Microb Drug Resist, 1996 Summer, 2(2), 225 - 9 Peptidoglycan composition of vancomycin-resistant Enterococcus faecium; de Jonge BL et al.; Muropeptide composition of peptidoglycan isolated from isogenic vancomycin-resistant and sensitive Enterococcus faecium strains was analyzed by reverse-phase high-performance liquid chromatography combined with amino acid and fast atom bombardment mass spectrometric analyses . Peptidoglycan of the sensitive and resistant strains was the same and was composed of tri- and tetrapeptides stem peptide subunits with or without aspartate or asparagine substitutions on the epsilon-amino group of the lysine residue . Thus, the synthesis of lactate-terminating peptidoglycan precursors in vancomycin-resistant E . faecium did not affect the chemical composition of peptidoglycan. Microb Drug Resist, 1996 Summer, 2(2), 219 - 23 Genetics of glycopeptide resistance in enterococci; Evers S et al.; Glycopeptide resistance in enterococci is phenotypically and genotypically heterogeneous . The genes responsible for inducible resistance to high levels of vancomycin and teicoplanin (VanA phenotype) are carried by the 10,851-bp Tn1546 transposon . Transposition of Tn1546 into self-transferable plasmids and subsequent transfer by conjugation appears to be responsible for the dissemination of this type of resistance . Nine polypeptides are encoded by Tn1546 that belong to five functional groups: transposition functions (ORF1 and ORF2), regulation of resistance gene expression (VanR and VanS), synthesis of depsipeptide D-Ala-D-lactate (VanH and VanA), hydrolysis of D-Ala-D-Ala-containing peptidoglycan precursors (VanX and VanY), and low-level teicoplanin resistance (VanZ) . VanB-type resistance (various levels of resistance to vancomycin and susceptibility to teicoplanin) is also due to production of D-Ala-D-Lac . The VanB ligase of VanB-type strains is structurally and functionally similar to VanA . The vanB gene was found on composite transposon Tn1547, which, in turn, was part of larger conjugative chromosomally located elements (90 to 250 kb) . In contrast to acquired VanA- and VanB-type resistance, VanC-type resistance (low level of resistance to vancomycin and susceptibility to teicoplanin) is an intrinsic property of motile enterococci . Resistance in these species is due to synthesis of dipeptide D-Ala-D-Ser by VanC ligases leading to production of cell wall precursors with reduced vancomycin affinity. Diagn Microbiol Infect Dis, 1996 Jul, 25(3), 127 - 31 Comparative in vitro activity of quinupristin/dalfopristin against multidrug resistant Enterococcus faecium; Bonilla HF et al.; The in vitro susceptibilities of 82 strains of vancomycin resistant Enterococcus faecium (VREF), (49 vanA and 33 vanB) from over 13 hospitals in Europe and United States were studied . The MIC for several antibiotics showed high levels of resistance to vancomycin, ampicillin, gentamicin, and imipenem . All VREF strains were highly susceptible to quinupristin/dalfopristin with a MIC 90% of 0.5 microgram/ml for both vanA and vanB phenotypes . Time-kill and synergy studies of VREF for quinupristin/dalfopristin alone and quinupristin/dalfopristin in combination with several antibiotics (ampicillin, gentamicin, ciprofloxacin, rifampin and novobiocin) did not show bactericidal activity . In induction experiments using SF6550, (VREF, a vanA strain), quinupristin/dalforpristin showed a delay in the expression of vancomycin resistance by 2.5 hours . The results of this study show quinupristin/dalfopristin to have excellent in vitro activity versus multiple resistant E . faecium. Electrophoresis, 1996 Jul, 17(7), 1234 - 41 Purification of glycopeptide antibiotics by isoelectric focusing in multicompartment electrolyzers with immobiline membranes; Bossi A et al.; The purification of small glycopeptides (a Hepta-Tyr of the teicoplanin family, exhibiting broad activity against highly glycopeptide-resistant enterococci) by isoelectric focusing in multicompartment electrolyzers with buffering, isoelectric membranes, is described . The main obstacle to such a preparative technique, in common with all focusing methodologies, is the poor solubility of the analyte at the pI value with resultant precipitation and coprecipitation of all impurities with the main fraction . A good solubilizing power was obtained in hydro-organic solvents, particularly a mixture of 6 M urea and 20-25% trifluoroethanol . Best results, however, were obtained with mixtures of 8 M urea and zwitterionic detergents, notably the 3-{(3-cholamidopropyl)dimethylammonia}-1-propanesulfonate (CHAPS) family . A unique behavior of the peptide was found in concentration gradients of CHAPS: solubility increases up to 3.5% CHAPS, but the curve shows a maximum and then solubility decreases again at 5% CHAPS . In mixtures of 8 M urea and 3.5% CHAPS, sample loads of 500 up to 1000 mg Hepta-Tyr could be purified in a single run, with recoveries > 90% and purity in excess of 99% . The main glycopeptide fraction (pI 8.56) was collected into an isoelectric trap delimited by pI 8.46 and pI 8.65 membranes . Attempts at purifying the glycopeptide by most known reversed phase-high performance liquid chromatography (RP-HPLC) techniques failed completely. Pharmacotherapy, 1996 Jul-Aug, 16(4), 584 - 92 Treatment of vancomycin-resistant enterococci, with a focus on quinupristin-dalfopristin; Fuller RE et al.; Enterococci are the second most common cause of hospital-acquired infections, and drug resistance among these organisms is a growing problem . Vancomycin-resistant enterococci (VRE) now account for 7.9% of the nosocomial enterococcal infections . There is no standard therapy for VRE . Although some agents have shown in vitro activity alone or in combination, including ciprofloxacin, doxycycline, novobiocin, teicoplanin, chloramphenicol, and rifampin, treatment options are limited to combinations of drugs with marginal efficacy against the pathogens . Quinupristin-dalfopristin is a new investigational agent with activity against gram-positive cocci, including VRE. J Hosp Infect, 1996 Jul, 33(3), 191 - 200 Investigation of an outbreak of vancomycin-resistant Enterococcus faecium by random amplified polymorphic DNA (RAPD) assay; Issack MI et al.; In this study, 122 isolates of vancomycin-resistant Enterococcus faecium (VREF) obtained from 103 patients over a four-year period in a London teaching hospital, were typed by a random amplified polymorphic DNA method . All the isolates exhibited high-level resistance to vancomycin (MIC 128-1024 mg/L), and were resistant to teicoplanin (32-256 mg/L) . Nine RAPD types were distinguished by using a single primer . Clustering of certain types in time and space was noted . These results suggest that although several different strains of VREF were involved in this outbreak, cross-infection with individual types occurred on some wards . RAPD is a useful technique for the investigation of the epidemiology of VREF. Am J Health Syst Pharm, 1996 Jul 1, 53(13), 1570 - 5 Limiting vancomycin use to combat vancomycin-resistant Enterococcus faecium; Belliveau PP et al.; The implementation, monitoring, and impact of a program to restrict vancomycin use are described . A vancomycin restriction program was implemented in February 1995 at an acute care teaching hospital after guidelines for vancomycin use were established by a multidisciplinary group . Pharmacists reviewed each vancomycin order, suggested alternative treatments when vancomycin use did not comply with the guidelines, and set duration limits for all orders . Orders requiring greater clinical experience for review were referred to a pharmacist or a physician in the division of infectious diseases . The program was monitored by an infectious diseases pharmacist . Data collected after the program was established showed that the volume of vancomycin use decreased substantially . Problems in enforcing the restriction program included administration of vancomycin for surgical prophylaxis before the order reached the pharmacy, continuing use of vancomycin for initial empirical treatment of febrile neutropenic and immunocompromised patients, inadequate tracking of the evaluations, and deficiencies in the evaluations related to a need for continuing education of the pharmacists about the program . Use of vancomycin decreased after a pharmacy-enforced restriction program was implemented. Antimicrob Agents Chemother, 1996 Jul, 40(7), 1745 - 7 In vitro activities in new oxazolidinone antimicrobial agents against enterococci; Eliopoulos GM et al.; The comparative in vitro activities of two new oxazolidinone antimicrobial agents, U-100592 and U-100766, against 180 isolates of enterococci representing several resistance profiles were examined by using an agar dilution technique . The two oxazolidinones inhibited all isolates, including strains resistant to vancomycin, ampicillin, and minocycline, at concentrations between 1 and 4 micrograms/ml. Antimicrob Agents Chemother, 1996 Jul, 40(7), 1645 - 8 Induction signals for vancomycin resistance encoded by the vanA gene cluster in Enterococcus faecium; Lai MH et al.; The induction of vancomycin resistance in enterococci containing the vanA gene cluster is thought to be controlled by a two-component sensor-response regulator system encoded by vanR and vanS . Eight inducing compounds were identified by screening a panel of more than 6,800 antibiotics and synthetic compounds including the three tested glycopeptides (vancomycin, avoparcin, and ristocetin), two other cell wall biosynthesis inhibitors (moenomycin and bacitracin), two cyclic peptide antibiotics (antibiotic AO341 beta and polymyxin B), and a macrocyclic lactone antibiotic (moxidectin) . Induction activity by structurally unrelated antibiotics suggests that the induction signal is not a structural feature of vancomycin. Gene, 1996 Jun 26, 172(2), 239 - 43 The Drosophila melanogaster gene vha14 encoding a 14-kDa F-subunit of the vacuolar ATPase; Guo Y et al.; A Drosophila melanogaster (Dm) cDNA (vha14) encoding the 14-kDa F-subunit of the vacuolar H(+)-ATPase (V-ATPase) has been cloned via homology with the corresponding Manduca sexta (Ms) gene . Its deduced translation product is a 124-amino-acid polypeptide sharing 90% identity with the Ms polypeptide and 50% identity with an analogous polypeptide of Saccharomyces cerevisiae, and a more distant similarity to a subunit of the Na(+)-transporting ATPase of Enterococcus hirae . Homology was also found with expressed sequence tags from man, Arabidopsis thaliana, Caenorhabditis elegans and C . briggsiae, Oryza sativa and Plasmodium falciparum, indicating that the subunit is phylogenetically conserved . The Dm gene (vha14) is present as a single copy at cytological position 52B on the second chromosome, and gives rise to an mRNA species of 0.65 kb . Expression of the latter shows relatively little variation during development, or between adult head, thorax and abdomen, suggesting that the F-subunit is a relatively ubiquitous component of the V-ATPase. Ann Pharmacother, 1996 Jun, 30(6), 680 - 2 Vancomycin resistance: when failure becomes an opportunity for leadership; Edmiston CE Jr; OBJECTIVE: To discuss the emergence of the enterococci as significant nosocomial pathogens and reports of glycopeptide resistance as demonstrating the failure of healthcare professionals to limit the clinical impact of these organisms . BACKGROUND: The enterococci have long occupied a peculiar position in medical and surgical patients . A component of the normal gastrointestinal tract, these organisms exhibit little overt pathogenicity in healthy hosts, but are frequently recovered in patients with severe debilitative or immunosuppresive disorders . While the enterococci have always demonstrated intrinsic resistance to a broad range of antiinfective agents, recent findings of moderate to high-level glycopeptide resistance potentially threaten the limited therapeutic options for methicillin-resistant gram-positive cocci . FINDINGS: The emergence and dissemination of vancomycin-resistant enterococci are signs of much greater problems, which include incomplete success of formulary controls, unreliable detection and identification of resistant microorganisms within the hospital environment, and poor fundamental infection control practices by all healthcare professionals . CONCLUSIONS: The Hospital Infection Control Practices . Advisory Committee Recommendations for the Prevention and Spread of Vancomycin Resistance are an important step in resolving these issues through the elements of collegiality and shared leadership. J Antibiot (Tokyo), 1996 Jun, 49(6), 575 - 81 Reductive alkylation of glycopeptide antibiotics: synthesis and antibacterial activity; Cooper RD et al.; Reductive alkylation of the A82846 family of glycopeptide antibiotics has the potential of producing seven products . N-Alkylation of the disaccharide amino function can be accomplished selectively, and offers the greatest increase in antibacterial activity . Products resulting from N-alkylation of LY264826 (A82846B) provide the most potent derivatives as compared to other members of this class of antibiotics . Two of these derivatives, LY307599 and LY333328 are approximately 500 times more active than vancomycin against vancomycin-resistant enterococci. Lett Appl Microbiol, 1996 Jun, 22(6), 417 - 9 Purification and N-terminal amino acid sequence of Enterocin CRL 35, a 'pediocin-like' bacteriocin produced by Enterococcus faecium CRL 35; Farias ME et al.; Enterocin CRL 35, a bacteriocin produced