Pvl Genes Research Articles

Staphylococcus aureus nasal isolates may have the same genetic profile in atopic dermatitis paediatric patients and their close contacts.

Atopic dermatitis (AD) is a chronic skin disease that affects up to 20 % of the paediatric population worldwide. Staphylococcus aureus colonizes anterior nares and can be transmitted in the home environment, aggravating AD. This study aimed to detect S. aureus from nares of AD patients and their family contacts, as well as to evaluate the antimicrobial resistance, virulence and clonality of these isolates. Among the 48 family groups investigated, 30 groups were selected, as both the child and his/her respective contact had methicillin-sensitive S. aureus (MSSA) (24 cases; 54 MSSA isolates) or methicillin-resistant S. aureus (MRSA) isolates (6 cases; 13 MRSA isolates). All MRSA isolates carried SCCmec IV. S. aureus carrying PVL genes were detected in 60 % of patients. Pulsed-field gel electrophoresis (PFGE) analysis was performed for 31 isolates from 15 family groups: all 6 with MRSA and 9 with MSSA isolates. Similar genotypic profiles between isolates from patients and their family contacts were noted in 10 (66.6 %) family groups, 5 (83.3 %) of the MRSA family groups and 5 (55.5 %) of the MSSA family groups, indicating that the pathogen was transmitted through family contacts.

Prevalence, Antimicrobial Resistance Profiles, Virulence and Enterotoxins-Determinant Genes of MRSA Isolated from Subclinical Bovine Mastitis in Egypt.

Subclinical mastitis caused by Staphylococcus aureus has worldwide public health significance. Here, we aimed to determine the prevalence of S. aureus, antimicrobial resistance profiles, and the virulence and enterotoxins determinant genes of MRSA strains that caused subclinical bovine mastitis. Milk samples were collected from 120 lactating animals (50 buffaloes and 70 dairy cattle) from different farms located in Ismailia Province (Egypt). The collected samples were investigated for subclinical mastitis using a California mastitis test. The total prevalence of S. aureus was 35.9% (84/234) with 36.3% (53/146) in cattle and 31% (31/88) in buffaloes. Antimicrobial susceptibility testing showed that 35.7% (30/84) of the isolated strains were resistant to cefoxitin, defined as methicillin-resistant S. aureus (MRSA), with 37.7% (20/53) in cattle and 32.2% (10/31) in buffaloes. Using PCR, 100% of the tested strains harbored coa and mecA genes, while 86.6% were positive for spa gene, with remarkable gene size polymorphism. Additionally, 10% of the tested strains contained the pvl gene. Further, using multiplex PCR, 26.6% of the tested samples had sea gene, two strains had sec gene and only one strain had sea and sec genes. The seb and sed genes were absent in the tested strains. In conclusion, mecA, coa and spa virulence genes were widely distributed in MRSA strains isolated from bovine milk, whereas the sea gene was the most predominant enterotoxin gene. Notably, this is the first report that emphasizes the prevalence of pvl gene of MRSA isolated from bovine milk in Egypt.

Characterization of biofilm formation and virulence factors of Staphylococcus aureus isolates from paediatric patients in Tehran, Iran.

Objective(s): Staphylococcus aureus can cause several infections. Its capability to form biofilm has been reported to be a vital property involved in the bacteria’s pathogenesis. Various genes contributing to biofilm formation have not yet been completely clarified. This study was designed to evaluate the factors influencing adherence and biofilm formation in S. aureus isolated from paediatric patients.Materials and Methods:One hundred and ninety-seven S. aureus isolates were obtained from pediatric patients and confirmed with phenotypic and molecular examinations. Antimicrobial susceptibility testing and biofilm formation were evaluated using standard methods. The genes encoding adhesion and virulence factors were investigated by the PCR method. Results:The most efficient antibiotics against S. aureus isolates were vancomycin and linezolid. Approximately, 54.2% of MSSA and 85.6% of MRSA isolates were biofilm producers according to the microtiter test. Our analysis indicated that MRSA isolates are better able to form biofilm compared with MSSA isolates. All isolates harbored clfA, fnbpA, icaA, icaB, icaC, and icaD, while clfB, fnbB, hlg, and pvl were detected in 99.5%, 42.1%, 97.5%, and 5.6% of isolates, respectively. In addition, a significant difference was found in fnhB gene and biofilm formation.Conclusion:Our findings showed a significant correlation between mecA and pvl genes and MRSA and biofilm formation in S. aureus isolates. Additionally, this study indicated the significant role of the fnhB gene as a major marker for S. aureus biofilm formation. Therefore, further experiments are warranted to exactly elucidate the function of the fnhB gene in the formation of biofilm.

