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PubMed2004

Influence of protein binding under controlled conditions on the bactericidal activity of daptomycin in an in vitro pharmacodynamic model.

OBJECTIVE: Daptomycin exhibits bactericidal activity against clinically significant Gram-positive bacteria despite being highly bound to human proteins. Evaluations characterizing the effect of protein on daptomycin pharmacodynamics are warranted. METHODS: We utilized an in vitro pharmacodynamic model to simulate daptomycin regimens of 6 mg/kg/day under controlled conditions of pH, calcium and/or protein. Free concentrations were simulated in broth, whereas total concentrations were simulated in broth supplemented with human albumin. Bacterial density was profiled over 48 h for two methicillin-resistant Staphylococcus aureus (MRSA) and two vancomycin-resistant Enterococcus faecium (VREF) clinical isolates. RESULTS: Daptomycin exhibited bactericidal activity against both MRSA isolates, with time to 99.9% killing occurring at 0.5 h and 8 h in broth and in albumin-supplemented broth, respectively. Initial kill was observed against both VREF isolates followed by regrowth. There was no statistical difference (P>0.05) in extent of bacterial kill at 24 or 48 h between the different media. CONCLUSIONS: Although delayed, the extent of kill for daptomycin was unaltered against all isolates in albumin-supplemented broth. Further antimicrobial studies that incorporate protein are warranted to assess the influence of protein in the pharmacodynamic evaluation of antimicrobials.

Anti-Bacterial Agents↗

PubMed1998

Conditional adherence of Enterococcus faecalis to extracellular matrix proteins.

The adherence of 44 clinical isolates of Enterococcus faecalis, a common cause of endocarditis, and 13 Enterococcus faecium to substrates of six extracellular matrix (ECM) proteins was examined using 35S-labeled bacteria. One E. faecalis strain, isolated from a patient with endocarditis, adhered to collagen types I and IV and another E. faecalis strain adhered to laminin and to collagen types I and IV. However, most isolates showed little adherence ( < 5% of added cells adhered) when grown at 37 degrees C regardless of their source (endocarditis, urine or fecal sample). When grown at 46 degrees C (but not when grown in CO2 or nutrient limited media), most isolates of E. faecalis increased their adherence to immobilized laminin, collagen types I and IV but not to fibronectin, fibrinogen or bovine serum albumin, whereas none of the E. faecium increased adherence when grown at 46 degrees C or 50 degrees C. The adherence of E. faecalis was eliminated by digestion with trypsin, suggesting that a protein is somehow important, directly or indirectly, for adherence to occur. Pre-incubation of bacteria with soluble collagen types I and IV inhibited the adherence to these ECM proteins. These results demonstrate that in E. faecalis, adherence to ECM proteins is produced during routine in vitro growth conditions by occasional isolates and can be produced during certain stressful growth conditions by others. Whether this adherence relates to the propensity of E. faecalis to cause endocarditis remains to be determined.

Animals↗

PubMed2005

An alcohol hand rub containing a synergistic combination of an emollient and preservatives: prolonged activity against transient pathogens.

A new alcohol-based hand antiseptic (Octoxy hand rub) containing a synergistic combination of an emollient (Octoxyglycerine) and preservatives was developed and evaluated for immediate and prolonged activity against transient bacteria. The in vitro and in vivo antimicrobial efficacy was compared with other alcohol hand rubs containing preservative/antimicrobial (Prevacare and Avagard). In vitro evaluation was carried out using a tube-dilution method and a pig-skin model. Rapid and prolonged efficacy in vivo was evaluated against Staphylococcus epidermidis on the hands of volunteers. Octoxy hand rub was 100% effective in rapidly killing pathogens including methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococcus faecium in vitro. In volunteers, all three hand rubs gave a significant reduction in microbial count within 15s. Octoxy hand rub showed significantly higher efficacy against S. aureus and Escherichia coli than Avagard and Prevacare 15 min after application to the pig-skin model, and against S. epidermidis in both the pig-skin model and in volunteers. Use of Octoxy hand rub with broad-spectrum immediate and prolonged antimicrobial activity may be a very effective way of improving hand hygiene without exposing the hands to higher concentrations of antimicrobials.

Administration, Cutaneous↗

PubMed1998

Heterogeneity of plant-associated streptococci as characterized by phenotypic features and restriction analysis of PCR-amplified 16S rDNA.