by Enterococcus faecium CRL 35 that inhibits food-borne pathogens, was purified by precipitation with (NH4)2SO4, gel filtration, ion exchange and reverse phase chromatography . The partial N-terminal amino acid sequence indicated a strong homology with other 'pediocin-like bacteriocins' previously described. Planta Med, 1996 Jun, 62(3), 275 - 7 Comparison of antimicrobial properties of monoterpenes and their carbonylated products; Naigre R et al.; Some monoterpenes and their carbonylated products were evaluated for their antibacterial and antifungal properties . The carbonylation of tested monoterpenes was shown to increase the bacteriostatic and fungistatic activities specifically by the contact method . Concerning the killing effects, only (1R,2S,5R)-isopulegol, its carbonylated products, and (R)-carvone showed significant bactericidal activities, particularly against Enterococcus faecium and Escherichia coli above a concentration of 10 microliters/ml . A fungicidal efficiency of (1R,2S,5R)-isopulegol and (R)-carvone against Aspergillus niger was also noted . It seems that the presence of an oxygenated function in the framework increases the antimicrobial properties . However, monoterpenes were more active using a micro-atmosphere method. Diagn Microbiol Infect Dis, 1996 May, 25(1), 15 - 20 Selection of Enterococcus faecium strains with stable and unstable resistance to the streptogramin RP 59500 using stepwise in vitro exposure; Millichap J et al.; Therapy of vancomycin-resistant enterococcal infections is an increasing problem for many large medical centers . One promising agent for use against Enterococcus faecium is RP 59500 (Quinupristin/Dalfopristin) . To assess the potential for emergence of resistant strains during clinical trials with this new compound, we collected and tested 11 vancomycin-resistant and three vancomycin-susceptible E . faecium strains . The strains were exposed to doubling dilutions of RP 59500, beginning at drug concentrations ranging from 0.25 to 16 micrograms/ml agar . A saturated swab with approximately 3 x 10(7) organisms/ml was spread as a lawn on agar containing RP 59500 and incubated at 35 degrees C for 48 h . Growth on the highest drug-containing plate was used to prepare the inoculum for the next series of resistance-selection experiments . After the final passage, the range of the highest RP 59500 plate concentration with growth was 32-512 micrograms/ml (initial resistance frequency ranging from 1 x 10(-6) to > 1 x 10(-4) . After the selection of resistant strains, the organisms were passed twice weekly on antimicrobial agent-free media to determine resistance stability in the absence of drug . Resistance in strains with an RP 59500 MIC of > or = 16 micrograms/ml was stable after repeated passage for 4 weeks . These results indicate that stable resistance to RP 59500 can be selected in E . faecium when the organism is exposed to increasing drug concentrations only slightly above the minimum inhibitory concentration (MIC) . This stable resistance is seen in strains exhibiting an MIC > or = 16 micrograms/ml, and unstable in those with resistance where the RP 59500 MIC is less than eight times the original drug MIC . Our observation suggests more than one mechanism of resistance is likely present when stable resistance to RP 59500 develops. Hepatogastroenterology, 1996 May-Jun, 43(9), 586 - 9 Bacteremia following postoperative choledochofiberscopy--a prospective study; Chen MF et al.; BACKGROUND/AIMS: The following investigation was undertaken in order to determine the frequency and clinical consequences of bacteremia after postoperative choledochofiberscopy . MATERIALS AND METHODS: A total of 100 patients were prospectively studied for the frequencies of bacteremia after postoperative choledochoscopy . RESULTS: Positive blood cultures were obtained in 15%; at 5-minutes period in seven patients, at 15-minutes in eight patients and at 30-minutes in two patients . There were two patients with positive cultures at 5 minutes and 10 minutes periods . All the bacteria species cultured were aerobes . Enterococcus, E-coli and Klebsilla were the most commonly cultured bacteria . The frequencies of occurrence of bacteremia seemed not be influenced by the existence of residual stones, session of the endoscopy and duration of the procedure . Six of the 15 bacteremic patients developed cholangitis within 24 hours of the procedure . They all recovered with antibiotic treatment . Patients with negative blood cultures were not found with cholangitic symptoms after the postoperative choledochoscopy . CONCLUSIONS: The results of our study indicated that fifteen percent of patients undergoing postoperative choledochofiberscopy are associated with bacteremia . We believe that with adequate aseptic preparation and meticulous, gentle manipulation, routine prophylactic antibiotics may not be necessary for postoperative choledochoscopy in selected conditions. Jpn J Antibiot, 1996 May, 49(5), 456 - 64 {Isolation rate of E . coli from surgical infections and their susceptibilities}; Shinagawa N et al.; Escherichia coli isolated from surgical infections during the period from July 1983 to June 1995 were investigated in a multicenter study involving 19 hospitals in Japan, and the following results were obtained . 1 . Although the isolation rate of E . coli was not high from postoperative infections, it was most frequently isolated from primary infections throughout the study period . E . coli, Klebsiella spp . and anaerobic bacteria were predominant from fresh infections . From the cases that had previous antibiotics treatment, Enterococcus spp . were the most predominant isolates followed by MRSA and Pseudomonas spp . in this order . 2 . Against E . coli, cefozopran, carumonam and aztreonam had the strongest activity, followed by cefmenoxime, imipenem, latamoxef, gentamicin and ofloxacin . Recently, we have noticed that antibiotic resistant E . coli strains particularly against cefazolin are increasing year by year. J Clin Microbiol, 1996 May, 34(5), 1096 - 9 Random amplified polymorphic DNA typing versus pulsed-field gel electrophoresis for epidemiological typing of vancomycin-resistant enterococci; Barbier N et al.; Sixty vancomycin-resistant vanA mutant Enterococcus faecium (VRE) isolates, collected during a 40-month period from 48 patients hospitalized in a French Cancer Referral Center, were typed by using random amplified polymorphic DNA (RAPD), and the results were compared with those previously obtained by typing with SmaI pulsed-field gel electrophoresis (PFGE), which is currently recognized as the "gold standard." The discriminating power of RAPD typing, with seven primers and 11 combinations of primers, was tested on 18 strains, and only the most discriminating combination was further tested on the whole collection . We compared the epidemiological usefulness of RAPD typing of 60 clinical VRE isolates with that of SmaI PFGE typing . With primers AP4 and ERIC1R, RAPD generated 30 patterns versus the 36 patterns generated by SmaI PFGE . However, this did not hamper the epidemiologically correct clustering of 15 related strains and the detection of multiple colonization in nine patients . We conclude that this simple RAPD technique is well suited to the epidemiological typing of VRE and the monitoring of its nosocomial spread. Antimicrob Agents Chemother, 1996 May, 40(5), 1263 - 5 Plasmid-borne high-level resistance to gentamicin in Enterococcus hirae, Enterococcus avium, and Enterococcus raffinosus; Straut M et al.; Enterococcus hirae, E . avium, and E . raffinosus isolated in Romania, Tunisia, and Portugal harbored plasmids pICC8, pIP1700, and pIP1701, respectively, encoding resistance to high levels of gentamicin (Gmr) . The Gmr marker was carried on pIP1700 by a Tn4001-like element and on pICC8 and pIP1701 by Tn4001-truncated structures . pICC8 carried, in addition to Gmr, chloramphenicol, erythromycin, and tetracycline-minocycline (TetM) resistance determinants . The gene tetM of pICC8 was carried on a Tn916-like element. Arch Microbiol, 1996 May, 165(5), 297 - 305 The Bradyrhizobium japonicum fixGHIS genes are required for the formation of the high-affinity cbb3-type cytochrome oxidase; Preisig O et al.; We report structural and functional analyses of the Bradyrhizobium japonicum fixGHIS genes, which map immediately downstream of the fixNOQP operon for the symbiotically essential cbb3-type heme-copper oxidase complex . Expression of fixGHIS, like that of fixNOQP, is strongly induced in cells grown microaerobically or anaerobically . A fixGHI deletion led to the same prominent phenotypes as those known from a fixNOQP deletion: defective symbiotic nitrogen fixation (Fix-) and decreased cytochrome oxidase activity in cells grown under oxygen deprivation . Only traces, if any, of cytochrome cbb3 subunits were present in membranes isolated from the delta fixGHI strain, as revealed by Western blot analysis with subunit-specific antibodies . This effect was not due to lack of fixNOQP transcription . The results suggested a critical involvement of the fixGHIS gene products in the assembly and/or stability of the cbb3-type heme-copper oxidase . On the basis of sequence similarities between the FixI protein and a Cu-transporting P-type ATPase (CopA) of Enterococcus hirae, and between FixG and a membrane-bound oxidoreductase (RdxA) of Rhodobacter sphaeroides, we postulate that a membrane-bound FixGHIS complex might play a role in uptake and metabolism of copper required for the cbb3-type heme-copper oxidase. Postgrad Med, 1996 May, 99(5), 60 - 5, 69-71 Vancomycin-resistant enterococci . The 'superbug' scourge that's coming your way; Hagman HM et al.; Strains of vancomycin-resistant enterococci (VRE) have emerged and spread widely throughout the United States during the last few years . Multiply-resistant strains of Enterococcus faecium are especially troublesome because they are often resistant to all commercially available antimicrobial agents . At present, VRE infections occur most often in hospitalized patients with severe underlying disease who have undergone invasive procedures and received prolonged courses of broad-spectrum antimicrobial therapy . Because therapeutic options are limited, prevention of spread from patients with known cases to other vulnerable patients is essential. J Biol Chem, 1996 Apr 26, 271(17), 10042 - 7 The ntpJ gene in the Enterococcus hirae ntp operon encodes a component of KtrII potassium transport system functionally independent of vacuolar Na+-ATPase; Murata T et al.; The ntpJ gene, the tail end in the vacuolar type Na+-ATPase (ntp) operon of Enterococcus hirae, encodes a putative 49-kDa hydrophobic protein resembling K+ transporter protein in Saccharomyces cerevisiae (Takase, K., Kakinuma, S., Yamato, I., Konishi, K., Igarashi, K., and Kakinuma, Y . (1994) J . Biol . Chem . 269, 11037-11044) . Northern blotting experiment revealed that the ntpJ gene was transcribed as a cistron in the ntp operon . We constructed an Enterococcus strain in which the ntpJ gene was disrupted by cassette mutagenesis with erythromycin resistance gene . The growth of this mutant was normal at low pH . However, the mutant did not grow at high pH in K+-limited medium (less than 1 mM), while the wild type strain grew well; the internal K+ concentration of this mutant was as low as 7% of that of the wild type strain, suggesting that the K+ accumulation at high pH was inactivated by disruption of the ntpJ gene . Potassium uptake activity via the KtrII system, which had been proposed as the proton potential-independent, Na+-ATPase-coupled system working at high pH (Kakinuma, Y., and Harold, F . M . (1985) J . Biol . Chem . 