the presence of leaf extract and fruit extract of Syzygium cumini for observing the antibacterial potential. Result: A total of 12 isolates of S. aureus were found to be non-susceptible to methicillin, 34%, 25%, 17% out of these were from pus, blood and urine respectively and 8% isolates were from wound swab, conjunctiva and tracheal aspirates each. Out of methicillin non-susceptible isolates, 25% and 16% were HA-MRSA and CAMRSA respectively, as seen from PCR analysis of the SCCmec gene cassette of the S. aureus genome. The rest of the 59% of the isolates were untypable. Overall, higher concentration of leaf and fruit extract reduced the optical density of MRSA culture and reduced bacterial growth in drop plate significantly. Conclusion: Dhaka population has S. aureus with varying sensitivity against methicillin, which needs further characterization by molecular epidemiology methods. Bangladesh Med Res Counc Bull 2021; 47(1): 82-89

Prevalence of Staphylococcus aureus and Staphylococcus sciuri Isolated from Apparently Healthy and Hospitalized Patients in Ekiti State, Nigeria

The emergence of microbial resistance to antibiotics on daily basis has become a major global challenge. The increasing prevalence of methicillin resistant Staphylococcus aureus (MRSA) infections among hospitalized and non-hospitalized patients due to emergence of unique community-associated strains has become a great problem in Nigeria in particular and the entire world in general. In this study, a total of four hundred (400) clinical specimens were collected from hospitalized patients at Ekiti State University Teaching Hospital (EKSUTH), Ado-Ekiti and Federal Teaching Hospital (FETHI), Ido-Ekiti and from apparently healthy individuals from both communities. Standard bacteriological procedures were employed to isolate and identify these organisms. Antibiotic susceptibility test was carried out using the modified Kirby Bauer disc diffusion method. Polymerase chain reaction (PCR) assay was used to identify S. aureus nuc gene, as well as mecA and aac resistance genes. Seventy seven (19.3%) S. aureus isolates were phenotypically identified; 43 (21.5%) hospital-associated (23 from EKSUTH and 20 from FETHI) and 34 (17.0%) community-associated (27 from Ado-Ekiti and 7 from Ido-Ekiti communities). No S. sciuri was phenotypically isolated in both locations. Fifty five (71.4 %) of the 77 S. aureus isolates were nuc gene positive, while 11 (20%) of the 55 isolates were mecA gene positive, implying that they were MRSA. Of the 11 isolates, 6 (19.4%) were hospital-associated MRSA while 5 (20.8 %) were community-associated MRSA. Phenotypic resistance of S. aureus to cefoxitin in the two locations ranged between 77.8 – 100%, but 80% of cefoxitin-resistant nuc gene positive S. aureus lacked mecA gene. S. aureus isolates exhibited high phenotypic resistance to tetracycline, erythromycin and fusidic acid but susceptible to clindamycin and gentamycin. The prevalence of clinical MRSA infection of 20% in this study is relatively high. However, there was 0% prevalence of S. sciuri colonization. Cefoxitin disc test demonstrated low specificity as a phenotypic marker of methicillin resistance.There is need to institute control measures for MRSA infections and colonization in this environment.

Drivers of methicillin-resistant Staphylococcus aureus (MRSA) lineage replacement in China

