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PubMed1979

Phage heterogeneity of coagulase-negative staphylococci isolated in the United States and Sweden from bovine milk.

A Swedish bovine and a Dutch human phage set for coagulase-negative staphylococci were used to phage type coagulase-negative staphylococci isolated from bovine milk from Minnesota dairy herds. A comparison was also made of the deoxyribonuclease activity of coagulase-negative staphylococci isolated from bovine milk in Sweden and Minnesota. Of 133 Minnesota isolates, only one could be typed with the Swedish set and one by the Dutch set, whereas of 218 Swedish strains, 49 could be typed with the Swedish set and 7 by the Dutch set. A larger number of coagulase-negative isolates from Sweden were deoxyribonuclease positive (35%) than were the similar isolates from Minnesota (12%). These findings substantiate the marked heterogeneity of coagulase-negative staphylococci isolated from bovine udders. Results presented point to the usefulness of establishing regional phage sets for epidemiological investigations of coagulase-negative staphylococci in cattle. It is anticipated that at a later stage the regional phage sets will be coordinated internationally.

Animals↗
PubMed1986

Persistent in vitro survival of coagulase-negative staphylococci adherent to intravascular catheters in the absence of conventional nutrients.

The in vitro survival of coagulase-negative staphylococci in media devoid of routine nutritional supplementation was assessed in the presence and absence of catheter materials to evaluate bacterium-device interactions. Strains of slime- and non-slime-producing coagulase-negative staphylococci were suspended in phosphate-buffered saline together with multiple segments of polyvinyl chloride (PVC), Teflon, Silastic, and polyurethane catheters and in control suspensions without catheters. Catheters were removed at 2 min and 24, 48, 72, and 96 h of incubation and washed thoroughly, and semiquantitative roll cultures were performed on blood agar. In addition, after 96 h catheters were introduced into tryptic soy broth (TSB), and roll cultures were performed after 18 h of incubation. Results demonstrated that after 96 h, 6 of 32 catheter specimens (4 PVC) had greater than 10 CFU of coagulase-negative staphylococci per catheter; after TSB addition, 18 of 32 catheter specimens had greater than or equal to 100 CFU per catheter (8 of 8 PVC catheters had greater than 1,000 CFU per catheter). In control suspensions, no growth was seen at 96 h or after TSB addition. No differences in the survival of slime- versus non-slime-producing strains were observed in control or catheter studies. These findings suggest that both slime- and non-slime-producing coagulase-negative staphylococci survive in vitro on catheters (especially PVC) in the absence of conventional nutrients and can proliferate on catheters when nutrients are added. Catheter-adherent coagulase-negative staphylococci appear to possess survival mechanisms under adverse conditions which may relate to the genesis of occult foreign-body-associated infections.

Bacterial Adhesion↗
PubMed1988

Evaluation of MicroScan MIC panels for detection of oxacillin-resistant staphylococci.

Clinical isolates of staphylococci (420 Staphylococcus aureus isolates and 248 coagulase-negative staphylococci) were tested by both MicroScan MIC panels (MicroScan, West Sacramento, Calif.) and an oxacillin agar screen (Mueller-Hinton agar [Difco Laboratories, Detroit, Mich.] containing 6 micrograms of oxacillin per ml and 4% NaCl) to evaluate the ability of MicroScan to detect oxacillin-resistant strains. MicroScan panels and oxacillin agar screen plates were incubated at 35 degrees C for 24 h and at 30 degrees C for an additional 24 h. Endpoints were recorded at 24 and 48 h. By MicroScan, 23 (5.5%) and 30 (7%) S. aureus isolates and 161 (65%) and 162 (65%) coagulase-negative staphylococci were oxacillin resistant at 24 and 48 h, respectively. At both 24 and 48 h, 23 (5.5%) S. aureus isolates and 162 (65%) coagulase-negative staphylococci were resistant by the oxacillin agar screen. Five strains for which the oxacillin MIC was 2 or 4 micrograms/ml and eight strains resistant to oxacillin only at 48 h were further evaluated by broth macrodilution testing for oxacillin with and without clavulanic acid, by oxacillin and amoxicillin-clavulanic acid disk diffusion, and by oxacillin agar screen comparing Mueller-Hinton agars purchased from Difco and BBL Microbiology Systems, Cockeysville, Md. By this additional testing, all 10 S. aureus isolates and 1 of 3 coagulase-negative staphylococci examined produced increased amounts of beta-lactamase. One coagulase-negative staphylococcus appeared to be truly intermediately oxacillin susceptible. There was no significant difference in the rate of detection of oxacillin resistance between MicroScan and the agar screen. MicroScan panels should be incubated for 24 h only, because prolonged incubation caused strains producing excessive amounts of beta-lactamase to appear to be falsely oxacillin resistant.

