Susceptibility to selected chemotherapeutics of Staphylococcus aureus strains resistant to methycyline isolated from clinical materials in the years 1991-1992 and 1997

The susceptibility to selected chemotherapeutic agents was determined in 100 strains of Staphylococcus aureus methicillin-resistant (MRSA) isolated from clinical materials in 1991-1992 (50 strains) and in 1997 (50 strains). Two methods were used for the determination: disc method and antibiotic dilution in agar. The minimal inhibitory concentration (MIC) was determined for vancomycin, teicoplanin, furazolidone, nitrofurantoin, ofloxacin, gentamicin, netilmicin and trimethoprim. The concentrations of the chemotherapeutics in the substrate ranged from 0.125 to 512 mg/l. The obtained results served for drawing of the following conclusions: all studied MRSA strains isolated in 1991-1992 and in 1997 were sensitive to glycopeptide antibiotics: vancomycin and teicoplanin, to nitrofurans: nitrofurantoin and furazolidone, and to fusidic acid. MRSA strains isolated in 1991-1992 were sensitive to ofloxacin, but in 1997 about 80% of the strains were resistant to that antibiotic, and this resistance was noted in S. aureus strains with homogeneous resistance to methicillin. Increasing frequency of resistance to mupirocin was found, in 1991-1992 4% of the strains were resistant, and in 1997 the resistance of MRSA to that antibiotic was found in 12%. No changes occurred in the sensitivity of staphylococci to trimethoprim/sulfamethoxazole (cotrimoxazole). About 94% of strains in 1991-1992 and 1997 were sensitive to that drug. The sensitivity to cotrimoxazole is connected with one of its components (trimethoprim), with 94% of MRSA strains sensitive to it.     

Antibiotic resistance of staphylococci isolated from outpatients

The aim of the study was to determine susceptibility of 587 strains of S. aureus and 85 strains of coagulase-negative staphylococci isolated from outpatients in Poznań to co-trimoxazole, amoxycillin/clavulanic acid, erythromycin, gentamycin, doxycycline, ampicillin, oxacillin, cephradine, clindamycin and neomycin. Also methicillin-resistant strains were determined as well as strains ability to produce beta-lactamases. Susceptibility testing and examination of methicillin-resistant strains were performed by the disc diffusion techniques according to recommendation of NCCLS. Methicillin-resistant strains were additionally examined to their sensitivity to vankomycin and teicoplanin. beta-lactamase production was detected using nitrocefin impregnated discs and iodometric method. Amoxacillin/clavulanic acid, gentamycin, co-trimoxazole, cephradin, oxacillin and clindamycin occurred to be very active against both, S. aureus and coagulase-negative staphylococci. 84.7% to 100% of examined strains were sensitive to these drugs. Doxycyclin, erythromycin and ampicillin were less effective. Nine strains (1.5%) of 587 strains of S. aureus as well as 7 strains (8.7%) of coagulase-negative staphylococci were methicillin-resistant. All of methicillin-resistant strains were sensitive to vancomycin and teicoplanin. More than 75% of S. aureus and close to 50% of coagulase-negative staphylococci were able to produce beta-lactamases.     

Antibiotic sensitivity of staphylococci isolated from the surface of the skin of the breasts in pregnant women

Nine hundred and fourty coagulase-positive and coagulase-negative strains of staphylococci isolated from the skin surface of the mammary glands of 94 pregnant women were tested by the disc agar diffusion method for their sensitivity to five antibiotics. The highest number of the isolates were sensitive to erythromycin and lincomycin (87.7 and 89.7 per cent, respectively). The highest number of moderately resistant strains were detected with respect to methicillin. 19 out of 42 cultures of Staph. aureus were resistant to benzylpenicillin and 24 cultures were resistant to tetracycline. Among staphylococci 130 strains or 13.8 per cent were polyresistant.     

Antimicrobial sensivity and ability to slime production of koagulaze-negative staphylococci

The aim of this study was to assess the ability of slime production ofcoagulase-negative staphylococci (CONS) and evaluate the susceptibility of bacteria to antibiotics. Strains were isolated from clinical specimens obtained from hospitalized patients. The most frequently isolated species were S. epidermidis (51%), S. hominis (18%), S. haemolyticus (13%). The result of this study shows that 61% of S.epidermidis produce slime on CRA (Congo red agar), whereas none of the tested S. haemolyticus strains has this ability. All examined strains were susceptible to vancomycin, linezolid and quinupristin/ dalfopristin. The majority of strains were susceptible to minocycline, fusid acid, nitrofurantoin and rifampicin. Sixty six percent of isolates were determined as methicillin-resistant coagulase-negative staphylococci.     