BackgroundMethicillin-resistant Staphylococcus aureus (MRSA) is a major nosocomial pathogen subdivided into lineages termed sequence types (STs). Since the 1950s, successive waves of STs have appeared and replaced previously dominant lineages. One such event has been occurring in China since 2013, with community-associated (CA-MRSA) strains including ST59 largely replacing the previously dominant healthcare-associated (HA-MRSA) ST239. We previously showed that ST59 isolates tend to have a competitive advantage in growth experiments against ST239. However, the underlying genomic and phenotypic drivers of this replacement event are unclear.MethodsHere, we investigated the replacement of ST239 using whole-genome sequencing data from 204 ST239 and ST59 isolates collected in Chinese hospitals between 1994 and 2016. We reconstructed the evolutionary history of each ST and considered two non-mutually exclusive hypotheses for ST59 replacing ST239: antimicrobial resistance (AMR) profile and/or ability to colonise and persist in the environment through biofilm formation. We also investigated the differences in cytolytic activity, linked to higher virulence, between STs. We performed an association study using the presence and absence of accessory virulence genes.ResultsST59 isolates carried fewer AMR genes than ST239 and showed no evidence of evolving towards higher AMR. Biofilm production was marginally higher in ST59 overall, though this effect was not consistent across sub-lineages so is unlikely to be a sole driver of replacement. Consistent with previous observations of higher virulence in CA-MRSA STs, we observed that ST59 isolates exhibit significantly higher cytolytic activity than ST239 isolates, despite carrying on average fewer putative virulence genes. Our association study identified the chemotaxis inhibitory protein (chp) as a strong candidate for involvement in the increased virulence potential of ST59. We experimentally validated the role of chp in increasing the virulence potential of ST59 by creating Δchp knockout mutants, confirming that ST59 can carry chp without a measurable impact on fitness.ConclusionsOur results suggest that the ongoing replacement of ST239 by ST59 in China is not associated to higher AMR carriage or biofilm production. However, the increase in ST59 prevalence is concerning since it is linked to a higher potential for virulence, aided by the carriage of the chp gene.

Clinical and Microbiological Characteristics of Hospital-Acquired Methicillin-Resistant Staphylococcus aureus Bacteremia Caused by a Community-Associated PVL-Negative Strain.

BackgroundST72-SCCmecIV, a community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strain in Korea, originated in the community and has been spreading in health care settings. Herein, we describe the clinical and microbiological characteristics of patients with hospital-acquired MRSA bacteremia (MRSAB) caused by community-associated strains.MethodsWe analyzed hospital-acquired MRSAB cases caused by ST72-SCCmecIV using a prospective cohort of patients with SAB in a tertiary hospital in Korea from July 2008 to December 2018. We compared the clinical and microbiological characteristics of ST72-SCCmecIV with ST5-SCCmecII, a representative hospital-associated genotype strain.ResultsOf the 1782 S. aureus bacteremia (SAB) cases, 628 (35.2%) were hospital-acquired MRSAB. Of the 628 isolates, 431 (68.6%) were ST5-SCCmecII and 152 (24.2%) were ST72-SCCmecIV. Patients with ST72-SCCmecIV were younger than those with ST5-SCCmecII and less likely to have a history of recent surgery, antibiotic treatment, nasal MRSA colonization, and central venous catheter placement. Compared with ST5-SCCmecII, ST72-SCCmecIV isolates were more likely to have vancomycin MICs ≤1.0 mg/L (P < .001). Osteoarticular infection as the site of infection (7.2% [11/152] vs 1.4% [6/431]) was more common in patients with ST72-SCCmecIV. There were no significant differences in the rate of recurrence (≤90 days), persistent bacteremia (≥7 days), or 30- and 90-day mortality rates between the 2 groups.ConclusionsOsteoarticular infections were more prevalent in ST72-SCCmecIV MRSAB. Mortality rates between the ST72-SCCmecIV and ST5-SCCmecII groups were not significantly different.

Nasal Methicillin-Resistant Staphylococcus aureus Colonization in Patients with Type 1 Diabetes in Taiwan.

Nasal methicillin-resistant Staphylococcus aureus (MRSA) colonies are an essential reservoir of infection, especially for patients with diabetes. However, data on MRSA colonization in patients with type 1 diabetes are limited. We investigated the epidemiology of MRSA colonization in patients with type 1 diabetes. This prospective cross-sectional study was conducted in a medical center (Chang Gung Memorial Hospital) in Taiwan from 1 July to 31 December 2020. Nasal sampling and MRSA detection were performed. The molecular characteristics of MRSA isolates were tested, and factors associated with MRSA colonization were analyzed. We included 245 patients with type 1 diabetes; nasal MRSA colonization was identified in 13 (5.3%) patients. All isolates belonged to community-associated MRSA genetic strains; the most frequent strain was clonal complex 45 (53.8%), followed by ST59 (30.8%) (a local community strain). MRSA colonization was positively associated with age ≤ 10 years, body mass index < 18 kg/m2, and diabetes duration < 10 years; moreover, it was negatively associated with serum low-density lipoprotein cholesterol ≥ 100 mg/dL. No independent factor was reported. The nasal MRSA colonization rate in type 1 diabetes is approximately 5% in Taiwan. Most of these colonizing strains are community strains, namely clonal complex 45 and ST59.

