Methicillin-Resistant Staphylococcus aureus ST80 Induce Lower Cytokine Production by Monocytes as Compared to Other Sequence Types

Methicillin-resistant Staphylococcus aureus (MRSA) remains an important cause of nosocomial and community-associated infections due to its ability to produce toxins and evade host’s immune responses. The aim of the present study was to investigate the association of monocytes immune response in terms of cytokines produced after inoculation with different MRSA clones. Thirty-one clinical MRSA strains were selected on the basis of clonal types, accessory gene regulator (agr) groups and toxin genes carriage. Isolates were identified as S. aureus by Gram stain, catalase, coagulase production and PCR for nuc gene. The presence of mecA, lukS/lukF-PV (Panton-Valentine Leukocidin) and tst (Toxic Shock Syndrome Toxin-1) genes, as well as, the determination of agr groups was performed by PCR. Clonality was investigated by means of multi-locus sequence typing (MLST). Peripheral blood mononuclear cells were stimulated with live bacterial cells for 45 min at a ratio of 1:10. Cells were incubated for 10 h and supernatants were collected. The levels of Tumor Necrosis Factor alpha (TNFa), IL-1b, IL-8, IL-6, IL-12p40, IL-10, interferon-gamma (IFN-γ) and IL-2, were measured by Human Cytokine Multiplex Immunoassay kit. Thirteen strains were tst and 12 lukS/lukF-PV-positive. Seven strains belonged to ST80 and ST225, five to ST30 and ST239, while the remaining seven isolates were grouped together as “other.” Strains belonging to ST80 induced statistically lower levels of TNFa, IL-1b, IL-8, IL-6, IL-10, IFN-γ, and IL-2. PVL-positive strains classified into ST80 clone induced statistically lower concentrations of most cytokines as compared to PVL-positive strains belonging to other clones, tst-positive strains and toxin-negative ones. Strains of agr3 group belonging to ST80 induced statistically lower concentrations of most tested cytokines as compared to agr3 strains not-belonging to ST80, agr2 or agr1. This low induction of immune response by MRSA ST80 cannot be attributed to the presence of neither lukS/lukF-PV nor agr3

Early KPC-Producing Klebsiella pneumoniae Bacteremia among Intensive Care Unit Patients Non-Colonized upon Admission

Among 140 patients colonized by KPC-producing Klebsiella pneumoniae (KPC-Kp) between fourth and seventh day of Intensive Care Unit stay, 24 developed bacteraemia immediately after colonization. Colistin-resistance of the colonizing isolate was the factor significantly associated with early KPC-Kp bacteraemia (P < 0.001; OR 6.6, 95% CI 2.4–18.4), a worrisome finding since infections by colistin-resistant isolates is associated with increased mortality due to limited remaining therapeutic options

Biofilm synthesis and its relationship with genetic characteristics in clinical methicillin-resistant staphylococci

Staphylococcus aureus can cause a broad range of infections, including skin infections, pneumonia and bacteraemia. Coagulase-negative staphylococci (CNS), mainly S. epidermidis, have also emerged as important pathogens, especially in immunocompromised patients or those with prosthetic devices, such as intravascular catheters or biomaterials. Of great importance in the initiation of these infections is the ability of staphylococci to adhere to various surfaces, such as host tissues and prosthetic devices and to form biofilm. The staphylococcal adhesins are encoded by a number of genes such as fnbA (S. aureus fibronectin binding protein A), sasG (S. aureus surface protein G), aap (S. epidermidis accumulation associated protein), bhp (Bap homologue protein) and fbe (fibrinogen binding protein epidermidis). In this study, 106 methicillin-resistant S. aureus (MRSA), 145 methicillin-resistant S. epidermidis (MRSE) and 70 non-epidermidis methicillin-resistant CNS (MR-CNS; 58 S. haemolyticus, 10 S. hominis and two S. lugdunensis) were compared in terms of biofilm formation, antimicrobial resistance, clonal distribution and adhesin genes carriage. Isolates were classified into pulsotypes by PFGE and assigned to sequence types by MLST. In total, 121/321 isolates (37.7%) produced biofilm and 219 (68.2%) carried ica operon. The majority was multidrug resistant (94.7%) and carried one or more adhesin genes. MRSE and all other MR-CNS prevailed in biofilm formation (P < 0.001) and antimicrobial resistance (P < 0.05) as compared to MRSA. MRSE also prevailed in ica carriage compared to the other methicillin-resistant staphylococci (P ≤ 0.007) Among MRSE, isolates from bacteraemias prevailed in biofilm formation (P = 0.031), whereas, strains from prosthetic device-associated infections carried more frequently aap (P = 0.003). Even though PFGE showed genetic diversity among MRSE, MLST revealed three major clones (ST2, ST5, ST16). MRSA isolates were less diverse, with five PFGE types and, among them, one major PFGE type (C) consisting of 77/106 strains (72.6%). MLST identified five sequence types: ST5, ST30, ST80, ST225 and ST239. One major PFGE type (h) was identified in S. haemolyticus. A clonal relationship was found concerning fnbA carriage in MRSA, ica carriage in MRSE, and antimicrobial susceptibility in both groups reinforcing the aspect of clonal expansion in hospital settings

Spread of Tst–Positive Staphylococcus aureus Strains Belonging to ST30 Clone among Patients and Healthcare Workers in Two Intensive Care Units