Staphylococcus argenteus isolated from retail foods in China: Incidence, antibiotic resistance, biofilm formation and toxin gene profile

Staphylococcus argenteus is a novel species of coagulase-positive staphylococci which was separated from Staphylococcus aureus in 2014. It can threaten human health like S. aureus but can not identify with conventional biochemical or other phenotypic testing. From 2011 to 2016, 1581 S. aureus strains were isolated from 4300 samples from retail foods covering most provincial capitals in China. According to multilocus sequence typing (MLST) and PCR confirmation, 7.2% of isolates (114/1581) were confirmed as S. argenteus. The pathogen was distributed in 22 of 39 sampled cities and all food types. Interestingly, most S. argenteus positive samples were collected from coastal cities in South China. MLST detected 8 different sequence types (STs), including five new STs. CC2250 was the predominant lineage of S. argenteus, followed by CC1223. To further characterize the isolates, their antibiotic resistance, virulence genes, biofilm formation and biofilm-related genes were examined. The pvl gene was not detected in S. argenteus, and only 1 isolate (0.9%) was positive for the tsst-1 gene. For 18 enterotoxin genes, 16.7% (19/114) of isolates harboured more than three genes, whereas 70.2% (80/114) of isolates had none of the investigated genes. Penicillin and ampicillin were the major antibiotics to which the S. argenteus isolates were resistant, followed by tetracycline, kanamycin and fusidic acid. A total of 94.7% of isolates had the ability to produce biofilms and all isolates harboured icaA, fnbA, and fib genes. Other biofilm-related genes, such as eno, clfB, fnbB, and icaC, were also found in 99.1%, 92.1%, 88.6%, and 74.6% of isolates, respectively. This study is the first systematic investigation of the prevalence of S. argenteus in retail foods in China and shows their ubiquity in food. We also provide comprehensive surveillance of the incidence of S. argenteus in retail foods and information to enable more accurate and effective treatment of infections of this new species.

Correction: Prevalence and Characterization of Oxacillin Susceptible mecA-Positive Clinical Isolates of Staphylococcus aureus Causing Bovine Mastitis in India.

[This corrects the article DOI: 10.1371/journal.pone.0162256.].

Prevalence, clinical expression, invasiveness and outcome of Staphylococcus aureus containing Panton-Valentine leukocidin in children treated in a university hospital of Lithuania

Background: There is a high prevalence of Staphylococcus aureus virulence factor Panton-Valentine leukocidin (PVL) in North-East parts of Europe. The aim was to evaluate data regarding the PVL occurrences in Lithuania, determine the relationship with Methicillin resistant Staphylococcus aureus (MRSA), association with demographic and clinical conditions, invasiveness and severity of the disease in children treated in hospital Kauno klinikos (KK).Methods: We performed a prospective case-cohort single-center study on paediatric patients hospitalized from 2012 to 2015 to KK. We compared characteristics in PVL positive [SA-PVL(+)] and PVL negative [SA-PVL(–)] groups among non-invasive and invasive infections. Logistic regression was performed to detect PVL predicting factors and Cox regression was presented to define factors associated with admission to intensive care unit (ICU).Results: PVL was detected in 51.5%, MRSA in 7.0% and MRSA-PVL(+) in 4.8% of cases. In general, PVL was associated with older age comparing with SA-PVL(–) (median 8.5 vs. 4.0 years, p < .001). Skin and soft tissue infections were presented in 87.9% of all SA-PVL(+) cases. Invasive infections (44.7% vs. 12.1%, p < .001) and co-morbidities (20.5% vs. 2.9%, p < .001) were associated with SA-PVL(–) infections compared to SA-PVL(+), but ICU admission number was higher in invasive SA-PVL(+) cases comparing to invasive SA-PVL(–) cases (41.2% vs. 10.2%, p = .007).Conclusions: There was a high prevalence of pvl gene in patients treated in KK. SA-PVL(+) infections were associated with SSTI and were not common in invasive infections, but the invasive infections caused by SA-PVL(+) were related to severe disease progression and admission to ICU.