Clonal diversity and spatial dissemination of multi-antibiotics resistant Staphylococcus aureus pathotypes in Southwest Nigeria

Spread of genetically diverse Staphylococcus aureus characterized with multi-antibiotic resistance and regulated by high level agr functionalities in several communities in southwest Nigeria was investigated and evaluated for infection control. Staphylococcus aureus pathotypes recovered from 256 cases including purulent pus from skin infections, soft tissue aspirates, wounds, otorrhea, eye, throat and endocervical infections were assayed for biofilm and antibiogram. Further genotyped with micro-array, mapped for geospatial distribution and evaluated for clonal diversity and functional accessory gene regulators (agr). Significant Staphylococci infection among the ages (OR:0.021, CI:0.545–1.914) and female gender with prevalence rate of MSSA (53.0%) and MRSA (1.5%) (OR:1.021, CI:0.374–1.785) were observed. More than 52.5% resistance rates to tetracycline and amoxicillin with significant median resistance were observed in all the infection cases (p = 0.001). Resistance rate of 78.8% at MIC50 32μg/ml and MIC90 128μg/ml to amoxicillin-clavulanate, and more than 40% resistance to ceftazidime, ciprofloxacin and tetracycline of MIC90 and MIC50 at 32 μg/ml were observed. Strains with multi-antibiotic resistance index above 0.83, high beta-lactamase and strong biofilm clustered into separate phylo-group. Heterogeneous t442 (wound and pus), t657 (wound), t091 (ear) and t657 (ear and wound) revealed high phylogenetic diversity. Only 4.6% pvl+ MSSA-CC1 agrI, pvl+ MSSA-CC5 (13.6%) and pvl+ MRSA-CC7 agrII (4.6%), expressed enterotoxin, leukocidins, proteases and resistance gene determinants. Livestock clonal types clustered with identified community-associated strains. Clonal dissemination of resistant pvl+ MSSA-CC1 and MRSA-CC5 encoding agr were predominant in several peri-urban communities where adequate geno-surveillance, population-target antimicrobial stewardship, extensive community structured infection control programs are needed to prevent further focal dissemination.

Comparative genomic and transmission analysis of Clostridioides difficile between environmental, animal, and clinical sources in China

ABSTRACT Clostridioides difficile is the most common pathogen causing antibiotic-associated diarrhea. Previous studies showed that diverse sources, aside from C. difficile infection (CDI) patients, played a major role in C. difficile hospital transmission. This study aimed to investigate relationships and transmission potential of C. difficile strains from different sources. A prospective study was conducted both in the intensive care unit (ICU) and six livestock farms in China in 2018–2019. Ninety-eight strains from CDI patients (10 isolates), asymptomatic hospitalized carriers (55), the ICU environment (12), animals (14), soil (4), and farmers (3) were collected. Sequence type (ST) 3/ribotype (RT) 001, ST35/RT046, and ST48/RT596 were dominant types, distributed widely in multiple sources. Core-genome single-nucleotide polymorphism (cgSNP) analysis showed that hospital and farm strains shared several common clonal groups (CGs, strains separated by ≤ 2 cgSNPs) (CG4/ST3/RT001, CG7/ST35/RT046, CG11/ST48/RT596). CDI patients, asymptomatic carriers, and the ICU environment strains also shared several common CGs. The number of virulence genes was not statistically different between strains from different sources. Multi-source strains in the same CG carried identical virulence gene sequences, including pathogenicity genes at the pathogenicity locus and adhesion-related genes at S-layer cassette. Resistance genes (ermB, tetM, etc.) were widespread in multiple sources, and multi-source strains in the same CG had similar resistance phenotypes and carried consistent transposons and plasmid types. The study indicated that interspecies and cross-regional transmission of C. difficile occurs between animals, the environment, and humans. Community-associated strains from both farms and asymptomatic hospitalized carriers were important reservoirs of CDI in hospitals.

