Study of virulence factors in coagulase-negative staphylococci isolated from newborns

Coagulase-negative staphylococci (CNS) have been identified as the etiological agent in various infections and are currently the microorganisms most frequently isolated in nosocomial infections. However, little is known about the virulence factors produced by CNS that contribute to the pathogenesis of infections caused by these microorganisms. The study of CNS isolated from infectious processes of newborns hospitalized in the Neonatal Unit of the Hospital of the Botucatu Medical School, UNESP, indicated Staphylococcus epidermidis as the most frequently isolated species (77.8%), which was also associated with clinically significant situations. The analysis of virulence factors revealed the production of slime in 20 (17.1%) of all CNS samples isolated and the synthesis of a broad spectrum of enzymes and toxins, including hemolysins (19.6%), lipase (17.1%), lecithinase (3.4%), DNAse (15.4%), thermonuclease (7.7%), and enterotoxin A, B or C (37.6%). Taking into consideration that the etiological importance of CNS has often been neglected, the present investigation confirmed that these microorganisms should not be ignored or classified as mere contaminants.     

Susceptibility to selected coagulase-negative chemotherapeutic agents of methicillin resistant staphylococci isolated from clinical materials in 1997-1998

The study was aimed at assessment of the sensitivity of methicillin-resistant coagulase-negative staphylococci isolated from clinical material in 1997/1998 to selected chemotherapeutic agents. The investigated material comprised 96 methicillin-resistant coagulase-negative staphylococci from hospital and ambulatory infections isolated during the period from April 1997 to May 1998. Species affiliation was determined by classical identification methods and commercial diagnostic tests for identification of staphylococci. Methicillin resistance was determined by agar disk-diffusion method and screening. Sensitivity to chemotherapeutics was determined by agar disk-diffusion method and agar dilution methods. All the investigated strains were sensitive to nitrofurantoin, furazolidone and vancomycin. To teicoplanin--the second glycopeptide antibiotic--84% strains were sensitive, whereas the percentages of resistant and moderately sensitive strains amounted to 5.2% and 10.4%, respectively. 85% and 82% of coagulase-negative staphylococcal strains were sensitive to fusidic acid and mupirocin. Considerable differences were noted with respect to sensitivity to aminoglycoside group antibiotics. About 35% of strains were sensitive to gentamicin, and 90% sensitive to netilmicin. Ca. 40% of coagulase-negative staphylococci were resistant both to cotrimoxazole and trimethoprim, which, in view of 98% resistance to the second component of cotrimoxazole, may be associated with the activity of only one of the components of the drug--trimethoprim.     

Genetic similarity of vancomycin resistant strains of Enterococcus faecium isolated from clinical specimens

Twenty vancomycin resistant E. faecium strains (VRE) isolated from patients of three different hospital wards in 2005-2008 were examined. The strains originated from patients of intensive therapy, urological and internistic wards. The chosen wards differ significantly in their specificity. In all cases the presence of o vanA and lack of vanB, vanD, vanE and vanG genes and were found. Strains were compared by using RFLP-PFGE, the reference method for molecular typing of VRE. One group including fourteen strains showing similarity higher than 79.5% was distinguished. This group was divided into subgroups. The greatest similarity was found among strains from patients of intensive therapy ward. Two subgroups of strains showing similarity more than 93.3%, of four strains each were identified. The similarity between these two subgroups was 79.5%. Most strains from other two wards showed less than 79.5% similarity and they could be recognised as not related. Only one strain from internal ward and two strains from urologic ward were similar in 82.1 - 86.4% to one of subgroups of strains originated from intensive therapy.     

Proposed virulence factors among coagulase-negative staphylococci isolated from two healthy populations

Proposed virulence factors, including multiple antibiotic resistance and slime-mediated adherence, among coagulase-negative staphylococci colonizing healthy individuals from two different study populations were examined. Resistance to methicillin was more commonly seen than initially anticipated. In addition, adherence characteristics, as quantitated by a microtiter plate method, were very similar to those of strains of coagulase-negative staphylococci causing nosocomial infections.     

