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Seasonal persistence and population characteristics of Escherichia coli and enterococci in deep backshore sand of two freshwater beaches.

We studied the shoreward and seasonal distribution of E. coil and enterococci in sand (at the water table) at two southern Lake Michigan beaches-Dunbar and West Beach (in Indiana). Deep, backshore sand (approximately 20 m inland) was regularly sampled for 15 months during 2002-2003. E. coli counts were not significantly different in samples taken at 5-m intervals from 0-40 m inland (P = 0.25). Neither E. coli nor enterococci mean counts showed any correlation or differences between the two beaches studied. In laboratory experiments, E. coli readily grew in sand supplemented with lake plankton, suggesting that in situ E. coil growth may occur when temperature and natural organic sources are adequate. Of the 114 sand enterococci isolates tested, positive species identification was obtained for only 52 (46%), with E. faecium representing the most dominant species (92%). Genetic characterization by ribotyping revealed no distinct genotypic pattern (s) for E. coli, suggesting that the sand population was rather a mixture of numerous strains (genotypes). These findings indicate that E. coli and enterococci can occur and persist for extended periods in backshore sand at the groundwater table. Although this study was limited to two beaches of southern Lake Michigan, similar findings can be expected at other temperate freshwater beaches. The long-term persistence of these bacteria, perhaps independent of pollution events, complicates their use as indicator organisms. Further, backshore sand at the water table may act as a reservoir for these bacteria and potentially for human pathogens.

Bathing Beaches↗

[Improvement of the membrane filter technique for enumeration of enterococci in water].

The membrane filter technique with AC agar medium supplemented with 0.04% NaN3 and 0.00015% 2,3,5-triphenyltetrazolium chloride for enumeration of enterococci in water is described. An appropriate volume of a water sample was filtered through the membrane filter. The membrane filter was put on an AC agar plate (designated as the AC.MF technique), which was incubated at 37 C for 18 hr and further at 45 C for 24 hr. By this AC.MF technique, all the colonies grown on the membrane filters were identified as enterococci, and the count of enterococci obtained by the AC.MF technique was similar to that by the AC.MPN technique. The AC.MF technique may be useful for accurate and rapid enumeration of enterococci in water and serve as a simple method for determining the sanitary quality of water.

Agar↗

Vancomycin resistance and antibiotic susceptibility of enterococci in raw meat.

The purpose of this study was to investigate antimicrobial resistance, in particular to vancomycin, of enterococci in samples (100) of meat (beef, chicken, turkey, lamb, and pork) sold in retail outlets of Catanzaro (Italy). Enterococci were identified to the species level. Antimicrobial susceptibility tests for a large spectrum of antibiotics including glycopeptides were performed by the disk diffusion method. Kappa statistic was used to evaluate associations of resistance to vancomycin with other antimicrobials. Enterococci were isolated from 45% of the samples, mostly from chicken meat (65.4%). Overall, 29% of samples were contaminated by vancomycin-resistant enterococci (VRE), whereas among those positive they represented 64.4% of isolates. Higher prevalence of vancomycin resistance was found in chicken samples (76.5%). The overall resistance to teicoplanin (TRE) was 30%, whereas among those positive, TRE represented 66.7% of isolates. The most frequent isolates were Enterococcus faecium (35.6%) and Enterococcus faecalis (33.3%). Resistance to vancomycin and teicoplanin was observed in 75% and 78.5% of E. faecium, and in 40% and 46.7% of E. faecalis, respectively. Most strains were susceptible to ampicillin (80%), while 88.9% were resistant to methicillin. The most effective antimicrobials were imipenem (73.3% susceptible) and rifampin (80%). The highest prevalence of resistance was for streptomycin (88.9%), tetracycline (84.4%), and erythromycin (75.6%). Resistance to vancomycin was significantly associated to methicillin, teicoplanin, erythromycin, tetracycline, and chloramphenicol. Further investigations about enterococcal colonization and infections in community and hospital subjects are needed.

Animals↗

Vancomycin-resistant enterococci isolates from the rumen content of deer.

The total enumeration of enterococci from the rumen content of deer, their species determination, resistance to antibiotics with the predominant importance of vancomycin resistance, and their metabolic properties, were examined. Enterococci (2.0 x 10(2) CFU per ml) encountered from the rumen of deer were phenotypically allotted to the species Enterococcus casseliflavus (26.6%), E. gallinarum (26.6%) and E. faecium (6.6%). The other isolates were not phenotypically specified. Although the resistance to the other antibiotics was recorded, the emergence of vancomycin-resistant enterococci in a high percentage (60%) was revealed. The majority of strains (80%) were multiresistant with the predominant resistance shown by vancomycin. These strains produced lactic acid in the range from 0.258 mol l-1 up to 0.489 mol l-1, and most of the isolates (53.3%) manifested medium urease activity. Vancomycin-resistant enterococci were selected for other studies such as plasmid determination and bacteriocin production, and to assess their value as indicators of faecal contamination.