Coagulase↗
PubMed1988

Salt-supplemented medium for testing methicillin-resistant staphylococci with newer beta-lactams.

The addition of 2% NaCl to cation-supplemented Mueller-Hinton broth (CSMHB) was evaluated for microdilution testing of the susceptibility of staphylococci to five cephalosporins, imipenem, amoxicillin-clavulanate, and ticarcillin-clavulanate. With Staphylococcus aureus, NaCl improved the recognition of methicillin (oxacillin) resistance to cefamandole, imipenem, or ticarcillin-clavulanate. Resistance to amoxicillin-clavulanate was readily determined, irrespective of the presence of added salt. The addition of 2% NaCl to CSMHB did not significantly improve detection of resistance to any of the beta-lactams among coagulase-negative staphylococci. Since the addition of NaCl did not have significant adverse effects on tests with coagulase-negative staphylococci, the routine addition of 2% NaCl to oxacillin or methicillin tests with staphylococci may be justifiable on the basis of convenience or standardization. However, addition of NaCl to susceptibility tests of other beta-lactams does not consistently improve recognition of resistance among staphylococci and thus cannot be recommended for routine use.

Amoxicillin↗
PubMed1989

Comparative virulence of human isolates of coagulase-negative staphylococci tested in an infant mouse weight retardation model.

Human infections caused by coagulase-negative staphylococci have steadily increased in numbers and severity. Causes may be the use of artificial prostheses, immunocompromising chemotherapy and radiation therapy, and sophisticated surgical techniques, to name a few. Although the infectivity of coagulase-negative staphylococci as a group has been well documented for humans, attempts to study the pathogenesis of infections caused by individual species of coagulase-negative staphylococci have been hampered by the lack of an animal model that is not refractory to infection by these organisms. In the study reported here, a 2-day-old-mouse weight retardation test was used to assay the virulence of 60 clinical and reference strains of coagulase-negative staphylococci. These strains represented eight species of coagulase-negative staphylococci. The most virulent strains were demonstrated to be of the species Staphylococcus haemolyticus, S. saprophyticus, and S. epidermidis. The data further suggest that production of slime is a marker of virulence in S. epidermidis and that intraspecies differences in virulence occur.

Animals↗
PubMed1993

Two percent sodium chloride is required for susceptibility testing of staphylococci with oxacillin when using agar-based dilution methods.

The need to add NaCl to agar media to ensure accuracy of results when testing staphylococci with oxacillin was investigated. The results of four antimicrobial susceptibility testing methods (agar and broth dilution, E test, and disk diffusion) in which the growth medium contained 0, 2, 4, or 5% NaCl were compared with the results of a hybridization assay using a mec gene probe. We tested 223 strains of staphylococci, 128 of which were mec gene positive. A total of 7 of the 128 positive strains were coagulase-negative staphylococci with 24-h oxacillin MICs of < or = 2 micrograms/ml. Ninety-five isolates were mec gene negative, including seven strains of Staphylococcus aureus with oxacillin MICs of > or = 4 micrograms/ml. The oxacillin MICs for mec gene-positive, oxacillin-resistant strains of staphylococci increased two- to fourfold with the addition of NaCl to the test medium, while the MICs for mec gene-negative strains did not change in the presence of added salt. Very major error rates for the agar dilution and E test methods in the absence of salt ranged from 18.2 to 20.2%. Major error rates for mec gene-negative S. aureus isolates were > 17% for all test methods when 4 or 5% NaCl was added to the test medium. The addition of 2% NaCl to Mueller-Hinton agar for testing of oxacillin resulted in very major error rates of < 1% for the agar dilution and E test methods although the major error rates for the two methods with added NaCl were 8.5 and 6.9%, respectively. The disk diffusion test did not perform well in this study, showing essential error rates of > or = 18.3%. We recommended the addition of 2% NaC1 to Mueller-Hinton agar when testing staphylococci with oxacillin by either the agar dilution or E test method. NaC1 should not be added for the disk diffusion test.

