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.     

Molecular Epidemiology of Methicillin-Resistant Staphylococcus aureus Isolated from Australian Veterinarians

This work investigated the molecular epidemiology and antimicrobial resistance of methicillin-resistant Staphylococcus aureus (MRSA) isolated from veterinarians in Australia in 2009. The collection (n = 44) was subjected to extensive molecular typing (MLST, spa, SCCmec, dru, PFGE, virulence and antimicrobial resistance genotyping) and antimicrobial resistance phenotyping by disk diffusion. MRSA was isolated from Australian veterinarians representing various occupational emphases. The isolate collection was dominated by MRSA strains belonging to clonal complex (CC) 8 and multilocus sequence type (ST) 22. CC8 MRSA (ST8-IV [2B], spa t064; and ST612-IV [2B], spa variable,) were strongly associated with equine practice veterinarians (OR = 17.5, 95% CI = 3.3–92.5, P < 0.001) and were often resistant to gentamicin and rifampicin. ST22-IV [2B], spa variable, were strongly associated with companion animal practice veterinarians (OR = 52.5, 95% CI = 5.2–532.7, P < 0.001) and were resistant to ciprofloxacin. A single pig practice veterinarian carried ST398-V [5C2], spa t1451. Equine practice and companion animal practice veterinarians frequently carried multiresistant-CC8 and ST22 MRSA, respectively, whereas only a single swine specialist carried MRSA ST398. The presence of these strains in veterinarians may be associated with specific antimicrobial administration practices in each animal species.     

Characterization of Staphylococcus aureus from Distinct Geographic Locations in China: An Increasing Prevalence of spa-t030 and SCCmec Type III

Staphylococcus aureus belongs to one of the most common bacteria causing healthcare and community associated infections in China, but their molecular characterization has not been well studied. From May 2011 to June 2012, a total of 322 non-duplicate S. aureus isolates were consecutively collected from seven tertiary care hospitals in seven cities with distinct geographical locations in China, including 171 methicillin sensitive S. aureus (MSSA) and 151 MRSA isolates. All isolates were characterized by spa typing. The presence of virulence genes was tested by PCR. MRSA were further characterized by SCCmec typing. Seventy four and 16 spa types were identified among 168 MSSA and 150 MRSA, respectively. One spa type t030 accounted for 80.1% of all MRSA isolates, which was higher than previously reported, while spa-t037 accounted for only 4.0% of all MRSA isolates. The first six spa types (t309, t189, t034, t377, t078 and t091) accounted for about one third of all MSSA isolates. 121 of 151 MRSA isolates (80.1%) were identified as SCCmec type III. pvl gene was found in 32 MSSA (18.7%) and 5 MRSA (3.3%) isolates, with ST22-MSSA-t309 as the most commonly identified strain. Compared with non-epidemic MRSA clones, epidemic MRSA clones (corresponding to ST239) exhibited a lower susceptibility to rifampin, ciprofloxacin, gentamicin and trimethoprim-sulfamethoxazole, a higher prevalence of sea gene and a lower prevalence of seb, sec, seg, sei and tst genes. The increasing prevalence of multidrug resistant spa-t030 MRSA represents a major public health problem in China.     

Matched-Cohort DNA Microarray Diversity Analysis of Methicillin Sensitive and Methicillin Resistant Staphylococcus aureus Isolates from Hospital Admission Patients

