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PubMed1990

Phage-typing of coagulase-negative staphylococci. Factors influencing typability.

Factors influencing the phage-typability of coagulase-negative Micrococcaceae have been studied in 2,778 clinical isolates comprising A) 209 consecutive isolates from one laboratory, B) 2,107 clinical strains submitted for phage-typing for epidemiological reasons, and C) 462 strains representing all isolates of presumed clinical significance found in two laboratories during one month. The reproducibility was acceptable at duplicate repeated typing of the same strains as well as by typing epidemiologically-related pairs of strains from the same patient. Strains of Staphylococcus haemolyticus were seldom typable, whereas strains of S. epidermidis and S. hominis had a higher typability. Methicillin-resistant strains and other multiple-resistant strains were rarely typable (11-13%). The typability was higher among susceptible strains (36%) and strains resistant to penicillin only (43-50%). The typability of strains of the same species and antibiotic-resistance pattern differed between hospitals compared and decreased markedly over the years for multiple-resistant S. epidermidis isolates.

Bacteriophage Typing↗

PubMed1988

In-vitro induction of resistance in coagulase-negative staphylococci to vancomycin and teicoplanin.

Twenty strains of coagulase-negative staphylococci (18 strains of Staphylococcus haemolyticus and two of S. epidermidis) were serially subcultured in broth media containing subinhibitory concentrations (half the MIC) of either vancomycin or teicoplanin. The MIC of the antibiotic was again measured after five passages in antibiotic-containing broth. The organisms were then subcultured in broth containing antibiotic concentrations half of the new MIC value. The experiment was terminated after 25 passages. Only one strain developed a four-fold increase in vancomycin MIC. On the other hand, 16 strains developed four-fold MIC increases to teicoplanin.

Anti-Bacterial Agents↗

PubMed1984

Bacterial and mycoplasmal flora of the middle ear of laboratory rats with otitis media.

Purulent otitis media was confirmed at necropsy in 35 laboratory rats, 14 of which had shown clinical signs of middle ear infection. The bacterial flora of both the affected and grossly normal middle ears was similar, often mixed, and comprised mainly of Staphylococcus haemolyticus, Pasteurella pneumotropica and Corynebacterium spp. Twenty-one isolates of Mycoplasma spp. were recovered from the middle ears of 11 rats with purulent otitis media and one normal rat. Fourteen of the isolates were further identified as either Mycoplasma arthriditis (eight isolates) or Mycoplasma pulmonis (six isolates). In ears affected with otitis media, mycoplasmas were isolated significantly more often from subclinical than from clinical cases, while Pasteurella pneumotropica was isolated significantly more often from clinical than subclinical cases.

Animals↗

PubMed1992

Phage pattern and antibiotic resistance pattern of coagulase-negative staphylococci obtained from immunocompromised patients.

A total of 152 coagulase-negative staphylococcal strains were isolated from clinical samples of 14 patients hospitalized after bone-marrow transplantation in a specialized hospital ward in Hungary, during an 18-month period between 1987 and 1989. Two species, Staphylococcus epidermidis and Staphylococcus haemolyticus, predominated (each, 45%). Using Pulverer and co-workers' phage set for typing, 68% of the isolates were typable; 16 phage patterns were observed. A characteristic long pattern with phages Ph10/Ph13/Ph15/U4/U15/U16/U20/U33 /U46 appeared only in S. epidermidis, among 5 of 11 colonized patients (8.5% of all strains). Single lysis with phage Ph13 was observed in 7 of the 14 patients (49% of all strains), in species S. capitis, S. epidermidis, S. haemolyticus, S. hominis, and S. warneri. In S. haemolyticus, non-typable strains predominated (66%); this character occurred only in 2% among other species. The strains colonizing the immunocompromised patients differed from each other in phage pattern, antibiotic resistance pattern, and/or slime production. No hospital infection was suggested. On the other hand, high incidence of two well-definable phage patterns raises some relationship between phage receptors or some regulatory systems in phage multiplication and factors responsible for special colonization as common surface properties.

