In order to characterize the immunogenicity and immunoprotection of the Staphylococcus aureus (S. aureus) surface protein Clumping factor A (ClfA), we amplified clfa genes from S. aureus Newman strain, Wood46 strain and HLJ23-1. The clfa gene from Newman strain was subsequently inserted into pQE-30 vector and the recombinant plasmid was transformed into Escherichia coli strain M15 (pREP4). The recombinant ClfA protein was expressed and purified. Then, we immunized mice with the purified recombinant protein. The antibody level and the concentration of cytokines were measured by enzyme-linked immunosorbent assay. Finally, immunized mice were challenged with S. aureus Newman, Wood46 and HLJ23-1. These results suggested that clfa gene sequences were highly conserved, and the recombinant ClfA was expressed correctly with good antigenicity. The antibody titer and the concentration of cytokines in the immunized groups increased significantly (P < 0.05) compared with control, and the mice in the immunized groups were protected against the challenge strains to some extent. These results showed that the ClfA had high immunogenicity and immunoprotective potential.
Animals ; Coagulase ; genetics ; immunology ; metabolism ; Escherichia coli ; genetics ; metabolism ; Immunization ; Mice ; Recombinant Proteins ; genetics ; immunology ; metabolism ; Staphylococcus aureus ; metabolism ; pathogenicity

Necrotizing fasciitis (NF) is a deep infection of the subcutaneous tissue that progressively destroys fascia and fat; it is associated with systemic toxicity, a fulminant course, and high mortality. NF most frequently develops from trauma that compromises skin integrity, and is more common in patients with predisposing medical conditions such as diabetes mellitus, atherosclerosis, alcoholism, renal disease, liver disease, immunosuppression, malignancy, or corticosteroid use. Most often, NF is caused by polymicrobial pathogens including aerobic and anaerobic bacteria. NF caused by Staphylococcus aureus as a single pathogen, however, is rare. Here we report a case of NF that developed in a healthy woman after an isolated shoulder sprain that occurred without breaking a skin barrier, and was caused by Staphylococcus aureus as a single pathogen.
*Arm ; Coagulase/metabolism ; Fasciitis, Necrotizing/*etiology/microbiology/pathology/surgery ; Female ; Humans ; Middle Aged ; Shoulder Joint/*injuries ; Sprains and Strains/*complications ; Staphylococcal Infections/*etiology/microbiology ; Staphylococcus aureus/enzymology/isolation & purification

To compare immunological characteristics of Extracellular fibrinogen-binding protein (Efb) and Clumping factor A (CfA) of Staphylococcus aureus, we constructed two prokaryotic expression vector pET28a-Efb and pET28a-ClfA. After prokaryotical expression and purification, Efb and ClfA were used to immunize experimental animal. After the second immunization the antisera were collected and the antibody titers, the bacteria binding activity and adhesion inhibition activity of these antisera were detected by enzyme linked immunosorbent assay, adhesion inhibition assay and challenge. Both Efb and ClfA had Fibrinogen binding activity whereas the former had better Fibronectin binding activity. The bacteria binding capability of antisera of rabbits immunized with ClfA was better than that with Efb (P < 0.01). Both antisera of Efb and ClfA could inhibit adherence activity of Staphylococcus aureus to Fibrinogen and Fibronectin adherence compare to the control group (P < 0.01), and Efb had better adhesion inhibition activity than ClfA. The antibody titer of immunized group could reach 1:40 500. After the second immunization, both Efb and ClfA had good protective efficacy. This result constitutes a good foundation for Staphylococcus aureus subunit vaccine development.
Animals ; Antibodies, Bacterial ; blood ; Bacterial Adhesion ; Bacterial Proteins ; immunology ; Cattle ; microbiology ; Coagulase ; immunology ; Enzyme-Linked Immunosorbent Assay ; Fibrinogen ; metabolism ; Genetic Vectors ; Immune Sera ; immunology ; Immunization ; Rabbits ; Staphylococcal Infections ; immunology ; Staphylococcus aureus