260, 2086-2091), was missing in this mutant strain . However, this mutant retained as high activities of Na+-ATPase and Na+ pumping as the wild type strain . From these results, we conclude that the NtpJ is a membraneous component of the KtrII K+ uptake system but not a functional subunit of vacuolar Na+-ATPase complex; the interplay between the KtrII system and the Na+-ATPase was discussed. Biochemistry, 1996 Apr 16, 35(15), 4732 - 40 Kinetic comparison of the specificity of the vancomycin resistance VanSfor two response regulators, VanR and PhoB; Fisher SL et al.; Induction of vancomycin resistance in the Gram-positive Enterococci requires a two-componet regulatory system, VanS and VanR, for transcriptional activation of three genes (vanH, A, X) that encode enzymes for a cell wall biosynthetic pathway that produces an altered peptidoglycan intermediate with lower affinity for the antibiotic . The catalytic efficiency (kcat/KM) has been determined for phosphotransfer from the phosphohistidyl form of VanS to both its homologous partner VanR and the heterologous (Escherichia coli) response regulator Phob . The rate of formation of the phosphoaapartyl forms of VanR and PhoB were determined as well as the rate of appearance of inorganic phosphate . Using PhoB in excess of P-VanS, a pseudo-first-order rate constant (kxfer) of 0.2 min-1 for phosphotransfer and a KM for PhoB of 100 microM were readily determined . The corresponding kxfer of 96 min-1 for phosphotransfer from P-VanS to VanR required quench kinetics . A KM of 3 microM was estimated for VanR, leading to a 10(4)-fold preference in kxfer/KM for phosphotransfer to VanR compared to PhoB . No phosphotransfer was detachable to three other E . coli response regulators, OmpR, ArcB, or CreB, providing some sense of the selectivity against two-component regulatory system cross-talk . In the phosphotransfer from P-VanS to PhoB and VanR, there was evidence of competition between water, to give Pi, and the specific aspartyl beta-COO- moiety of either PhoB or VanR with about 25% of the initial flux generating inorganic phosphate . The kinetics of phosphotransfer from P-VanS to VanR were complicated by inhibition by free VanS but, the inhibition pattern could be modeled to yield at KD of 30 nM for VanR binding to free VanS, an affinity similar to that of the CheA-CheY pair in E . coli chemotaxis. MMWR Morb Mortal Wkly Rep, 1996 Apr 12, 45(14), 289 - 91 Assessment of testing for and completeness of reporting of vancomycin-resistant enterococci--Connecticut, 1994; Penicillin resistance and autolysis in enterococci; Department of Microbiology and Immunology, Temple University, School of Medicine, Philadelphia, Pennsylvania 19140, USAComparison of several cell wall-related properties of the ATCC 9790 strain and the R40 strain, a penicillin-resistant, PBP5 overproducing strain, and Rev14, a penicillin-hypersensitive, PBP5-deficient strain, is consistent with a role of the genetic element, psr, in the global regulation of lysozyme sensitivity, autolytic capacity, and wall-rhamnose-containing polysaccharide content . These parameters appear to be independently regulated by a system that involves psr in a currently unknown manner. Microb Drug Resist, 1996 Spring, 2(1), 95 - 8 Bacterial walls, peptidoglycan hydrolases, autolysins, and autolysis; Shockman GD et al.; Knowledge of the chemistry, ultrastructure, biosynthesis, assembly, and function of bacterial cell walls has expanded enormously since the opening of this field of research approximately 40 years ago, primarily by the early work of Milton Salton . It has become abundantly clear that, in most environments, walls are essential to the survival and growth of bacteria and in many ways are structurally and functionally unique . A common but not universal feature of bacterial walls is the presence of peptidoglycan (PG; murein, or in the case of certain Archae the analogous structure-pseudomurein) . PGs are considered to be primarily responsible for the protective and shape-maintaining properties of walls . They are a biologically unique class of macro-molecules in that they are not linear or even branched macromolecules . Instead they are two- or three-dimensional net like polymers that are linked together by three different chemical bonds (glycosidic, amide, and peptide) . In addition, they contain the D-isomers of some amino acids and therefore may possess DL, LD, and DD linkages . Furthermore, the exact chemical structure of a PG may vary depending on environmental factors, however, retaining the essential protective and shape maintaining properties of the wall . Thus, the overall architectural plan of the wall may be more important than the exact shape of the bricks used for the construct . Another somewhat unique feature of PGs (and walls) is their final assembly in situ on the outside of the cellular permeability barrier . A broad variety of bacteria have been shown to possess enzymes that can hydrolyze bonds in the wall PG . Hydrolysis of a sufficient number of bonds can result in the weakening of, or serious damage to, the protective properties of the PG . Frequently, a bacterial strain may possess more than one PG hydrolase activity . A commonly believed, but as yet unproven, hypothesis is that PG hydrolases play one or more roles in PG assembly and/or surface growth and cell division . At a minimum, such potentially suicidal activities must be exquisitely well regulated . Currently we know little concerning the regulation of these activities, or how they communicate with, and integrate with, chromosome replication, synthesis of cytoplasmic macromolecules, cell growth, and division, although such, probably two-way, communications must occur in growing and dividing cells . Recent data indicate that the psr element in Enterococcus hirae described by Fontana and collaborators as a genetic element that is involved in the regulation of the synthesis of PBP 5, also is involved in the regulation of several other surface properties . These properties include (1) autolysis rates of exponential phase . cells, (2) the retention of this property after cells enter the stationary phase, (3) lysozyme sensitivity, and (4) the ratio of rhamnose-containing wall polysaccharide to PG in the walls . Thus the psr element may be a part of a "global" regulation and communication system in E . hirae. Enferm Infecc Microbiol Clin, 1996 Apr, 14(4), 240 - 4 {Risk factors associated with the development of surgical wound infection in a general surgery service}; Dierssen T et al.; BACKGROUND . To evaluate incidence, etiology and risk factors of surgical wound infection (SWI) in a service of general surgery in a tertiary hospital . METHODS . Retrospective cohort study . The relative risk (RR) and its 95% confidence interval (CI) have been used as a measure of association between risk factors and SWI . Multiple logistic regression has been selected as multivariate analysis . RESULTS . Of 619 surgical patients, 60 (9.7%) developed SWI . The most frequently isolated microorganism was Enterococcus (26%), but a higher prevalence of gram negative was also found . On admission, the factors associated with SWI were: diabetes mellitus (RR = 2.5, CI95% = {1.0-6.3}), age older than 65 years (RR = 2.66, CI95% = {0.8-9.0}) and contaminated and dirty surgery (linear trends chi square, p = 0.044); among the amendable medical care factors, the duration of the surgery is the unique to be pointed out with an increment of 5 /1000 per minute (p = 0.011) . The admission an emergency unit presented a non significant adjusted RR of SWI near to 3 (CI95% = {0.9-9.6}) . CONCLUSIONS . SWI is related most importantly to risk factors at admission not amendable by the physician . Our results showed that the only factor susceptible to be changed is the duration of the surgical intervention. Antimicrob Agents Chemother, 1996 Apr, 40(4), 886 - 90 Intracellular activities of RP 59500 (quinupristin-dalfopristin) and sparfloxacin against Enterococcus faecium; Herrera-Insua I et al.; RP 59500, a combination of the streptogramins quinupristin and dalfopristin, and sparfloxacin are new antibiotics with good in vitro activities against Enterococcus faecium, which is an increasingly important nosocomial pathogen with resistance to multiple antimicrobials . Since fluoroquinolones and related macrolides have displayed high intracellular concentrations inside host cells, we evaluated the intracellular activities of these agents inside neutrophils against three strains each of vancomycin-susceptible E . faecium (VSEF) and vancomycin-resistant E . faecium (VREF) . At concentrations equal to four times the MIC, RP 59500 and sparfloxacin decreased the number of intracellular VSEF organisms, while both antibiotics were at best bacteriostatic against intracellular VREF strains . At concentrations equal to one-fourth of the MIC, both antibiotics were bacteriostatic against intracellular VSEF strains but were ineffective in inhibiting the growth of VREF strains . Despite their anticipated markedly higher intracellular human neutrophil (PMN) concentrations, RP 59500 and sparfloxacin activities in medium alone were equal to or greater than those inside PMNs against almost all strains . We conclude that the intracellular PMN concentrations of these antibiotics may not be directly related to their intracellular activities in our assay . The reason for the differences in their activities against VSEF versus VREF remains undefined. Clin Infect Dis, 1996 Apr, 22(4), 663 - 70 Differences in outcomes for patients with bacteremia due to vancomycin-resistant Enterococcus faecium or vancomycin-susceptible E . faecium; Linden PK et al.; To determine the differences in outcome in cases of enterococcal bacteremia due to vancomycin-resistant organisms, we compared consecutive patients on a liver transplant service who had clinically significant bacteremia due to vancomycin-resistant Enterococcus faecium (VREF) (n = 54) with a contemporaneous cohort of patients who had vancomycin-susceptible E . faecium (VSEF) bacteremia (n = 48) . VREF bacteremia occurred significantly later in the hospitalization than did VSEF bacteremia (43 days vs . 24 days, respectively; P < .01); in addition, VREF was more frequently the sole blood pathogen isolated (91% of patients) than was VSEF (56% of patients) (P = .0002) . Invasive interventions for intraabdominal and intrathoracic infection were required more often in the VREF cohort than in the VSEF cohort (34 of 45 patients vs . 20 of 41 patients, respectively; P = .01) . Vancomycin resistance more frequently resulted in recurrent bacteremia (22 of 54 patients infected with VREF vs . 7 of 48 patients infected with VSEF; P = .006), persistent isolation of Enterococcus species at the primary site (27 of 33 patients infected with VREF vs . 7 of 18 patients infected with VSEF; P = .005), and endovascular infection (4 patients infected with VREF vs . none infected with VSEF) . The decrement in patient survival, as measured from the last bacteremic episode, was greater in the VREF cohort (P = .02) . Vancomycin resistance, shock, and liver failure were independent risk factors for Enterococcus-associated mortality . Higher rates of refractory infection, serious morbidity, and attributable death occurred in the VREF cohort and were partially mediated by the lack of effective antimicrobial therapy. J Infect Dis, 1996 Apr, 173(4), 909 - 13 Effective treatment of multidrug-resistant enterococcal experimental endocarditis with combinations of cell wall-active agents; Brandt CM et al.; The efficacy of treatment with a combination of ampicillin, imipenem, and vancomycin was compared with that of two-drug combinations or monotherapy in a model of experimental endocarditis using a strain of Enterococcus faecium with high-level resistance to vancomycin and moderate intrinsic resistance to ampicillin and imipenem . In vitro time-kill synergy studies demonstrated bactericidal synergistic activity only for the triple combination . In vivo, monotherapy with vancomycin was not effective . Treatment with either ampicillin or imipenem alone or in combination with vancomycin resulted in <4 log10 reduction in colony-forming units (cfu) per gram of vegetation . The combination of ampicillin with imipenem was highly active (an additional 5 log10 reduction in cfu per gram of vegetation compared with the most active single agent), but efficacy was not increased by the addition of vancomycin to ampicillin and imipenem . Therapy with the combination of ampicillin and imipenem may be effective for some strains of multidrug-resistant enterococcal infections. Plasmid, 1996 Mar, 35(2), 71 - 80 A novel tetracycline-resistant determinant, tet(U), is encoded on the plasmid pKq10 in Enterococcus faecium; Ridenhour MB et al.; Nine tetracycline (Tc)-resistant clinical isolates of Enterococcus faecium were screened for plasmid content using agarose gel electrophoresis . pKQ10, a 1.9-kb plasmid carrying a novel Tc resistance determinant, was isolated from one of the isolates . The nucleotide sequence of this plasmid revealed an open reading frame corresponding to an 11.8-kDa protein and containing 105 amino acid residues . There was some limited similarity between this protein and tet(M), tet(O), tet(Q), tet(S), tetB(P), and otr(A), which overlapped, but did not include, the consensus GTP-binding sequences . The low-level, Tc-resistant determinant of pKQ10, named tet(U), does not appear to correspond to any other known Tc resistance determinant. J Bacteriol, 1996 Mar, 178(6), 1774 - 5 Importance of the E-46-D-160 polypeptide segment of the non-penicillin-binding module for the folding of the low-affinity, multimodular class B penicillin-binding protein 5 of Enterococus hirae; Mollerach ME et al.; Compared with the other class B multimodular penicillin- binding proteins (PBPs), the low-affinity PBP5 responsible for penicillin resistance in Enterococcus hirae R40, has an extended non-penicillin-binding module because of the presence of an approximately 110-amino-acid E-46(-)D-160 insert downstream from the membrane anchor . Expression of pbp5 genes lacking various parts of the insert-encoding region gives rise to proteins that are inert in terms of penicillin binding, showing that during folding of the PBP, the insert plays a role in the acquisition of a correct penicillin-binding configuration by the G-364(-)Q-678 carboxy-terminal module. Arch Surg, 1996 Mar, 131(3), 338 - 42 Antibiotic-resistant enterococci and the changing face of surgical infections; de Vera ME et al.; BACKGROUND: Enterococci have not been thought to play an important role in intra-abdominal infections because of their relatively low virulence . However, this notion is changing because of the recent emergence of these microbes as significant nosocomial pathogens . OBJECTIVES: To review the mechanisms of antibiotic resistance of enterococci and to discuss the significance of multidrug-resistant enterococci in surgical infections . DATA SOURCES: Medical and basic science literature relating to enterococci . DATA SYNTHESIS: In addition to having intrinsic resistance to a number of antibiotics, enterococci have the ability to acquire resistant genes through the exchange of plasmids or transposons from other bacterial species . Moreover, enterococci have been shown to transmit these genes to other bacterial species in turn . The extensive resistance of these microorganisms has led to their emergence as significant nosocomial pathogens, ranking second only to Escherichia coli in the number of pathogenic isolates recovered from patients in intensive care units . There has also been a marked increase in vancomycin-resistant enterococcal infections in surgical patients in the last 5 years . Some studies associate the prior use of vancomycin or third-generation cephalosporins with the emergence of these strains . Overall, enterococcal infections are associated with increased morbidity and mortality . CONCLUSIONS: In view of the marked resistance of enterococci to antibiotics and their ability to disseminate resistance genes, these microbes have become important pathogens . Enterococci pose a threat to surgical patients, often causing significant therapeutic dilemmas. Biochemistry, 1996 Feb 20, 35(7), 2380 - 7 The active-site histidine-10 of enterococcal NADH peroxidase is not essential for catalytic activity; Crane EJ 3rd et al.; In order to test the proposal {Stehle, T., Claiborne, A., & Schulz, G . E . (1993) Eur . J . Biochem . 