Monitoring of uropathogens and their susceptibility to antibiotics]

Samples of urine collected from patients with complicated urology infection and hospitalized to the Moscow Region Research Clinical Institute in 1986, 1991, 1995 and 1999 were analysed. Of 11,444 samples examined, bacteriuria was estimated in 7143 samples. 9786 strains (29 genus) of bacteria were isolated--56.9 per cent as mono culture and 43.1 per cent as associations. Susceptibility to 21 antibiotic was determined by disk diffusion method for 1607 strains; beta-lactamase production was determined in 198 strains, MIC was determined for 41 antibiotics. Gram-negative rods relative amount among pathogens decreased substantially (84.7 per cent in 1986 against 61.6 per cent in 1999), particularly Enterobacteriaceae (74.7 per cent in 1986 against 41.4 per cent in 1999). Nonfermenting Gram-negative rods (NFGNR) relative amount increased (10.8 per cent against 19.2 per cent), along with Gram-positive cocci (19.8 per cent against 64.2 per cent), particularly coagulasenegative staphylococci (CNS) (10.8 per cent against 35.9 per cent) and enterococci (5 per cent against 16.5 per cent) and candida and fungi (0.5 per cent in 1986 against 15.9 per cent in 1999). At the period 1986-1999 the main pathogens in urology infection were E. coli, Enterobacter spp., NFGNR (including P. aeruginosa), Staphylococcus, CNS, Enterococcus spp. The problem pathogens for urological department were the following: E. coli, Klebsiella spp., Enterobacter spp., Proteus spp., NFGNR including P. aeruginosa, CNS, Enterococcus spp., candida and fungi. At the period 1991-1997 Gram-negative pathogens susceptibility to amikacin, ofloxacin, ciprofloxacin, imipenem, ceftazidime, cefotaxime was not changed in general, Gram-positive cocci (staphylococci and enterococci) retained the same susceptibility to vancomicin, cefamandol and amoxyclave. Staphylococci were also susceptible to amikacin, imipenem, rifampicin, oxacillin, ciprofloxacin, and ofloxacin. Production of beta-lactamase was registered for 38.7 per cent of CNS, 26.5 per cent of E. coli, 38.5 per cent of K. pneumoniae, 25 per cent of P. mirabilis and 55.6 per cent of P. aeruginosa strains.     

Microflora of patients with purulent septic diseases in the Moscow region

The microflora of clinical material (blood, wound discharge, abdominal exudate, pleural exudate, milk in mastitis, as well as feces in dysbiosis of the large intestine), obtained from patients with purulent septic diseases in the Moscow region was studied. The study revealed the change of microflora from surface biotopes to closed cavities of the body. Gram-positive cocci, primarily coagulase-negative staphylococci, mainly epidermal, were shown to play the leading etiological role in the development of purulent septic diseases. The development of bacteriemia was found to be caused by the penetration of staphylococci from local pyoinflammatory foci (appearing in purulent mastitis, wounds, peritonitis, pleural empyema and pleuritis), as well as by the translocation of staphylococci and Escherichia coli from the large intestine in dysbiosis.     

Characteristics of some unclassifiable strains of staphylococci isolated from goats and sheep

Fourteen strains of catalase-positive, Gram-positive and coagulase-negative cocci that were sensitive to 20 micrograms/ml of furazolidone were isolated from goats and sheep. Two of the strains had glycerol with glucose teichoic acids whilst another possessed glycerol, glucose, glucosamine and acetylglucosamine teichoic acids. Six strains had peptidoglycan type L-Lys-Ala-Gly4 peculiar to Staphylococcus sciuri and Staph. lentus but other phenotypic characters were different from those of Staph. sciuri and Staph. lentus. The guanine plus cytosine (G + C) content of the DNA determined for three of the strains examined ranged from 32.9-34.6 mol %. The coagulase-positive staphylococcal strain of caprine origin examined had glycerol and glucosamine teichoic acids in addition to peptidoglycan type L-Lys-Gly5-6. The characteristics of the strains of staphylococci described herein are different from those already described in the literature.     

