In vitro activity of RP59500, an injectable streptogramin antibiotic, against vancomycin-resistant gram-positive organisms

The in vitro activity of RP59500, a streptogramin antibiotic, against 146 clinical isolates of vancomycin-resistant gram-positive bacteria was examined. Five strains of the species Enterococcus casseliflavus and Enterococcus gallinarum, for which the MIC of vancomycin was 8 micrograms/ml, were also studied. Twenty-eight vancomycin-susceptible strains of Enterococcus faecalis and Enterococcus faecium were included for comparison. The drug was highly active against Leuconostoc spp., Lactobacillus spp., and Pediococcus spp. (MICs, < or = 2 micrograms/ml). RP59500 was more active against vancomycin-susceptible strains of E. faecium than E. faecalis (MICs for 90% of the strains [MIC90s], 1.0 versus 32 micrograms/ml). Vancomycin-resistant strains of E. faecalis were as resistant to RP59500 as vancomycin-susceptible strains (MIC90, 32 micrograms/ml), but some vancomycin-resistant E. faecium strains were relatively more resistant to the new agent (MIC90, 16; MIC range, 0.5 to 32 micrograms/ml) than were vancomycin-susceptible organisms of this species.     

Are clinical laboratories in California accurately reporting vancomycin-resistant enterococci?

In order to determine whether hospital-based clinical laboratories conducting active surveillance for vancomycin-resistant enterococci in three San Francisco Bay area counties (San Francisco, Alameda, and Contra Costa counties) were accurately reporting vancomycin resistance, five vancomycin-resistant enterococcal strains and one vancomycin-susceptible beta-lactamase-producing enterococcus were sent to 31 of 32 (97%) laboratories conducting surveillance. Each strain was tested by the laboratory's routine antimicrobial susceptibility testing method. An Enterococcus faecium strain with high-level resistance to vancomycin (MIC, 512 microg/ml) was correctly reported as resistant by 100% of laboratories; an E. faecium strain with moderate-level resistance (MIC, 64 microg/ml) was correctly reported as resistant by 91% of laboratories; two Enterococcus faecalis strains with low-level resistance (MICs, 32 microg/ml) were correctly reported as resistant by 97 and 56% of laboratories, respectively. An Enterococcus gallinarum strain with intrinsic low-level resistance (MIC, 8 microg/ml) was correctly reported as intermediate by 50% of laboratories. A beta-lactamase-producing E. faecalis isolate was correctly identified as susceptible to vancomycin by 100% of laboratories and as resistant to penicillin and ampicillin by 68 and 44% of laboratories, respectively; all 23 (74%) laboratories that tested for beta-lactamase recognized that it was a beta-lactamase producer. This survey indicated that for clinically significant enterococcal isolates, laboratories in the San Francisco Bay area have problems in detecting low- to moderate-level but not high-level vancomycin resistance. Increasing accuracy of detection and prompt reporting of these isolates and investigation of cases are the next steps in the battle for control of the spread of vancomycin resistance.     

Enterococcus faecium: in vitro activity of antimicrobial drugs, singly and combined, with and without defibrinated human blood, against multiple-antibiotic-resistant strains

The minimal inhibitory (MICs) and bactericidal concentrations of 14 antimicrobial drugs were determined against 17 clinical isolates of Enterococcus faecium, including 4 glycopeptide-resistant strains. Both teicoplanin and vancomycin lacked bactericidal activity against all 13 susceptible isolates. Time-kill experiments served to test various antibiotic combinations chiefly against glycopeptide-resistant strains in Mueller-Hinton broth (MHB) and in MHB supplemented with 65% (v/v) fresh defibrinated human blood. Co-trimoxazole, fusidic acid, and novobiocin yielded bacteriostatic effects. Rifampin was bactericidally active against rifampin-susceptible strains (MICs = 0.125 micrograms/ml), but less so against low-level-rifampin-resistant (MICs = 2-8 micrograms/ml) strains in MHB. However, in the presence of human blood, rifampin (2 micrograms/ml) combined with co-trimoxazole (0.25/4.75 micrograms/ml) killed rifampin-susceptible and low-level-rifampin-resistant, but not moderate-level-rifampin-resistant (MICs = 16-32 micrograms/ml) strains of E. faecium. Of two topical drugs examined, mupirocin merely inhibited strains of E. faecium; conversely, taurolidine at 2,000 micrograms/ml was efficacious against all strains examined, although the kinetics of bactericidal activity were retarded somewhat in the presence of 65 vol% human blood.     