Bacterial Typing Techniques↗

PubMed1990

Evaluation of restriction endonuclease fingerprinting of chromosomal DNA and plasmid profile analysis for characterization of multiresistant coagulase-negative staphylococci in bacteremic neonates.

A procedure was developed for restriction endonuclease fingerprinting (REF) of the chromosomal DNA of coagulase-negative staphylococci. A total of 48 isolates comprising 29 Staphylococcus epidermidis and 19 Staphylococcus haemolyticus isolates from blood and mucocutaneous sites of 15 premature neonates were characterized by REF, plasmid profile (PP) analysis, antimicrobial susceptibility testing, biotyping, and slime production. On the basis of REF analysis of chromosomal DNA, the 48 coagulase-negative staphylococcal isolates were subdivided into 10 subgroups, whereas PP analysis subdivided the strains into 20 distinct subgroups. REF analysis of total DNA (i.e., chromosome plus plasmid) resulted in the same 20 subgroups as were subdivided by PP analysis. The high discriminatory power of PP analysis was associated with the variability of plasmid content in coagulase-negative staphylococcal strains isolated during the outbreak. REF patterns were found to be stable both in vitro and in vivo. Isolates carried from 2 to 10 plasmids that ranged in molecular size from 0.9 to 39.5 megadaltons. Plasmids were disseminated among the coagulase-negative staphylococci, regardless of the genetic relatedness of their chromosomal DNAs. Hence, a lack of correlation existed between the grouping of isolates by REF analysis of chromosomal DNA and the grouping by PP analysis. There were one and two distinct chromosomal patterns among 4 of 4 blood cultures and 15 of 15 mucocutaneous cultures of S. haemolyticus, respectively. In contrast, a higher proportion of distinct chromosomal patterns was found for S. epidermidis in blood cultures (7 of 11 cultures) compared with those identified for isolates in mucocutaneous cultures (6 of 18 cultures). In summary, REF analysis of chromosomal DNA, rather than total DNA, is a useful marker for epidemiological investigations of coagulase-negative staphylococci. PP analysis can also be used to provide additional epidemiological information regarding the most recent genetic events.

Coagulase↗

PubMed1998

In vivo presence of capsular polysaccharide in coagulase-negative staphylococci of ovine origin.

Three strains of coagulase-negative staphylococci identified as Staphilococcus warnery, Staphylococcus simulans and Staphylococcus haemolyticus were examine for evidence of capsular polysaccharide, in vitro by negative staining with India ink, and in vivo by transmission electron microscopy. In vitro, unstained materials surrounded clusters or single cocci. In vivo, capsula materials were surrounding phagocytosed bacteria cells. These capsular materials were either closely or loosely bound to bacterial cells, or projecting like "pili" from the surface of the cell wall. The evidence suggests that capsular polysaccharide is a common characteristic of coagulase-negative staphylococci causing prolonged intramammary infection of sheep.

Animals↗

PubMed2006

Clonality of slime-producing methicillin-resistant coagulase-negative staphylococci disseminated in the neonatal intensive care unit of a university hospital.

Methicillin-resistant coagulase-negative staphylococci (MR-CNS) (n = 132), isolated from pre-term neonates, were analysed to determine their antibiotic resistance patterns, clonal distribution, biofilm production and the presence of the ica operon. All MR-CNS were multiresistant, and 89% produced slime. A major clone was identified (77 isolates) among 115 Staphylococcus epidermidis isolates. Ten of 16 Staphylococcus haemolyticus isolates also belonged to a single clone. Most (80%) slime-positive isolates possessed all the ica genes tested, while the remaining 23 (20%) had a variety of gene combinations. The entire ica cluster was detected in three of 15 slime-negative isolates. One major and two minor slime-positive, multiresistant MR-CNS clones had disseminated among hospitalised pre-term neonates.

Adhesins, Bacterial↗

PubMed1997

Avoparcin, a glycopeptide used in animal foods: antimicrobial spectrum and potency tested against human isolates from the United States.

