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PubMed2006

Measurement of ampicillin, vancomycin, linezolid and gentamicin activity against enterococcal biofilms.

BACKGROUND: Enterococci frequently cause biofilm infections but susceptibility of clinical isolates growing in biofilms has not been investigated. The minimum biofilm eradicating concentration (MBEC) has been suggested as a guide to treatment of biofilm infections. We measured an alternative endpoint, the minimum biofilm inhibitory concentration (MBIC) and compared the results with MIC and MBC. OBJECTIVES: To compare the MIC, MBC and MBIC of ampicillin, vancomycin and linezolid against enterococcal biofilms, to assess the impact of additional gentamicin and correlate findings with clinical outcome. METHODS: MIC and MBC were measured using standard techniques. MBICs were measured using a modification of the Calgary biofilm device method. Fifty-eight enterococcal isolates from episodes of intravascular catheter-related bloodstream infection were tested. RESULTS: Tolerance to ampicillin, vancomycin and linezolid was seen in 93%, 100% and 93% of isolates, respectively. MIC(90)s of ampicillin, vancomycin and linezolid were all 4 mg/L for Enterococcus faecalis isolates. MBC(90)s of ampicillin, vancomycin and linezolid for E. faecalis isolates were 1024, >128 and 2048 mg/L, respectively. MBIC(90)s of ampicillin, vancomycin and linezolid for E. faecalis isolates were 8192, 4096 and 4096 mg/L, respectively. Results for Enterococcus faecium were similar for vancomycin and linezolid but this species was generally more resistant to ampicillin. Adding 10 mg/L gentamicin had a variable effect on MIC, MBC or MBIC, which was not predictable by gentamicin susceptibility on disc testing. CONCLUSIONS: Very high concentrations of ampicillin, vancomycin and linezolid are required to inhibit enterococcal biofilms in vitro. Combining these agents with gentamicin significantly reduced MIC, MBC and MBIC against only a proportion of enterococcal isolates. No correlation between MBIC and outcome was found.

Acetamides↗

PubMed1999

An easy and safe method to store and disinfect explanted skull bone.

In our department extensive decompression craniectomies became the treatment of choice for patients with massive cerebral oedema following either trauma or acute cerebral infarction. The remarkable survival rates of this neurosurgical technique created the problem of adequate vault defect reconstruction. To evaluate the biological safety of using stored autologous skull flaps for this purpose, we compared three different disinfection methods. Skull bone fragments stored at -21 degrees C for different periods of time were artificially contaminated with clinically relevant strains of Serratia marcescens, Enterococcus faecium and Staphylococcus aureus. As potential methods for disinfection we tested immersion in 3% H2O2, boiling in normal saline for 15 and 30 minutes and a special process of steam disinfection at a temperature of 75 degrees C for 20 minutes. We were able to demonstrate that only steam disinfection completely eliminated the bacterial strains tested. Refrigeration plus steam disinfection of autologous skull bone prior to re-implantation seems to offer reliable safety for its use for defect closure. It is available at reasonable cost in many hospitals and does not require a bone bank.

Bone Transplantation↗

PubMed2001

Inhibition of vancomycin-resistant enterococci by an in vitro continuous-flow competitive exclusion culture containing human stool flora.

An in vitro anaerobic continuous-flow competitive exclusion (CFCE) culture model was used to study the ability of human stool flora to inhibit the growth of vancomycin-resistant (VR) enterococci (VRE). The CFCE culture was established from a stool sample obtained from a healthy adult. When 10(3) or 10(6) cfu/mL of VR Enterococcus faecium were added to the CFCE culture, the VRE were eliminated within 6 or 9 days, respectively. When 10(7) cfu/mL of the CFCE culture was added to a continuous-flow culture that contained 10(7) cfu/mL of VRE, the density of VRE was reduced but not eliminated. These data support the hypothesis that the indigenous intestinal flora inhibit growth of VRE and suggest that CFCE cultures may be a useful means to study interactions between the indigenous flora and VRE.