As genotyping of S. aureus is important for epidemiologic research and for hygiene management, methods are required for standardized fast and easily applicable evaluation of closely related epidemic strains with high prevalence in hospitals. In this single centre matched control study we compared a new commercially available DNA microarray (IdentiBAC) with standard spa-typing for S. aureus genotyping. Included in the study was a subgroup of 46 MRSA and matched 46 MSSA nasal isolates of the Saarland University Medical Center collected during a state-wide admission prevalence screening. Microarray (MA) and also spa-typing could easily differentiate the genetically diverse MSSA group. However, due to the predominance of CC5/t003 in the MRSA group a sufficient subtyping required analysis of more complex genetic profiles as was shown here by the MA comprising a total number of 334 different hybridization probes. The genetic repertoire of the MRSA group was characterized by more virulence genes as compared to the MSSA group. The standard evaluation of MA results by the original software into CCs, agr-, SCCmec- and capsule-types was substituted in the present study by implementation of multivariate subtyping of closely related CC5 isolates using three different bioinformatic methods (splits graph, cluster dendrogram, and principal component analysis). Each method used was applicable for standardized and highly discriminative subtyping with high concordance. We propose that the identified S. aureus subtypes with characteristic virulence gene profiles are presumably associated also with virulence and pathogenicity in vivo; however, this remains to be analyzed in future studies. MA was superior to spa-typing for epidemiologic and presumably also provide functional respectively virulence associated characterization of S. aureus isolates. This is of specific importance for the hospital setting. In future, MA could become a new standard test for S. aureus typing in combination with multivariate bioinformatic analysis.     

Methicillin-Sensitive and Methicillin-Resistant Staphylococcus aureus Nasal Carriage in a Random Sample of Non-Hospitalized Adult Population in Northern Germany

Objective

The findings from truly randomized community-based studies on Staphylococcus aureus nasal colonization are scarce. Therefore we have examined point prevalence and risk factors of S. aureus nasal carriage in a non-hospitalized population of Braunschweig, northern Germany.

Methods

A total of 2026 potential participants were randomly selected through the resident''s registration office and invited by mail. They were requested to collect a nasal swab at home and return it by mail. S. aureus was identified by culture and PCR. Logistic regression was used to determine risk factors of S. aureus carriage.

Results

Among the invitees, 405 individuals agreed to participate and 389 provided complete data which was included in the analysis. The median age of the participants was 49 years (IQR: 39–61) and 61% were females. S. aureus was isolated in 85 (21.9%; 95% CI: 18.0–26.2%) of the samples, five of which were MRSA (1.29%; 95% CI: 0.55–2.98%). In multiple logistic regression, male sex (OR = 3.50; 95% CI: 2.01–6.11) and presence of allergies (OR = 2.43; 95% CI: 1.39–4.24) were found to be associated with S. aureus nasal carriage. Fifty five different spa types were found, that clustered into nine distinct groups. MRSA belonged to the hospital-associated spa types t032 and t025 (corresponds to MLST CC 22), whereas MSSA spa types varied and mostly belonged to spa-CC 012 (corresponds to MLST CC 30), and spa-CC 084 (corresponds to MLST CC 15).

Conclusion

This first point prevalence study of S. aureus in a non-hospitalized population of Germany revealed prevalence, consistent with other European countries and supports previous findings on male sex and allergies as risk factors of S. aureus carriage. The detection of hospital-associated MRSA spa types in the community indicates possible spread of these strains from hospitals into the community.     

Clonal Complex 398 Methicillin Susceptible Staphylococcus aureus: A Frequent Unspecialized Human Pathogen with Specific Phenotypic and Genotypic Characteristics

Clonal complex 398 livestok-associated-MRSA (CC398 LA-MRSA) clone is described as a major animal pathogen that can also colonize and infect humans. CC398 methicillin susceptible Staphylococcus aureus (CC398 MSSA) is less described. We identified 126 CC398 MSSA strains of human origin within 6380 S. aureus isolates gathered between 2009 and 2011, from the French National Reference Centre for Staphylococci. They were characterized using antimicrobial susceptibility testing, spa typing, DNA microarrays (Identibac S. aureus Genotyping ®, Alere), CC398-specific sequence PCR, ermT (encoding macrolides résistance) PCR. Fifty-three CC398 LA-MRSA collected from French pigs and veal were used as comparators, and phylogenetic relations between human CC398 MSSA and animal CC398 MRSA populations were explored on the basis of spa-typing and DNA microarrays. CC398 MSSA were able to induce a large spectrum of infections (especially skin, bloodstream, and pneumonias). The prevalence rate of this clone was high in MSSA population, i.e., 24.7% in a local prospective study on nasal colonization, and 7.5% in a national prospective study on infective endocarditis. CC398 MSSA isolates were frequently (89%) erythromycin resistant, due to the presence of the ermT gene, a gene not detected in erythromycin resistant CC398 LA-MRSA strains. Expression of staphylococcal complement inhibitor (scn) and the chemotaxis inhibitory protein (chp), was also specific to this population. The CC398 MRSA signature included also a panel of antibiotic resistance genes, especially a type IV or V cassette mec and tetM. CC398 MSSA and CC398 LA-MRSA populations were closely related based on spa-typing and DNA microarrays, with the MRSA strains forming the most derived lineage in phylogenic trees. Both MSSA and MRSA populations may come from common ancestors, which would have evolved in the settings of different selective pressures, explaining the acquisition of ermT, chp and scn for MSSA, and antibiotic resistance genes for MRSA.     