Culture Media↗
PubMed2001

Development of a PCR assay for identification of staphylococci at genus and species levels.

We have developed a PCR-based assay which allows the detection of staphylococci at the genus level by targeting the tuf gene, which encodes the elongation factor Tu. Degenerate PCR primers derived from consensus regions of several tuf genes were used to amplify a target region of 884 bp from 11 representative staphylococcal species. Subsequently, the entire nucleotide sequence of these amplicons was determined. The analysis of a multiple alignment of these sequences revealed regions conserved among staphylococci but distinct from those of other gram-positive bacteria genetically related to staphylococci. PCR primers complementary to these regions could amplify specifically and efficiently a DNA fragment of 370 bp for all of 27 different staphylococcal species tested. There was no amplification with genomic DNA prepared from 53 nonstaphylococcal species tested to verify the specificity of the assay (20 gram positive and 33 gram negative). Furthermore, this assay amplified efficiently all 27 American Type Culture Collection (ATCC) staphylococcal reference strains as well as 307 clinical isolates of staphylococci from the Québec City region. Analysis of the multiple sequence alignment for the 884-bp fragment for the 11 staphylococcal species as well as comparison of the sequences for the 370-bp amplicon from five unrelated ATCC and clinical strains for each of the species S. aureus, S. epidermidis, S. haemolyticus, S. hominis, and S. saprophyticus demonstrated sufficient interspecies polymorphism to generate genus- and species-specific capture probes. This sequence information allowed the development of Staphylococcus-specific and species-specific (targeting S. aureus, S. epidermidis, S. haemolyticus, S. hominis, or S. saprophyticus) capture probes hybridizing to the 370-bp amplicon. In conclusion, this PCR assay is suitable for detection of staphylococci at both genus and species levels.

DNA Primers↗
PubMed2005

Widespread distribution of disinfectant resistance genes among staphylococci of bovine and caprine origin in Norway.

We demonstrate here a widespread distribution of genes mediating efflux-based resistance to quaternary ammonium compounds (QACs) in staphylococci from unpasteurized milk from 127 dairy cattle herds and 70 dairy goat herds. QAC resistance genes were identified in 21% of the cattle herds (qacA/B, smr, qacG, and qacJ) and in 10% of the goat herds (qacA/B and smr). Further examination of 42 QAC-resistant bovine and caprine isolates revealed the following genes: qacA/B (12 isolates) was present in four different species of coagulase-negative staphylococci (CoNS), smr (27 isolates) was detected in eight different CoNS species and in Staphylococcus aureus on a previously reported plasmid (pNVH99), qacG (two isolates) was detected on two plasmids (pST94-like) in Staphylococcus cohnii and Staphylococcus warneri, and qacJ (two isolates) was found in Staphylococcus hominis and Staphylococcus delphini on a plasmid (pNVH01) previously found in equine staphylococci. Isolation of indistinguishable pulsed-field gel electrophoresis (PFGE) CoNS types from tank milk and mammary quarter milk samples in a dairy cattle herd suggested that these QAC-resistant staphylococci were of intramammary origin. Indistinguishable or closely related PFGE types of bovine QAC-resistant CoNS were observed in different herds. One particular bovine S. warneri PFGE type was isolated repeatedly from samples collected during a 30-month period in a herd, showing long-term persistence. In conclusion, it seems that the widespread distribution of staphylococci carrying QAC resistance genes in Norwegian dairy cattle and goat herds is the result of both the intra- and interspecies spread of QAC resistance plasmids and the clonal spread of QAC-resistant strains.

Animals↗
PubMed2002

A rapid ELISA for the diagnosis of intravascular catheter related sepsis caused by coagulase negative staphylococci.