Avoparcin is a glycopeptide antimicrobial that is widely used as a growth promoter in animal feeds in portions of Western Europe. Recently, concern has emerged about the possible contribution of avoparcin use to the emergence of glycopeptide resistance in enterococci. Relatively little information exists on the spectrum of activity and potency of avoparcin. We studied the activity of avoparcin compared to vancomycin, teicoplanin, and 3 other antimicrobials against 814 recent human clinical isolates, including Staphylococcus spp. (240 strains), Streptococcus spp. (90 strains), and Enterococcus spp. (60 strains), using reference susceptibility test methods. Our results indicate that avoparcin was less potent than either vancomycin or teichoplanin against staphylococci (MIC50, 4 micrograms/ mL). There was a good correlation of avoparcin MICs with the MICs for vancomycin and teichoplanin for most species; however, the avoparcin MICs for Enterococcus spp. of the vanB phenotype were quite variable (MIC range, 2 to > 16 micrograms/mL). For Staphylococcus haemolyticus, high avoparcin MICs (> or = 16 micrograms/mL) were associated with oxacillin resistance. These results are relevant to the debate concerning the appropriateness of continued use of avoparcin as a growth promoter in animal husbandry. In particular, avoparcin as a glycopeptide with limited potency against some staphylococci seems to represent a theoretically greater risk for selecting glycopeptide resistance among staphylococci, but may not represent any greater threat for the selection of resistance in enterococci.

Animal Feed↗

PubMed1996

A particle agglutination assay for rapid identification of heparin binding to coagulase-negative staphylococci.

The heparin-binding properties of six different species of coagulase-negative staphylococci were examined by a particle agglutination assay. Heparin (mol. wt 4000-6000), mildly treated with sodium periodate, was covalently coupled to amino-modified latex beads (0.72 micron diameter). The particle agglutination assay was validated by comparing results with the adhesion (percentage binding of adherent cells) of coagulase-negative staphylococcal strains to heparinised microtitration plates. Of 38 different coagulase-negative staphylococcal strains tested, 30 showed agglutination reactivity with heparin-coated latex beads. Strains of different coagulase-negative staphylococcal species agglutinated heparin-coated latex beads to various extents (e.g., cells of Staphylococcus haemolyticus strains reacted more strongly than cells of S. epidermidis strains). The agglutination reaction was significantly inhibited by fucoidan, suramin, lambda-carrageenan and other sulphated compounds, but not by non-sulphated carbohydrate polymers such as hyaluronic acid. Agglutination of staphylococcal cells with heparin-coated latex beads was completely blocked by a cell-surface extract. These results suggest that structures responsible for heparin binding are exposed on the cell surface.

Agglutination Tests↗

PubMed2006

DNA fingerprinting of a multi-resistant coagulase-negative staphylococci isolated from biomaterials in dialysis services.

BACKGROUND: Coagulase-negative staphylococci (CoNS) have emerged as an important opportunistic pathogen isolated in hospital. Several species of CoNS have been implicated in human infections and disease especially in patients with poor health status. METHODS: A total of 71 clinical strains of CoNS were isolated from dialysis fluid and needles in a dialysis unit and characterized. Susceptibility to antibiotics, biofilm production and molecular typing by pulsed-field gel electrophoresis (PFGE) were achieved. RESULTS: The main isolated CoNS strains were Staphylococcus epidermidis (45%), Staphylococcus hominis (14%) and Staphylococcus haemolyticus (12.7%). The susceptibility profile of all strains revealed high resistance level to penicillin and oxacillin. PCR detection of oxacillin resistance gene (mecA gene) revealed a higher percentage of positive strains than the classic test (ATB Staph). Slime production test was positive in 60.6% of CoNS strains. PFGE analysis showed the presence of 69 restriction profiles clustered in 56 patterns. CONCLUSIONS: Profiles of all isolates were generally heterogeneous, suggesting independent circulation with some evidence of cross-transmission.