Thirty-five strains of streptococci isolated from forage grasses were examined by restriction analysis of PCR-amplified 16S rDNA. Using a set of seven endonucleases, five 16S rDNA genotypes were obtained. The isolates could be identified as belonging to the species Enterococcus faecium, Ent. mundtii, Ent. faecalis, Ent. casseliflavus and Lactococcus lactis ssp. lactis, respectively. To assign the isolates to one of these species, digestion with the endonuclease HinfI was sufficient. Data obtained from whole-cell protein analysis were in accordance with the species identification. Additionally, three of the five species found were subdivided into two related subgroups. Both methods are suitable for a rapid and unequivocal identification of plant-associated streptococci. The phenotypic characters showed a high diversity and were in some cases contrary to the expected features usually given in genus and species descriptions. The results demonstrated the close relatedness of the determined species in contrast to their high phenotypic heterogeneity.

Bacterial Typing Techniques↗

PubMed2002

First outbreak of vancomycin-resistant enterococci in a tertiary hospital in Turkey.

Twenty multidrug-resistant vancomycin-resistant Enterococcus faecium strains of the VanA phenotype were isolated over a 1 year period from five patients in the intensive care unit at the University Hospital of Antalya, Turkey. Molecular investigation showed that these strains belonged to five different pulsotypes and that the vanA gene was carried by a Tn1546-like transposon inserted onto a self-transferable plasmid of approximately 200 kb. One patient was infected by two different strains, suggesting horizontal gene transfer within that patient. This is the first documented outbreak of VRE in Turkey with concomitant spread of plasmid and strains.

Adolescent↗

PubMed2006

Morphine withdrawal lowers host defense to enteric bacteria: spontaneous sepsis and increased sensitivity to oral Salmonella enterica serovar Typhimurium infection.

Understanding the consequences of drug withdrawal on immune function and host defense to infection is important. We, and others, previously demonstrated that morphine withdrawal results in immunosuppression and sensitizes to lipopolysaccharide-induced septic shock. In the present study, the effect of morphine withdrawal on spontaneous sepsis and on oral infection with Salmonella enterica serovar Typhimurium was examined. Mice were chronically exposed to morphine for 96 h by implantation of a slow-release morphine pellet. Abrupt withdrawal was induced by removal of the pellet. In the sepsis model, bacterial colonization was examined and bacterial species were identified by necropsy of various tissues. It was found that at 48 h postwithdrawal, morphine-treated mice had enteric bacteria that were detected in the Peyer's patches (4/5), mesenteric lymph nodes (4/5), spleens (4/10), livers (6/10), and peritoneal cavities (8/10). In placebo pellet-withdrawn mice, only 2/40 cultures were positive. The most frequently detected organisms in tissues of morphine-withdrawn mice were Enterococcus faecium followed by Klebsiella pneumoniae. Both organisms are part of the normal gastrointestinal flora. In the infection model, mice were orally inoculated with S. enterica 24 h post-initiation of abrupt withdrawal from morphine. Withdrawal significantly decreased the mean survival time and significantly increased the Salmonella burden in various tissues of infected mice compared to placebo-withdrawn animals. Elevated levels of the proinflammatory cytokines were observed in spleens of morphine-withdrawn mice, compared to placebo-withdrawn mice. These findings demonstrate that morphine withdrawal sensitizes to oral infection with a bacterial pathogen and predisposes mice to bacterial sepsis.

Animals↗

PubMed2005

Biofilm formation and esp gene carriage in enterococci.

AIMS: To investigate biofilm production and esp carriage in enterococci. METHODS: Biofilm production in vancomycin resistant enterococci (VRE) and vancomycin susceptible enterococci (VSE) was tested on a microtitre plate method, using both brain heart infusion (BHI) broth and human serum as media. Isolates were screened for the esp gene, which has been reported to be essential for biofilm formation in enterococci, by means of the polymerase chain reaction. RESULTS: None of seven VRE and nine of 28 VSE tested formed a biofilm. One initially negative VRE Enterococcus faecium isolate produced a strong biofilm after 21 weeks of dry starvation on a cotton swab. By Fisher's exact test, there was no significant difference in biofilm formation between VRE and VSE, E faecalis and E faecium, or isolates from different sites. Biofilm formation was independent of possession of the esp gene. One isolate produced a strong biofilm in human serum but only a weak biofilm in BHI, whereas another produced a moderate biofilm in human serum but a weak biofilm in BHI. CONCLUSIONS: The acquisition of vancomycin resistance may result in a lower ability to form biofilms, but a larger study using clinical isolates is needed to test this hypothesis. That one initially negative VRE isolate produced a strong biofilm after prolonged dry starvation suggests that biofilm formation may be an adaptive response. The esp gene does not appear to be necessary or sufficient for production of biofilms in enterococci.