Occurrence of high-level mupirocin resistant coagulase-negative staphylococci in hospitals and their characterization

Occurrence of high-level mupirocin resistant coagulase-negative staphylococci in 5 hospitals in the region of Gdansk was determined. The study was carried out on 192 staphylococcal strains isolated from patients and medical staff. The mupirocin resistant strains were detected by 5 microgram mupirocin disc. The minimal inhibitory concentration (MIC) for mupirocin was estimated by E-tests. The mupirocin resistant strains were characterised by antibiotic sensitivity, including MIC for glycopeptides and oxacillin. Biotypes of resistant strains were also determined. Eight high-level mupirocin resistant strains (4.7%) were found. Only one strain expressed low-level resistance. All but one of high-level mupirocin resistant strains were resistant to methicillin. Six of them belonged to the S. epidermidis species but differences in the biotypes and antibiotic resistance patterns of these strains suggest they did not have a common origin.     

嗜麦芽寡养单胞菌临床分布特征与耐药分析

目的了解湖州市中心医院嗜麦芽寡养单胞菌临床分布特征与耐药性。方法采用常规方法分离,用VITE-COMPACT2全自动微生物分析仪进行菌种鉴定,用K—B法进行药敏试验。结果分离到嗜麦芽寡养单胞菌810株,复方新诺明耐药菌株48株（分离率5．9％）。标本来源主要来自ICU室,其次呼吸科,大部分来自痰液标本（约占89．2％）,年龄段以中老年人比率最高。嗜麦芽寡养单胞菌对亚胺培南、美罗培南、头孢吡肟、哌拉西彬他坐巴坦、庆大霉素、妥布霉素、阿米卡星高度耐药;头孢他啶、替卡西林／克拉维酸、环丙沙星耐药率为33．7％～58．2％;头孢哌酮／舒巴坦、左氧氟沙星、米诺环素、复方新诺明耐药率低于30．0％。复方新诺明耐药菌株对头孢哌酮／舒巴坦、左氧氟沙星和米诺环素耐药率分别为60．4％、91．7％和2．0％,对其余抗菌药物耐药率达100．0％。复方新诺明耐药菌株与复方新诺明敏感菌株相比,耐药情况更严重,其中对三、四代头孢菌素、喹诺酮类耐药率显著高于复方新诺明敏感菌株（P〈0．01）;对碳青霉烯类、青霉素类、氨基糖苷类抗菌药物耐药率与复方新诺明敏感菌株相比,差异无统计学意义（P〉0．05）。结论嗜麦芽寡养单胞菌呈高度耐药,对头孢哌酮／舒巴坦、左氧氟沙星、米诺环素、复方新诺明尚敏感,但对复方新诺明耐药的嗜麦芽寡养单胞菌耐药现象更严重。应重视嗜麦芽寡养单胞菌引起的院内感染,尽量减少不必要的侵人性操作,加强抗菌药物的合理规范使用。     

Susceptibility of Coagulase-negative Staphylococci to Lysostaphin and Other Antibiotics

In general, coagulase-negative staphylococci were found to be relatively less susceptible to the lytic action of lysostaphin than coagulase-positive staphylococci. To achieve, arbitrarily, a lysis greater than 75%, it was necessary to use an increased concentration of enzyme or a longer incubation period than that usually required with coagulase-positive strains. For the most part, the cultures studied were sensitive to oxacillin, cloxacillin, dicloxacillin, nafcillin, ancillin, cephalothin, cephaloridine, fusidic acid, lincomycin, novobiocin, and neomycin [median minimal inhibitory concentrations (MIC) of 1.56 mug/ml or less]. Some degree of resistance (median MIC values of 12.5 mug/ml or greater) to benzylpenicillin, ampicillin, methicillin, tetracycline, chloretetracycline, erythromycin, ristocetin, and lysostaphin was found. Ten methicillin-resistant, coagulase-negative staphylococal strains were found to be cross-resistant to all nine of the penicillins tested, but much less resistant to the two cephalosporin analogues. In several instances, some of these strains seemed to be more sensitive to benzylpenicillin and to certain of the semisynthetic penicillins than to methicillin. Of the 18 antibiotics tested with the viable plate count method, the methicillin-resistant strains were found to be the most sensitive to lincomycin and novobiocin.     

Staphylococci in orthopaedic surgical wounds.