Plasmid profile and antibiotic resistance in coagulase-negative staphylococci isolated from polluted water

Four different species of coagulase-negative staphylococci (CNS) were isolated from polluted waters in Fez, Morocco and found to be Staphylococcus simulans, Staph. lenticus, Staph. hyicus and Staph. xylosus . Eight isolates belonging to these four species were analysed for their plasmid content. Southern blot hybridizations were performed to define the resistance determinants of the plasmids harboured by these species. These determinants were found to be carried mainly by Class I staphylococcal plasmids (1–5 kb). A plasmid (4·3 kb) carrying a tetracycline resistance gene was present in five isolates from all identified species. Plasmids carrying a chloramphenicol resistance gene were more frequently encountered and found to be of different sizes. Plasmids carrying erythromycin, neomycin, and streptomycin resistance genes were less frequent and were the same size. The results indicate that the occurrence of multi-resistant CNS in polluted waters may constitute a reservoir for disseminating antibiotic-resistance into the community.     

CPRMethicillin resistant coagulase-negative staphylococci isolated from South Korean ducks exhibiting tremor

Background

We describe coagulase-negative staphylococci (CoNS) isolates collected from ducklings exhibiting tremor in South Korea over the period of 2010 to 2011. Screening of antimicrobial susceptibility and analysis of SCCmec elements of CoNS were also investigated.

Results

Staphylococcus cohnii was the most frequent staphylococcus (9 isolates) and S. sciuri (4 isolates), S. lentus (3 isolate), S. simulans (1 isolate) and S. epidermidis (1 isolate) were also detected. Among the 15 antimicrobials tested in this study, resistance against oxacillin (15 isolates, 83.3%) was most frequently observed, but only one isolate (SNUDS-1) possessed mecA. This isolate was shown to possess SCCmec type III; the type 3 ccr complex and the class A mec complex.

Conclusions

Based on these results, isolate SNUDS-1 was shown to possess SCCmec type III; the type 3 ccr complex and the class A mec complex. Although the SCCmec type III is not predominant in human, MR-CoNS (Methicillin resistance Coagulase-negative staphylococci) in food animals should be monitored to prevent the dissemination of antimicrobial resistance genes and resistant pathogens to the community.

     

Antibiotic resistance and molecular characterization of clinical isolates of methicillin-resistant coagulase-negative staphylococci isolated from bacteremic patients in oncohematology

Polymerase chain reaction (PCR) amplification of antibiotic resistance genes as well as staphylococcal cassette chromosome mec (SCCmec) typing and pulsed-field gel electrophoresis (PFGE) of SmaI macrorestriction fragments of genomic DNA were used to characterize 45 methicillin-resistant coagulase-negative staphylococci (MRCoNS) isolates responsible of bacteremia recovered in patients at the Bone Marrow Transplant Centre of Tunisia in 1998–2007. Among the 45 MRCoNS isolates, Staphylococcus epidermidis was the most prevalent species (75.6%) followed by Staphylococcus haemolyticus (22.2%) and Staphylococcus hominis (2.2%). Extended susceptibility profiles were generated for MRCoNS against 16 antimicrobial agents. Out of 45 mecA-positive strains, 43 (95.6%) were phenotypically methicillin-resistant and two (4.4%) were methicillin-susceptible. The msr(A) was the most prevalent gene (13 isolates; 48.1%) among erythromycin-resistant isolates. The erm(C) was found alone in seven (25.9%) or in combination with both erm(A) and erm(B) in two (7.4%) isolates. The aac(6′)-Ie-aph(2″)-Ia was the most prevalent gene among aminoglycoside-resistant isolates, detected alone in 14 isolates (33.3%) isolates, in combination with ant(4′)-Ia in 18 (42.8%) isolates, in combination with aph(3′)-IIIa in four (9.5%) or with both ant(4′)-Ia and aph(3′)-IIIa in two (4.7%) isolates. The ant(4′)-Ia was detected in three (7.1%) isolates and the aph(3′)-IIIa in one (2.4%) isolate. Among tetracycline-resistant isolates, six (85.7%) strains harbored the tet(K) gene and one (14.3%) strain carried tet(K) and tet(M) genes. SCCmec types IV (31%) and III (24.5%), the most prevalent types detected, were found to be more resistant to non-β-lactam antibiotics. A wide diversity of isolates was observed by PFGE among MRCoNS.     