Animals↗

[Vancomycin-resistant enterococci: in vitro activity of quinupristin / dalfoprostin (RP 59500)].

BACKGROUND: The recent emergence of glycopeptide-resistant enterococci limits the treatment of enterococcal infections. The aim of this study was to evaluate the in vitro activity of a new streptogramin, quinupristin/dalfopristin, against 30 clinical isolates of vancomycin-resistant enterococci and compared with those of other 15 antimicrobials. MATERIAL AND METHODS: Enterococci were identified by using Rapid ID 32 Strep system. Genotyping of the isolates was performed by PCR. The MICs of quinupristin/dalfopristin were determined by the agar dilution technique recommended by the NCCLS. Susceptibilities to the rest of antibiotics tested (teicoplanin, ampicillin, penicillin, imipenem, doxycicline, chloramphenicol, gentamicin, streptomycin, rifampin, levofloxacin, fleroxacin, trovafloxacin, sparfloxacin, pefloxacin and clinafloxacin) were determined by using the E test. beta-lactamase production was examined with nitrocefin disks. RESULTS: Quinupristin/dalfopristin has demonstrated excellent activity against Enterococcus faecium (MIC90' 2 micrograms/ml). Enterococcus faecalis was considerably less susceptible than E. faecium, at concentration of 4 micrograms/ml inhibited only 31% of tested strains. For doxycicline 77% of strains were susceptible. Only five isolates were susceptible to clinafloxacin; the other quinolones tested displayed poor activity. Resistance to chloramphenicol was detected in 47% of isolates. None of the isolates produced beta-lactamase. CONCLUSIONS: This study indicates that vancomycin-resistant enterococci are often concomitantly resistant to multiple antibiotics. Quinupristin/dalfopristin was the most active agent tested against E. faecium strains. On the basis of these results quinupristin/dalfopristin could be a therapeutic option for the treatment of vancomycin-resistant E. faecium infections.

Anti-Bacterial Agents↗

Antibiotic susceptibilities of enterococci isolated from Turkish children.

To evaluate the antibiotic resistance rates of enterococci isolated at Hacettepe Children's Hospital, in vitro antibiotic susceptibility tests were performed in 77 enterococci (32 hospital, 45 nonhospital strains) isolated from various clinical specimens in 1994. Microbroth dilution tests against ampicillin, vancomycin, gentamicin and streptomycin were performed according to the NCCLS standards. High-level resistance to aminoglycosides was investigated. Ampicillin resistance rates were 21.9 percent and 2.2 percent for hospital and nonhospital strains, respectively (p < 0.01). The same rates were 46.9 and 13.3 percent for gentamicin (p < 0.01), and 15.6 and 13.3 percent for streptomycin (p = 0.25). No resistance was detected against vancomycin. Six strains (7.8%) showed high-level resistance to both aminoglycosides studied. Special attention should be paid to enterococci isolated from hospitalized patients. Appropriate antibiotic use in serious infections can only be achieved by choosing an appropriate regimen according to antibiotic susceptibility tests. Periodic evaluation of the antibiotic susceptibility patterns of enterococci is necessary for the empirical treatment of infections due to these microorganisms.

Child↗

[Combination therapy against vancomycin-resistant enterococci].

The therapeutic options for patients infected with vancomycin-resistant enterococci are limited. Optimal therapy for serious enterococcal infections requires the use of synergistic combinations of a cell wall-active agent plus an aminoglycoside. Enterococci have acquired aminoglycoside resistance genes that mediate production of aminoglycoside-modifying enzymes, which eliminate this synergistic bactericidal effect. Traditional therapy has been compromised due to the increasing prevalence of enterococci with beta-lactam, glycopeptide, and high-level gentamicin resistance. Arbekacin, a semi-synthetic aminoglycoside, shows excellent activity against a wide variety of bacteria that produce aminoglycoside modifying enzymes, including AAC(6')-APH(2"). In recent studies, the combination of ampicillin and arbekacin demonstrated inhibitory activity against vancomycin-resistant enterococci, and may prove useful in the treatment of enterococcal infections where treatment options are limited.

Aminoglycosides↗

Antimicrobial susceptibility of clinical isolates of enterococci.