Antimicrobial activity of quinupristin-dalfopristin (RP 59500, Synercid) tested against over 28,000 recent clinical isolates from 200 medical centers in the United States and Canada

A total of 200 medical center laboratories in the USA and Canada contributed results of testing quinupristin-dalfopristin, a streptogramin combination (formerly RP 59500 or Synercid), against 28,029 Gram-positive cocci. Standardized tests [disk diffusion, broth microdilution, Etest (AB BIODISK, Solna, Sweden)] were utilized and validated by concurrent quality control tests. Remarkable agreement was obtained between test method results for characterizing the collection by the important emerging resistances: 1) oxacillin resistance among Staphylococcus aureus (41.0 to 43.7%); 2) vancomycin resistance among Enterococcus faecium (50.0 to 52.0%); and 3) the penicillin nonsusceptible rate for pneumococci (31.1% overall, with 10.6% at MICs of > or = 2 micrograms/mL). The quinupristin-dalfopristin MIC90 for oxacillin-susceptible and -resistant S. aureus was 0.5 microgram/mL and 1 microgram/mL, respectively. The quinupristin-dalfopristin MIC90 for vancomycin-resistant E. faecium was 1 microgram/mL, and only 0.2% of isolates were resistant. Other Enterococcus species were generally not susceptible to the streptogramin combination but were usually inhibited by ampicillin (86 to 97% susceptible; MIC50, 1.0 microgram/mL) or vancomycin (86 to 95%; MIC50, 1.0 microgram/mL). Among all tested enterococci, the rate of vancomycin resistance was 16.2%. The quinupristin-dalfopristin MIC90 (0.75 microgram/mL) for 4,626 tested Streptococcus pneumoniae strains was not influenced by the penicillin or macrolide susceptibility patterns. When five regions in the USA and Canada were analyzed for significant streptogramin and other antimicrobial spectrum differences, only the Farwest region had lower numbers of streptogramin-susceptible E. faecium. Canadian strains were generally more susceptible to all drugs except chloramphenicol and doxycycline when tested against E. faecalis (73% and 89% susceptible, respectively). The U.S. Southeast region had S. pneumoniae strains less susceptible to macrolides (73%) but had more susceptibility among E. faecium isolates tested against vancomycin and ampicillin. The Northeast region of the USA had the greatest rate of vancomycin resistance among enterococci. Strains retested by the monitor because of quinupristin-dalfopristin resistance (MICs, > or = 4 micrograms/mL) were generally not confirmed (2.2% validation), and only 0.2% of E. faecium isolates were identified as truly resistant. The most common errors were: 1) species misidentification (28.0%); 2) incorrect susceptibility results (65.6%); and 3) mixed cultures (4.3%) tested by participants. Overall, quinupristin-dalfopristin was consistently active (> or = 90% susceptible) against major Gram-positive pathogens in North America, regardless of resistance patterns to other drug classes and geographic location of their isolation.     

Cytokine autoantibodies in multiple sclerosis, aseptic meningitis and stroke

BACKGROUND AND OBJECTIVE: To determine and then compare the time-kill profiles of Enterococcus to antibiotics used for intravitreal therapy. PATIENTS AND METHODS: The time-kill profiles of four endophthalmitis isolates of Enterococcus faecalis, one vancomycin-resistant E. faecalis isolate, and three vancomycin-resistant isolates of E. faecium were determined against vancomycin, amikacin, cefazolin, gentamicin, ampicillin, ciprofloxacin, ceftazidime, clindamycin, and the combinations of vancomycin and amikacin, vancomycin and ceftazidime, vancomycin and gentamicin, vancomycin and ampicillin, cefazolin and gentamicin, and ampicillin and gentamicin. RESULTS: No single antibiotic or combination was bactericidal (defined as 99.9% kill) to all isolates of Enterococcus. Gentamicin was bactericidal to all E. faecalis isolates. None of the tested antibiotics were bactericidal to vancomycin-resistant E. faecium. CONCLUSIONS: The time-kill profiles demonstrated that vancomycin and ceftazidime did not produce a 99.9% kill for E. faecalis in this small study. Gentamicin combined with either cefazolin or ampicillin had somewhat better bactericidal activity and should be considered as an alternative therapy. Novel therapy may be necessary to treat endophthalmitis because of vancomycin-resistant Enterococcus, depending on the susceptibility patterns of the individual isolate and the response to initial therapy.     