Prevalence, Colonization, Epidemiology, and Public Health Significance of Clostridioides difficile in Companion Animals.

Clostridioides difficile, previously Clostrdium difficile, is a major cause of antibiotic-associated enteric disease in humans in hospital settings. Increased incidence of C. difficile infection (CDI) in community settings raises concerns over an alternative source of CDI for humans. The detection of genetically similar and toxigenic C. difficile isolates in companion animals, including asymptomatic pets, suggests the potential role of household pets as a source of community-associated CDI. The close association between companion animals and humans, in addition to the use of similar antibiotics in both species, could provide a selective advantage for the emergence of new C. difficile strains and thus increase the incidental transmission of CDI to humans. Therefore, screening household pets for C. difficile is becoming increasingly important from a public health standpoint and may become a part of routine testing in the future, for the benefit of susceptible or infected individuals within a household. In this review, we analyze available information on prevalence, pathophysiology, epidemiology, and molecular genetics of C. difficile infection, focusing on companion animals and evaluate the risk of pet-borne transmission of CDI as an emerging public health concern. Molecular epidemiological characterization of companion animal C. difficile strains could provide further insights into the interspecies transmission of CDI. The mosaic nature of C. difficile genomes and their susceptibility to horizontal gene transfer may facilitate the inter-mixing of genetic material, which could increase the possibility of the emergence of new community-associated CDI strains. However, detailed genome-wide characterization and comparative genome analysis are warranted to confirm this hypothesis.

Staphylococcus aureus Alpha-Toxin Limits Type 1 While Fostering Type 3 Immune Responses.

Staphylococcus aureus can cause life-threatening diseases, and hospital- as well as community-associated antibiotic-resistant strains are an emerging global public health problem. Therefore, prophylactic vaccines or immune-based therapies are considered as alternative treatment opportunities. To develop such novel treatment approaches, a better understanding of the bacterial virulence and immune evasion mechanisms and their potential effects on immune-based therapies is essential. One important staphylococcal virulence factor is alpha-toxin, which is able to disrupt the epithelial barrier in order to establish infection. In addition, alpha-toxin has been reported to modulate other cell types including immune cells. Since CD4+ T cell-mediated immunity is required for protection against S. aureus infection, we were interested in the ability of alpha-toxin to directly modulate CD4+ T cells. To address this, murine naïve CD4+ T cells were differentiated in vitro into effector T cell subsets in the presence of alpha-toxin. Interestingly, alpha-toxin induced death of Th1-polarized cells, while cells polarized under Th17 conditions showed a high resistance toward increasing concentrations of this toxin. These effects could neither be explained by differential expression of the cellular alpha-toxin receptor ADAM10 nor by differential activation of caspases, but might result from an increased susceptibility of Th1 cells toward Ca2+-mediated activation-induced cell death. In accordance with the in vitro findings, an alpha-toxin-dependent decrease of Th1 and concomitant increase of Th17 cells was observed in vivo during S. aureus bacteremia. Interestingly, corresponding subsets of innate lymphoid cells and γδ T cells were similarly affected, suggesting a more general effect of alpha-toxin on the modulation of type 1 and type 3 immune responses. In conclusion, we have identified a novel alpha-toxin-dependent immunomodulatory strategy of S. aureus, which can directly act on CD4+ T cells and might be exploited for the development of novel immune-based therapeutic approaches to treat infections with antibiotic-resistant S. aureus strains.

Subinhibitory Concentrations of Mupirocin Stimulate Staphylococcus aureus Biofilm Formation by Upregulating cidA.

