Molecular characterization of methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> nasal isolates from Turkey

Methicillin-resistant Staphylococcus aureus (MRSA) colonize most frequently in the anterior nares of the nose and cause serious infections all over the world. The aim of this study was to determine the nasal carriage rate of S. aureus and MRSA strains in Turkish elementary school children. We also analyzed molecular characterizations of MRSA strains by using pulse field gel electrophoresis (PFGE), multi locus sequence typing (MLST), staphylococcal chromosomal cassette mec (SCCmec) typing, and detection of the Panton-valentine leucocidin (PVL) gene. The nasal swabs were obtained from 4,050 children during a 4 month period in Ankara. In vitro antimicrobial susceptibility testing to 1 μg oxacillin and 30 μg cefoxitin was determined by a disk diffusion method. We found that the 1,001 of 4,050 (24.7%) children were colonized with S. aureus. Three S. aureus strains were resistant to oxacillin and cefoxitin. The rate of MRSA among all children was 0.07%. The MRSA strains revealed three different PFGE pattern. All MRSA isolates by harbored the SCCmec type IV element, but not the PVL gene. The two MRSA isolate belonged to sequence type (ST) 30, whereas the other one was a unique type. The results of this study demonstrated that S. aureus nasal carriage rate was consistent with previous studies. However, MRSA carriage rate was low. This study also indicated that the ST30-type IV without PVL gene MRSA clone may be expected to spread in Turkish community.     

Combined Use of Ribotyping,PFGE Typing and IS431 Typing in the Discrimination of Nosocomial Strains of Methicillin-Resistant Staphylococcus aureus

We have previously reported the phenotypic characterization of methicillin-resistant Staphylococcus aureus (MRSA) clinical strains isolated in Malaya University Hospital in the period 1987 to 1989 using antibiogram, coagulase typing, plasmid profiles, and phage typing. Here, we report the analysis of the same strains with three genotyping methods; ribotyping, pulsed-field gel electrophoresis (PFGE) typing, and IS431 typing (a restriction enzyme fragment length polymorphism analysis using an IS431 probe). Ribotyping could discriminate 46 clinical MRSA strains into 5 ribotypes, PFGE typing into 22 types, and IS431 typing into 15 types. Since the differences of the three genotyping patterns from strain to strain were quite independent from one another, the combined use of the three genotyping methods could discriminate 46 strains into 39 genotypes. Thus, the powerful discriminatory ability of the combination was demonstrated.     

The Role of MRSA (Methicillin-Resistant Staphylococcus aureus) Adherence and Colonization in the Upper Respiratory Tract of Geriatric Patients in Nosocomial Pulmonary Infections

The mechanism of nosocomial respiratory infections caused by MRSA (methicillin-resistant Staphylococcus aureus) in geriatric patients was investigated. Seriously ill patients (SIP) undergoing naso-gastric tube feeding or intravenous hyperalimentation and moderately ill patients (MIP) who were orally fed, were examined for their colonization and infection by Staphylococcus aureus (S. aureus) in the respiratory tract. Colonization of MRSA in the upper respiratory tract in SIP was from six to ten times higher than that in MIP and was associated with a high incidence of MRSA pulmonary infections. In vitro S. aureus adherence to nasal or oropharyngeal cells demonstrated that bacteria binding to nasal cells was higher, which probably can be interpreted as an elevated occurrence of S. aureus colonization in the nasal cavity than in the throat. The binding activity of MRSA was not superior to that of MSSA (methicillin-sensitive S. aureus). Though MRSA binding to the nasal cells from SIP was not higher than those from MIP, MRSA colonization in the upper respiratory tract was more frequently seen in SIP (P < 0.01). A higher incidence of total infectious episodes (P < 0.02-0.001) and more frequent use of antibiotics (P < 0.02-0.001), which were potent against MSSA might be the basis for selection of MRSA in these patients. In fact, the rate of MRSA colonization on the skin (pressure sores) was also higher in SIP (P < 0.01). A low nutritional state in SIP (P < 0.01-0.02) might also be associated with MRSA colonization. The present results indicate that the high frequency of infections, antibiotic administration, MRSA skin colonization and low nutritional condition, are enhancing factors of MRSA acquisition in the respiratory tract for SIP undergoing artificial feeding, in a geriatric hospital.     