Heat tolerance of vancomycin resistant Enterococcus faecium

The heat tolerance of 27 Enterococcus faecium isolates in water was studied. Stationary phase cultures including vancomycin resistant and sensitive clinical and food isolates were exposed to heat at 60 degrees, 65 degrees, 71 degrees, and 80 degrees C for one, three, 10, and 30 minutes and the log10 reductions in bacterial counts were determined. Exposure at 71 degrees and 80 degrees C resulted in > 6 log10 reduction in viable counts for all isolates. Seven (24%) isolates survived (< 5 log10 reduction) heat at 65 degrees C for 10 minutes. The E faecium isolates were more resistant to heat than the two E faecalis reference strains. No differences in heat tolerance were observed between vancomycin sensitive and resistant strains or between isolates of human or food origin.     

Vancomycin resistant enterococci

Enterococci are normal intestinal flora in humans. Among enterococci, Enterococcus faecalis and Enterococcus faecium are frequently isolated and can become nosocomial pathogens in hospitals, especially in intensive care units and oncology wards. Recently, vancomycin-resistant enterococci (VRE) such as E. faecalis and E. faecium have caused a serious problem of hospital-acquired infections in Europe and the USA. VRE also has another aspect as a cause of community-acquired infections. Especially, avoparcin which had been used to enhance growth of food animals is documented as associated with the spread of VRE in European countries. In Japan, there have only been a few of reports about VRE so far. However, there evidence that VRE might become prevalent in many hospitals in Japan. In fact, we have already isolated another highly vancomycin-resistant E. faecium (VCM:MIC > 128 micrograms/ml) from a hospitalized diabetic patient. We should pay a careful attention to VRE and perform the following control measures: 1)re-education and re-training about hospital infection control procedures, 2) prudent use of vancomycin in clinical settings, 3)accurate report of VRE in clinical laboratories, and 4) good communications and collaborations among physicians, nurses and other health care personnel and laboratory technicians. We should learn more from countries in which VRE are already prevalent, and pursue further investigations, to prevent the spread of VRE in Japan.     

Multicenter trial of Enterococcus antibiotic susceptibility

Antibiotic susceptibility of 446 Enterococcus isolates from 9 medical centres of Moscow and St. Petersburg was tested. Among the isolates 386 belonged to E.faecalis, 48 to E.faecium and 12 to the other species. All the isolates were susceptible to vancomycin. As for E.faecalis 84 and 85 per cent of the isolates were susceptible to ampicillin and ampicillin/sulbactam respectively (no production of beta-lactamases), the frequency of high resistance to aminoglycosides amounted to 44 per cent with respect to streptomycin and to 25 per cent with respect to gentamicin, 75 per cent of the isolates was susceptible to ciprofloxacin. As for E.faecium and the rare species of Enterococcus more than 70 per cent of the isolates was resistant to ampicillin and ampicillin/sulbactam, the frequency of high resistance to aminoglycosides exceeded 60 per cent, 17 and 25 per cent of the isolates were susceptible to ciprofloxacin.     

Preferential hydrolysis of oxidized phosphatidylcholine in cholesterol-containing phosphatidylcholine liposome by phospholipase A2

From 125 separate cloacal cultures from three turkey flocks fed virginiamycin, 104 Enterococcus faecium and 186 Enterococcus faecalis isolates were obtained. As the turkeys aged, there was a higher percentage of quinupristin-dalfopristin-resistant E. faecium isolates, with isolates from the oldest flock being 100% resistant. There were no vancomycin-resistant enterococci. Results of pulsed-field gel electrophoresis (PFGE) indicated there were 11 PFGE types of E. faecalis and 7 PFGE types of E. faecium that were in more than one group of flock cultures.     