Bacterial Proteins↗

PubMed2001

Isolation and characterization of enterocin BC25 and occurrence of the entA gene among ruminal gram-positive cocci.

Enterocin BC25, a bacteriocin produced by Enterococcus faecium BC25 isolated from the rumen of cow was purified to homogeneity and sequenced. Twenty amino acids were identified in the peptide chain (TTHSGKYYGNGVYCT-KNKCT), identical to the N-terminal sequence of enterocin A. The DNA sequence of the enterocin BC25 structural gene and putative immunity protein exhibited high similarity to the entA gene. The occurrence of a 726 bp amplicon containing the enterocin A structural gene was studied among gram-positive ruminal cocci by PCR. Our results showed wide occurrence of the entA structural gene among ruminal enterococcal and streptococcal bacterial strains tested, and indicate variable ability to express bacteriocin production and resistance.

Amino Acid Sequence↗

PubMed2003

Direct-fed microbial supplementation on the performance of dairy cattle during the transition period.

The influence of a direct-fed microbial (DFM) on the prepartum period and the effects on production performance during the postpartum period was investigated using 64 multiparous Holstein cows. Two close-up dry cow diets were fed to two groups of 32 cows each starting 21 d precalving as follows: 1) no DFM and 2) DFM. Post-calving cows were fed a lactation ration with or without DFM supplementation.The direct-fed microbial (DFM) supplement contained 2 × 109 viable yeast cells and 5 × 109 cfu of bacteria (Enterococcus faecium) per cow per day, top dressed in a 90-g supplement [corrected].The DMI during the prepartum period was not affected by DFM supplementation. During the postpartum period, DMI, milk yield, and milk protein content were higher for cows receiving DFM supplementation compared with no DFM. Blood glucose and insulin levels were higher and NEFA levels were lower for cows receiving DFM during the postpartum period. These data suggest that targeted DFM supplementation increased DMI and milk production postpartum. Blood metabolite information would suggest this response was associated with more glucose being made available and less fatty acids being mobilized from lipid stores.

Animals↗

PubMed2004

[Vancomycin resistant enterococci in the Netherlands].

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

Disease Outbreaks↗

PubMed1993

Sugar uptake and involved enzymatic activities by yeasts and lactic acid bacteria: their relationship with breadmaking quality.

The uptake kinetics of sugars present in wheat doughs and alpha-glucosidase as well as beta-fructosidase activities were determined in different strains of yeasts and lactic acid bacteria. These strains were previously selected according to their breadmaking quality. Saccharomyces cerevisiae (P6), Candida guilliermondii (P40), Lactobacillus plantarum (B31 and La18) and L. brevis (B21) showed good performance, while Sacch. fructuum (P43), L. cellobiosus (B37) and Enterococcus faecium (B11) yielded bread of lower quality. Leuconostoc mesenteroides (B10), when used in combination with other strains led also to high quality starters. All yeast strains used assimilated glucose, fructose and maltose, exhibiting saturable kinetics. Lactic acid bacteria showed saturable kinetics only for hexoses, whereas disaccharide uptake was linear. Sacch. cerevisiae, Leuconostoc mesenteroides, L. brevis and L. plantarum (B31) displayed better sugar uptake properties. For all the strains used alpha-glucosidase and beta-fructosidase activities were detected. The highest specific activities were found for Sacch. cerevisiae, C. guilliermondii and L. plantarum (B31). These results indicate good correlation between the parameters evaluated and the breadmaking potential of the microorganisms.

Biological Transport↗

PubMed1996

Inhibition of peptidoglycan biosynthesis in vancomycin-susceptible and -resistant bacteria by a semisynthetic glycopeptide antibiotic.

LY191145 is a p-chlorobenzyl derivative of LY264826 (A82846B) with activity against both vancomycin-susceptible and -resistant enterococci. Incorporation of L-[14C]lysine into peptidoglycan of intact vancomycin-susceptible and -resistant Enterococcus faecium was inhibited by LY191145 (50% inhibitory concentrations of 1 and 5 microgram/ml, respectively). Inhibition was accompanied by accumulation of UDP-muramyl-peptide precursors in the cytoplasm. This agent inhibited late-stage steps in peptidoglycan biosynthesis in permeabilized E. faecium when either the UDP-muramyl-pentapeptide precursor from vancomycin-susceptible E. faecium or the UDP-muramyl-pentadepsipeptide precursor from vancomycin-resistant E. faecium was used as a substrate. Inhibition of late-stage steps led to accumulation of an N-acetyl-[14C]glucosamine-labeled lipid intermediate indicative of inhibition of the transglycosylation step. Inhibition of peptidoglycan polymerization without affecting cross-linking in a particulate membrane-plus-wall-fragment assay from Aerococcus viridans was consistent with this explanation. The fact that inhibition of peptidoglycan biosynthesis by LY191145 was not readily antagonized by an excess of free acyl-D-alanyl-D-alanine or acyl-D-alanyl-D-lactate ligands indicates that the manner in which this compound inhibits transglycosylation may not be identical to that of vancomycin.