Differences in Epidemiological and Molecular Characteristics of Nasal Colonization with Staphylococcus aureus (MSSA-MRSA) in Children from a University Hospital and Day Care Centers

Background

Clinical significance of Staphylococcus aureus colonization has been demonstrated in hospital settings; however, studies in the community have shown contrasting results regarding the relevance of colonization in infection by community-associated MRSA (CA-MRSA). In Colombia there are few studies on S. aureus colonization. The aim of this study was to determine the molecular and epidemiological characteristics of nasal colonization by S. aureus (MSSA-MRSA) in children from a university hospital and day care centers (DCCs) of Medellin, Colombia.

Methods

An observational cross-sectional study was conducted in 400 children (200 in each setting), aged 0 months to 5 years, during 2011. Samples were collected from each nostril and epidemiological information was obtained from the parents. Genotypic analysis included spa typing, PFGE, MLST, SCCmec typing, detection of genes for virulence factors and agr groups.

Results

Frequency of S. aureus colonization was 39.8% (n = 159) (hospital 44.5% and DCCs 35.0%) and by MRSA, 5.3% (n = 21) (hospital 7.0% and DCCs 3.5%). Most S. aureus colonized children were older than two years (p = 0.005), the majority of them boys (59.1%), shared a bedroom with a large number of people (p = 0.028), with history of β-Lactamase inhibitors usage (p = 0.020). MSSA strains presented the greatest genotypic diversity with 15 clonal complexes (CC). MRSA isolates presented 6 CC, most of them (47.6%) belonged to CC8-SCCmec IVc and were genetically related to previously reported infectious MRSA strains.

Conclusion

Differences in epidemiological and molecular characteristics between populations may be useful for the understanding of S. aureus nasal colonization dynamics and for the design of strategies to prevent S. aureus infection and dissemination. The finding of colonizing MRSA with similar molecular characteristics of those causing infection demonstrates the dissemination capacity of S. aureus and the risk of infection among the child population.     

Typing of Staphylococcus epidermidis Colonizing in Human Nares by Pulsed-Field Gel Electrophoresis

From the nares of 11 healthy adults, 253 strains of coagulase negative staphylococcus were isolated and 88% of them were identified as Staphylococcus epidermidis using the API STAPH system. Chromosomal DNA fingerprinting of the isolated strains revealed that each person carried multiple types of S. epidermidis in his or her nares. The colonization of the strains was not stable; the types of the isolates changed in the first and the second examinations 5 months apart. The results contrasted with previous findings in which only one strain of S. aureus colonized persistently in the nares of healthy adults.     

Characterization of Virulence Factors and Genetic Background of Staphylococcus aureus Isolated from Peking University People’s Hospital Between 2005 and 2009

The aim of this study was to investigate both the genetic features of MRSA strains and the occurrence of virulence factors produced by Staphylococcus aureus strains isolated from Peking University People’s Hospital in Beijing, China, between 2005 and 2009. A total of 179 S. aureus strains were isolated, 139 of which were MRSA. The MRSA strains were characterized epidemiologically by SCCmec typing, spa typing and agr typing, then were classified into different genetic groups. The prevalence of genes coding for 14 exotoxins and eight adhesion factors among the S. aureus samples was assessed via polymerase chain reaction. Cluster analysis based on virulence factors-encoding gene content was performed to divide the MRSA isolates into valid clusters. Correspondence analysis was done to analyze the correlation between virulence factors clusters and genetic groups. JCSC1716-agrI-t030 (67.6%), SCCmec-IIIA-agrI-t030 (14.4%), SCCmec-IIIA-agrI-t037 (8.6%) and SCCmecII-agrII-t002 (2.2%) were four predominant MRSA clones. PVL was positive only in MSSA strains, there were at least three superantigenic toxins in our HA-MRSA clones, the prevalence of 16 virulence factors genes (sea, seb, sec, sed, seg, sei, sej, pvl, lukE-lukD, eta, bbp, can, ebp, clfA, fib, fnbB) in MRSA and MSSA was found to be significantly different from MSSA (P < 0.05). Results of correspondence analysis among clusters based on virulence factors genes and groups based on genetic typing illustrated not only the correspondence relationship between groups and clusters overall (P < 0.001), but also the genetic diversity of MRSA strains with respect to virulence factors genes.     