Anaerobiosis↗

PubMed2001

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

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

Adult↗

PubMed2002

Bacterial bioinformatics: pathogenesis and the genome.

As the number of completed microbial genome sequences continues to grow, there is a pressing need for the exploitation of this wealth of data through a synergistic interaction between the well-established science of bacteriology and the emergent discipline of bioinformatics. Antibiotic resistance and pathogenicity in virulent bacteria has become an increasing problem, with even the strongest drugs useless against some species, such as multi-drug resistant Enterococcus faecium and Mycobacterium tuberculosis. The global spread of Human Immunodeficiency Virus (HIV) and Acquired Immune Deficiency Syndrome (AIDS) has contributed to the re-emergence of tuberculosis and the threat from new and emergent diseases. To address these problems, bacterial pathogenicity requires redefinition as Koch's postulates become obsolete. This review discusses how the use of bacterial genomic information, and the in silico tools available at present, may aid in determining the definition of a current pathogen. The combination of both fields should provide a rapid and efficient way of assisting in the future development of antimicrobial therapies.

Acquired Immunodeficiency Syndrome↗

PubMed2004

Isolation and characterisation of an equol-producing mixed microbial culture from a human faecal sample and its activity under gastrointestinal conditions.

Only about one third of humans possess a microbiota capable of transforming the dietary isoflavone daidzein into equol. Little is known about the dietary and physiological factors determining this ecological feature. In this study, the in vitro metabolism of daidzein by faecal samples from four human individuals was investigated. One culture produced the metabolites dihydrodaidzein and O-desmethylangolensin, another produced dihydrodaidzein and equol. From the latter, a stable and transferable mixed culture transforming daidzein into equol was obtained. Molecular fingerprinting analysis (denaturing gradient gel electrophoresis) showed the presence of four bacterial species of which only the first three strains could be brought into pure culture. These strains were identified as Lactobacillus mucosae EPI2, Enterococcus faecium EPI1 and Finegoldia magna EPI3, and did not produce equol in pure culture. The fourth species was tentatively identified as Veillonella sp strain EP. It was found that hydrogen gas in particular, but also butyrate and propionate, which are all colonic fermentation products from poorly digestible carbohydrates, stimulated equol production by the mixed culture. However, when fructo-oligosaccharides were added, equol production was inhibited. Furthermore, the equol-producing capacity of the isolated culture was maintained upon its addition to a faecal culture originating from a non-equol-producing individual.

Bacteria↗

PubMed2005

Dutch guideline for preventing nosocomial transmission of highly resistant microorganisms (HRMO).

Hospitals are faced with the increasingly rapid emergence and dissemination of antimicrobial-resistant microorganisms. US and European guidelines on the prevention of antimicrobial resistance in hospitals were, until recently, mainly directed at methicillin-resistant Staphylococcus aureus (MRSA). In 2004, the Dutch Working Party on Infection Prevention issued a guideline on the prevention of nosocomial transmission of highly resistant microorganisms (HRMO), in order to fulfill the growing need for additional guidance on the control of other pathogens with acquired resistance and the potential to spread within hospitals (such as glycopeptide-resistant Enterococcus faecium, penicillin-resistant Streptococcus pneumoniae, extendedspectrum beta-lactamase producing Enterobacteriaceae, and other (multi)drug-resistant gram-negatives). In addition to providing criteria for defining HRMO, the Dutch guideline provides recommendations on isolation of patients, active surveillance, and contact tracing. The guideline will enable the comparison of HRMO rates between hospitals, and may be used to evaluate the efficacy of programs to control antibiotic use and/or nosocomial transmission of resistant pathogens. The eventual success of nationwide implementation of this guideline remains to be established in the coming years.

Cross Infection↗

PubMed1998

[Vancomycin resistant enterococci].

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

Animals↗

PubMed1998

Stability of vancomycin-resistant enterococcal genotypes isolated from long-term-colonized patients.