211, 221-226} that the active-site His10 of NADH peroxidase functions as an essential acid-base catalyst, we have analyzed mutants in which this residue has been replaced by Gln or Ala . The k(cat) values for both H10Q and H10A peroxidases, and the pH profile for k(cat) with H10Q, are very similar to those observed with wild-type peroxidase . Both mutants, however, exhibit K(m)(H2O2) values much higher (50-70-fold) than that for wild-type enzyme, and stopped-flow analysis of the H2O2 reactivity of two-electron reduced H10Q demonstrates that this difference is due to a 150-fold decrease in the second-order rate constant for this reaction with the mutant . Stopped-flow analyses also confirm that reduction of the enzyme by NADH is essentially unaffected by His10 replacement and remains largely rate-limiting in turnover; the formation of an E.NADH intermediate in the conversion of E-->EH2 is confirmed by diode-array spectral analyses with H10A . Both H10Q and H10A mutants, in their oxidized E(FAD, Cys42-sulfenic acid) forms, exhibit enhanced long-wavelength absorbance bands (lambda(max) = 650 nm and 550 nm, respectively), which most likely reflect perturbations in a charge-transfer interaction between the Cys42-sulfenic acid and FAD . Combined with the 50-fold increase in the second-order rate constant for H2O2 inactivation (via Cys42-sulfenic acid oxidation) of the H10Q mutant, these observations support the proposal that His10 functions in part to stabilize the unusual Cys42-sulfenic acid redox center within the active-site environment. J Chemother, 1996 Feb, 8(1), 33 - 6 Vancomycin susceptibility in enterococcal blood isolates in Italy: a multicenter retrospective analysis; Venditti M et al.; A retrospective, multicenter survey was performed to evaluate the frequency of vancomycin resistance among enterococcal blood isolates in Italian hospitals during 1993 . Twenty-four laboratories, representing 21 cities, provided data on 177,623 blood cultures . Of 15,500 positive cultures, 778 (5%) yielded an Enterococcus . Of 362 evaluable cases of enterococcal bacteremia, a vancomycin resistant Enterococcus (VRE) was found in only 6 (1.6%) . Based on these results, VRE bacteremia did not appear to be a major problem in Italian hospitals in 1993. J Antimicrob Chemother, 1996 Feb, 37(2), 323 - 9 Treatment of experimental endocarditis caused by multidrug resistant Enterococcus faecium with ramoplanin and penicillin; Landman D et al.; Antibiotic resistant strains of enterococci are being isolated with increasing frequency . Effective treatment of infections caused by Enterococcus faecium resistant to ampicillin, vancomycin and aminoglycosides has not been established . We studied the activity of ramoplanin, a new lipoglycopeptide antibiotic, against two strains of multidrug resistant E . faecium . In time kill studies, ramoplanin was bactericidal against both strains, but not in the presence of 50% serum . The combination of ramoplanin and penicillin was bactericidal even in the presence of serum . In rabbits with experimental endocarditis neither penicillin nor ramoplanin significantly reduced vegetation colony counts when given alone, although ramoplanin significantly reduced spleen and kidney bacterial counts of both strains . The combination of ramoplanin plus penicillin resulted in a significant reduction of vegetation bacterial counts (-3.2 and -3.7 log10 cfu/g for strains VA3 and MMC3, respectively, P < 0.01) . All spleen cultures and 9 out of 10 kidney cultures from each strain were sterile following combination therapy . While ramoplanin will not be available for parenteral therapy, further research into the development of other lipoglycopeptide antibiotics is warranted. Biochem J, 1996 Feb 1, 313 ( Pt 3), 711 - 5 Peptidoglycan structure of Enterococcus faecium expressing vancomycin resistance of the VanB type; Billot-Klein D et al.; Resistance to glycopeptide antibiotics in enterococci is due to the synthesis of UDP-MurNAc-tetrapeptide-D-lactate (where Mur is muramic acid) replacing the normal UDP-MurNAc-pentapeptide precursor . The peptidoglycan structures of an inducible VanB-type glycopeptide-resistant Enterococcus faecium, D366, and its constitutively resistant derivative, MT9, were determined . Using HPLC, 17 muropeptides were identified and were present regardless of whether resistance was expressed or not . The structures of 15 muropeptides were determined using MS and amino acid analysis . The cross-bridge between D-alanine and L-lysine consisted of one asparagine . No monomer pentapeptide or tetrapeptide-D-lactate could be identified . These results obtained with D366 (non-induced) and MT9 indicate that, in the absence of vancomycin, the cell wall synthetic machinery of E . faecium can process the lactate-containing precursor as efficiently as the normal pentapeptide . In contrast, the presence of subinhibitory inducing concentrations of vancomycin interfered with the synthesis of oligomers. Med J Aust, 1996 Jan 15, 164(2), 116 - 20 Emerging resistance in Enterococcus spp; Heath CH et al.; Enterococcus spp . are becoming increasingly important nosocomial pathogens . They are intrinsically resistant to most antibiotics, and effective therapy depends primarily on the penicillins, vancomycin and the aminoglycosides . Under antibiotic selection pressure they have developed high level resistance to these agents, and the first vancomycin-resistant enterococcal infection in Australia was described recently . The vancomycin-resistance genes are of particular concern because of their potential to transfer to other gram-positive organisms . The prevention and control of resistant enterococci is a major challenge that is best met by a combination of active infection control measures and restriction of broad-spectrum antibiotic use. Nephrologie, 1996, 17(6), 327 - 8 {Acute interstitial nephropathy induced by vancomycin}; Azar R et al.; The authors report a case of tubulo-interstitial nephritis with acute renal failure due to vancomycin used to treat a patient with enterococcus endocarditis . Rechallenge with vancomycin several days after stopping the drug resulted in the appearance of a maculopapular rash and rapid onset of acute oligo-anuric renal failure . Renal biopsy revealed acute interstitial nephritis . This feature is suggestive of cellular mediated hypersensitivity. C R Seances Soc Biol Fil, 1996, 190(4), 467 - 9 {Molecular and epidemiologic aspects of the resistance to antibiotics: example of glycopeptides on enterococci}; Courvalin P; The emergence of glycopeptide resistance in enterococci results in a severe clinical problem . Efforts to limit the spread of glycopeptide-resistant enterococci are now considered essential . The many ways in which the resistant strains can disseminate, both in the community and in hospitals, are a source of difficulty in reaching that goal. Annu Rev Microbiol, 1996, 50, 753 - 89 Bacterial heavy metal resistance: new surprises; Silver S et al.; Bacterial plasmids encode resistance systems for toxic metal ions including Ag+, AsO2-, AsO4(3-), Cd2+, CO2+, CrO4(2-), Cu2+, Hg2+, Ni2+, Pb2+, Sb3+, TeO3(2-), Tl+, and Zn2+ . In addition to understanding of the molecular genetics and environmental roles of these resistances, studies during the last few years have provided surprises and new biochemical mechanisms . Chromosomal determinants of toxic metal resistances are known, and the distinction between plasmid resistances and those from chromosomal genes has blurred, because for some metals (notably mercury and arsenic), the plasmid and chromosomal determinants are basically the same . Other systems, such as copper transport ATPases and metallothionein cation-binding proteins, are only known from chromosomal genes . The largest group of metal resistance systems function by energy-dependent efflux of toxic ions . Some of the efflux systems are ATPases and others are chemiosmotic cation/proton antiporters . The CadA cadmium resistance ATPase of gram-positive bacteria and the CopB copper efflux system of Enterococcus hirae are homologous to P-type ATPases of animals and plants . The CadA ATPase protein has been labeled with 32P from gamma-32P-ATP and drives ATP-dependent Cd2+ uptake by inside-out membrane vesicles . Recently isolated genes defective in the human hereditary diseases of copper metabolism, Menkes syndrome and Wilson's disease, encode P-type ATPases that are more similar to the bacterial CadA and CopB ATPases than to eukaryote ATPases that pump different cations . The arsenic resistance efflux system transports arsenite, using alternatively either a two-component (ArsA and ArsB) ATPase or a single polypeptide (ArsB) functioning as a chemiosmotic transporter . The third gene in the arsenic resistance system, arsC, encodes an enzyme that converts intracellular arsenate {As (V)} to arsenite {As (III)}, the substrate of the efflux system . The three-component Czc (Cd2+, Zn2+, and CO2+) chemiosmotic efflux pump of soil microbes consists of inner membrane (CzcA), outer membrane (CzcC), and membrane-spanning (CzcB) proteins that together transport cations from the cytoplasm across the periplasmic space to the outside of the cell . Finally, the first bacterial metallothionein (which by definition is a small protein that binds metal cations by means of numerous cysteine thiolates) has been characterized in cyanobacteria. J Infect, 1996 Jan, 32(1), 11 - 6 Mechanisms of glycopeptide resistance in enterococci; Arthur M et al.; Inducible resistance to high levels of glycopeptide antibiotics in clinical isolates of enterococci is mediated by Tn1546 or related transposons . Tn1546 encodes the VanH dehydrogenase which reduces pyruvate to D-lactate (D-Lac) and the VanA ligase which catalyses synthesis of the depsipeptide D-alanyl-D-lactate (D-Ala-D-Lac) . The depsipeptide replaces the dipeptide D-Ala-D-Ala leading to production of peptidoglycan precursors which bind glycopeptides with reduced affinity . In addition, Tn1546 encodes the VanX dipeptidase and the VanY D,D-carboxypeptidase that hydrolyse the dipeptide D-Ala-D-Ala and the C-terminal D-Ala residue of the cytoplasmic precursor UDP-MurNAC-L-Ala-gamma-D- Glu-L-Lys-D-Ala-D-Ala, respectively . These two proteins act in series to eliminate D-Ala-D-Ala-containing precursors . VanX is required for resistance whereas VanY only slightly increases the level of resistance mediated by VanH, VanA and VanX. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 1996 Jan, 81(1), 50 - 2 Prevention of microbial contamination of the dental unit caused by suction into the turbine drive air lines; Ojajarvi J; OBJECTIVE: To determine whether a specially designed antisuction device can prevent the bacterial contamination of the drive air lines of the dental turbine that is caused by suction when the turbine is stopped . STUDY DESIGN: A dental unit with and without the antisuction device and three different types of sterilized handpieces were used in the tests . Each turbine was operated in air, then submerged into a bacterial suspension of E . coli and enterococci for 3 seconds, removed, and stopped . This procedure was repeated 10 times . Possible bacterial contamination of the drive air lines was examined by submersing the head of a sterilized handpiece with the turbine running into a nutrient broth for 30 seconds . The broth was incubated at 35 degrees C up to 2 days . RESULTS: After use of the conventional dental unit, bacterial growth of drive air lines was found in 10 of 150 broth samples . After the installation of the antisuction device no bacterial growth was found in any of the 138 samples . The difference in the contamination frequencies is statistically significant (p = 0.011, Fisher's two-sided exact test) . CONCLUSIONS: The drive air lines of the turbine in the dental unit may become contaminated despite the sterilization of handpieces . The antisuction device installed into the dental unit was found to prevent the contamination . With the exception of possibly immunocompromised patients, the transmission of microbes by exhaust air may be too small to cause infections . However, transmission of oral material between patients should be prevented in dental practice. Clin Infect Dis, 1996 Jan, 22(1), 136 - 7 A citywide survey of vancomycin-resistant Enterococcus--New York City, 1993; Washko R et al.; Vancomycin-resistant Enterococcus (VRE) has become an increasingly important nosocomial pathogen . Questionnaires were sent to all New York City (NYC)-licensed