Identification and methicillin resistance of coagulase-negative staphylococci isolated from nasal cavity of healthy horses

The aim of this study was an analysis of the staphylococcal flora of the nasal cavity of 42 healthy horses from 4 farms, along with species identification of CoNS isolates and determination of resistance to 18 antimicrobial agents, particularly phenotypic and genotypic methicillin resistance. From the 81 swabs, 87 staphylococci were isolated. All isolates possessed the gap gene but the coa gene was not detected in any of these isolates. Using PCR-RFLP of the gap gene, 82.8% of CoNS were identified: S. equorum (14.9%), S. warneri (14.9%), S. sciuri (12.6%), S. vitulinus (12.6%), S. xylosus (11.5%), S. felis (5.7%), S. haemolyticus (3.4%), S. simulans (3.4%), S. capitis (1.1%), S. chromogenes (1.1%), and S. cohnii subsp. urealyticus (1.1%). To our knowledge, this was the first isolation of S. felis from a horse. The species identity of the remaining Staphylococcus spp. isolates (17.2%) could not be determined from the gap gene PCR-RFLP analysis and 16S rRNA gene sequencing data. Based on 16S-23S intergenic transcribed spacer PCR, 11 different ITS-PCR profiles were identified for the 87 analyzed isolates. Results of API Staph were consistent with molecular identification of 17 (19.5%) isolates. Resistance was detected to only 1 or 2 of the 18 antimicrobial agents tested in the 17.2% CoNS isolates, including 6.9% MRCoNS. The mecA gene was detected in each of the 5 (5.7%) phenotypically cefoxitin-resistant isolates and in 12 (13.8%) isolates susceptible to cefoxitin. In total, from 12 horses (28.6%), 17 (19.5%) MRCoNS were isolated. The highest percentage of MRCoNS was noted among S. sciuri isolates (100%).     

Sensitivity to antibacterial preparations of the pathogenic staphylococci isolated from carriers]

After discontinuation of the use of antibiotics for sanation of the carriers of pathogenic staphylococci among the staff of a maternity unit, the number of the strains resistant to penicillin, streptomycin, chlortetracycline, tetracycline, levomycetin, erythromycin, monomycin and neomycin decreased 1.3, 2.9, 10,4.3,5.5,1.5,3.9 and 2.4 times respectively. The differences were statistically reliable. The causes of the antibiotic resistance decrease require further investigation.     

Adhesion of coagulase-negative staphylococci to biomaterials

The adhesion of two Staphylococcus epidermidis strains and one Staphylococcus saprophyticus strain on to poly(tetrafluorethylene-co-hexafluorpropylene) (FEP)-fluorocarbon and cellulose acetate was studied in vitro. Both S. epidermidis strains showed a more hydrophobic character than the encapsulated S. saprophyticus as determined by the bacterial affinity towards xylene. Staphylococcus epidermidis showed a significantly higher adhesion on to the hydrophobic FEP than S. saprophyticus. The adhesion of staphylococci on to the more hydrophilic cellulose acetate was always low. Treatment of S. epidermidis with pepsin or extraction with aqueous phenol yielded cells with a decreased hydrophobicity, which resulted in a decreased adhesion on to FEP. Cells with a decreased hydrophobicity showed a lower rate of reaggregation in suspension. The hydrophobicity and the adhesion on the FEP of S. epidermidis were not affected by exposure to a subminimal inhibitory concentration of penicillin. The strong interaction between S. epidermidis and FEP, which appeared not to be influenced by the age or the metabolic stage of the bacteria, is mainly caused by hydrophobic bonding.     

Identification of coagulase-negative staphylococci from farm animals

The species identity of 661 strains of coagulase-negative staphylococci isolated from the skin and nares of cattle, pigs, poultry, goats and sheep was determined. They belonged either to the novobiocin-sensitive species Staphylococcus hyicus, Staph. simulans, Staph. epidermidis, Staph. haemolyticus and Staph. warneri or to the novobiocin-resistant species Staph. sciuri, Staph. lentus, Staph. xylosus, Staph. cohnii. Staph. saprophyticus and Staph. gallinarum ; twenty-one strains remained unidentified. The staphylococcal flora of the farm animals studied differed markedly from that associated with man; several species which do not occur in man were isolated and novobiocin-resistant strains, which occur infrequently in man, were present in large numbers in animals. Two simplified schemes for the identification of staphylococci from farm animals and man are presented.     