We investigated the distribution of commensal staphylococcal species and determined the prevalence of multi-drug resistance in healthy cats and dogs. Risk factors associated with the carriage of multi-drug resistant strains were explored. Isolates from 256 dogs and 277 cats were identified at the species level using matrix-assisted laser desorption ionisation-time of flight mass spectrometry. The diversity of coagulase-negative Staphylococci (CNS) was high, with 22 species in dogs and 24 in cats. Multi-drug resistance was frequent (17%) and not always associated with the presence of the mecA gene. A stay in a veterinary clinic in the last year was associated with an increased risk of colonisation by multi-drug resistant Staphylococci (OR = 2.4, 95% CI: 1.1~5.2, p value LRT = 0.04). When identifying efficient control strategies against antibiotic resistance, the presence of mechanisms other than methicillin resistance and the possible role of CNS in the spread of resistance determinants should be considered.
Animals ; Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/genetics/metabolism ; Cat Diseases/epidemiology/*microbiology ; Cats ; Coagulase/genetics/metabolism ; Dog Diseases/epidemiology/*microbiology ; Dogs ; *Drug Resistance, Multiple, Bacterial ; Female ; Male ; Prevalence ; Risk Factors ; Seasons ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/veterinary ; Staphylococcal Infections/epidemiology/microbiology/*veterinary ; Staphylococcus/classification/genetics/*isolation & purification ; Switzerland/epidemiology

BACKGROUND: We evaluated the performance of the BD MAX StaphSR Assay (SR assay; BD, USA) for direct detection of Staphylococcus aureus and methicillin resistance not only in S. aureus but also in coagulase-negative Staphylococci (CNS) from positive blood cultures. METHODS: From 228 blood culture bottles, 103 S. aureus [45 methicillin-resistant S. aureus (MRSA), 55 methicillin-susceptible S. aureus (MSSA), 3 mixed infections (1 MRSA+Enterococcus faecalis, 1 MSSA+MRCNS, 1 MSSA+MSCNS)], and 125 CNS (102 MRCNS, 23 MSCNS) were identified by Vitek 2. For further analysis, we obtained the cycle threshold (Ct) values from the BD MAX system software to determine an appropriate cutoff value. For discrepancy analysis, conventional mecA/mecC PCR and oxacillin minimum inhibitory concentrations (MICs) were determined. RESULTS: Compared to Vitek 2, the SR assay identified all 103 S. aureus isolates correctly but failed to detect methicillin resistance in three MRSA isolates. All 55 MSSA isolates were correctly identified by the SR assay. In the concordant cases, the highest Ct values for nuc, mecA, and mec right-extremity junction (MREJ) were 25.6, 22, and 22.2, respectively. Therefore, we selected Ct values from 0-27 as a range of positivity, and applying this cutoff, the sensitivity/specificity of the SR assay were 100%/100% for detecting S. aureus, and 97.9%/98.1% and 99.0%/95.8% for detecting methicillin resistance in S. aureus and CNS, respectively. CONCLUSIONS: We propose a Ct cutoff value for nuc/mec assay without considering MREJ because mixed cultures of MSSA and MRCNS were very rare (0.4%) in the positive blood cultures.
Anti-Bacterial Agents/pharmacology ; Bacteremia/diagnosis/microbiology ; Coagulase/metabolism ; Humans ; Methicillin-Resistant Staphylococcus aureus/drug effects/genetics/*isolation & purification ; Microbial Sensitivity Tests ; Oxacillin/pharmacology ; Reagent Kits, Diagnostic ; Staphylococcus/drug effects/enzymology/genetics/isolation & purification ; Staphylococcus aureus/drug effects/genetics/*isolation & purification

Bacteremia ; etiology ; Catheterization ; adverse effects ; Child ; Coagulase ; metabolism ; Cross Infection ; etiology ; Drug Resistance, Bacterial ; drug effects ; Endocarditis, Bacterial ; etiology ; Humans ; Methicillin ; pharmacology ; Quinolones ; pharmacology ; Staphylococcal Infections ; complications ; drug therapy ; microbiology ; Staphylococcus ; classification ; drug effects ; pathogenicity ; Urinary Tract Infections ; etiology ; Vancomycin ; pharmacology

BACKGROUNDGram-positive bacteria such as Staphylococcus aureus have been a common cause of infection among liver transplant (LT) recipients in recent decades. The understanding of local epidemiology and its evolving trends with regard to pathogenic spectra and antibiotic susceptibility is beneficial to prophylactic and empiric treatment for LT recipients. This study aimed to investigate etiology, timing, antibiotic susceptibility and risk factors for multidrug resistant (MDR) Gram-positive coccal bacteremia after LT.