Comparison of Capsular Typing of Staphylococcus aureus with Bacteriophage Typing: A Study in Gulbarga, India

Staphylococcus aureus (S. aureus) is important both as a nosocomial and community acquired pathogen causing various degrees of infections. Typing S. aureus has been a question that is still being addressed. Bacteriophage typing is still used as a golden standard for typing though molecular methods are investigated. In developing countries where neither molecular typing nor the bacteriophage typing methods can be routinely used, the recently developed capsular typing method can be considered as screening method. We compared capsular typing with bacteriophage typing of the strains isolated in Gulbarga, India. We observed that the typeability of capsular typing was significantly higher (96%) among the phage typed strains, and the predominant capsular type in the region was type-8. The data so generated can be used to group S. aureus based on capsules both as a screening prior to bacteriophage typing, and to identify capsular candidate for developing prophylactic and therapeutic alternatives.     

2016-2017年宁夏地区牛源金黄色葡萄球菌的全基因组分析

[目的]了解宁夏地区奶牛乳腺炎金黄色葡萄球菌(Staphylococcus aureus,SA)代表菌株的基因组序列基本特征,进一步探究其耐药基因型、毒力及进化关系,为兽医临床防治提供理论依据.[方法]采用纸片法对97株金黄色葡萄球菌临床分离株进行抗菌药物敏感性试验,同时进行葡萄球菌蛋白A(Staphylococcus...     

Molecular epidemiological characterization of Staphylococcus aureus isolates originating from food poisoning outbreaks that occurred in Tokyo,Japan

Staphylococcal food poisoning (SFP), one of the commonest food‐borne diseases, results from the ingestion of one or more staphylococcal enterotoxins (SEs) produced in foods by Staphylococcus aureus. In the present study, 203 S. aureus strains originating from 83 outbreaks that had occurred in Tokyo were examined for their coagulase type and genotype of SEs to analyze their molecular epidemiological characteristics. The representative subsets of the 83 S. aureus isolates were analyzed by multilocus sequence typing (MLST) and S. aureus pathogenicity island (SaPI) scanning. The isolates were integrated into eight specific clonal complexes (CC) s; CC81, CC8, CC6, CC5, CC508, CC59, CC20 and CC30. The profiles of the coagulase type, SE/SEl genotype and the suspected type of enterotoxin‐encoding mobile genetic element (MGE) indicated a correlation with each CC. SaPI scanning showed fixed regularity between the distributions of genomic islands, including SaPIs, and the phylogenetic lineage based on MLST. These results indicate that the S. aureus isolates, which classified into eight CCs, have distinguishable properties concerning specific coagulase type, enterotoxin genotype and MGE type. Strains of S. aureus harboring these particular elements possess the potential to cause SFP.     

Epidemiology of Staphylococcus aureus Bacteraemia at a Tertiary Children’s Hospital in Cape Town,South Africa

Background

Staphylococcus aureus is an important pathogen in paediatric patients with bloodstream infections. The epidemiology of S. aureus bacteraemia, however, has not been well documented in children in South Africa.

Methods

A retrospective study was conducted at a children’s hospital in Cape Town, South Africa, to investigate the epidemiology of S. aureus bacteraemia from 2007-2011. The incidence, clinical presentation, risk factors, management and outcomes of methicillin sensitive S. aureus (MSSA) and methicillin resistant S. aureus (MRSA) bacteraemia were compared.