Molecular Characterisation of Methicillin-Resistant Staphylococcus aureus Isolated from Patients at a Tertiary Care Hospital in Hyderabad, South India

AbstractContext: Infections with methicillin-resistant Staphylococcus aureus (MRSA) greatly influence clinical outcome. Molecular characterisation of MRSA can help to predict their spread and to institute treatment and hospital protocols. Aim: The aim of this study is to understand the diversity of MRSA in a tertiary care hospital in Hyderabad, India. Settings and Design: Samples collected at Gandhi Medical College, Hyderabad, and designed to assess hospital-or community-associated MRSA (HA-MRSA or CA-MRSA). Subjects and Methods: MRSA were subjected to antibiotic susceptibility testing, pulsed-field gel electrophoresis (PFGE), spa typing, multi-locus sequence typing and staphylococcal cassette chromosome–mec (SCCmec) typing. Statistical Analysis Used: Discriminatory index and 95% confidence interval. Results: Of the 30 MRSA, (a) 18 and 12 were HA-MRSA and CA-MRSA, respectively, and (b) 23.3% and 6.6% displayed induced clindamycin and intermediate vancomycin resistance, respectively. Genetic diversity was evident from the presence of (a) 20 pulsotypes, (b) eight spa types, with the predominance of t064 (n = 9) and (c) seven sequence types (ST), with the preponderance of ST22 and ST8 (9 each). ST22 and ST8 were the most prevalent among HA-MRSA and CA-MRSA, respectively. SCCmec type IV was the most frequent (n = 8). 44.4% of HA-MRSA belonged to SCCmec IV and V, whereas 33.3% of CA-MRSA belonged to SCCmec I and III; 33.3% (5/15) of the isolates harbouring the pvl gene belonged to SCCmec IVC/H. Conclusions: ST8 was a dominant type along with other previously reported types ST22, ST239, and ST772 from India. The observations highlight the prevalence of genetically diverse clonal populations of MRSA, suggesting potential multiple origins.

Spa gene-based molecular typing of nasal methicillin-susceptible staphylococcus aureus from patients and health-care workers in a dialysis center in southeast Iran

ABSTRACTStaphylococcus aureus is an important infectious agent in hemodialysis patients. This study was conducted to determine the frequency of S. aureus nasal carriage in hemodialysis patients (HD) and health-care workers (HCW) at the main dialysis center of Bam city, located in southeast of Iran. In this cross-sectional study, a total of 52 nasal swabs were obtained from health-care workers and hemodialysis patients to detect methicillin-sensitive S. aureus (MSSA) isolates. The resistance to different antibacterial agents was determined by disk diffusion method. Also, Panton-Valentine Leukocidin (PVL) – encoding gene as well as Staphylococcal protein A (spa) type were determined. The nasal carriage rate of S. aureus was found to be 24.4% and 18.8% in patients on hemodialysis and health-care workers, respectively. Among identified isolates, no methicillin-resistant S. aureus (MRSA) was found. Only two MSSA isolates (16.7%) were resistant to trimethoprim/sulfamethoxazole. One isolate (8.6%) was positive for pvl gene. Moreover, 8 spa types were found. According to BURP analysis, six out of the 12 S. aureus isolates (50%) belonged to the same clone, indicating a prevalence of a major clone among MSSA in carriage, including patients and HCW. Mupirocin is still the appropriate drug for reducing nasal colonization in our setting. Accumulation of isolates from patients and staff in one spa clonal complex is alarming for the necessity of more serious infection control in this center. Therefore, it is necessary to screen patients and health-care workers as a health priority, in order to prevent cross transmission.

Study of susceptibility to antibiotics and molecular characterization of high virulenceStaphylococcus aureusstrains isolated from a rural hospital in Ethiopia.

We characterised 80 Staphylococcus aureus strains isolated from human patients with SSTIs at a rural hospital in Ethiopia. Susceptibility to antibiotic of all strains was tested. The MLST method was used to type and a phylogenetic analysis was conducted employing the sequences of 7 housekeeping genes. PCR amplification was used to investigate the presence of the following virulence genes in all strains: hla (α-haemolysin), tstH (toxic shock syndrome toxin), luk PV (Panton-Valentine leukocidin), fnbA (fibronectin binding protein A) and mecA (methicillin resistance). Most of the strains were resistant to penicillin and ampicillin, but only 3 strains were resistant to oxacillin, and 1 of them was a true MRSA. The MLST results showed a high diversity of sequence types (ST), 55% of which were new, and ST152 was the most prevalent. A phylogeny study showed that many of the new STs were phylogenetically related to other previously described STs, but bore little relationship to the only ST from Ethiopia described in the database. Virulence gene detection showed a high prevalence of strains encoding the hla, fnbA and pvl genes (98.77%, 96.3% and 72.84%, respectively), a low prevalence of the tst gene (13.58%) and a markedly low prevalence of MRSA (1.25%). S. aureus strains isolated from patients in a rural area in Ethiopia showed low levels of antibiotic resistance, except to penicillin. Moreover, this study reveals new STs in Eastern Africa that are phylogenetically related to other previously described STs, and confirm the high prevalence of the pvl gene and the low prevalence of MRSA on the continent.