METHODSA cohort analysis of prospectively recorded data was performed to investigate etiologies, timing, antibiotic susceptibility and risk factors for MDR Gram-positive coccal bacteremia in 475 LT recipients.

RESULTSIn 475 LT recipients in the first six months after LT, there were a total of 98 episodes of bacteremia caused by Gram-positive cocci in 82 (17%) patients. Seventy-five (77%) bacteremic episodes occurred in the first post-LT month. The most frequent Gram-positive cocci were methicillin-resistant coagulase-negative staphylococcus (CoNS, 46 isolates), methicillin-resistant Staphylococcus aureus (MRSA, 13) and enterococcus (34, E. faecium 30, E. faecalis 4). In all Gram-positive bacteremic isolates, 59 of 98 (60%) were MDR. Gram-positive coccal bacteremia and MDR Gram-positive coccal bacteremia predominantly occurred in patients with acute severe exacerbation of chronic hepatitis B and with fulminant/subfulminant hepatitis. Four independent risk factors for development of bacteremia caused by MDR Gram-positive coccus were: LT candidates with encephalopathy grades II - IV (P = 0.013, OR: 16.253, 95%CI: 1.822 - 144.995), pre-LT use of empirical antibiotics (P = 0.018, OR: 1.029, 95%CI: 1.002 - 1.057), post-LT urinary tract infections (P < 0.001, OR: 20.340, 95%CI: 4.135 - 100.048) and abdominal infection (P = 0.004, OR: 2.820, 95%CI: 1.122 - 10.114). The main infectious manifestations were coinfections due to gram-positive cocci and gram-negative bacilli.

CONCLUSIONSMethicillin-resistant CoNS and enterococci are predominant pathogens among LT recipients with Gram-positive coccal bacteremia. Occurrences of Gram-positive coccal bacteremia may be associated with the severity of illness in the perioperative stage.

Anti-Bacterial Agents ; pharmacology ; Bacteremia ; etiology ; microbiology ; Coagulase ; metabolism ; Drug Resistance, Multiple, Bacterial ; Enterococcus ; drug effects ; enzymology ; physiology ; Gram-Positive Bacterial Infections ; enzymology ; microbiology ; transmission ; Humans ; Liver Diseases ; microbiology ; Liver Transplantation ; adverse effects ; Risk Factors ; Staphylococcal Infections ; enzymology ; microbiology ; transmission ; Staphylococcus ; drug effects ; enzymology ; physiology

Neonatal sepsis remains one of the most important causes of death and co-morbidity in very-low-birth-weight (VLBW) infants. The aim of this study was to determine the current incidences of early-onset sepsis (EOS) and late-onset sepsis (LOS), the distribution of pathogens, and the impact of infection on co-morbidities in VLBW infants. We analyzed the data including sepsis episode from 2,386 VLBW infants enrolled in Korean Neonatal Network from January 2013 to June 2014. We defined EOS as a positive blood culture occurring between birth and 7 days of life and LOS after 7 days of life. Sepsis was found in 21.1% of VLBW infants. The risk of sepsis was inversely related to birth weight and gestational age. EOS was found in only 3.6% of VLBW infants, however the mortality rate was as high as 34.1%. EOS was associated with the increased odds for bronchopulmonary dysplasia and intraventricular hemorrhage. The vast majority of EOS was caused by Gram-positive organisms, particularly coagulase-negative staphylococci (30.6%). LOS developed in 19.4% of VLBW infants with a 16.1% mortality rate. Pathogens in LOS were dominated by coagulase-negative staphylococci (38.3%). Twenty-five percent and fifty percent of first LOS episode occurred after 12 days and 20 days from birth, respectively. Younger and smaller VLBW infants showed the earlier occurrence day for the 25% of first LOS episode. This study provides a recent nationwide epidemiology of sepsis in VLBW infants in Korea. Based on this study, successful strategies to reduce infections would improve survival and reduce morbidity.
Coagulase/metabolism ; Databases, Factual ; Gestational Age ; Gram-Negative Bacteria/isolation & purification ; Gram-Positive Bacteria/isolation & purification ; Humans ; Incidence ; Infant, Newborn ; *Infant, Very Low Birth Weight ; Kaplan-Meier Estimate ; Republic of Korea/epidemiology ; Risk Factors ; Sepsis/*epidemiology/microbiology/mortality ; Staphylococcus/enzymology/isolation & purification