Geographic Distribution of Staphylococcus aureus Causing Invasive Infections in Europe: A Molecular-Epidemiological Analysis

Background

Staphylococcus aureus is one of the most important human pathogens and methicillin-resistant variants (MRSAs) are a major cause of hospital and community-acquired infection. We aimed to map the geographic distribution of the dominant clones that cause invasive infections in Europe.

Methods and Findings

In each country, staphylococcal reference laboratories secured the participation of a sufficient number of hospital laboratories to achieve national geo-demographic representation. Participating laboratories collected successive methicillin-susceptible (MSSA) and MRSA isolates from patients with invasive S. aureus infection using an agreed protocol. All isolates were sent to the respective national reference laboratories and characterised by quality-controlled sequence typing of the variable region of the staphylococcal spa gene (spa typing), and data were uploaded to a central database. Relevant genetic and phenotypic information was assembled for interactive interrogation by a purpose-built Web-based mapping application. Between September 2006 and February 2007, 357 laboratories serving 450 hospitals in 26 countries collected 2,890 MSSA and MRSA isolates from patients with invasive S. aureus infection. A wide geographical distribution of spa types was found with some prevalent in all European countries. MSSA were more diverse than MRSA. Genetic diversity of MRSA differed considerably between countries with dominant MRSA spa types forming distinctive geographical clusters. We provide evidence that a network approach consisting of decentralised typing and visualisation of aggregated data using an interactive mapping tool can provide important information on the dynamics of MRSA populations such as early signalling of emerging strains, cross border spread, and importation by travel.

Conclusions

In contrast to MSSA, MRSA spa types have a predominantly regional distribution in Europe. This finding is indicative of the selection and spread of a limited number of clones within health care networks, suggesting that control efforts aimed at interrupting the spread within and between health care institutions may not only be feasible but ultimately successful and should therefore be strongly encouraged. Please see later in the article for the Editors'' Summary     

A Typical Hospital-Acquired Methicillin-Resistant Staphylococcus aureus Clone Is Widespread in the Community in the Gaza Strip

Epidemiological data on community acquired methicillin-resistant-Staphylococcus aureus (CA-MRSA) carriage and infection in the Middle-East region is scarce with only few reports in the Israeli and Palestinian populations. As part of a Palestinian-Israeli collaborative research, we have conducted a cross-sectional survey of nasal S. aureus carriage in healthy children and their parents throughout the Gaza strip. Isolates were characterized for antibiotic susceptibility, mec gene presence, PFGE, spa type, SCCmec-type, presence of PVL genes and multi-locus-sequence-type (MLST). S. aureus was carried by 28.4% of the 379 screened children-parents pairs. MRSA was detected in 45% of S. aureus isolates, that is, in 12% of the study population. A single ST22-MRSA-IVa, spa t223, PVL-gene negative strain was detected in 64% of MRSA isolates. This strain is typically susceptible to all non-β-lactam antibiotics tested. The only predictor for MRSA carriage in children was having an MRSA carrier-parent (OR = 25.5, P = 0.0004). Carriage of the Gaza strain was not associated with prior hospitalization. The Gaza strain was closely related genetically to a local MSSA spa t223 strain and less so to EMRSA15, one of the pandemic hospital-acquired-MRSA clones, scarcely reported in the community. The rapid spread in the community may be due to population determinants or due to yet unknown advantageous features of this particular strain.     