Panton Valentine Gene in both Community-acquired and Healthcare-acquired Methicillin Resistant Staphylococcus aureus Isolates from Ain Shams University Hospitals

Abstract Background Methicillin-resistant Staphylococcus aureus (MRSA) is a major pathogen that is associated with both hospital and community infections. Panton Valentine leukocidin (PVL) is an important virulence factor of S. aureus that is considered by many authors a marker of community acquired MRSA (CA-MRSA). Aim of the Work This study aimed to determine the prevalence of PVL genes among healthcare acquired methicillin-resistant Staphylococcus aureus (HA-MRSA) and CA-MRSA isolates, and to test the hypothesis that PVL is a reliable marker of CA-MRSA isolates. Material and Methods This comparative cross-sectional study was done on fifteen community acquired methicillin resistant Staphylococcus aureus (CA-MRSA) and fifteen hospital acquired methicillin resistant Staphylococcus aureus (HA-MRSA), obtained from patients attending outpatient clinics, presenting with community-acquired pyogenic infections and patients with healthcare acquired pyogenic infections in Intensive Care Units (ICUs), during the period from May 2017 till February 2018. Clinical specimens included pus and different body fluids. Staphylococcus aureus was isolated and identified using conventional microbiological methods3. Isolates were then tested for methicillin resistance by culture on mannitol salt agar (MSA) with cefoxitin4. The presence of mecA and pvl genes in all MRSA isolates was subsequently detected by PCR5,6. Results Among 15 HA-MRSA isolates, mecA gene was positive in 40% (6/15) of isolates, while pvl gene was positive in 53.3% (8/15) of isolates. Among 15 CA-MRSA isolates, mecA gene was positive in 46.7% (7/15) of isolates, while pvl gene was positive in 26.7% (4/15) of isolates. Conclusion We conclude that pvl gene is not a sole genetic marker for diagnosis CA-MRSA, as there was no significant correlation between mecA that encodes for methicillin resistance and pvl genes among fifteen CA-MRSA isolates (P value =1).

Coagulase-negative staphylococci carrying cfr and PVL genes, and MRSA/MSSA-CC398 in the swine farm environment

This work aimed at characterizing four Staphylococcus aureus and 68 coagulase-negative staphylococci (CoNS), recovered from the air and liquid manure tank of two swine farms with intensive- and semi-extensive-production types, for their antimicrobial resistance pheno-/genotypes and their virulence gene content. Molecular typing was performed by spa typing, MLST, agr typing, and SCCmec typing, where applicable. Conjugation experiments were performed to assess the transferability of the linezolid resistance gene cfr, and its genetic environment was determined by Whole-Genome-Sequencing. The four S. aureus (intensive-production farm, IP-farm) were typed as t011-agrI-CC398-ST398, were scn-negative and two of them were methicillin-resistant (MRSA) with the mecA gene (SCCmec-V). Multidrug resistance was seen in 87 % of the CoNS. Statistically significant differences among the antimicrobial resistance rates of CoNS from the two farms were observed for cefoxitin, aminoglycosides, tetracycline, ciprofloxacin and trimethoprim-sulfamethoxazole. Eight methicillin-resistant CoNS, which were recovered from the IP-farm, carried the mecA gene. One S. simulans isolate was PVL-positive and three S. cohnii eta-positive. One S. equorum and one S. arlettae showed linezolid resistance and carried the cfr gene (IP-farm), which was non-transferable by conjugation into S. aureus. The cfr genetic context in both isolates was identical, with the lsa(B) gene located upstream of cfr. The environment of swine farms might contribute to the dissemination of CoNS that show multidrug resistance and harbor important virulence factors.

Screening of antibiotic resistance genes and virulence determinants of Staphylococcus aureus from skin infections