To describe antimicrobial resistance patterns of Enterococcus spp., we measured minimal inhibitory concentrations (MICs) of five antimicrobial agents for 174 clinical isolates of enterococci collected in Taichung Veterans General Hospital from November, 1996 to January, 1998. Major sources included blood (48), the genitourinary tract (39), soft tissue and wounds (28), and the gynecological tract (16). The sensitivity test of vancomycin was performed using the broth microdilution method. The susceptibility of ampicillin, penicillin, gentamicin, and teicoplanin was tested by the agar dilution method. The MIC90 of penicillin, ampicillin, gentamicin, teicoplanin and vancomycin were 2, 1, > 2,048, 0.125 and 2 microg/mL, respectively. Ampicillin-resistant isolates represented only 4.6% of all enterococci tested. These results reveal that vancomycin-resistant enterococci were found in 1.7% of all enterococci isolates in this study. However, all isolates were sensitive to teicoplanin.

Adult↗

Invasive infections due to vancomycin-resistant enterococci in adult patients.

Since 1990, vancomycin-resistant enterococci have emerged as important nosocomial pathogens. Invasive infections caused by these organisms have challenged most physicians because they are resistant to multiple antibiotics. We analyzed the clinical characteristics of adult patients with invasive vancomycin-resistant enterococci infections in the National Taiwan University Hospital from January 1993 through December 2000. A total of 11 adult patients were identified, 9 of whom had bacteremia (7 caused by vancomycin-resistant Enterococcus faecalis and 2 by vancomycin-resistant Enterococcus faecium) and one each had thoracic empyema (vancomycin-resistant E. faecium) and peritonitis (vancomycin-resistant E. faecium). Five patients had rectal swab cultures positive for vancomycin-resistant enterococci; 4 of them had underlying malignancies. The majority (91%) of these patients had prolonged hospitalization and prior long-term use of broad-spectrum cephalosporins (ceftriaxone, ceftazidime, or cefepime) or anti-anaerobic agents (clindamycin or metronidazole). The crude mortality rate was 64%. In conclusion, invasive infection caused by vancomycin-resistant enterococci is an emerging problem among hospitalized patients in Taiwan, particularly those with severe underlying diseases and exposure to multiple antibiotics.

Adult↗

[Development of enterococci and coli bacteria in Bulgarian yogurt].

The effect of lactic acid fermentation in Bulgarian yoghourt was studied by the development and decline of hygiene level indicating microorganisms--enterococci and coli-bacteriae. It was established that for the first 6 h after production of Bulgarian yoghourt, the quantity of enterococci (Str. faecalis, resp. Str. faecium) increases 5--6 times on the average, after the 12th hour it begins slowly to decrease and the number is reduced two times during storage for 7 days at 7--10 degrees C temperature of 6 days at 18--22 degrees C. The reduction in enterococci number is not fully correlated with the stage of lactic acid fermentation. Coli-bacteriae (E. coli) do not grow in number at the time the milk is turned into yoghourt. Their initial quantity diminishes along with the advance of the lactic acid process and after 24 h of milk storage at 18--22 degrees C or after 48 h at 7--10 degrees C they cannot be detected at all. The conclusion is drawn that enterococci are more suitable as hygiene level indicating microorganisms for hygienic evaluation of Bulgarian yoghourt, as compared to coli-bacteriae, due to their greater viability and longer survival in the product at the time of its production and during storage. It is recommended to perform the investigations on coli-forms not later than the 24th hour after yoghourt productions begins.

Dairy Products↗

Significance of vancomycin resistant enterococci from urinary specimens at a tertiary care centre in northern India.

BACKGROUND & OBJECTIVES: There is a paucity of information on vancomycin resistance in enterococci from India. In the present study, enterococci isolated from urine specimen were screened for vancomycin resistance. METHODS: 144 enterococci isolates obtained in pure and significant numbers (>10(5) cfu/ml) from 9664 urine specimens were subjected to agar screen method of National Committee for Clinical Laboratory Standard (NCCLS) for presence/absence of vancomycin resistance. Minimum inhibitory concentration (MIC) was performerd by E-test and agar dilution method. RESULTS: Eight vancomycin resistant enterococci (VRE) were obtained, namely, 5 Enterococcus faecium, 1 each of Enterococcus faecalis, E. casseliflavus and E. pseudoavium. The MIC ranged from 8 to 32-microgm/ml. The records of these 8 patients were retrospectively reviewed for clinical details. Five patients had nosocomial urinary tract infection (UTI), while in 2 patients UTI was community acquired. One patient had asymptomatic bacteriuria. Two patients with nosocomial UTI developed clinical sepsis and died in spite of vancomycin treatment. Urinary catheterization, surgery on genitourinary tract, prior exposure to third generation cephalosporins and metronidazole were present in 6,5,3 and 2 patients respectively. INTERPRETATION & CONCLUSION: Although the frequency of isolation of VRE is not very high in our setting as compared to West, this may just be the beginning of the problem. Screening for vancomycin resistance is recommended in all patients with suspected UTI who have significant bacterial counts.