The Role of the Staphylococcal VraTSR Regulatory System on Vancomycin Resistance and vanA Operon Expression in Vancomycin-Resistant Staphylococcus aureus

Vancomycin is often the preferred treatment for invasive methicillin-resistant Staphylococcus aureus (MRSA) infection. With the increase in incidence of MRSA infections, the use of vancomycin has increased and, as feared, isolates of vancomycin-resistant Staphylococcus aureus (VRSA) have emerged. VRSA isolates have acquired the entercoccal vanA operon contained on transposon (Tn) 1546 residing on a conjugal plasmid. VraTSR is a vancomycin and β-lactam-inducible three-component regulatory system encoded on the S. aureus chromosome that modulates the cell-wall stress response to cell-wall acting antibiotics. Mutation in vraTSR has shown to increase susceptibility to β-lactams and vancomycin in clinical VISA strains and in recombinant strain COLVA-200 which expresses a plasmid borne vanA operon. To date, the role of VraTSR in vanA operon expression in VRSA has not been demonstrated. In this study, the vraTSR operon was deleted from the first clinical VRSA strain (VRS1) by transduction with phage harvested from a USA300 vraTSR operon deletion strain. The absence of the vraTSR operon and presence of the vanA operon were confirmed in the transductant (VRS1Δvra) by PCR. Broth MIC determinations, demonstrated that the vancomycin MIC of VRS1Δvra (64 µg/ml) decreased by 16-fold compared with VRS1 (1024 µg/ml). The effect of the vraTSR operon deletion on expression of the van gene cluster (vanA, vanX and vanR) was examined by quantitative RT-PCR using relative quantification. A 2–5-fold decreased expression of the vanA operon genes occured in strain VRS1Δvra at stationary growth phase compared with the parent strain, VRS1. Both vancomycin resistance and vancomycin-induced expression of vanA and vanR were restored by complementation with a plasmid harboring the vraTSR operon. These findings demonstrate that expression in S. aureus of the horizontally acquired enterococcal vanA gene cluster is enhanced by the staphylococcal three-component cell wall stress regulatory system VraTSR, that is present in all S. aureus strains.     

Evaluation of Multiple-Locus Variable Number of Tandem Repeats Analysis for Typing Livestock-Associated Methicillin-Resistant Staphylococcus aureus

Background

The increasing occurrence of livestock-associated (LA) methicillin-resistant Staphylococcus aureus (MRSA) associated with the clonal complex (CC) 398 within the past years shows the importance of standardized and comparable typing methods for the purposes of molecular surveillance and outbreak detection. Multiple-locus variable number of tandem repeats analysis (MLVA) has recently been described as an alternative and highly discriminative tool for S. aureus. However, until now the applicability of MLVA for the typing of LA-MRSA isolates from different geographic origin has not been investigated in detail. We therefore compared MLVA and S. aureus protein A (spa) typing for characterizing porcine MRSA from distinct Dutch and German farms.

Methodology/Principal Findings

Overall, 134 MRSA isolates originating from 21 different pig-farms in the Netherlands and 36 farms in Germany comprising 21 different spa types were subjected to MLVA-typing. Amplification and subsequent automated fragment sizing of the tandem repeat loci on a capillary sequencer differentiated these 134 isolates into 20 distinct MLVA types. Whereas overall MLVA and spa typing showed the same discriminatory power to type LA-MRSA (p  = 0.102), MLVA was more discriminatory than spa typing for isolates associated with the prevalent spa types t011 and t034 (Simpson’s Index of Diversity 0.564 vs. 0.429, respectively; p<0.001).

Conclusion

Although the applied MLVA scheme was not more discriminatory than spa typing in general, it added valuable information to spa typing results for specific spa types (t011, t034) which are highly prevalent in the study area, i.e. Dutch-German border area. Thus, both methods may complement each other to increase the discriminatory power to resolute highly conserved clones such as CC398 (spa types t011, t034) for the detection of outbreaks and molecular surveillance of zoonotic MRSA.     