BackgroundVirulent strains of Staphylococcus aureus (S. aureus) express a series of virulence factors which cause severe infections such as skin and soft tissue infections which can be life-threatening. Additionally, extensive antibiotic resistance among nosocomial pathogens has left limited choices for their eradication. Our objective was investigation of antibiotic resistance and virulence determinants of S. aureus from skin infections. Materials and methodsTwo-hundred non-duplicate S. aureus isolates were collected from skin infections. Antibiotic susceptibility profile was evaluated using disc diffusion and methicillin resistance was confirmed. Biofilm formation was assessed by microtiter tissue plate assay. Determinants of virulence factors including hlA-α, tsst-1, pvl, PSM's, eta and etb genes and also the mupirocin resistance mup A gene and class I integron were detected by PCR. MLST was performed for isolates containing the pvl gene. ResultsThe age range of patients included 12–76 years (mean ± SD = 56.34 ± 5.43). MRSA (70/200) were isolated significantly higher among ages>50 years (p < .001) but not significantly different between both genders (p = .112). Prior antibiotic consumption and hospitalization were significantly associated with MRSA (81.42%) and MDR (84.28%) isolation. The existence of int1 gene (44.5%) was significantly higher in multidrug-resistant (MDR) isolates (p < .001). Thirty-six (18%) MDR isolates carried the mupA gene. The existence of mupA gene was significantly associated with prior hospitalization (n = 33, 91.66%, p < .0001) and antibiotic consumption (n = 30, 83.33%, p < .001). Predominant virulence determinant included PSMα (61.5%), followed by tsst-1 (17.5%), hla-α (9.5%), pvl (2.5%), eta (2.5%) and etb (1%). The rate of strong biofilm producers (totally 25%) was not significantly different between MRSA and MSSA. The existence of PSM-α gene was significantly higher among strong biofilm producers compared to biofilm non-producers (p = .002). PFGE exhibited no genetic relation among strains. ConclusionVirulence factors of S. aureus from skin infections contributed in severity of infection and biofilm formation. MDR phenotype was more common among older patients with history of hospitalization and prior antibiotic consumption. Mupirocin resistance has emerged among MDR and MRSA isolates. Hence suitable control strategies must be performed to inhibit the spread of these strains in healthcare and community settings.

Pathological profiles of systemic infections by Panton-Valentine leukocidin-positive, methicillin-resistant Staphylococcus aureus strains in a murine model.

Staphylococcus aureus is one of the most common pathogens in hospital environment and community. Panton-Valentine leukocidin (PVL) production is clinically associated with skin abscesses, soft tissues infections, bacteraemia and sepsis. This study aimed to investigate the effects of the presence of genes lukF/S-PV coding for PVL, in histological and haematological features during systemic infection, using a Swiss mice experimental model. Experiments were performed using 25 mice distributed into five experimental groups, intravenously inoculated with 50µl suspensions at density 1·0×107 CFU per ml of strains: methicillin-susceptible (MSSA) and pvl-negative strains isolated from nasal colonization; MSSA pvl-positive strains isolated from nasal colonization; methicillin-resistant (MRSA) and pvl-positive strains isolated from peripheral blood of a patient with severe pulmonary infection; and a MRSA pvl-positive strains isolated from a peripheral blood culture of a patient with bacteraemia. Haematological analysis was performed at 24, 48, 72 and 96h post-infection. Morphoanatomy and histopathological analyses were performed at 96h post-infection. For all S. aureus strains tested, the capability of intravenous dissemination and survival into mice tissues was demonstrated. Inflammatory processes at different levels were related to the presence of pvl genes, and included alterations in the format, size and colour of the organs. Staphylococcus aureus pvl-positive strains were detected in greater numbers in the organs of the infected animals. The pvl-positive strains isolated from blood cultures were capable to induce the greatest modifications in both haematological and histopathological profiles, and seemed to aggravate the systemic infections. These findings are valuable in characterizing infections caused by S. aureus in humans and murine.

Prevalence, molecular characteristics and antimicrobial susceptibility patterns of MRSA in hospitalized and nonhospitalized patients in Barbados.

The epidemiology and clonality of methicillin-resistant Staphylococcus aureus (MRSA) has not been investigated, as not much research or surveillance has been undertaken to identify and characterize the circulating MRSA strains in Barbados. Prevalence rates, molecular characteristics and antimicrobial susceptibility pattern of MRSA infections in hospitalized and nonhospitalized patients were investigated. A total of 293 isolates were included in the study, with 100 from the hospital and 193 from the public health laboratory. Isolates were collected over a period of 1 (2015–2016) and 3 years (2013–2016) respectively. MRSA was identified using standard microbiologic techniques and was further analysed by multiplex PCR for the presence of the spa, mec gene complex typing and PVL genes (lukS-PV and lukF-PV). A prevalence rate of 19.7% was calculated for those hospitalized. All hospital isolates were sensitive to vancomycin, rifampin, linezolid and cotrimoxazole (trimethoprim/sulfamethoxazole), whilst 82% were sensitive to clindamycin. The PVL gene was detected in 76% of hospital isolates. In the community isolates, resistance was observed in erythromycin (100%), ciprofloxacin (97.4%), clindamycin (13%) and cotrimoxazole (5.7%). There was no resistance to vancomycin. The PVL gene was detected in 97.9% of the isolates, the mecA gene in only 2.1% and the mecC gene in 0%. Most MRSA isolates were community acquired in both settings, and the antimicrobial susceptibility profile was similar, suggesting transmission of community-associated MRSA into the hospital environment. Further harmonization of antimicrobial policy for the treatment of MRSA (and by extension other pathogens) should be implemented to quell ongoing transmission. We found that 93.4% of MRSA in Barbados treated in the primary healthcare system were sensitive to cotrimoxazole. By typing MRSA isolates and drawing interferences on transmission on the basis of genetic relatedness, transmission pathways may be tracked. Further studies must be performed for this high level of comprehensiveness so that with the surveillance of MRSA, effective strategies may be developed to prevent or limit transmission.