Vancomycin Resistance Among Strains of Staphylococcus epidermidis: Effects on Adherence to Silicone

Nosocomial device-related infections with Gram-positive cocci and their resistance to vancomycin are of increasing occurrence. We examined clinical isolates of relatively avirulent coagulase-negative staphylococci for their resistance to vancomycin and for their capabilities to adhere in vitro to medical grade silicone. Vancomycin resistance was found in 9 of 20 isolates, but there was no correlation between adherence capacity to silicone in the absence of vancomycin and vancomycin resistance for a given strain. Vancomycin in the medium, adsorbed to the surface of medical grade silicone or adsorbed on nongrowing cells, reduced adherence of representative Staphylococcus epidermidis to medical grade silicone. Received: 27 November 2000 / Accepted: 10 January 2001     

Opportunistic microorganisms in respiratory tract diseases in patients of the Moscow Province region

The respiratory tract microflora in patients with inflammatory processes of the upper and lower respiratory tracts in the Moscow Province region has been studied. Changes in the microflora were found to occur in patients with pyoinflammatory diseases (PID) of the upper and lower respiratory tracts have been found to occur. Gram-positive cocci, mainly staphylococci and streptococci, were shown to play the leading etiological role in the development of PID. As revealed in this study, the microorganisms detected in the inflammations of the upper and lower respiratory tracts are coagulase-negative staphylococci, mainly S. epidermidis, as well as enterococci and streptococci belonging to the group viridans.     

Characteristics of some unclassifiable strains of staphylococci isolated from goats and sheep

A degoke , G.O. 1985. Characteristics of some unclassifiable strains of staphylococci isolated from goats and sheep. Journal of Applied Bacteriology 59 , 257–262.
Fourteen strains of catalase-positive, Gram-positive and coagulase-negative cocci that were sensitive to 20 μg/ml of furazolidone were isolated from goats and sheep. Two of the strains had glycerol with glucose teichoic acids whilst another possessed glycerol, glucose, glucosamine and acetylglucosamine teichoic acids. Six strains had peptidoglycan type L-Lys-Ala-Gly₄ peculiar to Staphylococcus sciuri and Staph. lentus but other phenotypic characters were different from those of Staph. sciuri and Staph. lentus . The guanine plus cytosine (G + C) content of the DNA determined for three of the strains examined ranged from 32.9–34.6 mol %. The coagulase-positive staphylococcal strain of caprine origin examined had glycerol and glucosarnine teichoic acids in addition to peptidogylcan type L-Lys-Gly_5–6. The characteristics of the strains of staphylococci described herein are different from those already described in the literature.     

Polar lipid and isoprenoid quinone composition in the classification of Staphylococcus

Representatives of 13 species of Staphylococcus were examined using a small-scale procedure for the sequential extraction of isoprenoid quinones and polar lipids. Menaquinones were the only isoprenoid quinones found in the 77 test strains which were divided into three groups based upon the predominant isoprenologue detected: (i) S. hyicus subsp. hyicus, S. sciuri subsp. lentus and S. sciuri subsp. sciuri contained unsaturated menaquinones with six isoprene units; (ii) S. capitis, S. cohnii, S. epidermidis, S. haemolyticus, S. hominis, S. hyicus subsp. chromogenes, S. intermedius, S. saprophyticus, S. simulans, S. warneri and S. xylosus contained unsaturated menaquinones with seven isoprene units and (iii) S. aureus contained unsaturated menaquinones with eight isoprene units and varying amounts of the corresponding lower isoprenologue. All of the organisms contained very similar polar lipid patterns consisting of diphosphatidylglycerol, phosphatidylglycerol, beta-gentiobiosyl diacylglycerol and a number of glycolipids and phospholipids. One of the glycolipids was chromatographically indistinguishable from beta-gentiotriosyl diacylglycerol. Lysylphosphatidylglycerol was a major component in S. aureus and S. intermedius but was usually present in minor amounts in the coagulase-negative strains. The polar lipid data underline the homogeneity of the genus Staphylococcus and distinguish staphylococci from aerobic, Gram-positive cocci and from the phylogenetically related aerobic, endospore-forming bacteria. Menaquinone composition can also be used to separate staphylococci from other aerobic, Gram-positive cocci.     