laboratories to ask about testing procedures used, number of isolates identified, species identified, and vancomycin susceptibility for enterococcal isolates in 1993 . Of 127 laboratories, 118 (93%) responded . Fifty-three (45%) of the 118 laboratories reported both the number of enterococcal isolates tested and the number of VRE isolates identified during 1993; 15 (28%) of the 53 laboratories did not isolate VRE, and the remaining 38 laboratories identified 3,822 (8.1%) VRE isolates . VRE was first identified by a NYC-licensed (commercial) laboratory in 1988 . Among NYC hospital laboratories, 65 (97%) of 67 identified at least one VRE isolate during 1989-1993 . This survey demonstrates that there has been a marked increase in the number of VRE isolates identified in NYC laboratories. Chem Biol, 1996 Jan, 3(1), 37 - 44 Synthesis and evaluation of inhibitors of bacterial D-alanine:D-alanine ligases; Ellsworth BA et al.; BACKGROUND: D-Alanine:D-alanine ligase is essential for bacterial cell wall synthesis, assembling one of the subunits used for peptidoglycan crosslinking . The resulting aminoacyl-D-Ala-D-Ala strand is the Achilles' heel of vancomycin-susceptible bacteria; binding of vancomycin to this sequence interferes with crosslinking and blocks cell-wall synthesis . A mutant enzyme (VanA) from vancomycin-resistant Enterococcus faecium has been found to incorporate alpha-hydroxy acids at the terminal site instead of D-Ala; the resulting depsipeptides do not bind vancomycin, yet function in the crosslinking reaction . To investigate the binding specificity of these ligases, we examined their inhibition by a series of substrate analogs . RESULTS: Phosphinate and phosphonate dipeptide analogs (which, after phosphorylation by the enzyme, mimic intermediates in the ligation reaction) were prepared and evaluated as reversible inhibitors of the wild-type ligases DdlA and DdlB from Escherichia coli and of the mutant enzyme VanA . Ki values were calculated for the first stage of inhibitor binding according to a mechanism in which inhibitor competes with D-Ala for both substrate binding sites . DdlA is potently inhibited by phosphinates but not by phosphonates, while DdlB and VanA show little discrimination; both series of compounds inhibit DdlB strongly and VanA weakly . CONCLUSIONS: VanA has greatly reduced affinity for all the ligands studied . The relative affinities of the inhibitors in the reversible binding step are not, however, consistent with the substrate specificities of the enzymes . We propose a mechanism in which proton transfer from the attacking nucleophile to the departing phosphate occurs directly, without intervention of the enzyme. Chem Biol, 1996 Jan, 3(1), 21 - 8 Bacterial resistance to vancomycin: five genes and one missing hydrogen bond tell the story; Walsh CT et al.; A plasmid-borne transposon encodes enzymes and regulator proteins that confer resistance of enterococcal bacteria to the antibiotic vancomycin . Purification and characterization of individual proteins encoded by this operon has helped to elucidate the molecular basis of vancomycin resistance . This new understanding provides opportunities for intervention to reverse resistance. Scand J Infect Dis, 1996, 28(2), 191 - 3 First documented isolation of vancomycin-resistant Enterococcus faecium in Sweden; Melhus A et al.; In recent years enterococci, and Enterococcus faecium in particular, have emerged as important nosocomial pathogens . Of major concern is the increasing antimicrobial resistance to traditionally used agents such as ampicillin, gentamicin and vancomycin . We present a patient with prosthetic heart valves colonized with vancomycin-resistant E . faecium . This is the first reported isolation of vancomycin-resistant E . faecium in Sweden. Infect Control Hosp Epidemiol, 1996 Jan, 17(1), 36 - 41 Resistant enterococci: a prospective study of prevalence, incidence, and factors associated with colonization in a university hospital; Weinstein JW et al.; OBJECTIVE: To determine the prevalence of gastrointestinal tract colonization with antibiotic-resistant enterococci at ward entry and to study the incidence and risk factors for nosocomial acquisition of colonization with resistant enterococci . DESIGN: A prospective cohort study conducted between February 1 and March 15, 1993 . METHODS: Rectal cultures were obtained within 24 hours of admission or transfer onto the study wards and repeated at weekly intervals and at the time of discharge . Patients harboring antibiotic-resistant enterococci at the time of admission or after admission were compared to patients who were not colonized with these organisms . Clinical and epidemiologic risk factors for colonization were abstracted prospectively by daily chart review . Following a univariate analysis of risk factors associated with colonization, a multivariate statistical analysis using three separate models was done . SETTING: A 1,125-bed, tertiary-care teaching hospital in North Carolina . PATIENTS: A total of 350 patients admitted to two general medical wards and the medical intensive care unit during the study period . RESULTS: Antibiotic-resistant enterococci were isolated from 52 patients: 19 were colonized at admission to the study, and 33 later acquired resistant strains . At the time of admission, 5.4% of the patients were colonized with ampicillin-resistant enterococci (ARE), including 1.1% that were colonized with vancomycin-resistant enterococci . Prior hospitalization was associated with colonization with ARE at admission (P = .01) . Independent risk factors for nosocomial acquisition of ARE included treatment with more than three antibiotics, empiric use of antibiotics, use of third-generation cephalosporins, and the use of enteral tube feedings . Antibiotics used prophylactically were not associated with resistant enterococcal colonization . CONCLUSIONS: Our data help to elucidate the epidemiology of gastrointestinal tract colonization with resistant enterococci . We hypothesize that surveillance and control programs will be more likely to succeed if targeted at patients receiving more than three antibiotics, empiric antibiotics, and enteral tube feedings (Infect Control and Hosp Epidemiol 1996;17:36-41). Antimicrob Agents Chemother, 1996 Jan, 40(1), 257 - 9 Penicillin tolerance and modification of lipoteichoic acid associated with expression of vancomycin resistance in VanB-type Enterococcus faecium D366; Gutmann L et al.; Induction of vancomycin resistance in Enterococcus faecium D366, which exhibits a VanB-type resistance, as well as its constitutive expression in MT9, a derivative of D366, was associated with penicillin tolerance as shown by decreased lysis and killing of the cells . This phenomenon was linked neither to decreased expression of the different autolysins nor to their decreased lytic activity on the different cell walls . The only change observed was that almost twice the normal amount of D-alanine was attached to the lipoteichoic acid. Chemotherapy, 1996 Jan-Feb, 42(1), 37 - 46 High-level gentamicin-resistant enterococci: in vitro activity of double and triple combinations of antimicrobial drugs; Ferrara A et al.; The ability of double and triple combinations of antimicrobials with different mechanisms of action, such as teicoplanin, meropenem, gentamicin and sparfloxacin, to achieve synergisms was investigated in vitro on some moderate-level gentamicin-resistant (MLGR: 8 < or = MIC < or = 256 mg/l) and high-level gentamicin-resistant (HLGR: MIC > 500 mg/l) enterococci . On MLGR strains, a constant synergistic effect was achieved by a combination of teicoplanin with gentamicin or with meropenem, while generally addition, sometimes close to synergism, was exhibited by gentamicin-meropenem, gentamicin-sparfloxacin and teicoplanin-sparfloxacin associations . Triple combinations of teicoplanin, meropenem and gentamicin, or teicoplanin, sparfloxacin and gentamicin, always showed a remarkable advantage in terms of synergism over double combinations . On HLGR enterococci, the only double association showing an additive effect, sometimes close to synergism, was teicoplanin plus meropenem, while the triple combination of teicoplanin with gentamicin and meropenem always showed a marked synergistic effect . An effect very close to synergism was also shown by the combination of teicoplanin with sparfloxacin and gentamicin. J Clin Microbiol, 1996 Jan, 34(1), 210 - 2 Comparison of rectal and perirectal swabs for detection of colonization with vancomycin-resistant enterococci; Weinstein JW et al.; Patients whose gastrointestinal tracts are colonized with vancomycin-resistant enterococci (VRE) may serve as a reservoir for nosocomial transmission . We compared the sensitivities and concordance of several methods used to detect VRE colonization . Eighty-two paired rectal and perirectal swabs were obtained from 13 patients over a 9-day period . The sensitivity of both rectal and perirectal swabs was 79% . There was 100% concordance of culture results between simultaneously obtained rectal and perirectal swabs, and the quantities of growth were similar by these two methods of detection . Our data suggest that rectal and perirectal swabs are equally sensitive for the detection of VRE colonization. Postgrad Med J, 1996 Jan, 72(843), 51 - 2 Enterococcal endocarditis after extracorporeal shock wave lithotripsy for nephrolithiasis; Zimhony O et al.; We report a case of enterococcal endocarditis following extracorporeal shock wave lithotripsy (ESWL) for ureteral stone . Although endocarditis following ESWL is very rare, transient bacteraemia occurs during ESWL . This case is a reminder that enterococcal endocarditis may follow innovative genitourinary procedures without appropriate prophylaxis. Vet Microbiol, 1996 Jan, 48(1-2), 29 - 39 The humoral immune response of turbot to recently isolated pathogenic Enterococcus strains . Cross-reactivity with other Gram-positive bacteria; Leiro J et al.; An Enterococcus sp . causing severe mortalities among farmed turbot (Scophthalmus maximus) has recently been detected in northwest Spain . We found that specific turbot serum antibodies raised against isolate RA-99.1 of the new pathogen by intraperitoneal immunization, did not cross react in enzyme-linked immunosorbent assay (ELISA) with other enterococcal or non-enterococcal Gram-positive bacteria . In immunoblotting, antibodies raised against strain RA-99.1 recognized the same components in the homologous total soluble antigen preparation (TSA) as in TSA of two other isolates of the pathogen obtained from different farms . Anti-RA-99.1 serum also recognized some components of the TSA of several other Gram-positive bacteria . Immunogold labelling indicated that the antigens which provoke the humoral immune response to this pathogen are located mainly on the bacterial surface. J Antimicrob Chemother, 1996 Jan, 37(1), 127 - 32 Treatment of experimental endocarditis due to multidrug-resistant Enterococcus faecium with clinafloxacin and penicillin; Zaman MM et al.; Clinafloxacin, a new quinolone antibiotic with enhanced activity against Gram-positive bacteria, has demonstrated in-vitro activity against multidrug-resistant Enterococcus faecium, particularly when combined with penicillin . Rabbits with experimental endocarditis due to a multidrug-resistant strain of E . faecium were treated with clinafloxacin and/or penicillin . After three days of therapy, significant reduction of bacterial concentrations were found in vegetations, kidneys, and spleens of animals treated with clinafloxacin . The combination of clinafloxacin and penicillin was significantly better in reducing vegetation bacterial concentrations compared to the other groups (-4.4 log10 cfu/g compared with control) . Serum levels of clinafloxacin consistently exceeded the MIC of the strain, and clinafloxacin-resistant isolates could not be detected . Clinafloxacin demonstrated promising activity in vivo against multidrug-resistant E . faecium, and further studies are warranted. Proc Natl Acad Sci U S A, 1995 Dec 5, 92(25), 11603 - 7 Phosphinate analogs of D-, D-dipeptides: slow-binding inhibition and proteolysis protection of VanX, a D-, D-dipeptidase required for vancomycin resistance in Enterococcus faecium; Wu Z et al.; VanX is a D-Ala-D-Ala dipeptidase that is essential for vancomycin resistance in Enterococcus faecium . Contrary to most proteases and peptidases, it prefers to hydrolyze the amino substrate but not the related kinetically and thermodynamically more favorable ester substrate D-Ala-D-lactate . The enzymatic activity of VanX was previously found to be inhibited by the phosphinate analogs of the proposed tetrahedral intermediate for hydrolysis of D-Ala-D-Ala . Here we report that such phosphinates are slow-binding inhibitors . D-3-{(1-Aminoethyl)phosphinyl}-D-2-methylpropionic acid I showed a time-dependent onset of inhibition of VanX and a time-dependent return to uninhibited steady-state rates upon dilution of the enzyme/inhibitor mixture . The initial inhibition constant Ki after immediate addition of VanX to phosphinate I to form the E-I complex is 1.5 microM but is then lowered by a relatively slow isomerization step to a second complex, E-I*, with a final K*i of 0.47 microM . This slow-binding inhibition reflects a Km/K*i ratio of 2900:1 . The rate constant for the slow dissociation of complex E-I* is 0.24 min-1 . A phosphinate analog with an ethyl group replacing what would be the side chain of the second D-alanyl residue in the normal tetrahedral adduct gives a K*i