Staphylococcal species less frequently isolated from human clinical specimens - are they a threat for hospital patients?

BackgroundCoagulase-negative staphylococci belonging to S. haemolyticus, S. hominis subsp. hominis, S. simulans, and S. warneri are often described as etiological factors of infections. Staphylococci are a phylogenetically coherent group; nevertheless, there are differences among the species which may be important to clinicians.MethodsWe investigated selected virulence factors and antibiotic resistance that were phenotypically demonstrated, the presence and expression of genes encoding the virulence factors, and the type of the SCCmec cassette.ResultsThe differences between the tested species were revealed. A great number of isolates produced a biofilm and many of them contained single icaADBC operon genes. Clear differences between species in the lipolytic activity spectrum could be related to their ability to cause various types of infections. Our studies also revealed the presence of genes encoding virulence factors homologous to S. aureus in the analysed species such as enterotoxin and pvl genes, which were also expressed in single isolates of S. simulans and S. warneri. S. haemolyticus and S. hominis subsp. hominis isolates were resistant to all clinically important antibiotics including ß-lactams. The identified SCCmec cassettes belonged to IV, V, VII, and IX type but most of the detected cassettes were non-typeable. Among the investigated species, S. hominis subsp. hominis isolates accumulated virulence genes typical for S. aureus in the most efficient way and were widely resistant to antibiotics.ConclusionsOur results clearly indicated significant differences between the tested species, which might be a result of the horizontal gene transfer (HGT) and can lead to the formation and selection of multi-drug resistant strains as well as strains with new virulence features. Such strains can have a new clinical relevance.

Genome Sequence of a Highly Virulent pvl-positive Vancomycin-intermediate-resistant Staphylococcus aureus Sequence Type 30.

BackgroundStaphylococcus aureus isolates expressing the Panton-Valentine Leukocidin (PVL) have been related to a wide range of diseases. Recently, pvl-positive community-associated methicillin-resistant S. aureus belonging to USA1100 (ST30/CC30/SCCmec IV) lineage has emerged in Brazilian hospitals.Objective The aim of this work was to sequence the genome of a pvl-positive USA1100 Vancomycin-Intermediate-Resistant S. aureus (VISA) isolate from Rio de Janeiro, Brazil.Methods The 13420 genome was sequenced using the HiSeq 2500 platform. The draft genome, plasmids annotation, and genome analysis were performed using RAST. Comparison of the relative pvl gene expression of six S. aureus isolates was performed by qRT-PCR.Results The isolate presented the ϕPVL phage codifying for the H2b PVL protein isoform, and another prophage carrying a PVL variant named lukF and lukS-PV.2. The 13420 genome presented a high number of virulence determinants, such as genes codifying for serine-protease proteins, enterotoxins (egc), the immune evasion cluster (IEC), adhesion proteins, spermine/spermidine acetyltransferase gene (blt), superantigen-like proteins, as well as the ica operon. Point mutations at vraS, tcaA, and tcaB genes were detected. Moreover, the PVL mRNA relative expression of the 13420 isolate was five times higher than mRNA PVL levels of the USA300/ST8 reference strain.Conclusion We described for the first time the genome sequence of a VISA isolate harboring two pvl-associated genes and other virulence factors that may improve the USA1100/ST30 lineage fitness and impact its pathogenicity and spreading at Brazilian hospitals.