Antimicrobial spectrum of bacteriocin-like substances produced by rumen staphylococci

Five strains of rumen coagulase-negative adherent and ureolytic staphylococci were obtained as bacteria producting bacteriocin-like substances or, lantibiotics. All examined staphylococci produced inhibitory agents which showed a wide range of inhibition against Gram-positive and Gram-negative indicator organisms from different sources. Clear zones of inhibition (diameter 1–6 mm) dominated. Most bacteriocin-like substances produced by the strains were stable and sensitive to trypsin, susceptible to chloroform vapours and heat-sensitive.     

Limitations of Fibrinogen-Polymyxin Medium in Detecting Coagulase-Positive Staphylococci in Raw Milk

A fibrinogen-polymyxin medium and Staphylococcus Medium 110 were used in the isolation of coagulase-positive staphylococci in raw milk. Results indicated that both media allow the growth of some rods and of many coagulase-negative cocci. A significantly greater number of coagulase-positive staphylococci were identified by the tube test than were revealed by halo formation on fibrinogen-polymyxin medium.     

Flucloxacillin,a New Isoxazolyl Penicillin,Compared with Oxacillin,Cloxacillin, and Dicloxacillin

Flucloxacillin, a new isoxazole penicillin, is active against penicillinase-producing strains of Staphylococcus aureus and is well absorbed in man after oral and intramuscular administration. Compared with isoxazole penicillins in current clinical use—namely, oxacillin, cloxacillin, and dicloxacillin—flucloxacillin has proved as active against Gram-positive cocci, including penicillin-resistant staphylococci. The extent of binding of flucloxacillin to the protein of human serum was similar to that of oxacillin and cloxacillin and less than that of dicloxacillin. In man flucloxacillin given orally produced total and free serum levels higher than those obtained with oxacillin and cloxacillin; total serum levels similar to those of dicloxacillin, and free levels greater than those of dicloxacillin. Similarly, after intramuscular injection the free serum levels of flucloxacillin were higher than those of oxacillin, cloxacillin, and dicloxacillin.     

Composition of AME encoding gene in Gram positive Cocci--study on spread of aminoglycoside resistance

Aminoglycoside modifying enzymes (AMEs) are major factors which confer aminoglycoside resistance on bacteria. Composition of six genes encoding AMEs (including lately described aph 2"-genes) was investigated by PCR for 16 clinical isolates of Enterococcus faecalis, 16 clinical isolates of coagulase-positive (S. aureus) and 13 clinical isolates of coagulase negative staphylococci (S. haemolyticus, S. epidermidis) collected in Gdańsk region (Northern Poland) in the years 1998-2001. Diversity of AME encoding gene profiles (composition) was used to analyze spread of AME encoding gene among and within studied group of cocci. According to presence of particular genes we distinguish eleven different AME encoding gene profiles: seven profiles were unique for particular species while the most common was shared among S. aureus, coagulase negative staphylococci and enterococci. Regarding profile frequency statistical analysis (Fstat, AMOVA, cluster analysis UPGMA) shows: the difference between S. aureus and enterococci and coagulase-negative staphylococci, lack of difference between enterococci and coagulase-negative staphylococci, higher variability within than between studied species and presence of multispecies cluster. On the basis of the reports about ability of staphylococci to synthesis enterococcal pheromones, this finding lets assume that spread of aminoglycoside resistance gene among gram (+) cocci is limited only by the ability of stains to synthesis or induction of synthesis conjugation protein.     

Effects of a topical essential oil-containing formulation on biofilm-forming coagulase-negative staphylococci

AIMS: To evaluate the antimicrobial effects of Polytoxinol (PT), a topical essential oil-based formulation, against biofilm positive strains of coagulase-negative staphylococci. METHODS AND RESULTS: Using a microtitre plate assay we measured inhibitory effects for PT against a selection of biofilm-forming clinical isolates of coagulase-negative staphylococci. Susceptibility varied considerably (MIC = 0.6-20 000 ppm). For the most tolerant clinical isolate (Staphylococcus warneri) biofilm growth was inhibited by a 32-fold lower PT concentration than planktonic growth. This inhibition of biofilm development, which was not observed with the other test isolates, was related to an inhibition of the initial phase of S. warneri cell adherence to the polystyrene surface. CONCLUSION: The antimicrobial efficacy of PT was verified against clinical isolates of coagulase-negative staphylococci in vitro. PT was able to inhibit biofilm formation in the most tolerant isolate at sub-inhibitory concentrations. SIGNIFICANCE AND IMPACT OF THE STUDY: These observations indicate that an ability to prevent biofilm formation, independently of effects on cell viability may contribute to the in vivo topical efficacy of essential oils.     