value of 90 nM . Partial proteolysis of VanX reveals two protease-sensitive loop regions that are protected by the intermediate analog phosphinate, indicating that they may be part of the VanX active site. Infect Control Hosp Epidemiol, 1995 Dec, 16(12), 680 - 5 Natural history of colonization with vancomycin-resistant Enterococcus faecium; Montecalvo MA et al.; OBJECTIVE: To determine the incidence, duration, and genetic diversity of colonization with vancomycin-resistant Enterococcus faecium (VREF) . SETTING: Oncology unit of a 650-bed university hospital . METHODS: Surveillance perianal swab cultures were performed on admission and weekly . The molecular relatedness of VREF isolates was determined by pulsed-field gel electrophoresis and by the hybridization pattern of the vanA resistance determinant . RESULTS: During 8 months of surveillance, the VREF colonization rate was 16.6 patients per 1,000 patient-hospital days, which was 10.6 times greater than the VREF infection rate . Eighty-six patients with VREF colonization were identified . Colonization persisted for at least 7 weeks in the majority of patients . Of 36 colonized patients discharged from the hospital and then readmitted, an average of 2 1/2 weeks later, 22 (61%) patients still were colonized with VREF . Of the 14 patients who were VREF-negative at readmission, only three patients remained culture-negative throughout hospitalizations . PFGE demonstrated that colonization with the same VREF isolate may persist for at least 1 year, and patients may be colonized with more than one strain of VREF . CONCLUSION: VREF colonization is at least 10-fold more prevalent than infection among oncology patients . Colonization often persists throughout lengthy hospitalizations and may continue for long periods following hospitalization. Indian J Med Res, 1995 Dec, 102, 255 - 7 Determination of high level resistance to aminoglycosides among enterococci; Cherian BP et al.; Two hundred and ten strains of enterococci showing resistance to gentamicin (10 micrograms/disc) were tested for high level resistance by detecting the minimum inhibitory concentration and by using high content disc diffusion test . Only 67 per cent of these had high level resistance to gentamicin . High level kanamycin resistance in the group was 84 per cent, while high level streptomycin resistance was 61 per cent . Only 85 of the 140 strains with high level gentamicin resistance had similar streptomycin resistance . Results using locally made high content discs, correlated 100 per cent with MIC results . High level resistance to enterococci should be reported on a routine basis, especially when isolated from patients with serious infections. J Biol Chem, 1995 Nov 24, 270(47), 28392 - 6 An iron-regulated gene, magA, encoding an iron transport protein of Magnetospirillum sp . strain AMB-1; Nakamura C et al.; Magnetospirillum sp . AMB-1 is a freshwater magnetic bacterium which synthesizes intracellular particles of magnetite (Fe3O4) . A genomic DNA fragment required for synthesis of magnetic particles was previously isolated from a nonmagnetic transposon Tn5 mutant . We have determined the complete nucleotide sequence of this fragment . The 2975-base pair region contains two putative open reading frames . One open reading frame, designated magA, encodes a polypeptide which is homologous to the cation efflux proteins, the Escherichia coli potassium ion-translocating protein, KefC, and the putative Na+/H(+)-antiporter, NapA, from Enterococcus hirae . Northern hybridization demonstrated that the magA mRNA transcript is 1.3 kilobases in size, corresponding to the size of the magA gene . A functional promoter was located upstream from the magA gene, and the transcription in AMB-1 was regulated by environmental iron concentration . Vesicles isolated from E . coli in which the MagA protein was expressed exhibited iron accumulation ability . We consider that the MagA protein is an iron transport involved in the synthesis of magnetic particles in AMB-1. Diagn Microbiol Infect Dis, 1995 Nov, 23(3), 123 - 7 Activity of two novel fluoroquinolones (DU-6859a and DV-7751a) tested against glycopeptide-resistant enterococcal isolates; Jones RN et al.; Two novel fluoroquinolones, DU-6859a and DV-7751a, were compared with three peer compounds (ciprofloxacin, levofloxacin, and ofloxacin) by testing 150 strains of enterococci that were resistant to vancomycin . Standardized methods recommended by the National Committee for Clinical Laboratory Standards were used for all minimum inhibitory concentration tests, and DU-6859a was additionally tested by the standardized disk diffusion procedure (5-mu g disks) . The rank order of the fluoroquinolone spectrums against these Enterococcus spp . isolates was DU-6859a (71.3% of strains inhibited at < or = 2 mu g/ml) > DV-7751a (38.7%) > levofloxacin (33.3%) > ofloxacin (32.0%) > ciprofloxacin (3.3%) . Using previously proposed break point zone diameters of > or = 16 mm (susceptible) and < or = 12 mm (resistant), the DU-6859a disk diffusion tests for these glycopeptide-resistant organisms were without false-susceptible or false-resistant error (81.3% absolute agreement) . These results indicate that among the investigational and currently marketed fluoroquinolones, DU-6859a appears to have the greatest potential value in the therapy of Enterococcus spp . strains that are resistant to the glycopeptides and many other therapeutic agents. Diagn Microbiol Infect Dis, 1995 Nov, 23(3), 119 - 22 Selective media for detecting gastrointestinal carriage of vancomycin-resistant enterococci; Barton AL et al.; Nosocomial infection with vancomycin-resistant enterococci (VRE) has become a significant problem . Effective institution of infection control measures depends on rapid identification of carriage of the organism, especially in asymptomatic individuals . We compared two selective media for use in screening for the presence of VRE and found that an agar medium containing bile esculin azide supplemented with 8 mu g/ml of vancomycin was a useful and cost-effective means for primary screening for asymptomatic gastrointestinal carriage of VRE. Vet Med (Praha), 1995 Nov, 40(11), 337 - 9 {The effect of monensin on a rumen strain of Enterococcus faecium CCM 4231}; Laukova A et al.; Monensin from three different manufacturing companies (Eli Lilly Co., Indianapolis, USA; Pharmachim, Bulgaria; Spofa Prague, Czech Republic) were added to pure cultures of rumen strain Enterococcus faecium CCM 4231 at final concentrations 25 micrograms per ml and 50 microgram per ml . Enterococci represent a strong bacterial group colonizing the rumen regarding to lactic acid production . Ent . faecium CCM 4231 is own, lactic acid-producing isolate from the rumen content of calf with a broad antimicrobial activity (bacteriocin production) . This strain was obtained in the microbial preparation Inhicol to protect enteritis in young ruminants, especially . The growth of CCm 4231 strain was inhibited at both concentrations (25 microgram/ml; 50 micrograms/ml) using all three monensins in comparison with controls (Figs . 1 and 2) . The beginning of the growth inhibition was detected within 2 hours after ionophore addition . Monensin made in Bulgaria was the most effective of all when the three monensins were compared . No significant differences are found in the effect of monensins made in different production companies . Our results extend the knowledge about inhibition effect of monensin on Gram-positive bacteria, enterococci including . Moreover, the quality of monensins made by different companies is attested synchronous . In general, regarding the practical point of view, it is also contribution for the selection and application of the most suitable additives. Eur J Clin Microbiol Infect Dis, 1995 Nov, 14(11), 959 - 63 Epidemiologic analysis of glycopeptide-resistant Enterococcus strains in neutropenic patients receiving prolonged vancomycin administration; Plessis P et al.; Vancomycin-resistant enterococci have been isolated with increasing frequency since 1988 . Thus far, most of these resistant enterococci have belonged to the Enterococcus faecium species, and epidemiological studies have shown a wide diversity among interhospital and intrahospital isolates . This report presents an epidemiologic investigation of 25 vancomycin-resistant Enterococcus strains--24 Enterococcus faecium and one Enterococcus gallinarum--isolated from the stools or blood of adult patients receiving intravenous vancomycin prophylaxis during neutropenia and hospitalized in a single hematologic unit . Macrorestriction patterns of total DNA and of ribosomal DNA regions were used to analyze the strains . Strains produced different total DNA restriction fragment length polymorphism patterns after SmaI digestion . Ribotyping was less discriminative than pulsed-field gel electrophoresis . The results confirmed the genetic unrelatedness of the strains . Prolonged vancomycin administration, commonly used in hematologic units, could be involved in the selection of endogenous resistant enterococcal strains. J Antimicrob Chemother, 1995 Nov, 36(5), 821 - 5 Characterization of vancomycin resistance in Enterococcus durans; Cercenado E et al.; During investigation of an outbreak of vancomycin resistant Enterococcus faecium in a paediatric hospital, an isolate of Enterococcus durans resistant to vancomycin, teicoplanin, ampicillin and highly resistant to gentamicin and streptomycin was found in the stools of a patient also colonized with a strain of E . faecium with the same resistance pattern . Minimal inhibitory concentrations of vancomycin and teicoplanin were 512 and 64 mg/mL, respectively . Resistance to vancomycin as well as high-level resistance to gentamicin was transferable to an E . faecium recipient strain . Both multiresistant E . faecium and E . durans isolates as well as the transconjugant presented only one plasmid . The vanA gene was detected and localized to the high molecular weight plasmid by DNA hybridization with a vanA gene probe . Growth in vancomycin resulted in induction of an approximately 40 kDa protein visible in membrane preparations from these cells . Genetic linkage between vancomycin and gentamicin resistance genes in the same plasmid is suggested. Infect Control Hosp Epidemiol, 1995 Nov, 16(11), 634 - 7 Controlling vancomycin-resistant enterococci; Boyce JM et al.; After controlling an epidemic of vanB-type vancomycin-resistant Enterococcus faecium (VRE), we contained a subsequent vanA E faecium outbreak by using prospective laboratory-based surveillance, placing patients with VRE in private rooms, requiring the use of both gowns and gloves by all personnel entering the patients' rooms, and conducting prevalence surveys of patients on affected wards. Clin Infect Dis, 1995 Nov, 21(5), 1234 - 7 Clinical and molecular epidemiology of vancomycin-resistant Enterococcus faecium during its emergence in a city in southern Texas; Moreno F et al.; During a 19-month period from April 1993 to October 1994, 41 isolates of vancomycin-resistant Enterococcus faecium (VREF) were detected in seven different hospitals in a city in southern Texas . A case-control study to determine the risk factors for acquisition was done in the hospital in which the majority of isolates were detected . Pulsed-field gel electrophoresis (PFGE) of whole-cell DNA was used to determine strain identity . Thirty-five (85%) of the 41 VREF isolates were of the vanB phenotype . Of these, 32 (91%) of 35 were the same strain by PFGE typing . The same vanB strain was documented in five different hospitals in the city . In contrast, 4 (67%) of 6 of the vanA phenotype VREF isolates were distinct strains by PFGE typing . Significant risk factors for colonization or infection with VREF were prior exposure to antibiotics (P = .04), the previous use of third-generation cephalosporins (P = .03), and the previous use of parenteral vancomycin (P = .002) . Infection-control and antibiotic-utilization measures were implemented to control cross-transmission and selection of VREF isolates . During the emergence of VREF in our city, clonal dissemination of a single strain of vanB VREF among six hospitals was documented . Limited cross-transmission of vanA phenotype VREF isolates occurred, but most vanA VREF isolates were distinct strains selected in individual hospital environments. J Clin Microbiol, 1995 Nov, 33(11), 3008 - 18 Multilaboratory evaluation of screening methods for detection of high-level aminoglycoside resistance in enterococci . National Committee for Clinical Laboratory Standards Study Group on Enterococci; Swenson JM et al.; Since the early 1970s, the synergistic activity of an aminoglycoside with a cell wall-active agent has been predicted by determining the ability of an enterococcus to grow in the presence of high levels of the aminoglycoside (usually > or = 2,000 micrograms/ml) . However, a variety of media and concentrations of aminoglycosides has been used for this screening procedure . In the present study, we sought to optimize the agar dilution, broth microdilution, and disk diffusion tests used to detect high-level gentamicin and streptomycin resistance in enterococci . For dilution tests, brain heart infusion agar or broth gave the best growth and performance . For agar dilution, 500 micrograms of gentamicin per ml, 2,000 micrograms of streptomycin per ml, and an inoculum of 1 x 10(6) CFU/ml were optimal, while for broth microdilution, 500 micrograms of gentamicin per ml, 1,000 micrograms of streptomycin per ml, and an