MRSA Clonal Complex 22 Strains Harboring Toxic Shock Syndrome Toxin (TSST-1) Are Endemic in the Primary Hospital in Gaza,Palestine

Background

Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in both community and healthcare-related settings worldwide. Current knowledge regarding the epidemiology of S. aureus and MRSA in Gaza is based on a single community-based carriage study. Here we describe a cross-sectional analysis of 215 clinical isolates collected from Al-Shifa Hospital in Gaza during 2008 and 2012.

Methods

All isolates were characterized by spa typing, SCCmec typing, and detection of genes encoding Panton-Valentine leukocidin (PVL) and toxic shock syndrome toxin (TSST-1). Representative genotypes were also subjected to multilocus sequence typing (MLST). Antibiotic susceptibility testing was performed using VITEK2 and MicroScan.

Results

MRSA represented 56.3% of all S. aureus strains, and increased in frequency from 2008 (54.8%) to 2012 (58.4%). Aside from beta-lactams, resistance was observed to tetracycline, erythromycin, clindamycin, gentamicin, and fluoroquinolones. Molecular typing identified 35 spa types representing 17 MLST clonal complexes (CC), with spa 998 (Ridom t223, CC22) and spa 70 (Ridom t044, CC80) being the most prevalent. SCCmec types I, III, IV, V and VI were identified among MRSA isolates, while type II was not detected. PVL genes (lukF/S-PV) were detected in 40.0% of all isolates, while the TSST-1 gene (tst) was detected in 27.4% of all isolates, with surprisingly high frequency within CC22 (70.4%). Both PVL and TSST-1 genes were found in several isolates from 2012.

Conclusions

Molecular typing of clinical isolates from Gaza hospitals revealed unusually high prevalence of TSST-1 genes among CC22 MRSA, which is noteworthy given a recent community study describing widespread carriage of a CC22 MRSA clone known as the ‘Gaza strain’. While the latter did not address TSST-1, tst-positive spa 998 (Ridom t223) has been detected in several neighboring countries, and described as endemic in an Italian NICU, suggesting international spread of a ‘Middle Eastern variant’ of pandemic CC22 strain EMRSA-15.     

Staphylococcus aureus Nasal Carriage among Beefpacking Workers in a Midwestern United States Slaughterhouse

Occupational contact with livestock is an established risk factor for exposure to livestock-associated methicillin-resistant Staphylococcus aureus (MRSA), particularly among industrial swine workers. While S. aureus is known to infect cattle, livestock-associated S. aureus carriage among workers in the beef production chain has received limited attention. Beefpacking workers, who slaughter, butcher and process cattle, have intensified exposure to potentially infectious animal materials and may be at risk of livestock-associated S. aureus exposure. We conducted a cross-sectional study of beefpacking workers (n = 137) at an industrial slaughterhouse in the Midwestern United States to evaluate prevalence and characteristics of S. aureus nasal colonization, specifically the absence of the scn gene to identify putative association with livestock, antibiotic susceptibility, presence of Panton-Valentin leukocidin (PVL) genes lukS-PV and lukF-PV, and spa type. Overall prevalence of S. aureus nasal carriage was 27.0%. No workers carried livestock-associated MRSA. Methicillin-sensitive S. aureus isolates (MSSA) recovered from five workers (3.6%) lacked the scn gene and were considered putative livestock-associated S. aureus (pLA-SA). Among pLA-SA isolates, spa types t338, t748, t1476 and t2379 were identified. To our knowledge, these spa types have not previously been identified as associated with livestock. Prevalence of human-adapted MRSA carriage in workers was 3.6%. MRSA isolates were identified as spa types t002, t008 and t024, and four of five MRSA isolates were PVL-positive. To date, this is the first study to indicate that industrial beefpacking workers in the United States may be exposed to livestock-associated S. aureus, notably MSSA, and to spa types not previously identified in livestock and livestock workers. Occupational exposure to livestock-associated S. aureus in the beef production chain requires further epidemiologic investigation.     