Results

Over the five year study period, 365 episodes of S. aureus bacteraemia were identified. The annual incidence was 3.28 cases per 1000 hospital admissions. MRSA was responsible for 26% of S. aureus bacteraemia and 72% of nosocomial infections. Only six possible cases of community-acquired MRSA infections were described. MSSA bacteraemia was more likely to present as pulmonary and bone or joint infections, while bacteraemia without a source was the most common presentation with MRSA.  Infants, children with malnutrition, and residents of long-term care facilities were at highest risk for MRSA bacteraemia. The overall case fatality rate for S. aureus bacteraemia was 8.8% over five years, with MRSA being the only significant risk factor for mortality.

Conclusion

The incidence of S. aureus bacteraemia and MRSA bacteraemia in children has remained stable over the past five years. MRSA is a predominantly nosocomial pathogen in children with S. aureus bacteraemia in Cape Town, South Africa.     

Screening of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> nasal strains isolated from medical students for toxin genes

Three hundred twenty-one students (156 students with no clinical exposure and 165 students with clinical exposure) were screened for nasal colonization by Staphylococcus aureus; 20.9% of students were S. aureus nasal carriers, and 40.3% of S. aureus isolates harbored toxin genes. The most prevalent genes were tst (15.0 %) and sec (13.4 %). Isolates with multiple genes were only found among clinical students (p = 0.045). Six of 11 PFGE clones were positive for toxin genes. Methicillin-resistant (MRSA) isolates were only detected in the clinical students (4.5 %). The exposure of students to the hospital environment neither radically increased S. aureus nasal carriage, nor the frequency of clinically important toxin gene presence, but it could have influenced the positive selection of toxigenic MRSA strains.     

Changes in the nasal colonization with methicillin-resistant Staphylococcus aureus in children: 2004-2009

Background

Staphylococcus aureus is an important cause of infection, particularly in persons colonized with this organism. This study compared the annual prevalence and microbiological characteristics of methicillin-resistant S. aureus (MRSA) nasal colonization in Taiwanese children from 2004 through 2009. Risk factors for MRSA were determined for the overall study period.

Methods

Children from birth to ≤14 years of age presenting for health maintenance visits or attending 1 of 57 kindergartens were recruited. Nasal swabs were obtained, and a questionnaire was administered. The prevalence and microbiological characteristics of MRSA colonization were also calculated for two 3-year periods: 2004–2006 and 2007–2009.

Results

Cultures of the anterior nares were positive for S. aureus in 824 (25.8%) of the 3,200 children, and MRSA colonization was found in 371 (11.6%) children. The prevalence of S. aureus colonization decreased from 28.1% in 2004–2006 to 23.3% in 2007–2009 (p<0.01), whereas the prevalence of MRSA colonization increased from 8.1% to 15.1% during this period (p<0.0001). Multivariate analysis revealed that the independent risk factors for MRSA carriage were different for male and female children, and also among age groups. Most MRSA isolates belonged to sequence type 59 (ST59) (86.3%); however, a multiresistant MRSA clone with ST338 background emerged in 2007–2009. Ten (62.5%) of the 16 MRSA isolates expressed the genotypic profile ST338/staphylococcal cassette chromosome mec V_T/Panton-Valentine leukocidin-positive/staphylococcal enterotoxin B-positive, and differed only in their antimicrobial susceptibility patterns.

Conclusion

The prevalence of nasal colonization by MRSA increased among healthy Taiwanese children from 2004–2006 to 2007–2009, despite an overall decrease in the prevalence of nasal colonization by S. aureus. A multiresistant MRSA clone characterized as ST338 was identified from these children.     

Antimicrobial resistance and presence of <Emphasis Type="Italic">ileS-2</Emphasis> gene encoding mupirocin resistance in clinical isolates of methicillin-resistant <Emphasis Type="Italic">Staphylococcus</Emphasis> sp.