Genotypic variation and stability of isolates of vancomycin-resistant enterococci (VRE) were studied to determine genetic diversity and whether strain definition based on pulsed-field gel electrophoresis (PFGE) is applicable to an endemic setting. Twenty-two PFGE types were identified among 455 VRE isolates. One-on-one comparisons of 10 vanA Enterococcus faecium strain types all yielded > 10 band differences. Variations among vanA and vanB E. faecium isolates from individual long-term-colonized (4-160 days) patients yielded < 3 band differences for > 85% of comparisons. Comparison of all strains without grouping by vancomycin resistance types yielded two peaks of band differences: one with < 3 and one with > 10 band differences. These data show that VRE isolates were genetically closely related or very different; demonstrate that within individual patients, VRE isolates show little genetic variation; and provide empirical evidence that PFGE can be used to study the epidemiology of VRE endemicity.

Anti-Bacterial Agents↗

PubMed2000

Effects of probiotic bacteria on diarrhea, lipid metabolism, and carcinogenesis: a review of papers published between 1988 and 1998.

We reviewed the evidence from human intervention studies for the health effects of probiotic bacteria, ie, live bacteria that survive passage through the gastrointestinal tract and have beneficial effects on the host. Of the 49 studies reviewed, 26 dealt with the prevention or treatment of diarrheal disease, 9 with the prevention of cancer or of the formation of carcinogens, 7 with the lowering of serum cholesterol, and 7 with the stimulation of the immune system. The most widely studied probiotic bacteria were Lactobacillus GG (22 studies), Lactobacillus acidophilus (16 studies), Bifidobacterium bifidum (6 studies), and Enterococcus faecium (7 studies). Intake of Lactobacillus GG consistently shortened the diarrheal phase of rotavirus infection by 1 d. However, evidence for the prevention by Lactobacillus GG and other probiotics of diarrhea due to viral or bacterial infections was less strong. Effects of probiotics on the immune system are inconclusive because of the variety of outcome variables reported. Cholesterol lowering by L. acidophilus was shown in some but not all studies; cholesterol lowering by E. faecium seems to be transient. Two studies of one research group showed a smaller recurrence of bladder tumors in patients after treatment with Lactobacillus casei; these results await confirmation. The production of mutagens after a meal might be reduced by the concomitant intake of probiotics, but the relevance of this finding is unclear. In conclusion, consumption of foods containing Lactobacillus GG may shorten the course of rotavirus infection. Other health effects of probiotic bacteria have not been well established. Well-designed placebo-controlled studies with validated outcome variables are needed to determine the health effects of probiotics.

Bifidobacterium↗

PubMed2003

Antimicrobial resistance patterns of vancomycin-resistant Streptococcus equinus isolated from animal foods and epidemiological typing of resistant S. equinus by microbial uniprimer kit.

Raw milk samples, and cow and chicken intestines were tested to isolate vancomycin-resistant, gram-positive bacteria. From these samples, we isolated seven vancomycin-resistant Streptococcus equinus, two vancomycin-resistant viridans Streptococcus and two vancomycin-resistant Enterococcus faecium. The MICs of several antibiotics, including vancomycin, against these strains were tested. Seven isolates of S. equinus showed high level resistance to vancomycin and teicoplanin (>100 microg/mL). The cell wall thickness of these strains was compared with that of the sensitive strain by TEM and no differences were obserbed between these strains. We compared the strains of vancomycin-resistant Streptococcus equinus using PCR with Microbial Uniprimer Kit. We concluded that it is necessary to combine other methods in order to cluster and identify all isolates of S. equinus.

Animals↗

PubMedDate not supplied

Emergence of vancomycin-resistant enterococci.

Vancomycin and ampicillin resistance in clinical Enterococcus faecium strains has developed in the past decade. Failure to adhere to strict infection control to prevent the spread of these pathogens has been well established. New data implicate the use of specific classes of antimicrobial agents in the spread of vancomycin-resistant enterococci (VRE). Extended-spectrum cephalosporins and drugs with potent activity against anaerobic bacteria may promote infection and colonization with VRE and may exert different effects on the initial establishment and persistence of high-density colonization. Control of VRE will require better understanding of the mechanisms by which different classes of drugs promote gastrointestinal colonization.