Previous studies have shown that the administration of antibiotics at subinhibitory concentrations stimulates biofilm formation by the majority of multidrug-resistant Staphylococcus aureus (MRSA) strains. Here, we investigated the effect of subinhibitory concentrations of mupirocin on biofilm formation by the community-associated (CA) mupirocin-sensitive MRSA strain USA300 and the highly mupirocin-resistant clinical S. aureus SA01 to SA05 isolates. We found that mupirocin increased the ability of MRSA cells to attach to surfaces and form biofilms. Confocal laser scanning microscopy (CLSM) demonstrated that mupirocin treatment promoted thicker biofilm formation, which also correlated with the production of extracellular DNA (eDNA). Furthermore, quantitative real-time PCR (RT-qPCR) results revealed that this effect was largely due to the involvement of holin-like and antiholin-like proteins (encoded by the cidA gene), which are responsible for modulating cell death and lysis during biofilm development. We found that cidA expression levels significantly increased by 6.05- to 35.52-fold (P < 0.01) after mupirocin administration. We generated a cidA-deficient mutant of the USA300 S. aureus strain. Exposure of the ΔcidA mutant to mupirocin did not result in thicker biofilm formation than that in the parent strain. We therefore hypothesize that the mupirocin-induced stimulation of S. aureus biofilm formation may involve the upregulation of cidA.

233. The Epidemiology, Genomics, and Evolution of Staphylococcus aureus in Northeast Ohio

BackgroundInfections due to S. aureus result in significant morbidity, mortality, and healthcare expense. We sought to identify the strains of S. aureus causing infections in hospitalized patients in Northeast Ohio and determine whether they are reflective of the S. aureus strains present in the surrounding environment.MethodsThe study was approved by the Institutional Review Board at Cleveland Clinic Akron General. Clinical S. aureus isolates (n = 300) were cultured and PCR was used to amplify the staphylococcus protein A (spa), Panton–Valentine Leukocidin (PVL), and mecA genes. The clinical spa types were compared with ones from our data base of S. aureus strains previously collected and sequenced from the community and environment in Northeast Ohio.ResultsA total of 51 spa types were detected from 129 S. aureus clinical isolates (discriminatory index, 0.876; 95% confidence interval [CI], 0.827–0.925; Table 1). The most common spa types were t008 (42/129, 32.6%), t002 (16/129, 12.4%), and t334 (6/129, 4.7%). In comparison, the most frequently detected spa types from the environmental samples were t189 (40/257, 15.6%), t002 (16/257, 6.2%), and t008 (11/257, 4.3%). Among the S. aureus isolates (n = 146), 45 were PVL-positive (30.8%) and 94 (66.7%) carried mecA. Of the 42 t008 (ST8/USA300; a common community-associated strain) isolates, 35 (83.3%) were methicillin-resistant S. aureus (MRSA) (based on the presence of the mecA gene) and 25 (59.5%) were PVL-positive. Thirteen of the sixteen (81.2%) t002 (ST5/USA100; a common hospital-associated strain) were MRSA and only one (6.2%) was PVL-positive.ConclusionThere is considerable overlap of S. aureus strains present in clinical samples with those found in the environment. This finding should draw attention to the need for more effective prevention strategies to reduce the risk of transmission of S. aureus, including MRSA, in the environment to humans. Disclosures All authors: No reported disclosures.

Multidrug-resistant Staphylococcus aureus Colonization in Healthy Adults Is more Common in Bhutanese Refugees in Nepal than Those Resettled in Ohio.

Although studies have shown that human migration is one of the risk factors for the spread of drug-resistant organisms such as methicillin-resistant Staphylococcus aureus (MRSA), surveillance studies examining MRSA among refugee populations in the US are lacking. This study aimed to assess the prevalence and molecular characteristics of S. aureus among Bhutanese refugees living in Nepal and resettled in Northeast Ohio (NEO). One hundred adult Bhutanese refugees from each geographic location were enrolled between August 2015 and January 2016. The participants were interviewed to collect demographic information and potential risk factors for carriage. Nasal and throat swabs were collected for bacterial isolation. All S. aureus isolates were characterized by spa typing and tested for the presence of Panton-Valentine leukocidin (PVL) and mecA genes; selected isolates were tested by multilocus sequence typing (MLST). The overall prevalence of S. aureus was 66.0% and 44.0% in NEO and Nepal, respectively. In Nepal, 5.8% (3/52) of isolates were MRSA and 1.1% (1/88) in NEO. Twenty-one isolates in NEO (23.9%) were multidrug-resistant S. aureus (MDRSA), while 23 (44.2%) in Nepal were MDRSA. In NEO, 41 spa types were detected from 88 S. aureus isolates. In Nepal, 32 spa types were detected from 52 S. aureus isolates. spa types t1818 and t345 were most common in NEO and Nepal, respectively. The overall prevalence of PVL-positive isolates among S. aureus in Nepal and NEO was 25.0% and 10.2%. ST5 was the most common sequence type in both locations. Bhutanese refugees living in Nepal and resettled in NEO had high prevalence of S. aureus and MDRSA. The findings suggest a potential need for CA-MRSA surveillance among the immigrant population in the U S and among people living in Nepal, and a potential need to devise appropriate public health measures to mitigate the risk imposed by community-associated strains of S. aureus and 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.