inoculum of 5 x 10(5) CFU/ml were best . Growth of more than one colony in the agar dilution test was determined to be the best indicator of high-level resistance . For disk diffusion, Mueller-Hinton agar, 120-micrograms gentamicin disks, and 300-micrograms streptomycin disks with breakpoints of no zone for resistance and > or = 10 mm for susceptibility gave the best sensitivity and specificity if results for strains with zones of 7 to 9 mm are considered inconclusive, indicating that a broth or agar test should be performed to determine susceptibility or resistance. Enferm Infecc Microbiol Clin, 1995 Nov, 13(9), 516 - 21 {Detection of antibiotic resistance in Enterococcus sp . Comparison of GPS-TA (BioMérieux-Vitek), Uniscept MIC-3 (bioMérieux-Vitek) and conventional methods}; Alamo I et al.; BACKGROUND: The treatment of severe enterococcus infections requires synergism of a beta-lactamic or glycopeptide and a aminoglycoside, but when resistance to first one or high-level resistance to aminoglycosides are present, synergism would be lost . We compared the adequacy of two commercially available systems to detect antibiotic resistance . METHODS: We studied 158 isolates of Enterococcus sp., with high-level resistance to gentamicin (40 isolates) and streptomycin (89 isolates), resistance to ciprofloxacin (34 isolates), resistance to ampicillin (7 isolates) and with intermediate susceptibility to vancomycin (3 isolates) . No one was beta-lactamase producer by Cefinase disk method . We use disk diffusion as reference technique to detect high-level streptomycin resistance . The susceptibility to the remainder antibiotics was studied by agar dilution method, according to NCCLS . We studied the accuracy of GPS-TA cards and Uniscept MIC-3 in relation to the degree of agreement with conventional means, following FDA criteria . RESULTS: Essential agreement for MIC was less than 90 with MIC-3 for ampicillin (81.5%) and ciprofloxacin (71.3%) . Categorical agreement rate was less than 90% (76.4%) and major error rate was higher than 3% (10.9%) with the use of MIC-3 for ciprofloxacin . Very major errors for ampicillin, vancomycin and ciprofloxacin were not produced by any system . The very major error rates for high level resistance to gentamicin and streptomycin with GPS-TA card were 5 and 15.7%, respectively . CONCLUSIONS: We do not recommend the use of the Uniscept MIC-3 panel with visual reading to detect susceptibility to ciprofloxacin . Detection of high levels of aminoglucoside resistance by GPS-TA card should be supplemented with conventional techniques because of the high rate of major error . Due to the low number of strains that have been studied, we can not assure the suitability of these systems to detect ampicillin or vancomycin resistance. Biochemistry, 1995 Oct 31, 34(43), 14114 - 24 Analysis of the kinetic mechanism of enterococcal NADH peroxidase reveals catalytic roles for NADH complexes with both oxidized and two-electron-reduced enzyme forms; Crane EJ 3rd et al.; Anaerobic titrations of the two-electron-reduced NADH peroxidase (EH2) with NADH and 3-acetylpyridine adenine dinucleotide (AcPyADH) yield the respective complexes without significant formation of the four-electron-reduced enzyme (EH4) . Further analysis of the EH2/EH4 redox couple, however, yields a midpoint potential of -312 mV for the free enzyme at pH 7 . The catalytic mechanism of the peroxidase has been evaluated with a combination of kinetic and spectroscopic approaches, including initial velocity and enzyme-monitored turnover measurements, anaerobic stopped-flow studies of the reactions of both oxidized enzyme (E) and EH2 with NADH and AcPyADH, and diode-array spectral analyses of both the reduction of E-->EH2 by NADH and the formation of EH2.NADH . Overall, these results are consistent with rapid formation of an E.NADH complex with distinct spectral properties and a rate-limiting hydride transfer step that yields EH2, with no direct evidence for intermediate FADH2 formation . The EH2.NADH complex described previously {Poole, L . B., & Claiborne, A . (1986) J . Biol . Chem . 261, 14525-14533} is not catalytically competent and reacts relatively slowly with H2O2 . Stopped-flow analyses do, however, support the very rapid formation of an EH2.NADH* intermediate, with spectral properties that distinguish it from the static EH2.NADH form, and yield a first-order rate constant for the conversion between the two species that is smaller than kcat . The combined rapid-reaction and steady-state data are best accommodated by a limiting type of ternary complex mechanism very similar to that proposed previously {Parsonage, D., Miller, H., Ross, R.P., & Claiborne, A . (1993) J . Biol . Chem . 268, 3161-3167}. Microb Drug Resist, 1995 Fall, 1(3), 249 - 53 Successful treatment of persistent bacteremia due to vancomycin-resistant, ampicillin-resistant Enterococcus faecium; Mekonen ET et al.; 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. Microb Drug Resist, 1995 Fall, 1(3), 245 - 7 Quinupristin/dalfopristin (RP 59500) therapy for vancomycin-resistant Enterococcus faecium aortic graft infection: case report; Sahgal VS et al.; A 46-year-old woman was admitted to the hospital with severe peripheral vascular disease, requiring multiple vascular surgical procedures . During the sixth hospital week, after prior therapy with multiple antibiotics, Enterococcus faecium was isolated as the only organism from an operating room culture of an infected aortic graft . Histological examination of the graft showed infiltration with polymorphonuclear leukocytes . Subsequently, cultures of an infected inguinal wound yielded Enterococcus faecium with mixed bacterial growth . Both isolates of Enterococcus faecium were resistant to all available antimicrobials, including ampicillin, vancomycin, tetracycline, chloramphenicol, and ciprofloxacin . Compassionate use therapy with quinupristin/dalfopristin (RP59500) was administered for 25 days, the patient's clinical condition improved, and wound healing occurred . Transient elevation of serum alkaline phosphatase was noted . This case demonstrates successful eradication of deep VREF infection by quinupristin/dalfopristin with good tolerance of prolonged therapy. Microb Drug Resist, 1995 Fall, 1(3), 241 - 3 Serious infections caused by multiply-resistant Enterococcus faecium; Wade JJ et al.; Three liver transplant patients developed serious intraabdominal infections and recurrent bacteremias due to strains of Enterococcus faecium with high-level resistance to vancomycin . The enterococci were also resistant to all other antibacterials except pristinamycin, which, given orally, proved ineffective . One strain was sensitive to tetracycline . Increasingly, clinicians are likely to encounter infections caused by multiply-resistant enterococci, and these cases illustrate the seriousness of such infections in compromised patients. Ann Pharmacother, 1995 Oct, 29(10), 1022 - 7 Quinupristin/dalfopristin (RP 59500): a new streptogramin antibiotic; Chant C et al.; OBJECTIVE: To review the current knowledge on RP 59500 (quinupristin/dalfopristin, Synercid), a new streptogramin antibiotic, with respect to its pharmacology, pharmacokinetics, pharmacodynamics, mechanism of resistance, and in vitro inhibitory and bactericidal activity . DATA SOURCES: A MEDLINE search using keywords RP 59500, pristinamycin, virginiamycin, and streptogramin was performed . Relevant abstracts presented at recent scientific conferences also were consulted . STUDY SELECTION: Because RP 59500 is a relatively new investigational agent, relevant in vitro and animal studies were selected . All available human studies were included as well . DATA EXTRACTION: Data from in vitro and in vivo studies were included, with particular emphasis on human studies . DATA SYNTHESIS: RP 59500 is a new injectable streptogramin antibiotic consisting of a mixture o 2 synergistic pristinamycin compounds . RP 59500 possesses in vitro inhibitory and bactericidal activity against most isolates of gram-positive organisms including vancomycin-resistant Enterococcus faecium, selected gram-negative bacteria, and most anaerobic organisms . Based on preliminary data, the drug appears to be metabolized rapidly and extensively while exhibiting a significant postantibiotic effect . Data from ongoing clinical trials suggests that RP 59500 is well-tolerated except for mild injection site irritations . However, before the role of RP 59500 within the vast armamentarium of antimicrobials can be elucidated, additional studies need to be conducted to document its clinical efficacy . CONCLUSIONS: Based on in vitro susceptibility testing, in vivo studies, and preliminary clinical data, RP 59500 may be an alternative to the glycopeptides, especially for inherently resistant organisms . Further studies are needed to confirm this agent's in vitro activity and to establish its clinical efficacy. Clin Microbiol Rev, 1995 Oct, 8(4), 585 - 615 Current perspectives on glycopeptide resistance; Woodford N et al.; In the last 5 years, clinical isolates of gram-positive bacteria with intrinsic or acquired resistance to glycopeptide antibiotics have been encountered increasingly . In many of these isolates, resistance arises from an alteration of the antibiotic target site, with the terminal D-alanyl-D-alanine moiety of peptidoglycan precursors being replaced by groups that do not bind glycopeptides . Although the criteria for defining resistance have been revised frequently, the reliable detection of low-level glycopeptide resistance remains problematic and is influenced by the method chosen . Glycopeptide-resistant enterococci have emerged as a particular problem in hospitals, where in addition to sporadic cases, clusters of infections with evidence of interpatient spread have occurred . Studies using molecular typing methods have implicated colonization of patients, staff carriage, and environmental contamination in the dissemination of these bacteria . Choice of antimicrobial therapy for infections caused by glycopeptide-resistant bacteria may be complicated by resistance to other antibiotics . Severe therapeutic difficulties are being encountered among patients infected with enterococci, with some infections being untreatable with currently available antibiotics. Antimicrob Agents Chemother, 1995 Oct, 39(10), 2341 - 4 In vitro interactions between different beta-lactam antibiotics and fosfomycin against bloodstream isolates of enterococci; Pestel M et al.; The effects of 16 different beta-lactam-fosfomycin combinations against 50 bloodstream enterococci were compared by a disk diffusion technique . Cefotaxime exhibited the best interaction . By checkerboard studies, the cefotaxime-fosfomycin combination provided a synergistic bacteriostatic effect against 45 of the 50 isolates (MIC of cefotaxime at which 90% of the isolates were inhibited, >2,048 micrograms/ml; MIC of fosfomycin at which 90% of the isolates were inhibited, 128 micrograms/ml; mean of fractional inhibitory concentration indexes, 0.195) . By killing curves, cefotaxime (at 64 micrograms/ml) combined with fosfomycin (at > or = 64 micrograms/ml) was bactericidal against 6 of 10 strains tested. Thorac Cardiovasc Surg, 1995 Oct, 43(5), 302 - 4 Aortic insufficiency and enterococcal endocarditis complicating systemic lupus erythematosus; Demircin M et al.; A case of aortic valve replacement for aortic insufficiency complicated with enterococcal endocarditis in a patient with systemic lupus erythematosus (SLE) is described . The cardiac complications of SLE may involve the endocardium, myocardium, pericardium, and coronary vessels . Steroids which are used for treatment probably increase the incidence of valve incompetence . Aortic insufficiency or infective endocarditis complicating Libman-Sacks endocarditis is rarely observed and may require valve replacement . Echocardiography is an important diagnostic tool . Renal function, pulmonary status, and cerebral complications require special attention in these patients. Eur J Clin Microbiol Infect Dis, 1995 Oct, 14(10), 878 - 82 Detection of aminoglycoside-penicillin synergy against Enterococcus faecium using high-content aminoglycoside disks; Torres C et al.; Thirty-seven Enterococcus faecium strains were screened for high-level aminoglycoside resistance with an agar diffusion test using high-content aminoglycoside disks (300 micrograms of streptomycin and 120 micrograms of gentamicin, tobramycin, kanamycin or amikacin) . The inhibition zones obtained were correlated with results of time-kill penicillin-aminoglycoside synergy studies . An 11 mm breakpoint differentiated strains susceptible or resistant to the synergy of streptomycin plus penicillin . Irrespective of the inhibition zones obtained with tobramycin and kanamycin disks, Enterococcus faecium strains never showed synergy with penicillin in combination with these aminoglycosides . Penicillin-amikacin synergy cannot be predicted by the amikacin disks . Nevertheless, even though kanamycin disks do not predict penicillin-kanamycin synergy, they can be used to predict penicillin-amikacin synergy . In summary, high-content streptomycin, gentamicin and kanamycin disks can be used to predict the susceptibility of Enterococcus faecium strains to the synergistic combination of penicillin plus one of the aminoglycosides (streptomycin, gentamicin or amikacin, respectively). FEMS Immunol Med Microbiol, 1995 Oct, 12(2), 97 - 112 Cytokine-inducing glycolipids in the lipoteichoic acid fraction from Enterococcus hirae ATCC 9790; Suda Y et al.; Five high molecular weight glycolipids capable of stimulating human peripheral whole-blood cell cultures to cause interleukin 6 (IL-6) and tumor necrosis factor (TNF)-alpha induction were isolated from one of the lipoteichoic acid fractions (LTA-2) extracted from Enterococcus hirae ATCC 9790 (Tsutsui et al., (1991) FEMS Microbiol . Immunol . 