Spa typing of Staphylococcus aureus Isolated from Clinical Specimens from Outpatients in Iraq

Methicillin-resistant Staphylococcus aureus (MRSA) is notorious as a hospital superbug and a problematic pathogen among communities. The incidence of MRSA has substantially increased over time in Iraq. The aim of this study was to determine the prevalence and spa types of MRSA isolates from outpatients or patients upon admission into hospitals. Various biochemical tests identified S. aureus isolates, and then this identification was confirmed by PCR using species-specific 16S rRNA primer pairs. Antibiotic susceptibility was determined against methicillin, oxacillin, and vancomycin using the disk diffusion method. Vancomycin MIC was detected by VITEK 2 compact system. All the identified isolates were screened for the presence of mecA and lukS-PV-lukF-PV genes; 36 of them were subjected to spa typing-based PCR. Out of 290 clinical samples, 65 (22.4%) were S. aureus, of which 62 (95.4%) strains were resistant to oxacillin and methicillin. Except for two isolates, all MRSA isolates were mecA positive. One of the three MSSA isolates was mecA positive. Five strains were resistant to vancomycin. Fourteen (21.5%) isolates were positive for the presence of lukS-PV-lukF-PV genes. Spa typing of 36 S. aureus isolates revealed eleven different spa types, t304 (30.3%), t307 (19.4%), t346 (8.3%), t044 (8.3%), t15595 (8.3%), t386 (5.5%), t5475 (5.5%), t17928 (2.8%), t14870 (2.8%), t021 (2.8%), and t024 (2.8%). These findings could be useful for assessing the genetic relatedness of strains in the region for epidemiological and monitoring purposes, which would be essential to limiting the spread of MRSA.     

EnzyBase: a novel database for enzybiotic studies

Background

Staphylococcus aureus (S. aureus) is a major nosocomial pathogen that causes a variety of infections and toxicoses. In recent years, the percentage of rifampicin-resistant S. aureus has increased rapidly in China. The aims of this study were to analyze 1) the level of rifampicin resistance in S. aureus and its correlation with mutations in the rpoB gene, and 2) the molecular characterization of rifampicin-resistant S. aureus isolates.

Results

88 rifampicin-resistant S. aureus isolates were collected for this study. Of the 88 isolates, 83 (94.3%) were high-level rifampicin resistant (MIC??8 mg/L) while the remaining 5 isolates (5.7%) had a low-level resistance to rifampicin (MIC, 2 to 4 mg/L). Four amino acid substitutions were found in the 88 isolates, which were 481His/Asn (95.5%), 466Leu/Ser (87.5%), 477Ala/Asp (6.8%) and 486Ser/Leu (4.5%) respectively. All mutations were found to be present in cluster I of the rpoB gene. The low-level resistant isolates were found to have only one mutation, while the high-level resistant isolates had at least two or more mutations. The most common multiple mutations were 481His/Asn+466Leu/Ser(92.8%,77/83). The other multiple mutations found were 481His/Asn+477Ala/Asp (6.0%,5/83), and 481His/Asn+466Leu/Ser+477Ala/Asp (1.2%,1/83). Out of 28 high-level rifampicin-resistant S. aureus isolates, three molecular types were found, namely, ST239-MRSA-III-spa t030 (25/28, 89.3%), ST239-MRSA-III-spa t021 (2/28, 7.1%), and ST239-MRSA-III-spa t045 (1/28, 3.6%).

Conclusions

Rifampicin resistance in S. aureus was closely associated with mutations in the rpoB gene. High-level rifampicin-resistant S. aureus is one of the most important features in Anhui Provincial Hospital, and high-level rifampicin resistance in S. aureus is associated with multiple mutations of rpoB gene. The prevalence of high-level rifampicin-resistant S. aureus in Anhui may be associated with the spread of the ST239-MRSA III-spa t030 clone.     