Adult↗

[Vancomycin resistant enterococci in the Netherlands].

Enterococci (Enterococcus faecalis and Enterococcus faecium) are relatively avirulent enteric bacteria that usually only cause infections in immunocompromised patients. Antimicrobial treatment, however, is hampered as enterococci are intrinsically resistant to many antibiotics. For years, vancomycin was considered the last available antibiotic. Plasmid-mediated resistance against vancomycin among enterococci was first described in the nineteen-eighties and since then incidences of infection caused by vancomycin-resistant enterococci (VRE) have increased dramatically, especially in the United States. In 2000, three outbreaks of VRE occurred in hospitals in the Netherlands and a set of infection-control measures was proposed to limit further transmission. These measures were based on the simultaneous isolation of VRE from multiple patients. All three outbreaks were controlled by these measures and no new outbreaks in Dutch hospitals have been reported since then. Epidemiological studies have shown that hospital outbreaks on three continents were caused by a subpopulation of E. faecium, which is characterized by the presence of a potential virulence gene (variant esp) and resistance to amoxicillin. This 'hospital strain' of E. faecium has probably been prevalent within hospital settings for some time, but only became clinically relevant when it had acquired vancomycin-resistance. Current advice is to implement the set of infection control measures formulated in 2000, only in those patients colonized by amoxicillin-resistant VRE. The potential dangers of VRE were recently underlined by the proven transmission of the vancomycin-resistance gene from VRE to methicillin-resistant Staphylococcus aureus (MRSA) in two patients in the United States. It is in the interest of the patients that prevalence of VRE and MRSA in Dutch hospitals should be kept as low as possible.

Disease Outbreaks↗

[Comparative evaluation of membrane filter methods for enumeration of enterococci in water].

Many laboratory methods for the assay of viable counts of enterococci in water have been reported, but the evaluation or verification of the methods have yet to be reported. The three most commonly used membrane filter methods were selected and compared in terms of the verification of enterococci counts in environmental water. The verified percentages, i.e. identified enterococci count versus apparent enterococci count, assayed by each method, were as follows: KF.MF (membrane filter cultured on KF agar) method 64.5%; mENT.MF (membrane filter cultured on mENT agar) method 80.7%; and mE.MF (membrane filter cultured on mE agar) method 63.4%. The AC.MF (membrane filter cultured on AC agar) method which we reported previously (Koujima I, et al. Jap J Bact, 1989; 44: 813-816) consistently gives 100% verification and therefore is the most consistent method and superior in accuracy to the other three methods.

Bacteriological Techniques↗

Isolation, distribution and prevalence of various species of enterococci isolated from clinical specimens in a tertiary care hospital.

Various clinical specimens were processed to find the prevalence rate of enterococci and to identify the species of clinical isolates of enterococci. Screening of various clinical specimens revealed that enterococci were prevalent in 2% of the total specimens, with urine and wound to be the major site of isolation. Conventional test scheme proposed by Facklam and Collins and commercially available systems Rapid ID 32 Strep (biomereiux) were successfully used to speciate enterococcal strains. Five species of enterococci were identified in the study from a set of 396 cultures, with E. faecalis (79.79%), and E. faecium (11.11%) predominating. E. hirae (3.03%), E. gallinarum (3.03%), and E. casseliflavus (3.03%), were the other members of Enterococcus species identified.

Cross Infection↗

[Studies of bacterial indicators for water pollution--growth of Escherichia coli and enterococci in limited nutrient conditions].

Bacteria, that would be an ideal indicator for fecal contamination in environmental water, should not proliferate but at the same time should survive a little longer than enteropathogenic bacteria in the environmental water. While conventional methods have recommended Escherichia coli as an indicator, enterococci is recommended as a preferable indicator for the estimation of fecal contamination based on the following observation. The relationship between the specific growth rate of organism and the concentration of peptone was determined and the value of Ks, a parameter analogous to Michaelis-Menten constant, was estimated. Ks of Escherichia coli, Enterococcus faecalis and Enterococcus faecium were 3.8, 11.1 and 12.5 mg/ml respectively, indicating that Escherichia coli will grow at lower concentration of peptone than enterococci. Escherichia coli also grew in human fecal suspension and in filtrated (0.22 microns) river water, and while in those conditions enterococci did not grow or survive longer than Escherichia coli, survival time was longer than for Salmonella typhi and Shigella sonnei. It appears that enterococci reflect more accurately actual fecal contamination.