Anti-Bacterial Agents↗

PubMed2003

In vivo efficacy of the ketolide ABT-773 (cethromycin) against enterococci in a mouse peritonitis model.

Using six Enterococcus faecalis and five Enterococcus faecium strains, the ketolide ABT-773 (ABT), now known as cethromycin, was found to have in vivo efficacy against both erythromycin (ERY)-susceptible (Ery(s)) and -intermediate (Ery(i)) enterococci (ABT 50% protective doses [PD(50)s], 0.5 to 4.1 and 10.3 to 16.2 mg/kg of body weight, respectively). Against four highly Ery-resistant (Ery(r)) strains for which ABT MICs were low, ABT showed much greater activity (PD(50), 6.3 to 32.5 mg/kg) than ERY (PD(50), >200 mg/kg) but was not protective for strains for which ABT MICs were high. In conclusion, ABT-773 showed in vivo efficacy and considerably greater activity than ERY in a mouse peritonitis model.

Animals↗

PubMed2000

In vitro action of a combination of selected antimicrobial agents and chondroitin sulfate.

Chondroitin sulfate (CS), a highly anionic polymer and the most predominant sulfated glycosaminoglycan in connective tissues, was investigated regarding to its interaction with cationic disinfectants, which are used as antiinfectives in humans. Combinations of cetylpyridiniumchloride (CPC), chlorhexidine (CHex), and polyhexamethylene biguanide (PHMB) with CS, respectively, were prepared and the resulting microbicidal activity of the mixtures was tested in the quantitative suspension test without organic matter. Polyvidone-iodine and Ringer's solution were used as controls. Even precipitated, the resulting test combinations behave differently against Staphylococcus aureus, Enterococcus faecium, Escherichia coli, Pseudomonas aeruginosa, and Candida albicans. CPC/CS demonstrated only microbicidal activity against Gram-positive bacteria, and CHex/CS was more active against Gram-negative bacteria and C. albicans. PHMB/CS, especially in combination with CS-A, only revealed an antimicrobial effect against P. aeruginosa after 60 min action. The interaction of cationic disinfectants with CS showed depending on the investigated microorganism a more or less controlled sustained release manner of the microbicidal agent from the precipitated complex, with the only exception of PHMB in combination with CS-C, which is completely neutralized. Polyvidone-iodine and Ringer's solution were not affected by CS.

Anti-Infective Agents, Local↗

PubMed2003

Antimicrobial resistance in swine and chickens fed virginiamycin for growth promotion.

In a prospective controlled study, we evaluated pigs (5-month period) and chickens (11-week period) fed subtherapeutic levels of virginiamycin. A total of 13 Enterococcus faecium were isolated from 10 pigs and 17 from 8 chickens. There were 8 pulsed-field gel electrophoresis (PFGE) patterns in E. faecium isolates from pigs and 17 from chickens. Resistance to quinupristin/dalfopristin resistance occurred in 2 of 13 E. faecium from pigs and 2 of 17 E. faecium from chickens. There were no strains exhibiting high-level gentamicin (MIC> or =2000 microg/ml) or vancomycin resistance. There was no relative weight gain in animals that received virginiamycin. The mean weight increase for the pigs in the group fed virginiamycin was 107.6 lb vs. 126.4 lb in the group that did not receive virginiamycin (P=n.s.). Chickens fed virginiamycin had a mean weight increase of 1672 g vs. 1886 g in the group that did not receive virginiamycin (P=n.s.). There was no correlation between receipt of virginiamycin or weight gain and presence of quinupristin/dalfopristin-resistant strains.

Animals↗

PubMed1994

Clinical and molecular epidemiology of enterococcal bacteremia in a pediatric teaching hospital.