AIM: To develop and evaluate a rapid enzyme linked immunosorbent assay (ELISA) for the diagnosis of intravascular catheter related sepsis caused by coagulase negative staphylococci. METHODS: Forty patients with a clinical and microbiological diagnosis of intravascular catheter related sepsis and positive blood cultures, caused by coagulase negative staphylococci, and 40 control patients requiring a central venous catheter as part of their clinical management were recruited into the study. Serum IgG responses to a previously undetected exocellular antigen produced by coagulase negative staphylococci, termed lipid S, were determined in the patient groups by a rapid ELISA. RESULTS: There was a significant difference (p = < 0.0001) in serum IgG to lipid S between patients with catheter related sepsis and controls. The mean antibody titre in patients with sepsis caused by coagulase negative staphylococci was 10 429 (range, no detectable serum IgG antibody to 99 939), whereas serum IgG was not detected in the control group of patients. CONCLUSIONS: The rapid ELISA offers a simple, economical, and rapid diagnostic test for suspected intravascular catheter related sepsis caused by coagulase negative staphylococci, which can be difficult to diagnose clinically. This may facilitate treatment with appropriate antimicrobials and may help prevent the unnecessary removal of intravascular catheters.

Adolescent↗
PubMedDate not supplied

Methicillin-resistant staphylococci and ofloxacin-resistant bacteria from clinically healthy conjunctivas.

We evaluated the existence of methicillin (DMPPC)-resistant staphylococci and ofloxacin (OFLX)-resistant bacteria among preoperative patients at an eye clinic. Bacterial growth was studied in 194 conjunctival specimens of 125 preoperative patients without signs of ocular infection. Specimens were evaluated for growth of bacteria on Drigalski's plates, blood agar plates, or chocolate agar plates for 2 days at 37 degrees C. Susceptibilities of the strain to DMPPX, OFLX, minocycline (MINO), gentamicin (GM), erythromycin (EM), cefmenoxime (CMX), chloramphenicol (CP), and sulbenicillin (SBPC) were determined by the disk diffusion method. When DMPPC-resistant Staphylococcus aureus (MRSA) or DMPPC-resistant coagulase- negative staphylococci (MRCNS) were isolated, the susceptibility of the isolated strains to vancomycin (VCM) was additionally examined by the disk diffusion method. Of the 125 preoperative patients (194 eyes), 109 patients (159 eyes) had positive bacterial growth. Methicillin-resistant Staphylococcus aureus (MRSA) were positive in 1 patient. Methicillin-resistant coagulase-negative staphylococci (MRCNS) were positive in another patient. Two eyes with MRSA or MRCNS growth were treated with topical instillation of VCM, which was more sensitive than MINO and GM. They underwent the planned surgery after two subsequent tests showing negative growth. Of 120 isolated coagulase-negative staphylococci (CNS) 8 (6.7%) were resistant to OFLX. OFLX-resistant CNS were sensitive to MINO, CMX, and CP, but also resistant to SBPC. Of 114 isolated aerobic gram-positive rods (GPR), 55 (48.2%) were resistant to OFLX. OFLX-resistant aerobic GPR were sensitive to MINO, CMX, SBPC, and DMPPC. The 157 eyes with positive bacterial growth other than MRSA were treated with topical instillation of sensitive antibiotics. The present findings indicate that DMPPC-resistant staphylococci and OFLX-resistant bacteria exist in conjunctivas without signs of infection.

Adult↗
PubMed2002

Microbiological quality and somatic cell count of ewe milk with special reference to staphylococci.

A total 1502 useful half udders of 762 Churra ewes from eight herds were aseptically sampled in midlactation to study both the bacteriological isolates and the SCC of milk. Corynebacteria, enterococci, micrococci, staphylococci, and streptococci represented 11.2, 2.9, 1.4, 78.9, and 3.1% of all isolates, respectively. Within staphylococci, novobiocin-sensitive species (71.1%) were much more frequently isolated than novobiocin-resistant ones (7.8%). Staphylococcus epidermidis was the most prevalent species (53.2% of the isolates). Log SCC of uninfected half udder milk was 4.86. Isolates of novobiocin-resistant coagulase-negative staphylococci, micrococci, and corynebacteria were associated to low values of log SCC (4.85 to 5.20). In contrast, infection by novobiocin-sensitive coagulase-negative staphylococci, streptococci, and enterococci organisms was related to a sharp inflammatory response with log SCC means between 5.92 and 6.32. The species that showed the highest log SCC were Pasteurella haemolytica (7.62), Streptococcus agalactiae (7.28), and Staphylococcus aureus (6.68). High prevalence of infections by novobiocin-sensitive staphylococci together with high SCC related to such infections show a relevant role of these organisms in ewe mastitis. Consequently, implementation of staphylococcal mastitis control programs would be of great interest in dairy ewe herds to improve microbiological and hygienic quality of milk.