Seventy-eight staphylococcal strains were isolated from surgical-site, blood-stream and other hospital-acquired infections. Eighteen isolates were determined as methicillin (MET)-resistant S. aureus (MRSA), while the remaining were MET-resistant coagulase-negative staphylococci (CoNS). Fifty percent of CoNS strains were multiresistant, while 10 % of isolates were resistant only to β-lactams. Clinical isolates of CoNS were generally more resistant to antimicrobial agents than S. aureus strains. Thirty-nine % of S. aureus strains were resistant only to β-lactams. None of the MRSA strains carried ileS-2 gene; this gene was found in two strains of S. epidermidis.     

Comparative Evaluation of Five Culture Media with Triplex PCR Assay for Detection of Methicillin-Resistant Staphylococcus aureus

The emergence of methicillin-resistant Staphylococcus aureus (MRSA) is responsible for nosocomial and community-acquired infections. Hence, rapid and accurate laboratory diagnosis of MRSA is a vital constituent of control measures. The present study evaluated five different methods for the identification of MRSA. A total of 207 S. aureus clinical isolates that consisted of 89 MRSA and 118 methicillin-susceptible S. aureus (MSSA) strains confirmed by PCR were tested. MRSA strains were evaluated by five different methods: chromogenic MRSA agar (CMRSA), oxacillin resistance screening agar base (ORSAB), mannitol salt oxacillin agar (MSO), mannitol salt cefoxitin agar with two different concentrations of cefoxitin [4 μg/ml (MSC-4) and 6 μg/ml (MSC-6)]. The results of the different methods were compared to mecA PCR as the gold standard. MSC-6 showed only six false-positive MRSA in comparison with PCR. The sensitivities and specificities of MSC-6, MSC-4, MSO-4, ORSAB, and CMRSA were as follows: 98.9/94.9%, 100/83.1%, 89.9/87.3%, 97.8/96.6%, and 95.5/94.9%, respectively. In comparison with PCR, it was found that both MSC-6 and ORSAB were relatively the least expensive screening tests (0.70 and0.70 and 1.00, respectively). In conclusion, all methods were comparable, but MSC-6 was the least expensive medium for MRSA screening.     

A multi-center blinded study on the efficiency of phenotypic screening methods to detect glycopeptide intermediately susceptible Staphylococcus aureus (GISA) and heterogeneous GISA (h-GISA)

Background

Staphylococcus aureus, one of the most frequently isolated pathogens in both hospitals and the community, has been particularly efficient at developing resistance to antimicrobial agents. In developed countries, as methicillin-resistant S. aureus (MRSA) has prevailed and, furthermore, as S. aureus with reduced susceptibility to vancomycin has emerged, the therapeutic options for the treatment of S. aureus infections have become limited. In developing countries and especially African countries very little is known concerning the resistance of S. aureus to antibiotics. In Madagascar no data exist concerning this resistance.

Objective

To update the current status of antibiotic resistance of S. aureus in Antananarivo, Madagascar.

Methods

Clinical S. aureus isolates were collected from patients at the Institut Pasteur of Madagascar from January 2001 to December 2005. Susceptibility tests with 18 antibiotics were performed by the disk diffusion method.

Results

Among a total of 574 isolates, 506 were from community-acquired infections and 68 from nosocomial infections. There was no significant difference in the methicillin resistance rate between community-acquired strains (33 of 506; 6.5%) and nosocomial strains (3 of 68, 4.4%). Many MRSA isolates were resistant to multiple classes of antibiotics. Resistance to tetracyclin, trimethoprim-sulfamethoxazole and erythromycin was more common. Among MRSA isolates resistance rates to rifampicin, fusidic acid, gentamicin and ciprofloxacin were lower than that observed with other drugs easily available in Madagascar. No isolates were resistant to glycopeptides.

Conclusion

The rate of methicillin-resistant S. aureus is not different between community-acquired and nosocomial infections and is still rather low in Madagascar.     