Whole-genome sequencing of Staphylococcus aureus L401, a mecA-negative community-associated methicillin-resistant strain isolated from a healthy carrier.

Methicillin-resistant Staphylococcus aureus (MRSA) is a human pathogen of great concern owing to its antimicrobial resistance and virulence properties. Here we report the first draft genome sequence of a mecA-negative community-associated MRSA strain isolated from a healthy young Mexican paediatric carrier in order to reveal the genomic structure underlying the multidrug-resistant phenotype and to discover the virulence properties of this strain. The draft genome sequence of S. aureus L401 was obtained using an Ion Torrent™ PGM platform. De novo assembled contigs were annotated, and antimicrobial resistance genes and virulence factors were identified using ResFinder and VirulenceFinder, respectively. In addition, a mutational survey of native pbp, gdpP and yjbH genes was performed. In silico multilocus sequence typing (MLST) and spa typing were also performed. S. aureus L401 has a genome size of 2 831 587 bp with 2799 protein-coding sequences. Various antimicrobial resistance genes conferring resistance to aminoglycosides, β-lactams, fluoroquinolones and macrolide-lincosamide-streptogramin B antimicrobials were found. Although both mecA and staphylococcal cassette chromosome mec (SCCmec) elements were absent, a missense mutation in PBP3 was identified. Moreover, genes encoding exfoliative toxin A, γ- and β-haemolysin, and several enterotoxins were also identified. S. aureus L401 belongs to ST109 and spa type t209. The availability of this genome will allow an insight into S. aureus resistance and virulence determinants as well as its epidemiology, lineage, evolution and genomic features involved in the paediatric commensal carriage.

Prevalence of Antibiotic and Heavy Metal Resistance Determinants and Virulence-Related Genetic Elements in Plasmids of Staphylococcus aureus.

The use of antibiotics on a mass scale, particularly in farming, and their release into the environment has led to a rapid emergence of resistant bacteria. Once emerged, resistance determinants are spread by horizontal gene transfer among strains of the same as well as disparate bacterial species. Their accumulation in free-living as well as livestock and community-associated strains results in the widespread multiple-drug resistance among clinically relevant species posing an increasingly pressing problem in healthcare. One of these clinically relevant species is Staphylococcus aureus, a common cause of hospital and community outbreaks. Among the rich diversity of mobile genetic elements regularly occurring in S. aureus such as phages, pathogenicity islands, and staphylococcal cassette chromosomes, plasmids are the major mean for dissemination of resistance determinants and virulence factors. Unfortunately, a vast number of whole-genome sequencing projects does not aim for complete sequence determination, which results in a disproportionately low number of known complete plasmid sequences. To address this problem we determined complete plasmid sequences derived from 18 poultry S. aureus strains and analyzed the prevalence of antibiotic and heavy metal resistance determinants, genes of virulence factors, as well as genetic elements relevant for their maintenance. Some of the plasmids have been reported before and are being found in clinical isolates of strains typical for humans or human ones of livestock origin. This shows that livestock-associated staphylococci are a significant reservoir of resistance determinants and virulence factors. Nevertheless, nearly half of the plasmids were unknown to date. In this group we found a potentially mobilizable plasmid pPA3 being a unique example of accumulation of resistance determinants and virulence factors likely stabilized by a presence of a toxin–antitoxin system.