Biocompatible Materials↗

PubMed1998

Determination of the chromosomal relationship between mecA and gyrA in methicillin-resistant coagulase-negative staphylococci.

mecA, the gene that mediates methicillin resistance, and its accompanying mec locus DNA, insert near the gyrA gene in Staphylococcus aureus. To investigate whether there is a similar relationship between mecA and gyrA in coagulase-negative staphylococci (CNS), mecA- and gyrA-specific DNA fragments were used to probe methicillin-resistant isolates of Staphylococcus epidermidis (MRSE) (n = 11) and Staphylococcus haemolyticus (MRSH) (n = 11). The gyrA probe hybridized to the same SmaI DNA fragment as the mecA probe in all strains tested. However, since the size of the SmaI fragments containing mecA and gyrA varied from 73 to 600 kb, the distance between the two genes was determined more precisely. Cloned mecA or gyrA fragments plus vector sequences each containing a SmaI site were introduced into the chromosome of three isolates each of MRSE and methicillin-resistant S. aureus (MRSA), and the sizes of the generated SmaI fragments were determined by pulsed-field gel electrophoresis. The distance between gyrA and mecA was found to be between 38 and 42 kb in both MRSE and MRSA, and the two genes were in the same relative orientation in all strains. Restriction fragment length polymorphism (RFLP) patterns around the gyrA gene in CNS were identical, but species specific, for all 10 MRSE and 10 MRSH isolates examined. In contrast, 8 of 11 methicillin-susceptible S. epidermidis isolates and 7 of 7 methicillin-susceptible S. haemolyticus isolates had different gyrA RFLP patterns. These data show that mecA is site and orientation specific, relative to gyrA, in both MRSE and MRSA. In addition, the local environment around gyrA in methicillin-resistant CNS, in contrast to methicillin-susceptible isolates, is similar, suggesting clonality or the requirement for specific DNA sequences with which the mec complex must interact for chromosomal integration to occur.

Bacterial Proteins↗

PubMed2000

Patterns of multidrug resistance among methicillin-resistant hospital isolates of coagulase-positive and coagulase-negative staphylococci collected in the international multicenter study RESIST in 1997 and 1998.

The primary purpose of the multicenter international study "RESIST" was to obtain an update on the degree of multidrug resistance among methicillin-resistant staphylococci collected from a geographically diverse sample. A total of 3,307 staphylococcal isolates were recovered from single patients and primarily from clinical specimens that were collected at 20 collaborating regional health centers located in several countries in Europe, Asia, and Latin America during a 3- to 4-month period each in 1997 and 1998. All strains were deposited at the Laboratory of Molecular Genetics at ITQB/UNL in Oeiras, Portugal, for quality control and for testing by microbiological and molecular typing techniques; the Laboratory of Microbiology at The Rockefeller University serving as organizational center. The majority of strains, 3,100, were methicillin-resistant, of which 1,749 were coagulase positive (methicillin-resistant Staphylococcus aureus, MRSA), and 1,351 were coagulase negative (methicillin-resistant coagulase negative staphylococci, MRCNS). The overall frequency of drug resistance traits among the 1,749 MRSA strains was high (over 70% and up to and over 90% of the strains) to ciprofloxacin, erythromycin, clindamycin, gentamicin, and tetracycline, and was somewhat less frequent to sulfamethoxazole-trimethoprim (45%), chloramphenicol (30%), and rifampin (38%). None of the 3,307 staphylococcal isolates showed reduced susceptibility to vancomycin except for a single methicillin-resistant coagulase-negative isolate. The great majority of staphylococci were also susceptible to the new antimicrobial Synercid. In contrast, resistance to teicoplanin was significant among methicillin-resistant strains of coagulase-negative staphylococci, particularly among Staphylococcus haemolyticus. MRSA isolates showed marked geographic variation in their patterns of multiresistance, most likely reflecting the properties of unique multiresistant MRSA clones dominant in the hospitals that provided the MRSA isolates from the various geographic areas. The multiresistance patterns of MRSA strains and strains of methicillin-resistant coagulase-negative staphylococci originating at the same country source also showed striking differences, suggesting that resistance to antimicrobial agents emerged under different antibiotic pressures in these bacterial species.

Anti-Bacterial Agents↗

PubMed2004

Comparison of the BBL CHROMagar Staph aureus agar medium to conventional media for detection of Staphylococcus aureus in respiratory samples.