Molecular Analysis of Human and Canine Staphylococcus aureus Strains Reveals Distinct Extended-Host-Spectrum Genotypes Independent of Their Methicillin Resistance

Staphylococcus aureus causes a wide range of infectious diseases in humans and various animal species. Although presumptive host-specific factors have been reported, certain genetic lineages seem to lack specific host tropism, infecting a broad range of hosts. Such Extended-Host-Spectrum Genotypes (EHSGs) have been described in canine infections, caused by common regional human methicillin-resistant S. aureus (MRSA) lineages. However, information is scarce about the occurrence of methicillin-susceptible S. aureus (MSSA) EHSGs. To gain deeper insight into EHSG MSSA and EHSG MRSA of human and canine origin, a comparative molecular study was carried out, including a convenience sample of 120 current S. aureus (70 MRSA and 50 MSSA) isolates obtained from infected dogs. spa typing revealed 48 different spa types belonging to 16 different multilocus sequence typing clonal complexes (MLST-CCs). Based on these results, we further compared a subset of canine (n = 48) and human (n = 14) strains, including isolates of clonal complexes CC5, CC22, CC8, CC398, CC15, CC45, and CC30 by macrorestriction (pulsed-field gel electrophoresis [PFGE]) and DNA-microarray analysis. None of the methods employed was able to differentiate between clusters of human and canine strains independently of their methicillin resistance. In contrast, DNA-microarray analysis revealed 79% of the 48 canine isolates as carriers of the bacteriophage-encoded human-specific immune evasion cluster (IEC). In conclusion, the high degree of similarity between human and canine S. aureus strains regardless of whether they are MRSA or MSSA envisions the existence of common genetic traits that enable these strains as EHSGs, challenging the concept of resistance-driven spillover of MRSA.     

Detection of methicillin-resistant <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> using double duplex real-time PCR and dye Syto 9

A screening method for methicillin-resistant Staphylococcus aureus (MRSA) using real-time polymerase chain reaction (PCR) and dye Syto 9 was developed and evaluated. The assay was based on the two duplex reactions run simultaneously. The detection reaction amplified staphylococcal cassette chromosome mec (SCCmec) right extremity sequences and S. aureus-specific 442-bp DNA (Sa442). The control reaction amplified S. aureus-specific nuclease gene nuc and a marker of methicillin resistance, mecA. The method was evaluated by analyzing 214 clinical S. aureus isolates yielding 98.7 % sensitivity, 100 % specificity, 100 % positive predictive value and 96.6 % negative predictive value for detection of MRSA. The detection limit was determined to be 15–80 genome copies per real-time PCR. It was able to discriminate between MRSA, methicillin resistant coagulase negative staphylococci and methicillin susceptible S. aureus (MSSA) isolates containing only small fragments of the right extremity of the SCCmec (MSSA revertants).     

An evaluation of survival and detection of Campylobacter jejuni and C. coli in broiler caecal contents using culture‐based methods

Aims: (i) To cultivate methicillin‐resistant Staphylococcus aureus (MRSA) from a full‐scale wastewater treatment plant (WWTP), (ii) To characterize the indigenous MRSA‐flora, (iii) To investigate how the treatment process affects clonal distribution and (iv) To examine the genetic relation between MRSA from wastewater and clinical MRSA. Methods: Wastewater samples were collected during 2 months at four key sites in the WWTP. MRSA isolates were characterized using spa typing, antibiograms, SSCmec typing and detection of Panton–Valentine leukocidin (PVL). Conclusions: MRSA could be isolated on all sampling occasions, but only from inlet and activated sludge. The number of isolates and diversity of MRSA were reduced by the treatment process, but there are indications that the process was selected for strains with more extensive antibiotic resistance and PVL+ strains. The wastewater MRSA‐flora had a close genetic relationship to clinical isolates, most likely reflecting carriage in the community. Significance and Impact of the Study: This study shows that MRSA survives in wastewater and that the WWTP may be a potential reservoir for MRSA.