Prevalence and characterization of Staphylococcus aureus isolates from subclinical bovine mastitis in southern Xinjiang, China

Staphylococcus aureus is one of the major pathogens causing mastitis in dairy herds. The colonization of dairy cows and subsequent contamination of raw milk by S. aureus, especially strains exhibiting multidrug resistance and biofilm-forming and toxin-producing abilities, remains an important issue for both dairy farmers and public health. In this study, we investigated the prevalence, antimicrobial susceptibility, biofilm formation, and genetic diversity of S. aureus from subclinical bovine mastitis in dairy farms located in southern Xinjiang, China. Sixty-five isolates from 84 subclinical mastitic milk samples were identified as S. aureus. The resistance rates to penicillin, erythromycin, clindamycin, tetracycline, gentamicin, linezolid, rifampicin, quinupudin-dafupudin, ciprofloxacin, norfloxacin, and chloramphenicol were 58.5, 44.6, 40.0, 18.5, 12.3, 10.8, 9.2, 6.2, 4.6, 4.6, and 1.5%, respectively. All isolates were susceptible to cefoxitin, sulfamethoxazole-trimethoprim, and vancomycin. Isolates from farm A showed a significantly higher resistance rate to tetracycline (16.9%) than those from farm B (1.5%). The most frequently detected virulence factors were hla (96.9%, 63/65) and hlb (100.0%, 65/65). The percentage rates of the staphylococcal enterotoxin genes sea, sec, sed, seg, seh, sei, and sej in S. aureus isolates were 4.6, 33.8, 27.7, 3.1, 41.5, 41.5, and 7.7%, respectively. The percentage rate of the sec gene in isolates from farm B (30.8%) was significantly higher than that of farm A (3.1%). The percentage rates of the tsst and pvl genes in S. aureus isolates were 26.2 and 40.0%. The percentage rate of the pvl gene in isolates from farm B (32.3%) was significantly higher than that of farm A (7.7%). The adhesion molecules fnbA, fnbB, clfA, clfB, and cna were detected in 21 (32.3%), 23 (35.4%), 65 (100.0%), 65 (100.0%), and 65 (100.0%) isolates, respectively. The percentage rates of the icaA, sarA, tcaR, ccp, luxS, and sigB genes in S. aureus isolates were 69.2, 100.0, 86.2, 95.4, 84.6, and 100.0%, respectively. The fnbB and icaA genes were more frequently detected in isolates from farm A (29.2 and 40.0%, respectively) than those from farm B (6.2 and 29.2%, respectively). The luxS gene was more often found in isolates from farm B (50.8%) than those from farm A (33.8%). Using the microplate method, 61.5, 26.2, and 10.8% of the isolates showed weak, moderate, and strong biofilm-forming abilities, respectively. Different clonal complex (CC) and spa-types were identified, including CC81, CC398, CC88, CC5405, and CC5406. Importantly, in this study we report for the first time 41 new sequence types (ST) among 44 distinct ST. These results indicated high genetic diversity of S. aureus involved in subclinical bovine mastitis in southern Xinjiang, China. The results also showed that S. aureus from subclinical bovine mastitis cases in southern Xinjiang, China, were mainly resistant to β-lactams, erythromycin, and clindamycin. Also, biofilm- and adhesion-related genes, which are increasingly known as important virulence factors in the pathogenesis of S. aureus infections, were detected at a high rate. This study could help identify predominant clones and provide surveillance measures to decrease or eliminate S. aureus contamination in raw milk of dairy cows with subclinical mastitis.

Characteristics of community-acquired methicillin-resistant Staphylococcus aureus associated with wound infections in Tehran, Iran: High prevalence of PVL+ t008 and the emergence of new spa types t657, t5348, and t437 in Iran

Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) is a major concern in Iran. The present study aimed to determine characteristics and the distribution of spa types of CA-MRSA strain isolated in wound infections. We investigated 120 CA-MRSA isolates by spa typing, virulence gene detected and in vitro antimicrobial susceptibility was assayed. Of the 120 CA-MRSA clinical isolates, 89.1% were confirmed as multidrug resistance (MDR). Virulence genes pvl, tst, eta, and etb were detected in 33.3%, 12.5%, 4.2%, and 1.7% of the isolates, respectively. All strains were distributed across 14 distinct spa types including t008 (21.7%), t790 (11.7%), t030 (9.1%), t044 (8.3%), t021 (7.5%), t019 (6.7%), t015 (6.7%), t852 (5.8%), t091 (5%), t437 (4.2%), t084 (4.2%), t005 (3.3%), t5348 (3.3%) and t657 (2.5%). PVL-carrying strains belonged to t008 (21.6%), t044 (5%), t021 (1.7%), (t019 1.7%), t005 (0.8%) and t852 (2.5%). t008 spa type was found to be associated with high-level mupirocin-resistant phenotype while t790 and t084 spa type were related to high-level mupirocin-resistant phenotype. Importantly, spa types t030 (n = 2) and t790 (n = 1) displayed resistance to fusidic acid. Mupirocin resistance was found for 20% of isolates which of these, 6.7% and 13.3% demonstrated high- and low-level mupirocin resistance phenotype respectively. Out of 120 strains examined, MS, iMLSB and cMLSB phenotypes were detected in 16 (13.3%), 20 (16.7%) and 39 (32.5%) isolates, respectively. The present findings clarified a genetic diversity of CA-MRSA strains in our area with the majority of t008 spa type highlighting the need for more effective surveillance/infection control strategies to keep track and prevention of the emerging spa types.