Eradication of Methicillin-Resistant Staphylococcus aureus and of Enterobacteriaceae Expressing Extended-Spectrum Beta-Lactamases on a Model Pig Farm

Colonization of livestock with bacteria resistant to antibiotics is considered a risk for the entry of drug-resistant pathogens into the food chain. For this reason, there is a need for novel concepts to address the eradication of drug-resistant commensals on farms. In the present report, we evaluated the decontamination measures taken on a farm contaminated with methicillin-resistant Staphylococcus aureus (MRSA) and Enterobacteriaceae expressing extended-spectrum β-lactamases (ESBL-E). The decontamination process preceded the conversion from piglet breeding to gilt production. Microbiological surveillance showed that the decontamination measures eliminated the MRSA and ESBL-E strains that were detected on the farm before the complete removal of pigs, cleaning and disinfection of the stable, and construction of an additional stable meeting high-quality standards. After pig production was restarted, ESBL-E remained undetectable over 12 months, but MRSA was recovered from pigs and the environment within the first 2 days. However, spa (Staphylococcus aureus protein A gene) typing revealed acquisition of an MRSA strain (type t034) that had not been detected before decontamination. Interestingly, we observed that a farmworker who had been colonized with the prior MRSA strain (t2011) acquired the new strain (t034) after 2 months. In summary, this report demonstrates that decontamination protocols similar to those used here can lead to successful elimination of contaminating MRSA and ESBL-E in pigs and the stable environment. Nevertheless, decontamination protocols do not prevent the acquisition of new MRSA strains.     

The prevalence of Staphylococcus aureus and methicillin resistant Staphylococcus aureus in milk and dairy products in Riyadh,Saudi Arabia

In terms of life- menaced contagion, methicillin resistant Staphylococcus aureus (MRSA) is known to be one of which and it is truly notable in the contaminated food causing a community health anxiety. However, the occurrence of S. aureus and MRSA in diverse kinds of dairy products have been tested in this study. Samples from: raw milk (unpasteurized) from horse, goat, camel, and cow origins and unpacked cheese were checked for the recovered strains of such bacterium and MRSA. Wholly, MRSA isolates were verified for antimicrobial susceptibility and further characterized by mecA and staphylococcal cassette chromosome mec (SCCmec) typing. Also, Panton-Valentine Leukocidin (PVL), Staphylococcus aureus protein A (spa), and Staphylococcal enterotoxins (SEs) were also tested between all positive MRSA isolates in order to discover the virulence factors. Consequently, 70% of the 100 collected dairy products samples were contaminated by S. aureus bacteria and 72.9% of them were defined as MRSA. 9.8% of MRSA isolates contained mecA genes with SCCmec type II (80%) as the most common SCCmec type. Moreover, large number of MRSA isolates were identified as multidrug resistance and 28.6% of MRSA-mecA positive isolates were also carried vancomycin resistance genes (i.e., vanB). Too, spa gene was detected between 9.8% of MRSA isolates but PVL gene was not spotted at all. Additionally, the existing of SEs was variable between MRSA isolates and the most common type was SEH (51%). In general, our results confirmed that raw milk and unpacked cheese in the Kingdom of Saudi Arabia (Riyadh) is a potential vehicle for multidrug resistant MRSA transmission. It is a critical civic health menace and stresses, thus; the need of applying well cleanliness practices is essential.     

Molecular Characterization of Staphylococcus aureus from Patients with Surgical Site Infections at Mulago Hospital in Kampala,Uganda

Background

The prevalence of Methicillin resistant Staphylococcus aureus (MRSA) is progressively increasing globally with significant regional variation. Understanding the Staphylococcus aureus lineages is crucial in controlling nosocomial infections. Recent studies on S. aureus in Uganda have revealed an escalating burden of MRSA. However, the S. aureus genotypes circulating among patients are not known. Here, we report S. aureus lineages circulating in patients with surgical site infections (SSI) at Mulago National hospital, Kampala, Uganda.

Methods

A cross-sectional study involving 314 patients with SSI at Mulago National Hospital was conducted from September 2011 to April 2012. Pus swabs from the patients’ SSI were processed using standard microbiological procedures. Methicillin sensitive Staphylococcus aureus (MSSA) and MRSA were identified using phenotypic tests and confirmed by PCR-detection of the nuc and mecA genes, respectively. SCCmec genotypes were determined among MRSA isolates using multiplex PCR. Furthermore, to determine lineages, spa sequence based-genotyping was performed on all S. aureus isolates.