An apparent increase in the incidence of enterococcal bacteremias from 7 to 48/1000 bacteremias during 1986 to 1991 (p < 0.01) prompted this descriptive clinical and molecular epidemiologic study of 83 episodes occurring in 80 children between 1986 and 1992. Most community-acquired cases were in infants, in comparison with nosocomial episodes (24/26 and 34/57; p < 0.01); many of them were neonates (10/26 and 6/57; p < 0.01). Nosocomial cases were associated with underlying conditions including major surgery 56%, immunosuppression 49%, organ and tissue transplants 30%, and cardiac 32%, pulmonary 25%, renal 21%, and hepatic 21% disorders. Nosocomial episodes developed after a median of 32 days. There were 58 primary and 25 secondary bacteremias. Thirty-two episodes were polymicrobial and 44 organisms were involved. Twenty-six percent of the patients died. In 15%, death was preceded by septic shock, disseminated intravascular coagulation, and polymicrobial bacteremia (p < 0.01). Of 75 isolates, 82% were Enterococcus faecalis and 14% were Enterococcus faecium. Fourteen E. faecalis strains produced hemolysin; none produced beta-lactamase. Three had high-level resistance to gentamicin and 13 to streptomycin; two E. faecium and none of the E. faecalis strains were vancomycin resistant at a low level (p < 0.01) and one was ampicillin resistant. Pulsed-field gel electrophoresis of whole-cell DNA digested with restriction enzymes Sma I and Eag I showed five isolates with a homogeneous pattern, two with another homogeneous pattern, and 68 with distinct heterogeneous patterns. The increase in enterococcal bacteremias was not due to a clonal strain dissemination but to an increase in cases of heterogeneous enterococcal strains. We conclude that enterococcal septicemia is now an important cause of serious morbidity and death in critically ill children.

Anti-Bacterial Agents↗

PubMed2006

[Molecular analysis of vancomycin-resistant enterococci isolated from clinical samples].

Enterococcus species are the inhabitants of gastrointestinal flora and can cause endocarditis, urinary tract infections, and bacteremia. In recent years, infections caused by vancomycin-resistant enterococci (VRE) have increased in our country. The infections caused by VRE are treated with glycopeptide antibiotics like vancomycin and teicoplanin. Although seven different resistant genes have been described in VRE, VanA is the most frequently detected one in Turkey. The aim of this study was (i) to genotype four vancomycin-resistant Enterococcus faecium strains isolated in our hospital, by using pulsed field gel electrophoresis (PFGE); and (ii) to detect the type of Van gene by using polymerase chain reaction. Three of the strains were isolated from blood cultures and one from cerebrospinal fluid of the hospitalized patients between the years 2000-2004, and identified by both conventional methods and commercial kits. By using PFGE, we detected that isolates were different according to Tenover criteria and they had VanA genes. As a result, VanA carrying VRE cause sporadic infections in our hospital. The molecular epidemiologic investigation of resistant microorganisms is important in terms of infection control and epidemiology.

Bacteremia↗

PubMed1998

Evaluation of in vitro activity of quinupristin/dalfopristin and comparator antimicrobial agents against worldwide clinical trial and other laboratory isolates.

This report summarizes the activities of quinupristin/dalfopristin (Q/D) and appropriate comparator antibiotics, including ciprofloxacin, erythromycin, gentamicin, rifampin, teicoplanin, and vancomycin, against selected gram-positive pathogens, including Enterococcus faecium, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus pneumoniae, Streptococcus agalactiae, and Streptococcus pyogenes. The study pathogens were obtained from 2 sources: (1) clinical isolates taken from patients participating in Q/D worldwide Phase III comparative and noncomparative (emergency-use program) clinical trials; and (2) other isolates collected from the laboratories of 45 geographically distinct medical centers around the world. Q/D was highly active, with minimum inhibitory concentrations (MICs) < or = 1.0 microg/mL against most isolates, including those known to be resistant to methicillin, vancomycin, or erythromycin. Q/D was active (MICs < or = 1 microg/mL) against 95% of the vancomycin-resistant E. faecium strains, for example, whereas ciprofloxacin was active against 6%. Q/D was equally active against methicillin-susceptible or -resistant S. aureus strains (MIC90=1 microg/mL), as was vancomycin (MIC90=2 microg/mL), whereas ciprofloxacin was much less active against methicillin-resistant strains than against methicillin-susceptible strains (MIC90=32 vs 1 microg/mL). Given its spectrum of activity, Q/D may provide a viable option for the treatment of severe respiratory and skin and skin-structure infections caused by gram-positive bacteria, especially when strains with known or suspected resistance to other commonly used antibiotics are present.

Anti-Bacterial Agents↗