Genetically programmed development of salivary gland abnormalities in the NOD (nonobese diabetic)-scid mouse in the absence of detectable lymphocytic infiltration: a potential trigger for sialoadenitis of NOD mice

In a study designed to gain data on the in vitro transferability of vancomycin resistance from enterococci of the VanA phenotype to listeriae of different species, three clinical Enterococcus isolates-Enterococcus faecium LS10, Enterococcus faecalis LS4, and Enterococcus faecalis A3208, all harboring a plasmid that strongly hybridized with a vanA probe-were used as donors in transfer experiments. Strains of five Listeria species were used as recipients. From Enterococcus faecium LS10, glycopeptide resistance was transferred to Listeria monocytogenes, Listeria ivanovii, and Listeria welshimeri recipients, whereas no transfer occurred to Listeria seeligeri or Listeria innocua strains. From the two Enterococcus faecalis isolates, no transfer occurred to any Listeria recipient. MICs of both vancomycin and teicoplanin were > or = 256 mg/l for all transconjugants tested. Furthermore, all transconjugants harbored a plasmid that strongly hybridized with the vanA probe, with vanA consistently located in an EcoRI fragment of about 4 kb. Exposure of Listeria transconjugants to vancomycin resulted in synthesis of a membrane protein similar in size (39 kDa) to a vancomycin-induced membrane protein of Enterococcus faecium LS10. In retransfer experiments with Listeria transconjugants used as donors, glycopeptide resistance was transferred to all Listeria recipients tested, including strains of Listeria innocua and Listeria seeligeri, which were unable to receive the resistance from Enterococcus faecium LS10. The frequency of vanA transfer to listerial recipients was greater in retransfer experiments than in the primary matings. These findings suggest that the vanA resistance determinant might spread to the established pathogen Listeria monocytogenes, both directly from a resistant enterococcus and through strains of nonpathogenic Listeria species acting as intermediate resistance vehicles.     

Unstable atherosclerotic plaque. Pathophysiology and therapeutic guidelines

One hundred fifty clinical isolates of Enterococcus faecalis (88 isolates) and Enterococcus faecium (62 isolates) were tested in vitro for their susceptibility to vancomycin and high-level aminoglycosides (HLA). Remel's Synergy Quad Plates (RSQ) were used as the reference method and compared to Kirby-Bauer disc diffusion test, Vitek GPS-TA card, MicroScan Panel (GP-6), and Etest. Streptomycin susceptibility results for MicroScan GP-6 and RSQ were recorded at 24 and 48 h and all other methods and antibiotics were read at 24 h or less. When compared with the agar screen method, all of the methods demonstrated > 99% agreement. One isolate was falsely sensitive to gentamicin at 24 h, but resistant at 48 h, when tested on both MicroScan and RSQ agar screen. Thirty-nine isolates showed resistance to vancomycin with all methods. These isolates were from three different local hospitals and were identified as E. faecium. Pulse-field gel electrophoresis demonstrated that all of the vancomycin-resistant isolates were derived from the same clone. Of interest is the observation that high-level resistance to aminoglycosides varied between the clonally related isolates.     

Successful treatment of persistent bacteremia due to vancomycin-resistant, ampicillin-resistant Enterococcus faecium

Emergence of vancomycin-resistant enterococci has become an increasing problem in many medical centers. We report a liver transplant recipient with vancomycin-resistant Enterococcus faecium bacteremia who was successfully treated using very high dose continuous infusion ampicillin/sulbactam, plus gentamicin after he remained bacteremic on high dose ampicillin and gentamicin. At our institution, 83% of E. faecium isolates from 1994 were inhibited by ampicillin/sulbactam compared to 66% for ampicillin at an MIC < or = 64 micrograms/ml. None of these strains produced beta-lactamase, suggesting sulbactam may have an unexplained beneficial effect against some enterococci. Although an MIC of < or = 8 micrograms/ml is required for ampicillin to be considered active against enterococci, much higher levels of ampicillin or ampicillin/sulbactam are safely achievable. The response of our patient and the reported in vivo data have implications for future treatment of this pathogen, and may necessitate a reevaluation of susceptibility interpretation guidelines by clinical laboratories, and therapeutic drug dosing by clinicians.     