Molecular epidemiology and resistance profiles among healthcare- and community-associated Staphylococcus aureus keratitis isolates.

PurposeTo characterize the molecular, epidemiological, and resistance profiles of methicillin-resistant (MRSA) and methicillin-susceptible (MSSA) keratitis isolates.Patients and methodsWe used a combination of standard microbiological techniques and DNA microarray analysis to characterize the molecular and antibiotic resistance profiles of 75 Staphylococcus aureus keratitis isolates collected over an 11-year period (2006–2016).ResultsTwo major USA clonal complexes (CC), CC5 (n=30, 40%) and CC8 (n=28, 37.3%), accounted for 77.3% of the collected S. aureus isolates. USA100, traditionally healthcare associated (n=18/47, 38.3%), and USA300, traditionally community associated (n=12/47, 25.5%), were the dominant MRSA strains. Four (22.2%) of the USA100 MRSA isolates were recovered from patients with no prior healthcare exposure. Eleven (91.7%) of the USA300 isolates were recovered from patients with documented healthcare risk factors. MSSA isolates were polyclonal (n=13). Ninety-three percent of MSSA infections were of healthcare origin. Thirty-seven of 61 (60.6%) healthcare- and 11 of 14 (78.6%) community-associated strains were resistant to three or more antibiotic classes. Sixty-eight percent (n=51) of isolates harbored three of more resistance determinants (genes). The Panton-Valentine Leucocidin gene was detected in 11 (14.7%) of the study isolates. The majority (72.7%) of the strains were members of the USA300 MRSA clone.ConclusionClonal complexes CC5 and CC8 were the most frequent clones detected among both the MSSA and the MRSA keratitis isolates. USA100 and USA300 clones were the dominant MRSA genotypes. The USA300 MRSA clone has become a leading cause of healthcare-associated keratitis in South Florida. The USA100 MRSA clone has emerged as an increasing cause of community-associated corneal infections in our outpatient population. This shifting epidemiology coupled with the increasing prevalence of multidrug resistance among both MSSA and MRSA keratitis is a cause of concern.

Prevalence and Molecular Characterization of Staphylococcus aureus and Methicillin-resistant S. aureus on Children's Playgrounds.

Staphylococcus aureus is a major public health concern due to the emergence of virulent and drug-resistant strains such as methicillin-resistant Staphylococcus aureus (MRSA). Although numerous studies have been conducted to assess the environmental contamination of S. aureus in health care and household settings, little is known about the prevalence and epidemiology of S. aureus, including MRSA, on environmental surfaces of children's playgrounds. This study investigated the prevalence and molecular epidemiology of S. aureus and MRSA at playgrounds in northeast Ohio. A total of 280 environmental samples were collected from 10 playgrounds in northeast Ohio in July 2016. Sampling sites were selected based on playground size and availability of equipment located in both small and large cities and their suburbs. Samples were analyzed using established microbiology methods, and resulting S. aureus isolates were typed by spa typing. Polymerase chain reaction was used to detect the presence of the Panton-Valentine leukocidin and mec A genes. Antibiotic susceptibility was tested via the Vitek-2 System. The overall prevalence of S. aureus and MRSA was 31.8% (89/280) and 3.9% (11/280), respectively. A total of 43 spa types were detected from 257 S. aureus isolates. Overall, t189 was the most common spa type, accounting for 15.6% (40/257) of the isolates. Sixteen isolates (6.2%) were t002 (ST5/USA100), a common hospital-associated strain, and 11 isolates (4.3%) were t008 (ST8/USA300), a common community-associated strain. Five livestock-associated strain (t571/ST398) were also identified. Twenty-nine (11.3%) isolates were resistant to oxacillin, and 66 (25.7%) were multi-drug resistant S. aureus. The results of this study indicate that environmental surfaces of playgrounds in northeastern Ohio were contaminated with S. aureus and MRSA. These data reinforce the need for implementing effective prevention strategies to mitigate the risk imposed to children by environmental contamination of MRSA.