76, 211-218) by a combination of hydrophobic interaction and anion-exchange chromatographies . This purification procedure resulted in a remarkable increase in the cytokine-inducing activities on the weight basis of isolated glycolipids (a maximum of 36- and 17-fold increases of IL-6 and TNF-alpha induction, respectively) . The total yield of these bioactive glycolipids amounted to 6 wt% of the parent LTA-2 fraction, while the recovery rate in terms of the cytokine-inducing activities was estimated to be sufficient . The chemical composition and the profile, using SDS-PAGE, revealed that all of the isolated bioactive components were high molecular weight glycolipids, which were distinct from each other and from the parent LTA-2 fraction . These findings suggest that the IL-6 and TNF-alpha-inducing activities previously noted in the parent LTA-2 fraction are not attributable to a chemical entity, the structure of which had been proposed elsewhere (Fischer, W . (1990) in Glycolipids, Phosphoglycolipids and Sulfoglycolipids (Kates, M . ed.) pp . 123-234, Plenum Press, New York), but to the other high molecular weight glycolipids described here. J Clin Microbiol, 1995 Oct, 33(10), 2770 - 3 Vancomycin-dependent Enterococcus faecium isolated from stool following oral vancomycin therapy; Dever LL et al.; The isolation of clinical strains of enterococci requiring vancomycin for growth has only recently been reported . We describe the isolation of Enterococcus faecium requiring vancomycin for growth from the stool of a patient who had completed oral vancomycin therapy . Growth of the vancomycin-dependent E . faecium was supported by ristocetin and D-alanyl-D-alanine but not by daptomycin, teicoplanin, or D,L-alanine . Spontaneous revertants not requiring vancomycin occurred at a rate of 1 in 10(6) . Both the vancomycin-dependent E . faecium and the revertant hybridized with a vanB gene probe and had identical contour-clamped homogeneous electrophoresis patterns . The majority of revertant colonies were resistant to teicoplanin, suggesting constitutive production of the vanB ligase . We believe the vancomycin-dependent E . faecium evolved from a vancomycin-resistant, vancomycin-independent E . faecium in the presence of high concentrations of vancomycin in the intestine. J Bacteriol, 1995 Oct, 177(20), 5912 - 7 Identification of a gene (arpU) controlling muramidase-2 export in Enterococcus hirae; Lleo MM et al.; Muramidase-2 of Enterococcus hirae is a 74-kDa peptidoglycan hydrolase that plays a role in cell wall growth and division . To study its regulation, we isolated a mutant defective in muramidase-2 release under certain growth conditions . This mutant had cell walls which apparently lacked 74-kDa muramidase-2 but which accumulated two proteolytic fragments of 32 and 43 kDa, which exhibited muramidase-2 activity in the membrane fraction . By complementation cloning, we identified a 2.6-kb fragment of the E . hirae chromosome containing a gene cluster coding for proteins of 58 to 137 amino acids . One of these genes (arpU), which encoded a 15.9-kDa protein, was shown to complement the defect of the A9 mutant in trans . We propose that this gene may be involved in the regulation of muramidase-2 export. FEMS Microbiol Lett, 1995 Oct 1, 132(1-2), 107 - 14 Induction of vancomycin resistance in Enterococcus faecium by non-glycopeptide antibiotics; Allen NE et al.; Bacitracin and other antibiotics that inhibit late stages in peptidoglycan biosynthesis induce vancomycin resistance in a high-level, inducibly vancomycin-resistant strain of Enterococcus faecium . Exposure to bacitracin led to synthesis of the lactate-containing UDP-MurNAc-pentadepsipeptide precursor required for vancomycin resistance . These findings indicate that inhibition of peptidoglycan biosynthesis can lead to induction of vancomycin resistance and raise the possibility that multiple signals may serve to induce resistance. Surgery, 1995 Oct, 118(4), 716 - 21; discussion 721-3 Definition of the role of enterococcus in intraabdominal infection: analysis of a prospective randomized trial; Burnett RJ et al.; BACKGROUND . The role of enterococcus in intraabdominal infection is controversial . This study examines the contribution of enterococcus to adverse outcome in a large intraabdominal infection trial . METHODS . A randomized prospective double-blind trial was performed to compare two different antimicrobial regimens in combination with surgical or percutaneous drainage in the treatment of complicated intraabdominal infections . A total of 330 valid patients was enrolled from 22 centers in North America . RESULTS . In 330 valid patients, 71 had enterococcus isolated from the initial drainage of an intraabdominal focus of infection . This finding was associated with a significantly higher treatment failure rate than that of patients without enterococcus (28% versus 14%, p < 0.01) . In addition, only Acute Physiology and Chronic Health Evaluation II score and presence of enterococcus were significant independent predictors of treatment failure when stepwise logistic regression was performed (p < 0.01 and < 0.03) . Risk factors for the presence of enterococcus include age, Acute Physiology and Chronic Health Evaluation II, preinfection hospital length of stay, postoperative infections, and anatomic source of infection . There was no difference between the clinical trial treatment regimens with regard to overall failure, failure associated with enterococcus, or frequency of enterococcal isolation . CONCLUSIONS . This study is the first to report enterococcus as a predictor of treatment failure in complicated intraabdominal infections . This trial also identifies several significant risk factors for the presence of enterococcus in such infections. Orv Hetil, 1995 Oct 1, 136(40), 2161 - 4 {Experience with microbiological studies of the diabetic foot}; Kajetan M et al.; In order to determine the microbiological characteristics of diabetic foot infection, 60 diabetic patients (21 women, 39 men; age between 43 and 89 years with a duration of diabetes from 0.5 to 37 years) were investigated . Immediately after the hospitalisation specimens from infected foot lesions were taken using Port-A-Cul special transport medium . Aerobic cultures were done in all cases according to conventional methods while anaerobic cultures were carried out when clinical signs indicated to perform it . Out of 60 lesions only 2 proved to be sterile . In the remaining 58 patients a total of 138 isolates were found resulting in an average of 2.3 organisms per lesions . Only aerobic isolates were identified in 45 patients whereas anaerobic species were also found in 12 patients . Only Candida was found in 1 patient while Candida in combination with bacterial strains was observed in 3 patients . In antimicrobial susceptibility testing beta-lactamase-stable antibiotics with broad spectrum, covering enterococcus and anaerobic organisms proved to be most effective . Diabetic foot infections have a polymicrobial nature . Antibiotic treatment of infections should be based on the results of microbiological investigation of diabetic foot. J Infect Dis, 1995 Oct, 172(4), 993 - 1000 Epidemiology and mortality risk of vancomycin-resistant enterococcal bloodstream infections; Shay DK et al.; Risk factors for vancomycin-resistant enterococcal (VRE) bloodstream infection (BSI) were studied at a tertiary-care hospital by comparing 46 patients with VRE-BSI with 46 randomly selected patients with vancomycin-susceptible enterococcal (VSE) BSI . Among patients with an enterococcal BSI, risk factors for mortality were determined . Independent risk factors for VRE-BSI were increasing APACHE II score (odds ratio {OR}, 2.3/5-point increase; 95% confidence interval {CI}, 1.4-3.9), receipt of vancomycin (OR, 11; 95% CI, 5.5-21), or diagnosis of hematologic malignancy (OR, 8.4; 95% CI, 3.9-18) . After controlling for APACHE II score and gender, patients with VRE- versus VSE-BSI did not have a significantly elevated risk of mortality (OR, 3.3; 95% CI, 0.7-15) . Five of 28 VRE blood isolates typed using pulsed-field gel electrophoresis had identical banding patterns . These data suggest that increasing severity of illness, underlying disease, and receipt of vancomycin are major risk factors for VRE-BSI. J Biol Chem, 1995 Sep 29, 270(39), 23143 - 9 Cross-talk between the histidine protein kinase VanS and the response regulator PhoB . Characterization and identification of a VanS domain that inhibits activation of PhoB; Fisher SL et al.; VanS is a two-component transmembrane sensory kinase that, together with its response regulator VanR, activates the expression of genes responsible for vancomycin resistance in Enterococcus faecium BM4147 . In this report, we demonstrate that the cytoplasmic domain of VanS (including residues Met95 to Ser384) is capable of high level activation (> 500 fold) of the Escherichia coli response regulator PhoB in vivo in the absence of its signaling kinases PhoR, CreC (PhoM), or acetyl phosphate synthesis . In vitro experiments carried out on the purified proteins confirmed that the activation is due to efficient cross-talk between VanS and PhoB, since phospho-VanS catalyzed transfer of its phosphoryl group to PhoB with approximately 90% transfer in 5 min at a 1:4 VanS/PhoB stoichiometry . However, the rate of transfer was at least 100-fold slower than that observed between phospho-VanS and VanR . The in vivo activation of PhoB was used as a reporter system to identify peptide fragments of VanS capable of interfering with activation by VanS(Met95-Ser384), in order to identify an interaction domain . A library of plasmids encoding fragments of VanS(Met95-Ser384) was constructed using transposon mutagenesis, and a subpopulation of these plasmids encoded peptides that interfered with activation of PhoB by VanS(Met95-Ser384) . A minimal size fragment (Met95-Ile174) was shown to be both necessary and sufficient for potent inhibition (85%) of this activation. Antimicrob Agents Chemother, 1995 Sep, 39(9), 2112 - 5 Characterization of antimicrobial resistance in enterococci of animal origin; Thal LA et al.; Among 97 enterococci cultured from animals, gentamicin MICs were > or = 2,000 micrograms/ml for 9 isolates and between 250 and 1,024 micrograms/ml for 6 isolates . For two isolates tested (gentamicin MICs, 256 and 512 micrograms/ml, respectively), there was no in vitro synergy with penicillin plus gentamicin, resistance was transferable, and there was no hybridization with a probe specific for 6'-aminoglycoside acetyltransferase-2"-aminoglycoside phosphotransferase . The results of the study indicate the presence of a unique gentamicin resistance genotype in enterococci of animal origin. Antimicrob Agents Chemother, 1995 Sep, 39(9), 2068 - 72 Identification of daptomycin-binding proteins in the membrane of Enterococcus hirae; Boaretti M et al.; Daptomycin, a lipopeptide antibiotic active against gram-positive bacteria, was preliminarily shown to inhibit lipoteichoic acid (LTA) synthesis as a consequence of membrane binding in the presence of Ca2+ (P . Canepari, M . Boaretti, M . M . Lleo, and G . Satta, Antimicrob . Agents Chemother . 34:1220-1226, 1990) . In the present study, it is shown that, along with binding bacterial-membrane components, daptomycin binds the protein fraction with a noncovalent bond, as suggested by the instability of the bond in the presence of ionic detergents such as sodium dodecyl sulfate . Analysis of membrane proteins by isoelectric focusing electrophoresis reveals that five bands with isoelectric points ranging from 5.9 to 6.2 bind radioactive daptomycin . These proteins are therefore called daptomycin-binding proteins . In an attempt to correlate these proteins to the main inhibition observed during LTA synthesis, two-dimensional thin-layer chromatography of lipids synthesized during daptomycin treatment was performed . A threefold increase in diglucosyl diacylglycerol is demonstrated, while the compounds phosphatidyl-alpha-kojibiosyldiacylglycerol, glycerophospho-phosphatidyl-alpha-kojibiosyldiacylglycerol, and glycerophospho-kojibiosyldiacylglycerol, which follow diglucosyl diacylglycerol in LTA synthesis, decrease progressively with time during the course of daptomycin treatment. Clin Infect Dis, 1995 Sep, 21(3), 516 - 22 Enterococcal arthritis: case report and review; Raymond NJ et al.; We report a case of septic arthritis due to Enterococcus species and review 18 additional cases reported in the literature from 1966 through 1993 for which clinical or treatment data were available . In 11 of the 19 cases, prosthetic joints were affected (9 knees, 2 hips) and in 8 cases, native joints were affected . Of those patients with prosthetic joint infections, 6 had preexisting osteoarthritis and 3 had rheumatoid arthritis; only one patient with native joint infection had a recognized (although unspecified), preexisting joint abnormality . Pain, fever (temperature, > 37 degrees C), and tenderness were the most common clinical findings in patients with native joint infections . The microbiological diagnosis was made by culture of synovial fluid or synovial tissue (16 of 19), blood (1 of 19), or an u