From 50 infected surgical wounds of orthopaedic patients, 43 (86%) staphylococcal strains were isolated. 34 of all these staphylococci belonged to Staphylococcus aureus species (i.e. 68 % of all isolates from surgical wounds; 79% of staphylococcal isolates); 9 were coagulase-negative staphylococci (i.e. 21% of all isolates from surgical wounds; 18% of staphylococcal isolates). Among microorganisms isolated from the wounds we also found 2 (4%) of the Enterobacteriaceae family; 2 (4%) of the Pseudomonas genus; 3 (6%) of the Streptococcus genus. Thus, orthopaedic surgical wounds were infected by staphylococci (mainly S. aureus) more frequently than by other micro-organisms. All the staphylococcal strains were screened for methicillin resistance by agar disk diffusion testing and for the presence of mecA gene responsible for methicillin resistance by PCR. 32% of the S. aureus and 33% of the S. epidermidis strains resulted methicillin resistant and mecA-positive. The data confirm the diffusion of methicillin resistant S. aureus in surgical site infections and shows that the so-called "new pathogens", i.e. S. epidermidis and other coagulase-negative staphylococci, also exhibit a frequent and hazardous methicillin-resisting ability.     

Methicillin-resistant Staphylococcus aureus (MRSA) with high resistance to mupirocin in hospitals of the Gdańsk region

Occurrence of high-level mupirocin resistance in methicillin-resistant Staphylococcus aureus (MRSA) strains isolated from 18 hospitals in Gdańsk area was determined. The study was carried out on 190 MRSA isolated in 1997-2000 from various clinical samples. The strains were tested for high-level mupirocin resistance by 200 micrograms mupirocin disc. The minimum inhibitory concentrations (MIC) for methicillin were estimated by agar dilution. Sensitivity to other antibiotics was determined in disc-diffusion method and to vancomycin in agar dilution method additionally. The strains were typed by set of 10 experimental phages and compared by the method of PCR-RFLP analysis of coagulase gen restriction fragment length polymorphism. There were low frequency of high-level mupirocin resistance in MRSA strains (4.7%) that were found only in 3 hospitals, in 6 patients. All of them were high-resistant also to methicillin and resistant to doxycyclin, gentamycin, erytromycin, klindamycin, ciprofloksacin, rifampicin, resistant or intermediate sensitive to fusidic acid but sensitive to vancomycin, teikoplanin and bacitracin. The origin all of the MRSA strains high-resistant to mupirocin probably was the same, except one strain, because they were belonged to one genetic type and possessed the same phage pattern.     

Role of Staphylococcus saprophyticus in urinary infections]

A proportion of S. saprophyticus in other coagulase-negative staphylococcal isolates from the urine of patients with urinary infections and healthy individuals has been investigated. Certain diagnostic aspect of the urinary infections with S. saprophyticus have also been considered. Hundred four coagulase-negative staphylococcal strains isolated from patients in S?upsk and Gdańsk area and 72 strains of the coagulase-negative staphylococci isolated from the urine of healthy women have been divided into 9 species, according to Kloos and Schleifers' classification. Bacteriologic tests have shown that S. saprophyticus produced 20.4% of the urinary tract infections in S?upsk area holding the second place after S. haemolyticus (27.3%). This species was the most infrequent in the urine of patients in Gdańsk area (3.3%). Its sensitivity to antibiotics did not differ from that of other coagulase-negative staphylococci. In contrast to the majority of other strains, S. saprophyticus has not been isolated from the urinary tract of healthy women and has been encountered most frequently in low bacteriuria. Test of resistance to novobiocin which is considered as a simplified identification method of this species proved to be not very precise as other species have also been resistant to this antibiotic.     

Slime production and antibiotic susceptibility in staphylococci isolated from clinical samples

A total of 187 isolates from several clinical specimens were identified to species level as 129 Staphylococcus aureus strains and 58 coagulase-negative staphylococci (CNS) strains by the API Staph System (Biomerieux). Slime production was detected both by the conventional Christensen's method as well as by the Congo red agar method. Seventy-two strains of staphylococci isolates (38.5%) were found to be slime producers by Christensen's test tube method whereas 58 strains (31%) were slime positive with Congo red agar method. There was no statistically significant difference between the two methods for the detection of slime production (P > 0.05). Susceptibility of isolates against antimicrobial agents was tested by the disk diffusion method. Staphylococcal species had resistance to one or more antibiotics. Among the various antimicrobial agents, oxacillin (71.1%) and erythromycin (47.1%) showed higher resistance than most of the agents used against all isolates. Oxacillin resistant S. aureus (ORSA) and oxacillin resistant coagulase-negative staphylococci (ORCNS), 97 (75.2%) and 36 (62.1%) respectively were frequently observed in strains isolated from clinical materials. Among the ORSA strains, two strains were resistant to vancomycin. Moreover, 96 (74.4%) of 129 S. aureus strains were positive for beta-lactamase enzyme. However, 78 (81.25%) of 96 beta-lactamase positive S. aureus strains were beta-lactamase positive ORSA isolates, but none of them had vancomycin resistance.     