Staphylococcus aureus is a major cause of infections. Toxic shock syndrome toxin (TSST-1) and Panton–Valentine leukocidin (PVL) are associated with severe clinical syndromes. S. aureus colonizing isolates recovered from healthcare workers and patients in the intensive care unit (ICU) of a university hospital comprising Group A were compared with those from adult non-ICU carriers (Group B). mecA, lukS/lukF-PV (Panton–Valentine leukocidin, PVL), and tst (toxic shock syndrome toxin) gene carriage was detected by PCR. Clones were identified in all methicillin-resistant S. aureus (MRSA) and toxin-positive methicillin-susceptible strains (MSSA) by pulsed-field gel electrophoresis (PFGE), agr groups, and multi locus sequencing typing (MLST). Group A included 90 S. aureus isolates, whereas Group B 53. PVL was more frequently found among MRSA vs. MSSA (p < 0.001) and in strains of Group B as compared to Group A (p < 0.001), consistent with the spread of ST80-IV. Higher incidence of tst gene carriage was identified among MSSA vs. MRSA (P 0.005) belonging mainly to ST30, and Group A vs. Group B (P 0.002). The wide dissemination of ST80-IV mainly in the community is responsible for a high percentage of PVL-positive MRSA, while silent spread of tst-positive S. aureus clones among ICU patients and personnel poses a threat of hospital transmission and possible severe infections

Trends of Bloodstream Infections in a University Greek Hospital during a Three-Year Period: Incidence of Multidrug-Resistant Bacteria and Seasonality in Gram-negative Predominance

The aim of the study was to assess the epidemiology, the incidence of multidrug-resistant bacteria and bloodstream infections’ (BSIs) seasonality in a university hospital. This retrospective study was carried out in the University General Hospital of Patras, Greece, during 2011–13 y. Blood cultures from patients with clinical presentation suggestive of bloodstream infection were performed by the BacT/ALERT System. Isolates were identified by Vitek 2 Advanced Expert System. Antibiotic susceptibility testing was performed by the disk diffusion method and E-test. Resistance genes (mecA in staphylococci; vanA/vanB/vanC in enterococci; blaKPC/blaVIM/blaNDM in Klebsiella spp.) were detected by PCR. In total, 4607 (9.7%) blood cultures were positive from 47451 sets sent to Department of Microbiology, represent­ing 1732 BSIs. Gram-negative bacteria (52.3%) were the most commonly isolated, followed by Gram-positive (39.5%), fungi (6.6%) and anaerobes bacteria (1.8%). The highest contamination rate was observed among Gram-positive bacteria (42.3%). Among 330 CNS and 150 Staphylococcus aureus, 281 (85.2%) and 60 (40.0%) were mecA-positive, respectively. From 113 enterococci, eight were vanA, two vanB and two vanC-positives. Of the total 207 carbapenem-resistant Klebsiella pneumoniae (73.4%), 202 carried blaKPC, four blaKPC and blaVIM and one blaVIM. A significant increase in monthly BSIs’ incidence was shown (R2: 0.449), which may be attributed to a rise of Gram-positive BSIs (R2: 0.337). Gram-positive BSIs were less frequent in spring (P < 0.001), summer (P < 0.001), and autumn (P < 0.001), as compared to winter months, while Gram-negative bacteria (P < 0.001) and fungi (P < 0.001) were more frequent in summer months. BSIs due to methicillin resistant S. aureus and carbapenem-resistant Gram-negative bacteria increased during the study period. The increasing incidence of BSIs can be attributed to an increase of Gram-positive BSI incidence, even though Gram-negative bacteria remained the predominant ones. Seasonality may play a role in the predominance of Gram-negative’s BSI

Trends of Bloodstream Infections in a University Greek Hospital during a Three-Year Period: Incidence of Multidrug-Resistant Bacteria and Seasonality in Gram-negative Predominance

The aim of the study was to assess the epidemiology, the incidence of multidrug-resistant bacteria and bloodstream infections’ (BSIs) seasonality in a university hospital. This retrospective study was carried out in the University General Hospital of Patras, Greece, during 2011–13 y. Blood cultures from patients with clinical presentation suggestive of bloodstream infection were performed by the BacT/ALERT System. Isolates were identified by Vitek 2 Advanced Expert System. Antibiotic susceptibility testing was performed by the disk diffusion method and E-test. Resistance genes (mecA in staphylococci; vanA/vanB/vanC in enterococci; blaKPC/blaVIM/blaNDM in Klebsiella spp.) were detected by PCR. In total, 4607 (9.7%) blood cultures were positive from 47451 sets sent to Department of Microbiology, represent­ing 1732 BSIs. Gram-negative bacteria (52.3%) were the most commonly isolated, followed by Gram-positive (39.5%), fungi (6.6%) and anaerobes bacteria (1.8%). The highest contamination rate was observed among Gram-positive bacteria (42.3%). Among 330 CNS and 150 Staphylococcus aureus, 281 (85.2%) and 60 (40.0%) were mecA-positive, respectively. From 113 enterococci, eight were vanA, two vanB and two vanC-positives. Of the total 207 carbapenem-resistant Klebsiella pneumoniae (73.4%), 202 carried blaKPC, four blaKPC and blaVIM and one blaVIM. A significant increase in monthly BSIs’ incidence was shown (R2: 0.449), which may be attributed to a rise of Gram-positive BSIs (R2: 0.337). Gram-positive BSIs were less frequent in spring (P < 0.001), summer (P < 0.001), and autumn (P < 0.001), as compared to winter months, while Gram-negative bacteria (P < 0.001) and fungi (P < 0.001) were more frequent in summer months. BSIs due to methicillin resistant S. aureus and carbapenem-resistant Gram-negative bacteria increased during the study period. The increasing incidence of BSIs can be attributed to an increase of Gram-positive BSI incidence, even though Gram-negative bacteria remained the predominant ones. Seasonality may play a role in the predominance of Gram-negative’s BSI