Ampicillin Resistance↗

PubMed2006

Improvement of enterocin P purification process.

Purification and heterologous expression of enterocin P (EntP), a sec-dependent bacteriocin produced by Enterococcus faecium, in Escherichia coli is described. PCR-amplified product of the enterocin P structural gene entP was cloned into plasmid pET-32b under the control of the inducible T7lac promoter. The neo-synthesized EntP was genetically modified by an addition of 3 extra amino acids, leading to recombinant EntRP. Active EntRP was recovered from the cytoplasmic soluble fraction of E. coli harboring appropriate recombinant plasmid, characterized by ELISA and Western-blot analysis and purified by immunoaffinity chromatography. The use of E. coli as heterologous host and pET-32b as expressing vector offers promising tools for heterologous production of class IIa bacteriocin.

Bacteriocins↗

PubMed2003

Coexistence of vancomycin-resistant enterococci and Staphylococcus aureus in the intestinal tracts of hospitalized patients.

The potential for transfer of vancomycin-resistance genes from enterococci to Staphylococcus aureus exists when these organisms share an ecologic niche. We performed an 8-month prospective study to determine the frequency at which S. aureus and vancomycin-resistant enterococci (VRE) coexist in the intestinal tracts of VRE-colonized patients and evaluated whether antianaerobic antibiotic therapy promoted increased density of S. aureus colonization. Of 37 patients colonized with vancomycin-resistant Enterococcus faecium, 23 (62%) had S. aureus recovered from stool specimens and 20 (87%) had methicillin-resistant strains. There was no significant difference in the mean density (+/- standard deviation) of S. aureus during versus > or =1 month after discontinuation of antianaerobic antibiotic therapy (5.1+/-1.5 vs. 4.7+/-1.6 log10 colony-forming units per gram of stool; P=.34). No S. aureus isolates were resistant to vancomycin. S. aureus and VRE often coexist in the intestinal tract, providing a potential reservoir for the emergence of vancomycin-resistant S. aureus isolates.

Aged↗

PubMed2001

A simple model host for identifying Gram-positive virulence factors.

We demonstrate the use of the nematode Caenorhabditis elegans as a facile and inexpensive model host for several Gram-positive human bacterial pathogens. Enterococcus faecalis, Streptococcus pneumoniae, and Staphylococcus aureus, but not Bacillus subtilis, Enterococcus faecium, or Streptococcus pyogenes, kill adult C. elegans. Focusing our studies on the enterococcal species, we found that both E. faecalis and E. faecium kill C. elegans eggs and hatchlings, although only E. faecalis kills the adults. In the case of adults, a low inoculum of E. faecalis grows to a high titer in the C. elegans intestine, resulting in a persistent infection that cannot be eradicated by prolonged feeding on E. faecium. Interestingly, a high titer of E. faecium also accumulates in the nematode gut, but does not affect the longevity of the worms. Two E. faecalis virulence-related factors that play an important role in mammalian models of infection, fsr, a putative quorum-sensing system, and cytolysin, are also important for nematode killing. We exploit the apparent parallels between Gram-positive infection in simple and more complex organisms by using the nematode to identify an E. faecalis virulence factor, ScrB, which is relevant to mammalian pathogenesis.

Animals↗

PubMed1997

Emergence of vancomycin resistance in the genus Streptococcus: characterization of a vanB transferable determinant in Streptococcus bovis.