Selective isolation of vancomycin-resistant enterococci

Broth formulations of two media selective for enterococci, Enterococcel, M-Enterococcosel broths were supplemented with 6 micrograms of vancomycin per ml and evaluated for isolation of vancomycin-resistant enterococci (VRE). Each broth was challenged with various concentrations of Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and vancomycin-susceptible and vancomycin-resistant enterococci and with 193 perianal specimens obtained from patients at risk in our institution for VRE colonization. Both the Enterococcosel and M-Enterococcus broths with vancomycin detected as few as 1 to 9 CFU of VRE while inhibiting growth of the other organisms tested. Enterococcus faecium organisms (MIC, > 256 micrograms/ml) were recovered from 66 perianal swab cultures in the enterococcosel-vancomycin broth, and VRE were recovered from 62 perianal swab cultures in the M-Enterococcus-vancomycin broth. Enterococcosel-vancomycin broth detected VRE in perianal specimens 48 h earlier than did M-Enterococcus-vancomycin broth. Enterococcosel broth with 6 micrograms of vancomycin per ml can be used for the rapid and selective isolation of VRE from surveillance specimens.     

Lipoid proteinosis of the oral mucosa: case report and review of the literature

High-level resistance (minimum inhibitory concentration, MIC > 1,000 micrograms/ml) to gentamicin (HLGR) in enterococci is common in Taiwan. In this study, we investigated the distribution of gentamicin resistance elements in enterococci isolated at National Taiwan University Hospital in a 1-year period, and also examined the transfer and the genetic variability of the resistance elements of different isolates. Among 109 isolates tested, 43 (39%) HLGR isolates were identified. HLGR was most common in Enterococcus faecium isolates (7/15, 47%), followed by Enterococcus faecalis (34/80, 43%), Enterococcus avium (1/5, 20%), and Enterococcus casseliflavus (1/9, 11%). To understand the mechanism of resistance transfer, four isolates of E. faecalis and five isolates of E. faecium showing HLGR were studied. Transfer of resistance markers to a plasmid-free recipient strain of E. faecalis JH2-7 was observed, with transfer frequencies ranging from 10(-2) to 10(-8). All of the transconjugants contained plasmids, with sizes ranging from 45 kb to larger than 70 kb. At least three plasmid patterns were observed on digestion with HaeIII. Hybridization with a probe specific for the aac6'aph2" gentamicin resistance gene confirmed that all of these HLGR isolates carried a Gm(r) determinant, though the hybridization patterns of the plasmids from E. faecalis and E. faecium were different. Although many similarities exist among enterococcal Gm(r) determinants, the results suggest heterogeneity may occur in the flanking regions of resistance elements.     

Antibiotic susceptibility testing (agar disk diffusion and agar dilution) of clinical isolates of Enterococcus faecalis and E. faecium: comparison of Mueller-Hinton, Iso-Sensitest, and Wilkins-Chalgren agar media

Forty-two isolates of Enterococcus faecalis and 56 isolates of Enterococcus faecium, including 8 vancomycin-resistant strains, were examined for comparative susceptibility to 27 antimicrobial drugs with the agar dilution method, employing Mueller-Hinton (MHA), Iso-Sensitest (ISTA), and Wilkins-Chalgren (WCA) agar. The Bauer-Kirby agar disk diffusion method was used to comparatively test 24 of the agents in parallel. The enterococci yielded better growth on ISTA and WCA. However, WCA completely antagonized co-trimoxazole and, though less, fosfomycin. Importantly, WCA slightly reduced the activities of teicoplanin (minimal inhibitory concentrations, MICs, raised up to twofold) and vancomycin (MICs raised two- to fourfold) against enterococci and staphylococcal quality control strains. Therefore, WCA was judged unsuitable for susceptibility testing of enterococci. For E. faecalis no discrepancies between agar dilution MICs and inhibition zone diameters were encountered with augmentin, ampicillin, ampicillin-sulbactam, chloramphenicol, mupirocin, oxacillin, teicoplanin, and co-trimoxazole. Overall, MHA yielded fewer very major (category I) and major (category II) discrepancies than ISTA. However, numerous minor (category III), slight (category IV), minimal (category V), and/or negligible (category VI) discrepancies were encountered with ciprofloxacin, doxycycline, erythromycin, fosfomycin, fusidic acid, meropenem, ofloxacin and rifampin. With respect to E. faecium, only cefotaxime, mupirocin, oxacillin, and teicoplanin yielded nondiscrepant results. Several very major (I) and major (II) discrepancies were observed with augmentin, ampicillin, ampicillin-sulbactam, doxycycline, fusidic acid, imipenem, and penicillin G. Minor discrepancies (categories III-VI) were particularly numerous with augmentin, chloramphenicol, ciprofloxacin, doxycycline, and piperacillin. The largest numbers of negligible (VI) discrepancies were noted with fosfomycin, fusidic acid, and ofloxacin. It is recommended to test one cephalosporin (cefuroxime or the like) in parallel for educational purposes and to exclude fosfomycin, fusidic acid, and rifampin from test batteries because of the wide scatter of test results. The large number of minimal (V) discrepancies of ciprofloxacin against E. faecalis, the numerous minor (III) and slight (IV) discrepancies of chloramphenicol against E. faecium, and the not insignificant number of very major (I) and minor (III) discrepancies observed with meropenem against isolates of E. faecalis necessitated proposals for new disk intermediate susceptibility criteria.     