Detection of methicillin and mupirocin resistance in staphylococcal hospital isolates with a touchdown multiplex polymerase chain reaction

Staphylococcal hospital isolates (n = 166) were tested in a touchdown multiplex-polymerase chain reaction assay for the identification of methicillin and mupirocin resistance and discrimination of S. aureus (femA gene) from coagulase negative staphylococci and other bacteria. All isolates harbored the 16SrDNA (Staphylococcus genus specific internal control) gene, and 130 (78 %) the mecA (methicillin resistance) gene. Fifty-seven (44 %) of these were determined as methicillin-resistant S. aureus, while the remaining 73 (56 %) were methicillin-resistant coagulase-negative staphylococci. Seventy-five (45 %) isolates harbored the ileS-2 (high-level mupirocin resistance) gene and were determined as mupirocin-resistant. This assay represents a simple, rapid, reliable approach for the detection and discrimination of methicillin-and mupirocin-resistant staphylococci.     

Quantitative disk diffusion as a convenient method for determining minimum inhibitory concentrations of oxacillin for staphylococci strains

In this study the susceptibility of 58 coagulase-negative staphylococci (CoNS) strains and 58 Staphylococcus aureus strains to oxacillin was evaluated by a novel method called quantitative disk diffusion (DD) method. The results obtained were compared to phenotypic methods as agar dilution (AD) for oxacillin, disk diffusion (DD) for cefoxitin, and related to the presence of the mecA gene detected by PCR. Minimum inhibitory concentrations (MIC) determined by the quantitative DD method were equivalent to MICs determined in the AD method for S. aureus (Student's t test, p=0.99) and CoNS (Student's t test, p=0.97). Incongruent results between PCR mecA gene determinations and the quantitative DD method were obtained in 8 strains (5 S. aureus and 3 CoNS) where the mecA gene expression was blocked. However, oxacillin resistance was detected by the proposed method even in staphylococci strains showing low-level or heterogeneous resistance to the antibiotic while other phenotypic methods failed. The single quantitative DD method is not expensive, it can be performed in any laboratory and permits accurate identification of oxacillin resistant staphylococci.     

Co-trimoxazole resistance in methicillin-resistant staphylococci isolated from clinical material in 2001-2003

The aim of this study was to determine resistance to co-trimoxazole among 476 strains of methicillin-resistant Staphylococus aureus (MRSA) and 2137 strains of methicillin resistant coagulase-negative staphylococci (MRCNS) isolation during three years (2001-2003). Their susceptibility testing were performed by the disc-diffusion techniques according to recomendation of NCCLS (National Committe for Clinical Laboratory Standards). Co-trimoxazole resistance was demonstrated more frequently among MRCNS (52,8%, 51,4%, 63,9% in 2001, 2002, 2003), than among MRSA (23,4%, 19,5%, 16,8% adequately).     

Methicillin resistance in staphylococci isolated from subclinical mastitis in sheep

One hundred ovine milk samples were subjected to bacteriological analysis to detect staphylococci. Twenty-four staphylococcal strains isolated were characterised for methicillin resistance with disk diffusion test (DDT) after incubation at 24 and 48 h, oxacillin agar screen test, Minimal Inhibitory Concentration (MIC), nitrocefin test for beta-lactamase production and PCR for the mecA gene. Nine staphylococcal strains resulted resistant in DDT; some differences in the halo diameter at double incubation period were noted; eight of these strains were resistant in MIC test; just one strain was positive to oxacillin agar screen test. All strains were mecA negative by PCR and positive by nitrocefin test. On the basis of these results methicillin-resistant strains can be classified as beta-lactamase hyperproducers.     

Species characteristics of coagulase-negative staphylococci of animal origin

The specific identification of 271 strains of coagulase-negative staphylococci isolated from different animals (cows, sheep, swine, hens, monkeys, minks, sables, foxes, etc.) was carried out according to the scheme of Akatov--Devriese. Species could be determined in 77.5% of the strains. The representatives of S. sciuri (55.5%) and S. xylosus (11.4%), very seldom occurring in humans, prevailed among the identified cultures. In 31.4% of coagulase-negative strains of animal origin the presence of protein A was established. The study of the time of glucose fermentation in the cultures and the type of colonies formed on agar with crystalline violet permitted the additional characterization of the majority of the strains of coagulase-negative staphylococci. Only in 4.8% of cases the strains under study could be lyzed by typing phages Holmberg and belonged to 6 phage types; of these, 117 A was the most numerous one (7 out of 13 typed cultures). No relationship between the phage type and the species of the strains was established.     