Molecular Characterization of Staphylococcus aureus Isolates Associated with Nasal Colonization and Environmental Contamination in Academic Dental Clinics.

Aim: To determine the genetic makeup of methicillin-sensitive/methicillin-resistant Staphylococcus aureus (MSSA/MRSA) from nasal colonization and environmental contamination in dental clinics. Materials and Methods: Nasal swabs from students and health care workers and environmental swabs were obtained at two academic dental clinics in the United Arab Emirates. The StaphyType DNA microarray-based assay was used for molecular characterization. Results: Forty-eight S. aureus isolates were identified phenotypically (nasal: n = 43; environmental: n = 5), but 6 of these were assigned to S. argenteus by genotyping. These were CC(argenteus)2596, CC(arg)2250-MSSA, CC(arg)2250-MSSA-(Panton Valentine leukocidin [PVL]+) (n = 2), and CC(arg)2198-MSSA (n = 2). MRSA nasal colonization rate was 5.4% (n/N = 8/146) with the following strain affiliations: CC5-MRSA-[IV+fus+ccrAB], "Maltese Clone"; CC6-MRSA-IV, "WA MRSA-51"; CC22-MRSA-IV (PVL+/tst+); CC22-MRSA-[IV+fus+ccrAA/(C)]; and two each of CC5-MRSA-[VI+fus] and CC97-MRSA-[V/VT+fus]. The SCC-borne fusidic acid resistance (fusC) gene was detected in MRSA (n = 5) and MSSA (n = 1). Some MSSA strains, CC1-MSSA-[fus+ccrAB1] and ST1278-MSSA-[ccrA1], harbored recombinase genes. A CC30-MSSA harbored ACME locus/arc-genes, while ST1278-MSSA-[ccrA1] had an ACME-III element. Enterotoxin genes were commonly carried, but tst-1 gene was found in only CC22, CC30, and CC34 strains, while pvl genes were identified in CC(arg)2250 and CC22-MRSA-IV. Of the 51 noncoagulase staphylococci (CoNS) identified, 18 were mecA positive. Conclusion: The findings demonstrate the first report of rare strains (ST1278 MSSA, CC(arg)2198, CC(arg)2596, and PVL+CC(arg)2250) in our region. Detection of MSSA with recombinase genes and ACME loci alongside mecA-positive CoNS is of clinical significance as this could provide a milieu for acquisition and transfer of SCC-elements, either with different ACME types, with fusC or the mecA gene resulting in conversion of MSSA into MRSA.

Should we leave the paper currency? A microbiological examination.

ABSTRACTObjetivesPathogens can be transmitted to banknotes due to the personal unhygienic habits. The aim of study was to find the possible pathogens on the banknotes circulating in the market and also to present their antibacterial resistance and their various virulence factors using genotypic and phenotypic methods.Material and methodsA total of 150 samples of banknotes were randomly collected between August 2017 and March 2018. VITEK systems were used for identification and antimicrobial susceptibility testing respectively. Antimicrobial resistance genes (mecA, van, extended-spectrum β-lactamase [ESBL] and carbapenemases) and staphyloccoccal virulence genes (staphyloccoccal enterotoxins [SEs], pvl, and tsst-1) were determined using with real-time PCR.ResultsStaphylococcus aureus, coagulase-negative staphylococci (CoNS), Enterococcus spp., Gram-negative enteric bacteria, non-fermentative Gram-negative bacteria and Candida spp. were detected 48%, 54.7%, 56%, 21.3%, 18.7%, and 4%, respectively. Methicillin-resistant S. aureus, vancomycin-resistant enterococci and ESBL producing Gram-negative were found 46.8%, 1.3%, and 28.7%, respectively. Pvl, tsst-1, and SEs genes were found in a 2.8/4.9%, 1.4/1.2%, and 100/ 87.8% of the S. aureus/CoNS strains, respectively. The sea gene was found the most common enterotoxigenic gene. blaTEM, blaSHV, blaCTX-M-2, blaCTX-M-1, blaKPC, and blaOXA-48 were found 55.8%, 46.5%, 41.2%, 18.6%, 18.6%, and 18.6%, respectively in Gram-negative strains.ConclusionThese results is very important to highlight hygienic status of paper currencies. This can be considered as an indication that banknotes may contribute to the spread of pathogens and antimicrobial resistance. Therefore, we may need to start using alternative products instead of banknotes.