Evidence that ferritin is UV inducible in human skin: part of a putative defense mechanism

The low-affinity penicillin-binding protein (PBP) 5 is the main beta-lactam target and is responsible for resistance to this class of antibiotics in Enterococcus faecium. The PBP 5 variants of 15 clinical isolates (including 8 resistant to vancomycin) with different levels of beta-lactam resistance were analyzed. Most of the highly beta-lactam-resistant isolates produced small quantities of PBP 5 of low affinity. This was associated with particular amino acid substitutions: an Ala or Ile for Thr-499, a Glu for Val-629, and a Pro for Ser-667. A change of Met-485 to Thr or Ala (adjacent to the conserved SDN box) was observed in isolates with MICs of ampicillin of 64 or 128 microg/mL, respectively. In the 2 most resistant isolates, with MICs of ampicillin of 256 microg/mL, an additional Ser was present just after Ser-466. Thus, particular point mutations in PBP 5 and combinations thereof may lead to high-level beta-lactam resistance in E. faecium.     

Characterization of staphylococci with reduced susceptibilities to vancomycin and other glycopeptides

During the last several years a series of staphylococcal isolates that demonstrated reduced susceptibility to vancomycin or other glycopeptides have been reported. We selected 12 isolates of staphylococci for which the vancomycin MICs were > or =4 microg/ml or for which the teicoplanin MICs were > or =8 microg/ml and 24 control strains for which the vancomycin MICs were < or =2 microg/ml or for which the teicoplanin MICs were < or =4 microg/ml to determine the ability of commercial susceptibility testing procedures and vancomycin agar screening methods to detect isolates with reduced glycopeptide susceptibility. By PCR analysis, none of the isolates with decreased glycopeptide susceptibility contained known vancomycin resistance genes. Broth microdilution tests held a full 24 h were best at detecting strains with reduced glycopeptide susceptibility. Disk diffusion did not differentiate the strains inhibited by 8 microg of vancomycin per ml from more susceptible isolates. Most of the isolates with reduced glycopeptide susceptibility were recognized by MicroScan conventional panels and Etest vancomycin strips. Sensititre panels read visually were more variable, although with some of the panels MICs of 8 microg/ml were noted for these isolates. Vitek results were 4 microg/ml for all strains for which the vancomycin MICs were > or =4 microg/ml. Vancomycin MICs on Rapid MicroScan panels were not predictive, giving MICs of either < or =2 or > or =16 microg/ml for these isolates. Commercial brain heart infusion vancomycin agar screening plates containing 6 microg of vancomycin per ml consistently differentiated those strains inhibited by 8 microg/ml from more susceptible strains. Vancomycin-containing media prepared in-house showed occasional growth of susceptible strains, Staphylococcus aureus ATCC 29213, and on occasion, Enterococcus faecalis ATCC 29212. Thus, strains of staphylococci with reduced susceptibility to glycopeptides, such as vancomycin, are best detected in the laboratory by nonautomated quantitative tests incubated for a full 24 h. Furthermore, it appears that commercial vancomycin agar screening plates can be used to detect these isolates.     