Screening for Staphylococcus aureus has become routine in certain patient populations. This study is the first clinical evaluation of the BBL CHROMagar Staph aureus agar (CSA) medium (BD Diagnostics, Sparks, Md.) for detection of S. aureus in nasal surveillance cultures and in respiratory samples from cystic fibrosis (CF) patients. S. aureus colonies appear mauve on CSA. Other organisms are inhibited or produce a distinctly different colony color. S. aureus was identified from all media by slide coagulase, exogenous DNase, and mannitol fermentation assays. Susceptibility testing was performed using the agar dilution method. A total of 679 samples were evaluated. All samples were inoculated onto CSA. Nasal surveillance cultures were inoculated onto sheep blood agar (SBA) (BD Diagnostics), and samples from CF patients were inoculated onto mannitol salt agar (MSA) (BD Diagnostics). Of the 679 samples cultured, 200 organisms produced a mauve color on CSA (suspicious for S. aureus) and 180 were positive for S. aureus on SBA or MSA. Of 200 CSA-positive samples 191 were identified as S. aureus. Nine mauve colonies were slide coagulase negative and were subsequently identified as Staphylococcus lugdunensis (one), Staphylococcus epidermidis (three), Staphylococcus haemolyticus (one), and Corynebacterium species (four). CSA improved the ability to detect S. aureus by recovering 12 S. aureus isolates missed by conventional media. Of the 192 S. aureus isolates recovered, 122 were methicillin susceptible and 70 were methicillin resistant. Overall, the sensitivity and specificity of CSA in this study were 99.5 and 98%, respectively. There was no difference in the performance of the slide coagulase test or in susceptibility testing performed on S. aureus recovered from CSA compared to SBA or MSA. Our data support the use of CSA in place of standard culture media for detection of S. aureus in heavily contaminated respiratory samples.

Bacteriological Techniques↗

PubMedDate not supplied

Acute conjunctivitis associated with biofilm formation on a punctal plug.

BACKGROUND: Punctal plugs are used for the treatment of tear-deficient type dry eye. We recently examined a case of acute conjunctivitis associated with bacterial biofilm formation on a punctal plug. CASE: A 63-year-old woman diagnosed as having tear-deficient type dry eye came to our hospital with a complaint of soreness in her right eye. Punctal plugs had been inserted into this eye 5 1/2 months previously. On the day of her visit, she presented with acute conjunctivitis. OBSERVATIONS: In biomicroscopical examination, the top of the punctal plug was seen to be covered with a whitish soft material. Microbiological analysis performed on a part of this material was positive for Staphylococcus haemolyticus and Candida tropicalis. Scanning electron microscopy of the removed punctal plug revealed widespread bacterial colonization embedded within an extensive extracellular matrix. Treatment consisted of the replacement of the plug, and administration of a combination of antibacterial eyedrops and preservative-free artificial solution. As a result, the acute conjunctivitis cleared up within 1 month. CONCLUSIONS: This case suggests that a punctal plug poses a potential risk of causing the formation of bacterial biofilm. In such a case, replacement of the plug and/or removal of the accumulated materials should be considered.

Acute Disease↗

PubMed1999

Topoisomerase sequences of coagulase-negative staphylococcal isolates resistant to ciprofloxacin or trovafloxacin.

Coagulase-negative staphylococcal isolates (n = 188) were screened for susceptibility to oxacillin, ciprofloxacin, and trovafloxacin, a new fluoroquinolone. At an oxacillin concentration of >/=4 microg/ml, 43% were methicillin resistant; of these, 70% were ciprofloxacin resistant (MIC, >/=4 microg/ml). Of the methicillin-resistant, ciprofloxacin-resistant isolates, 46% were susceptible to </=2 microg of trovafloxacin per ml and 32% were susceptible to </=1 microg of trovafloxacin per ml. Sixteen isolates, including twelve that expressed fluoroquinolone resistance, were chosen for detailed analysis. Identification of species by rRNA sequencing revealed a preponderance of Staphylococcus haemolyticus and S. hominis among fluoroquinolone-resistant strains. Segments of genes (gyrA and grlA) encoding DNA gyrase and DNA topoisomerase IV were sequenced. Considerable interspecies variation was noted, mainly involving noncoding nucleotide changes. Intraspecies variation consisted of coding changes associated with fluoroquinolone resistance. As for S. aureus, ciprofloxacin resistance (MIC, >/=8 microg/ml) and increased trovafloxacin MICs (0.25 to 2 microg/ml) could be conferred by the combined presence of single mutations in each gyrA and grlA gene. Trovafloxacin MICs of >/=8 microg/ml also occurred, but these required an additional mutation in grlA.