Identification of coagulase-negative staphylococci from farm animals

The species identify of 661 strains of coagulase-negative staphylococci isolated from the skin and nares of cattle, pigs, poultry, goats and sheep was determined. They belonged either to the novobiocin-sensitive species Staphylococcus hyicus, Staph. simulans, Staph. epidermidis, Staph. haemolyticus and Staph. warneri or to the novobiocin-resistant species Staph. sciuri, Staph. lentus, Staph. xylosus, Staph. cohnii, Staph. saprophyticus and Staph. gallinarum; twenty-one strains remained unidentified. The staphylococcal flora of the farm animals studied differed markedly from that associated with man; several species which do not occur in man were isolated and novobiocin-resistant strains, which occur infrequently in man, were present in large numbers in animals. Two simplified schemes for the identification of staphylococci from farm animals and man are presented.     

Sensitivity to antibiotics of bacterial strains isolated from clinical specimens during the years 1985-1986

Sensitivity of 312 strains of staphylococci, 386 strains of streptococci and 1193 strains of aerobic gram-negative bacilli to the selected antibiotics was tested. These strains were isolated from the clinical material at the Clinical Hospital No. 1 in Warsaw within 1985-1986. Staphylococci were sensitive to pristinamycin, cefazolin, fusidic acid, oxacillin, and clindamycin. In 1986, a decrease in the number of strains sensitive to these antibiotics, except cefazolin, was seen. In case of streptococci the most active proved chloramphenicol and gentamicin but a significant decrease in the percentage of sensitive strains was also noted in 1986. The highest number of gram-negative bacilli was sensitive to amikacin, colistin, nalidixic acid, pipemidic acid, and gentamicin. In 1986, a decrease in the percentage of sensitive strains was noted. Amikacin and colistin were the most active against Pseudomonas spp. while amikacin and nalidixic and pipemidic acids--against Proteus spp. Comparison of the results with those obtained in 1981-1984 has shown that the sensitivity of staphylococci changed the most significantly and this change was unfavourable. Gentamicin and amikacin remained the most active against gram-negative bacilli while amikacin and colimycin against Pseudomonas spp. In case of anaerobes the majority of strains was sensitive to chloramphenicol, tetracycline and clindamycin. Metronidazole was active against high percentage of Clostridium spp. and all gram-negative bacilli while the percentage of gram-positive bacilli and cocci was sensitive to metronidazole.     

Antimicrobial susceptibility of coagulase-negative staphylococci isolated from bovine mastitis between 2003 and 2008 in Korea

A total of 1,444 coagulase-negative staphylococci (CNS) isolates from bovine mastitic milk samples collected during 2003-2008 in Korea were identified to the species level. Of 14 species identified, S. simulans, S. haemolyticus, and S. sciuri accounted for over 60% of the isolates. All the CNS isolates were tested for susceptibility to eight antimicrobials commonly used in dairy cattle. With a few exceptions, similar resistance patterns were observed among the CNS species: penicillin and ampicillin showed the lowest activity while amikacin, cephalothin, and gentamycin were highly effective. About 39% (557/1,444) of the CNS isolates were pan-susceptible, while 12% (175/1,444) showed resistance to four or more antimicrobials tested.     

Antibiotic resistance in coagulase-negative staphylococci isolated from Cope''s gray treefrogs (Hyla chrysoscelis)

Microcin J25 (MccJ25) is a cyclic peptide of 21 unmodified amino acid residues produced by a fecal strain of Escherichia coli. It has previously been shown that the antibiotic activity of this peptide is mainly directed to Enterobacteriaceae, including several pathogenic E. coli, Salmonella and Shigella strains. In this paper we show that MccJ25 acts on the cytoplasmic membrane of Salmonella newport cells producing alteration of membrane permeability, and the subsequent gradient dissipation, that initiate the inhibition of process, such as oxygen consumption. These results, taken together with our in vitro observations [Rintoul et al. (2000) Biochim. Biophys. Acta 1509, 65-72], strongly suggest that the disruption of the cytoplasmic membrane gradient is closely related to the bactericidal activity of MccJ25 in S. newport.