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.     

Environmental microbial contamination in a stem cell bank

AIM: The aim of this study was to evaluate the main environmental microbial contaminants of the clean rooms in our stem cell bank. METHODS AND RESULTS: We have measured the microbial air contamination by both passive and active air sampling and the microbial monitoring of surfaces by means of Rodac plates. The environmental monitoring tests were carried out in accordance with the guidelines of European Pharmacopeia and US Pharmacopeia. The micro-organisms were identified by means of an automated system (VITEK 2). During the monitoring, the clean rooms are continually under good manufacturing practices specifications. The most frequent contaminants were Gram-positive cocci. CONCLUSIONS: The main contaminants in our stem cell bank were coagulase-negative staphylococci and other opportunistic human pathogens. In order to assure the levels of potential contamination in both embryonic and adult stem cell lines, a continuous sampling of air particles and testing for viable microbiological contamination is necessary. SIGNIFICANCE AND IMPACT OF THE STUDY: This study is the first evaluation of the environmental contaminants in stem cell banks and can serve as initial evaluation for these establishments. The introduction of environmental monitoring programmes in the processing of stem cell lines could diminish the risk of contamination in stem cell cultures.     

Synthesis and antibacterial activity of N-[2-[5-(methylthio)thiophen-2-yl]-2-oxoethyl] and N-[2-[5-(methylthio)thiophen-2-yl]-2-(oxyimino)ethyl]piperazinylquinolone derivatives

A number of N-substituted piperazinylquinolone derivatives were synthesized and evaluated for antibacterial activity against Gram-positive and Gram-negative bacteria. Preliminary results indicated that most compounds tested in this study demonstrated comparable or better activity against Staphylococcus aureus and Staphylococcus epidermidis than their parent piperazinylquinolones as reference drugs. Among these derivatives, ciprofloxacin derivative 5a, containing N-[2-[5-(methylthio)thiophen-2-yl]-2-oxoethyl] residue, showed significant improvement of potency against staphylococci, maintaining Gram-negative coverage.     

Antibiotic resistance dynamics and isolation rate of staphylococci and enterococci from patients of reconstructive surgery units

The dynamics of isolation of staphylococci and enterococci from clinical material of patients and their antibiotic susceptibility within a 5-year period (2005-2009) was analysed. 5990 isolates were tested: 1250 isolates of Staphylococcus aureus, 3268 isolates of S. epidermidis, 1005 isolates of Enterococcus faecalis and 467 isolates of E. faecium. Grampositive infections were shown to be prevailing within the last 2-3 years, the nosocomial epidermal staphylococci more and more replacing S. aureus (the ratio of S. epidermidis and S. aureus in 2009 was 3.3). The isolation rate of E. faecalis significantly increased (by 3.5 times) and the ratio of E. faecalis and E. faecium in 2009 was 4.3. The microflora composition with respect to the isolation source was analysed and its clinical significance was estimated. The study of the antibiotic susceptibility showed that oxacillin had its own specific niche, while antibiotics active against resistant grampositive cocci, such as rifampicin, fusidic acid, fluoroquinolones (moxifloxacin), cefoxitin, as well as amoxicillin/clavulane in infections due to E. faecalis, might be considered as the drugs of choice. In the treatment of nosocomial infections, when the etiological role of MRSA or VRE is suspected or confirmed, the complex therapy should obligatory include the most active antibiotics (vancomycin or linezolid among them).     