Anti-Infective Agents↗

PubMed1999

Distribution of insertion sequence-like element IS1272 and its position relative to methicillin resistance genes in clinically important Staphylococci.

The distribution of insertion sequence-like element IS1272 was analyzed for clinical isolates of Staphylococcus aureus, Staphylococcus epidermidis, and Staphylococcus haemolyticus. In each of the staphylococcal species, IS1272 was detected in both methicillin-resistant (MR) and methicillin-susceptible strains of different genetic types. In MR isolates, IS1272 was generally located downstream of the truncated mecR1 gene (DeltamecR1), with an identical junction sequence occurring between DeltamecR1 and IS1272, although insertion of an additional gene sequence in the junction sequence was detected in one S. epidermidis isolate. These findings suggest that the mec element with the rearranged form of mecR1 (DeltamecR1-IS1272) has been transmitted to multiple clones of staphylococci.

Bacterial Proteins↗

PubMed2003

[Yearly changes in antibacterial activities of cefozopran against various clinical isolates between 1996 and 2001--I. Gram-positive bacteria].

The in vitro antibacterial activities of cefozopran (CZOP), an agent of cephems, against various clinical isolates obtained between 1996 and 2001 were yearly evaluated and compared with those of other cephems, oxacephems, carbapenems, and penicillins. A total of 1,274 strains in 15 species of Gram-positive bacteria were isolated from the clinical materials annually collected from January to December, and consisted of methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant Staphylococcus aureus (MRSA), Staphylococcus epidermidis, Staphylococcus haemolyticus, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus pneumoniae, Enterococcus faecalis, Enterococcus faecium, Enterococcus avium, and Peptostreptococcus spp. CZOP possessed stable antibacterial activities against all strains tested throughout 6 years. The MIC90 of CZOP against MRSA and S. haemolyticus tended to decrease while against S. pneumoniae and Peptostreptococcus spp., tended to increase year by year. However, the MIC90 just changed a little and were consistent with the results from the studies performed until the new drug application approval. Increases in the MIC90 against S. pneumoniae were also observed with ceftazidime (CPR), cefepime (CFPM), flomoxef (FMOX), sulbactam/cefoperazone (SBT/CPZ), and imipenem (IPM). Increases in the MIC90 against Peptostreptococcus spp. were also observed with FMOX, SBT/CPZ, and IPM. In conclusion, the annual antibacterial activities of CZOP against the Gram-positive bacteria did not considerably change. It, therefore, was suggested that CZOP had maintained high antibacterial activity during 6 years of post marketing.

Cefepime↗

PubMed2004

Methicillin-resistant staphylococci isolated from animals.

Staphylococci isolated from animals (n=311) were screened for methicillin resistance by oxacillin agar screening. Oxacillin-resistant strains were tested for the presence of the mecA gene by PCR. Isolates were identified by standard techniques and 16S rDNA analysis, and their antimicrobial susceptibilities were tested using an agar diffusion method. MecA-positive strains were further analyzed using pulsed-field gel electrophoresis (PFGE). From 11 multidrug-resistant staphylococci, 6 were mecA-positive: 2 methicillin-resistant Staphylococcus aureus (MRSA) and 4 Staphylococcus haemolyticus. Screening of 300 staphylococci (100 S. aureus, 100 S. intermedius and 100 coagulase-negative staphylococci (CNS)) randomly chosen from the strain collection of the Veterinary Microbiological Diagnostic Center yielded five oxacillin-resistant coagulase-negative staphylococci, four of which were mecA-positive. PFGE showed that all mecA-positive staphylococci isolated from animals had distinct patterns. However, one MRSA isolated from a flank fistula of a dog showed homology to a human epidemic MRSA cluster, suggesting that transfer of MRSA between humans and dogs might occur.

Animal Diseases↗