Enterococcus faecalis↗

Rapid immunochemical identification of enterococci with use of a centrifugal analyzer after a brief antigen-extraction step.

The precipitin reaction between antigens from enterococci and group-specific antisera was monitored turbidimetrically with a centrifugal analyzer. Linear standard curves (absorbance vs antigen concentration) were developed for studying optimal conditions for enzymic extraction of the antigen by use of an enzyme-containing filtrate from Streptomyces albus. This extraction was optimized by adding to the filtrate 1 mg of lysozyme per milliliter and incubating the bacteria for 40 min at 52 degrees C. The assay was adjusted so that detection of enterococci was indicated by positive absorbance changes near 0.1 A. When the concentration of bacteria from which the antigen was extracted was within a standardized range, non-enterococci produced no positive absorbance changes. Reproducibility of standard antigen and antisera solutions was excellent during three months. The procedure provides an objective, well-controlled means for identifying clinically important enterococci from pure cultures in about 1 h. In contrast, current methods involving biochemical media require as long as 48 h.

Antigens, Bacterial↗

[Determination of the susceptibility of staphylococci and enterococci to plycopeptides by the E-test method].

E-test was evaluated by comparison with the reference agar dilution method on 15 enterococci and 15 staphylococci with variable resistance to glycopeptides and isolated from clinical samples: 15 enterococci with 3 glycopeptide-resistant (2 E. facalis and 1 E. faecium), 1 glycopeptide intermediate E. gallinarum and 11 glycopeptide-sensible E. faecalis; 15 staphylococci with 4 teicoplanine-resistant (1 S. aureus, 1 S. epidermidis and 2 S. hominis), 3 teicoplanin-intermediate (1 S. epidermidis, 2 S. haemolyticus) and 8 teicoplanin sensible strains (6 S. aureus, 1 S. epidermidis and 1 S. hominis). Repeatability (10 determinations by strain) and exactitude were tested for all strains. Our data shows very good results for repeatability and exactitude with the reference method except in the case of coagulase negative staphylococci and teicoplanin. Some technical problems appear in our experience: for staphylococci, a 48 h incubation is necessary to detect teicoplanin-intermediate strains but 24 h is sufficient for enterococci. E-test is expensive, require a strict methodology, but is a good method to detect glycopeptide resistance for staphylococci and enterococci except for coagulase negative staphylococci and teicoplanine.

Bacteriological Techniques↗

[Enterococci isolated from blood (1989-1993): evolution of antibiotic susceptibility].

BACKGROUND: The incidence of enterococci infection is on the rise. Treatment is ever more difficult due to the intrinsic resistance to many antimicrobial and their ability to acquire new resistance. The aim of this study was to: a) determine the antibiotic sensitivity of 80 enterococci strains isolated from blood in 1989-1993 and, b) detect the possible changes in the resistance patterns over this period. METHODS: Eighty enterococci strains isolated from blood cultures, since 1989 to 1993 were studied. Sixty-nine were Enterococcus faecalis and 11 Enterococcus faecium. The minimum inhibitory concentrations (MIC) of 11 antimicrobial was determined by the agar dilution method. The Mantel-Haenszel and Fisher tests were used for statistical analysis of the results. RESULTS: All the strains were sensitive to ampicillin and vancomycin. A high level of resistance (HLR) to gentamycin and streptomycin was observed in 18.7% and 23.7%, respectively of the strains. Fourteen strains (17.7%) presented HLR to both antibiotics. Approximately one third of the strains were resistant to the fluoroquinolones tested. Analysis of the results by year only demonstrated a significant lineal trend of resistance for ofloxacin (p = 0.014) and a significant difference between the years for ciprofloxacin (p = 0.017). Strains with an MIC > or = 16 mg/l of fluoroquinolones presented HLR to gentamycin (p < 0.001) and to all the aminoglucosides (p < 0.001). CONCLUSIONS: The incidence of antibiotic resistance of enterococci isolated from blood in the authors hospital is generally low and, with the exception of the fluoroquinolones, did not demonstrate any significant differences during the period studied. Periodic updating and analysis of sensitivity results are required to monitor any changes which may occur over time.

Drug Resistance, Microbial↗