Streptococcus bovis NEM760 was isolated from a stool swab collected on admission from a patient as surveillance for vancomycin-resistant enterococci. Strain NEM760 was identified as S. bovis by conventional biochemical methods and partial sequence analysis of its 16S rRNA. This strain was resistant to a low level of vancomycin (MIC, 64 micrograms/ml) but was susceptible to teicoplanin (MIC, 1 micrograms/ml), and vancomycin induced resistance to both glycopeptides. The presence of a vanB-related gene in NEM760 was demonstrated in a PCR assay which enabled specific amplification of a 635-hp internal segment of vanB. Sequence analysis of the corresponding PCR product revealed that it was highly homologous (96% identity) to the prototype vanB sequence of Enterococcus faecalis V583. The VanB resistance of determinant of S. bovis NEM760 was transferred by conjugation to E. faecalis and Enterococcus faecium at a similar frequency of 2 x 10(-5) per donor. SmaI-digested genomic DNAs of independently obtained transconjugants of E. faecalis and E. faecium were analyzed by pulsed-field gel electrophoresis and Southern hybridization with a vanB DNA probe. The electrophoretic and hybridization patterns obtained with all transconjugants of the same species were indistinguishable and revealed vanB-containing chromosomal insertions of approximately 100 kb. These results suggest that the genes mediating VanB-type resistance in S. bovis NEM760 are part of large transferable genetic elements. The results presented in the report demonstrate for the first time the role of streptococci in the dissemination of vancomycin resistance among gram-positive bacteria.

Anti-Bacterial Agents↗

PubMed2004

[Test schemes for evaluation of the effect of a probiotic feed additive on the gut microflora of young calves and newly hatched turkeys].

Probiotic feed additives have to undergo a strict assessment according to EU regula-tions, including the evaluation of the influence on the gut microflora of the host animal. A probiotic strain of the genus Enterococcus faecium and two different host animal species, young calves and newly hatched turkeys, were chosen as a test model for the establishment of schemes for feeding trials. Investigations included faeces samples and pooled swab samples of faeces as well as samples from gut sections derived from diagnostic slaughtering. The samples were tested for aerobic total count, for enterococci count and for Enterobacteriaceae and E. coli. The test schemes were appropriate for the evaluation of the influence of the probiotic feed additive on the physiological gut microflora of the host animals. Additional information could be gathered concerning safety aspects and effectiveness.

Animal Feed↗

PubMed2003

Baseline study to determine in vitro activities of daptomycin against gram-positive pathogens isolated in the United States in 2000-2001.

The activity of daptomycin was assessed by using 6,973 gram-positive bacteria isolated at 50 United States hospitals in 2000 and 2001. Among the isolates of Streptococcus pneumoniae (n = 1,163) collected, the rate of penicillin resistance was 16.1%; rates of oxacillin resistance among Staphylococcus aureus isolates (n = 1,018) and vancomycin resistance among Enterococcus faecium isolates (n = 368) were 30.0 and 59.5%, respectively. Multidrug-resistant (MDR) phenotypes (isolates resistant to three or more different chemical classes of antimicrobial agents) accounted for 14.2% of S. pneumoniae isolates, 27.1% of S. aureus isolates, and 58.4% of E. faecium isolates. For all gram-positive species tested, MICs at which 90% of the isolates tested were inhibited (MIC(90)s) and MIC ranges for directed-spectrum agents (daptomycin, quinupristin-dalfopristin, and linezolid) were identical or highly similar for isolates susceptible or resistant to other agents or MDR. Daptomycin had a MIC(90) of 0.12 micro g/ml for both penicillin-susceptible and -resistant isolates of S. pneumoniae. Against oxacillin-resistant S. aureus daptomycin had a MIC(90) of 0.5 micro g/ml, and it had a MIC(90) of 4 micro g/ml against both vancomycin-susceptible and -resistant E. faecium. The MIC(90)s for daptomycin and other directed-spectrum agents were unaffected by the regional or anatomical origin of isolates or patient demographic parameters (patient age, gender, and inpatient or outpatient care). Our results confirm the gram-positive spectrum of activity of daptomycin and that its activity is independent of susceptibility or resistance to commonly prescribed and tested antimicrobial agents. This study may serve as a baseline to monitor future changes in the susceptibility of gram-positive species to daptomycin following its introduction into clinical use.

Anti-Bacterial Agents↗