Antimicrobial susceptibility of staphylococci isolated from otitis externa in dogs

Samples were obtained from 65 unmedicated adult dogs, processed for isolation of Staphylococcus species and tested for susceptibility to penicillin G, gentamicin, oxacillin, tetracycline, trimethoprim-sulphamethoxazole, streptomycin, ampicillin and rifampin. Forty-four isolates were obtained, which represents 67.7% of samples. Coagulase-negative species were most commonly found, and the most frequently isolated staphylococcus species were Staph. epidermidis and Staph. aureus. Other species, such as Staph. simulans, Staph. haemolyticus, Staph. saprophyticus and Staph. intermedius were also isolated. Resistance to antibiotics was frequently observed, with 90.9% of the isolates showing resistance to at least one drug. The most active antimicrobial agents against staphylococci isolated from otitis externa of dogs were rifampin and oxacillin. Multidrug resistance was a common finding, and one strain of Staph. haemolyticus species, was resistant to all tested antimicrobial agents. Resistance to three or more different drugs was a common finding, observed in 16 strains (36.4%) of both coagulase-positive and coagulase-negative staphylococci. This study highlights the emergence of cases of otitis externa determined by coagulase-negative staphylococcus strains and once more emphasizes the need for bacterial culture with species identification and susceptibility testing of swab specimens from the ear canal in order to choose appropriate antimicrobial agents.     

Sensitivity to vancomycin and teicoplanin of coagulase-negative staphylococci isolated from clinical specimens

The frequency of resistance and elevated resistance to teicoplanin and vancomycin among 689 strains of coagulase-negative staphylococci isolated in one year from clinical specimens was determined. Using ATB.STAPH test, a resistance was shown mainly among strains of S. epidermidis and S. haemolyticus. The elevated resistance to teicoplanin was much more frequently observed than to vancomycin. About 27% of isolated strains of S. haemolyticus and 6.8% of S. epidermidis were classified as resistant. Among other species only single strains were recognised as resistant: one strain of S. xylosus, one of S. cohni and one of S. intermedius. 94.7% of S. epidermidis and 100% of S. haemolyticus strains classified as resistant to teicoplanin in ATB showed MIC values 14 mg/l. Moreover it was shown that 26.3% of these strains of S. epidermidis and 33.3% of S. haemolyticus had MBC of teicoplanin values equal to or higher than 32 mg/l.     

Effects of various types of Triton X on the susceptibilities of methicillin-resistant staphylococci to oxacillin

We examined the effect of six types of the nonionic detergent Triton X on the susceptibilities of methicillin-resistant staphylococci to oxacillin. We used five methicillin-resistant Staphylococcus aureus isolates and 17 methicillin-resistant coagulase-negative staphylococci isolates. All strains of S. aureus, S. epidermidis and S. sciuri had enhanced susceptibility to oxacillin following exposure to the types of Triton X having 7–13 polymerized ethylene oxides. These strains were altered from homogeneously resistant to heterogeneously resistant by Triton X-100. Those types of Triton X that affected the resistance level also promoted the release of lipoteichoic acid. These results and those of previous studies suggest that Triton X might act on factors other than the mecA or femA products.     

Expression of glycocalyx by coagulase-negative Staphylococcus species isolated from bovine milk

Two hundred and six strains of coagulase-negative Staphylococcus species were assessed for expression of glycocalyx on serum soft agar, india ink and adherence techniques. The organisms were maintained on trypticase soy agar plates at 4 degrees C for 30 d (120 strains) or stored at -80 degrees C in skim milk for 90 d (60 strains). Additionally, 26 milk samples from cows known to have excreted coagulase-negative staphylococci were used to inoculate serum soft agar directly. Nine of 26 direct culture samples and 43 of 180 strains maintained for an extended period had diffuse-type growth on serum soft agar. The proportion that exhibited an unstained halo by india ink was similar regardless of storage time. Slime production determined by in vitro adherence revealed a higher proportion of positive strains than had been predicted by serum soft agar or india ink techniques. More strains of Staphylococcus chromogenes, Staph. epidermidis, Staph. hominis, Staph. simulans and Staph. warneri expressed glycocalyx than other coagulase-negative Staphylococcus species. These results suggest that most coagulase-negative staphylococci produce slime rather than a capsule. However, evidence for classical encapsulation was demonstrated in several strains by india ink. The finding that Staphylococcus species other than Staph. aureus isolated from bovine milk are capable of glycocalyx production may be of importance in investigations on the relationship between staphylococci and host defence mechanisms.