Results

Of the 314 patients with SSI, S. aureus accounted for 20.4% (64/314), of which 37.5% (24/64) were MRSA. The predominant SCCmec types were type V (33.3%, 8/24) and type I (16.7%, 4/24). The predominant spa lineages were t645 (17.2%, 11/64) and t4353 (15.6%, 10/64), and these were found to be clonally circulating in all the surgical wards. On the other hand, lineages t064, t355, and t4609 were confined to the obstetrics and gynecology wards. A new spa type (t10277) was identified from MSSA isolate. On multivariate logistic regression analysis, cancer and inducible clindamycin resistance remained as independent predictors of MRSA-SSI.

Conclusion

SCCmec types I and V are the most prevalent MRSA mecA types from the patients’ SSI. The predominant spa lineages (t645 and t4353) are clonally circulating in all the surgical wards, calling for strengthening of infection control practices at Mulago National 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.     

Clonal distribution of bone sialoprotein-binding protein gene among Staphylococcus aureus isolates associated with bloodstream infections

Staphylococcus aureus is a leading cause of bloodstream infections (BSI) and diseases that may be caused by hematogenous spread. The staphylococcal adhesin, for which the association with the infections emerging as a complication of septicemia has been well documented, is a bone sialoprotein-binding protein (Bbp). The aim of the study was to assess the prevalence of a bbp gene in S. aureus bloodstream isolates associated with BSI and to investigate to what degree the distribution of this gene is linked to the clonality of the population. Spa typing, used in order to explore the genetic population structure of the isolates, yielded 29 types. Six spa clusters and seven singletons were identified. The most frequent was spa clonal complex CC021 associated with MLST CC30 (38 %). The bbp gene was found in 47 % of isolates. Almost all isolates (95 %) clustered in spa clonal complex CC021 were positive for this gene. All isolates carrying the bbp gene were sensitive to methicillin, and if clustered in the spa CC021, belonged to agr group III. Our study shows that Bbp is not strictly associated with BSI. However, one may conclude that for clonally related S. aureus strains most commonly causing BSI, the risk of Bbp-mediated complications of septicemia is expected to be higher than for other strains.     

Genetic Diversity and Ecological Success of Staphylococcus aureus Strains Colonizing Humans

The genetic determinants and phenotypic traits which make a Staphylococcus aureus strain a successful colonizer are largely unknown. The genetic diversity and population structure of 133 S. aureus isolates from healthy, generally risk-free adult carriers were investigated using four different typing methods: multilocus sequence typing (MLST), amplified fragment length polymorphism analysis (AFLP), double-locus sequence typing (DLST), and spa typing were compared. Carriage isolates displayed great genetic diversity which could only be revealed fully by DLST. Results of AFLP and MLST were highly concordant in the delineation of genotypic clusters of closely related isolates, roughly equivalent to clonal complexes. spa typing and DLST provided considerably less phylogenetic information. The resolution of spa typing was similar to that of AFLP and inferior to that of DLST. AFLP proved to be the most universal method, combining a phylogeny-building capacity similar to that of MLST with a much higher resolution. However, it had a lower reproducibility than sequencing-based MLST, DLST, and spa typing. We found two cases of methicillin-resistant S. aureus colonization, both of which were most likely associated with employment at a health service. Of 21 genotypic clusters detected, 2 were most prevalent: cluster 45 and cluster 30 each colonized 24% of the carrier population. The number of bacteria found in nasal samples varied significantly among the clusters, but the most prevalent clusters were not particularly numerous in the nasal samples. We did not find much evidence that genotypic clusters were associated with different carrier characteristics, such as age, sex, medical conditions, or antibiotic use. This may provide empirical support for the idea that genetic clusters in bacteria are maintained in the absence of adaptation to different niches. Alternatively, carrier characteristics other than those evaluated here or factors other than human hosts may exert selective pressure maintaining genotypic clusters.