Avoparcin used as a growth promoter is associated with the occurrence of vancomycin-resistant Enterococcus faecium on Danish poultry and pig farms

We determined the association between the use of the glycopeptide antibiotic avoparcin as a growth promoter and the occurrence of Enterococcus faecium (VREF) with high-level resistance to vancomycin (MIC > or = 64 micrograms ml-1) on poultry and pig farms. The investigations were conducted as retrospective cohort studies, where groups of farms exposed or not exposed to avoparcin between September 1994 and April 1995 were compared. In poultry, the association between the use of avoparcin and the occurrence of VREF was confounded by the use of broad-spectrum antibiotics, and the adjusted relative risk was 2.9 (1.4-5.9). In pigs, the association had a similar magnitude with a non-adjusted relative risk of 3.3 (0.9-12.3). The similar findings in the two studies provide evidence in favour of a causal association between the use of avoparcin and the occurrence of VREF on farms, and suggest that food animals constitute a potential reservoir of infection for VREF in humans.     

Influence of erythromycin resistance, inoculum growth phase, and incubation time on assessment of the bactericidal activity of RP 59500 (quinupristin-dalfopristin) against vancomycin-resistant Enterococcus faecium

RP 59500, a mixture of two semisynthetic streptogramin antibiotics (quinupristin and dalfopristin), is one of a few investigational agents currently in clinical trials with inhibitory activity against multiple-drug-resistant strains of Enterococcus faecium. We evaluated the bactericidal activity of this antimicrobial against 30 recent clinical isolates of vancomycin-resistant E. faecium, including 23 erythromycin-resistant (MIC, >256 microg/ml) and 7 erythromycin-intermediate (MIC, 2 to 4 microg/ml) strains. All isolates were inhibited by RP 59500 at 0.25 to 1.0 microg/ml. The bactericidal activity of RP 59500 was markedly influenced by the erythromycin susceptibility of the strains and by several technical factors, such as inoculum growth phase and time of incubation of counting plates. As determined by time-kill methods, RP 59500 at a concentration of 2 or 8 microg/ml failed to kill erythromycin-resistant organisms under any conditions. Bactericidal activity was observed against all seven erythromycin-intermediate isolates when log-phase inocula were used and the cells were counted after 48 h of incubation (mean reductions in viable bacteria for RP 59500 at concentrations of 2 and 8 microg/ml, 3.45 and 3.50 log10 CFU/ml, respectively), but killing was diminished when the plates were examined at 72 h (mean killing, 3.06 and 2.95 log10, CFU/ml, respectively). No bactericidal activity was observed when stationary-phase cultures were used. On the basis of these data, we expect that bactericidal activity of RP 59500 against the multiple-drug-resistant E. faecium strains currently encountered would be distinctly uncommon.     

Emergence of vancomycin-resistant enterococci in Australia: phenotypic and genotypic characteristics of isolates

Enterococci with resistance to glycopeptides have recently emerged in Australia. We developed multiplex PCR assays for vanA, vanB, vanC1, and vanC2 or vanC3 in order to examine the genetic basis for vancomycin resistance in Australian isolates of vancomycin-resistant Enterococcus faecium and E. faecalis (VRE). The predominant genotype from human clinical E. faecium isolates was vanB. The PCR van genotype was consistent with the resistance phenotype in all but six cases. One vanA E. faecalis isolate had a VanB phenotype, one vanB E. faecium isolate had a VanA phenotype, and four E. faecalis isolates were consistently negative for vanA, vanB, vanC1, and vanC2 or vanC3, even though they exhibited a VanB phenotype. These four isolates were subsequently examined for the presence of vanD by published methods and were found to be negative. No vancomycin-susceptible strains produced a PCR product. On the basis of our findings the epidemiology of VRE in Australia appears to be different from that in either the United States or Europe. Our multiplex PCR assays gave a rapid and accurate method for determining the genotype and confirming the identification of glycopeptide-resistant enterococci. Rapid and accurate methods are essential, because laboratory-based surveillance is critical in programs for the detection, control, and prevention of the transmission of glycopeptide-resistant enterococci.