Development of a real-time Staphylococcus aureus and MRSA (SAM-) PCR for routine blood culture

The notification of "Gram-positive cocci, possibly staphylococcus" in a blood culture drawn from a seriously ill patient is responsible for a large amount of vancomycin prescribing in institutions where methicillin-resistant Staphylococcus aureus (MRSA) is an important cause of bacteraemia. A duplex real-time TaqMan polymerase chain reaction targeting the species-specific nuc gene, and the mecA gene encoding methicillin-resistance, was developed as a tool for rapid identification and detection of S. aureus and methicillin-resistance, and optimised for immediate as-needs testing. Three different DNA extraction methods achieved varying DNA quality, with PCR inhibition the main problem. Serial blood cultures (n=120) identified as possible staphylococci on Gram stain from our clinical laboratory were examined. There was one false negative result for a methicillin-resistant Staphylococcus epidermidis, which was positive on repeat testing, and one false negative result due to DNA extraction failure for MRSA from peritoneal dialysate inoculated into blood culture medium. Sensitivity and specificity of 97% and 100%, respectively, were obtained for mecA; and sensitivity and specificity of 98% and 100%, respectively, for nuc. Detection of slow-growing coagulase-negative staphylococci as co-infecting strains may be reduced. The assay quickly and reliably identified S. aureus in mixed infection, and identified methicillin resistance in both S. epidermidis and S. aureus strains.     

Rifampicin in the treatment of infections of non-tuberculous etiology

Clinical efficacy of rifampicin, a semisynthetic broad spectrum antibiotic was estimated in 247 patients with purulent inflammations. It was shown advisable to use rifampicin intravenously in treatment of severe bronchopulmonary pathology, disorders of the bile excretion system, osteomyelitis, severe wound infections and in prophylaxis of postoperative purulent complications in cardiovascular surgery and other cases. High rifampicin sensitivity of staphylococci and streptococci belonging to various species was revealed. Rifampicin was found to be less active against gramnegative pathogens. The isolation frequency of rifampicin sensitive strains of E. coli, Proteus spp., Klebsiella spp. and P. aeruginosa amounted to 88.4, 52.1, 58.8 and 49.3 per cent respectively.     

Species-specific sensitivity of coagulase-negative Staphylococci to single antibiotics and their combinations

The activity of beta-lactam antibiotics (oxacillin, cloxacillin, cephalotin), vancomycin, gentamicin and rifampicin applied in vitro individually and in combination against 37 nosocomial methicillin-resistant strains of coagulase-negative staphylococci (CNS) was assessed to demonstrate the heterogeneity of this group of bacteria and estimate the chance of the efficacy of such therapy. The strains belonged to four species: Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus cohnii, Staphylococcus hominis. They originated from a hospital environment and from the skin of medical staff of the intensive care unit of a paediatric ward at a university hospital. All strains were methicillin-resistant, according to CLSI standards, but individual strains differed in MIC(ox) values. Susceptibility to other tested antibiotics was also characteristic for the species. The increased susceptibility to antibiotics in combinations, tested by calculating the fractional inhibitory concentration (FIC) index, concerned 26 out of 37 investigated strains and it was a feature of a particular species. Combinations of vancomycin and cephalotin against S. epidermidis and oxacillin with vancomycin were significant, as well as cephalotin and rifampicin in growth inhibition of multiresistant S. haemolyticus strains.     

Specific features of normal vaginal microflora in girls of preschool age

The study of the vaginal microbial cenosis in 20 healthy girls aged 3-7 years did not confirm the notion on the dominating role of cocci (including epidermal staphylococci). The associations of 2-5 different microorganisms represented by more than 20 species in an amount of 4-6 Ig PFC/g of discharge were established. In the overwhelming majority of the examinees (84.2%) the microbial associations of the vagina were found to contain bifidobacteria. Gram positive cocci (staphylococci and streptococci) took the 2nd and 3rd places in the isolation rates and were detected in vaginal associations in 78.9% of the girls. Staphylococci were represented by 5 coagulase-negative staphylococcal species with S. simulans and S. epidermidis prevailing. Hemolytic streptococci variants alpha and beta were isolated in the proportion of 2:1. The latter belonged to serogroups C and F. No S. aureus, Lactobacillus sp., streptococci of groups A and B, yeast-like fungi were detected. Genital mycoplasms (M. hominis) could rarely be found in the vaginal discharge of the girls aged 3-7 years (5.3%). No resident and transitory components could be isolated from the normal vaginal microflora and no quantitative domination of any bacterial species (genus) was shown. The concentrations of all organisms in this association were moderate or low.