Animals↗
PubMed1991

The mechanism of staphylococci resistance to methicillin: a critical analysis of dominant opinions.

Resistance to methicillin has been reported to be constantly associated with production of a novel penicillin-binding protein (PBP), indicated as PBP2a. This PBP is always present in strains that are homogeneously resistant to the drug, but is also constantly present in the strains heterogeneously resistant, where by far the vast majority of the cells are indeed sensitive to the antibiotic. This apparent lack of correlation between the presence of PBP2a and methicillin resistance has led to the idea, shared by most scientists, that the presence of PBP2a alone cannot explain resistance of staphylococci to methicillin. This confusion has implications also for clinical microbiology. In fact in in-vitro assays, not rarely, strains that are resistant to methicillin appear sensitive to other beta-lactams. However, clinical experience has demonstrated that when infections caused by methicillin-resistant staphylococci, which, in in vitro assays, become sensitive to other beta-lactams, are treated with the latter antibiotics, the incidence of failures is higher than that observed when the same antibiotics are used to treat infections caused by staphylococci sensitive to methicillin. Also in consideration of the fact that the mechanism of resistance to methicillin is not yet understood, many authors recommend to consider methicillin-resistant staphylococci as being in-vivo resistant also to the beta-lactams to which they may become sensitive in in-vitro assays. In the opinion of the authors, the mechanism of staphylococcal resistance to methicillin is reasonably explainable at least in its fundamental aspects. The present dominant opinions concerning methicillin-resistant staphylococci will be critically analyzed on the basis both of data presented in the literature and of data obtained in the authors' laboratory.

Anti-Bacterial Agents↗
PubMed1975

[Changes in the ultrastructure of staphylococci in kidney tissue in experimental infection].

A study of ultrathin sections fo the kidney tissue of mice infected with staphylococci showed intracellular localization of the bacteria. In vivo there occurred a morphological reconstruction of external layers of the cell wall of staphylococci, and lysis of individual staphylococci. The middle part of the cell wall and the cytoplasmic membrane proved to be the structures the most stable to lysis. Specific changes of bacteria similar to the changes noted with the action of penicillin on staphylococci in vitro followed effective penicillin therapy of mice infected with staphylococci.

Animals↗
PubMed1975

Staphylococcus aureus-strains of phage group II and their possible relation to animal staphylococci. 1. Antigenic properties, phage typing, and resistance to antibiotics.

Investigations carried out on 350 strains isolated from clinical materials and carriers revealed that among coagulase-positive staphylococci of phage group II two main serologic types predominated. The first (61,1%) was characterised by positive reaction with factor serum 263-2, the second (30,8%) one gave positive reaction with factor serum i1-2. The strains of type 263-2 contained as a rule polysaccharide A as well as polysaccharide 263, contrary to staphylococci belonging to type i1-2 which possessed polysaccharide A only. Both serologic types differed also epidemiologically, in phage typing and resistance to antibiotics. Staphylococci of serologic type 263-2 occurred in neraly all purulent lesions, they were more resistant to antibiotics than type i1-2 and prevailed in hospital environments. In phage-typing the differences between both serologic types were quantitative in character, with two being most outstanding, namely the prevalence of strains sensitive to phage 71 among staphylococci of serologic type 263-2 and of phage types 3A, and 3C as well as 3A, 3B, 3C among staphylococci of serologic type i1-2. There was no correlation between the susceptibility of strains to phage 71 and their virulence, the latter seemed to be connected with serologic type.

Anti-Bacterial Agents↗
PubMed1977

Establishment of a new bacteriophage set for typing avian staphylococci.

A new phage set was proposed for the typing of coagulase-positive staphylococci from chickens. The set was composed of 16 phages isolated from lysogenic staphylococci from chickens and 3 adapted phages derived from the international phage set. These phages were classified into 4 groups (I, II, III and IV) according to their host ranges. The 19 phages were used for typing 569 coagulase-positive staphylococci; 94.2% of the isolates were lysed by 1 or more phages, 88.0% at a routine test dilution, and 6.25% only at a routine test dilution X 100. The reacting ability of the phages seemed to be limited to staphylococci from chickens. It was proved that the phages were more accurate than the international phage set in typing staphylococci isolated from chicken in Japan.

Animals↗
PubMedDate not supplied

[Physico-chemical properties of staphylococci of different species in resistance to human thrombodefensins].

Testing 54 strains of staphylococci (Staphylococcus aureus, S. epidermidis, S. haemolyticus, S. warneri, S. hominis, S. capitis) revealed that S. aureus in contrast to coagulase-negative staphylococci (CNS) is more resistant to bactoriocidal action of human thrombodefensins (resistance index: 60.3 vs 25.6%), less hydrophilicolipophilic balance-HLB: -0.42 vs -0.64) and less charged (x-potential: -32.4 vs -35.6 mV). In groups of staphylococci (S. aureus and CNS) correlation links of bacterial resistance to human thrombodefensins with their HLB and x-potential (r=-0.32...-0.36). By In vitro experiments, it was shown that 5 passages of staphylococci in meat-peptonic broth with human thrombodefensins (50 mkg protein/ml) lead to adaptation of bacteria followed by the formation of resistance to cationic peptides from thrombocytes, a decrease of hydrophobicity and x-potencial. The role of physico-chemical properties in providing thrombodefensin-resistance of staphylococci as a developmental factor of infectious-and-inflammatory process and persistence of bacteria was confirmed with Salmonella infection.

Adaptation, Physiological↗
PubMed1989

Methicillin resistance in staphylococci: an evaluation of conditions for detection.

Ninety-four strains of coagulase positive staphylococci and 73 strains of coagulase negative staphylococci were tested for their susceptibility to methicillin, using agar dilution and disc diffusion techniques with Diagnostic Sensitivity Test agar (DST) and DST supplemented with salt, incubated at 30 degrees C and 37 degrees C. Disc diffusion on salt-incorporated DST at 30 degrees C was most reliable for detecting methicillin resistance in both types of staphylococci. The agar dilution methods were particularly poor for detecting resistance with coagulase negative staphylococci. The simultaneous testing of coagulase positive and negative staphylococci by this method indicated the need for both DST and salt-incorporated DST agar plates, incubated at 30 degrees C. However, sensitive strains should still be tested by disc diffusion for confirmation.

Coagulase↗
PubMed1979

[Comparative characteristics of the antibioticograms and biological properties of the staphylococci isolated from patients with various eye diseases].

A total of 1488 examinations of the conjunctival content of patients with various ophthalmic diseases were performed within 5 years (1973--1977). 989 (66.4 per cent) cultures were isolated. Staphylococci were isolated in 79.2 per cent of the patients irrespective of the diagnosis. The plasma-coagulating strains were isolated in 43.6 per cent of the patients with acute conjunctivitis. The plasma-negative staphylococci were more frequent in chronic conjunctivitis (83.5 per cent). A total of 188 strains were studied according to 18 tests. Determination of the antibiotic sensitivity showed that the number of the benzylpenicillin and tetracycline resistant strains among the plasma-coagulating staphylococci was higher in the patients with acute ophthalmic diseases. In chronic conjunctivitis no differences in the antibiotic sensitivity of the plasma-coagulating and plasma-negative strains were noted. Similar antibiotic sensitivity of all plasma-negative staphylococci irrespective of the diagnosis was found. High biological activity of the plasma-negative staphylococci was shown: 74.1 per cent of the isolates possessed phosphatase, 59.8 per cent possessed protease and 88.4 per cent a lysozyme-like enzyme. The strains isolated in chronic conjunctivitis produced protease more frequently that those isolated in acute ophthalmic diseases.

Anti-Bacterial Agents↗