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PubMed2001

Rhizobial survival and nodulation of chickpea as influenced by fungicide seed treatment.

The survival of Rhizobium ciceri on chickpea (Cicer arietinum cv. Myles) seed, treated separately with 1 of 4 commercial fungicides, i.e., Apron, Arrest 75W, Crown, or Captan, was examined under laboratory conditions using standard serial dilution and plate count techniques. The resulting effects of fungicide-Rhizobium interactions on nodulation, N2 fixation, and plant growth were assessed in a controlled environment. Fungicide treatment decreased the number of viable rhizobia on the seed. In general, the toxicity of the fungicides in terms of rhizobial viability increased in the following order: Control = Crown < Arrest = Apron < Captan. Although Crown had no effect on rhizobial viability assessed under laboratory conditions, it significantly reduced nodulation, percent N derived from the atmosphere (%Ndfa), and shoot dry matter. Seed treated with Arrest and Captan decreased nodule dry weight and %Ndfa, but only Arrest reduced dry matter yield. Apron had no effect on any of the parameters measured at the early pod-filling stage and was compatible with the chickpea inoculum used in this study.

Cicer↗

PubMed2003

High osmotic stress improves electro-transformation efficiency of fission yeast.

A preincubation of fission yeast cells with hyperosmotic solution improved the electro-transformation efficiency. The efficiency increased approximately five-fold when the cells were preincubated with 2.0 M sorbitol and 1.5 M NaCl at 30 degrees C for 60 min before an applied high electric pulse. Losses in the efficiency of the cells after hyperosmotic stress above 2.5 M sorbitol and 2.0 M NaCl were directly related to the marked reduction of viability. The efficiency at 2.0 M sorbitol gradually increased until 60 min of the preincubation period, but longer exposure resulted in a gradual decrease. On the other hand, when the cells of the osmotic-sensitive mutant were preincubated with isosmotic solution of 0.5 M sorbitol, the efficiency was also dramatically increased by approximately 15-fold. These improvements in efficiency were observed in sublethal conditions of osmotic stress regardless of osmoticums and strains.

Colony Count, Microbial↗

PubMed1999

Reducing bacterial resistance to antibiotics with ultrasound.

The effect of erythromycin on planktonic cultures of Psedomonas aeruginosa, with and without application of 70 kHz ultrasound, was studied. Ultrasound was applied at levels that had no inhibitory effect on cultures of Ps. aeruginosa. Ultrasound in combination with erythromycin reduced the viability of Ps. aeruginosa by 1-2 orders of magnitude compared with antibiotic alone, even at concentrations below the minimum inhibitory concentration (MIC). Electron-spin resonance studies suggest that ultrasound induces uptake of antibiotic by perturbing or stressing the membrane. This application of ultrasound may be useful for expanding the number of drugs available for treating localized infections by rendering bacteria susceptible to normally ineffective antibiotics.

Anti-Bacterial Agents↗

PubMed1997

A flow cytometric study of antibiotic-induced damage and evaluation as a rapid antibiotic susceptibility test for methicillin-resistant Staphylococcus aureus.

Flow cytometry using the anionic membrane potential-sensitive fluorescent probe, bis-(1,3-dibutylbarbituric acid) trimethine oxonol (DiBAC4(3)), enabled assessment of antibiotic-induced membrane perturbation in five clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and two antibiotic-sensitive reference strains, NCTC 6571 and 8325-4, after establishment of steady-state growth in liquid cultures inoculated from single colonies. Flow cytometric indications of the enhanced DiBAC4(3) uptake after treatment with vancomycin at 0.1, 1, 4 and 10 x MIC showed excellent comparison with viability losses quantified as cfu on solid agar in MRSA isolate QC. The antibiotic susceptibility patterns to benzylpenicillin, methicillin and vancomycin for all isolates used in this study could be determined in 2-4 h from an overnight plate culture. This technique thus provides a rapid and reproducible antibiotic sensitivity test which may be applicable in routine clinical practice.

Anti-Bacterial Agents↗

PubMed1994

Activity of eight antibacterial agents on Staphylococcus epidermidis attached to Teflon catheters.

The presence of a Teflon catheter had no effect on the in-vitro activity of a range of antibacterial agents against slime producing and non-producing Staphylococcus epidermidis strains as determined by a microdilution assay. The susceptibility of S. epidermidis attached to Teflon catheters for 6, 24 and 48 h was also evaluated. MICs for planktonic and attached bacteria were similar. When bacteria attached to Teflon for 6 h were used as inocula, MBC values increased 32-8192-fold for the antibacterial agents tested. Similar results were observed when bacteria attached for 24 and 48 h were used as inocula. The activity of a high concentration (16 x MBC) of these antimicrobial agents against S. epidermidis biofilms in Teflon catheters was evaluated; for five slime non-producing strains, the highest reduction (around 99%) in bacterial viability was produced by cloxacillin and teicoplanin; for the slime producers, the highest effect (99.5% reduction) was shown by amikacin, clindamycin cloxacillin and ciprofloxacin but all cases still showed bacterial counts higher than 10(3) cfu/catheter segment. It is concluded that adherence of S. epidermidis to Teflon catheters decreases the bactericidal activity of the antibacterial agents tested in vitro.

Anti-Bacterial Agents↗

PubMed1986

The in vitro antimicrobial effect of azelaic acid.

Various strains of cutaneous micro-organisms were tested in vitro for their survival rates in 0.5 mol/l (8.4% w/v) azelaic acid solution. All bacterial strains exhibited large reductions in viability (at least 40-fold) over a 24 h test period, but little response was noted with Pityrosporum ovale. The bactericidal effect of azelaic acid was reduced considerably in the presence of nutrients. Minimum inhibitory concentrations (MICs) and minimum bactericidal (or fungicidal) concentrations (MBCs) were also determined. MICs varied from 0.03 mol/l to 0.25 mol/l; MBCs were all either 0.25 mol/l or greater.

Acne Vulgaris↗

PubMed1991

Lansoprazole, a novel benzimidazole proton pump inhibitor, and its related compounds have selective activity against Helicobacter pylori.

The activities of various types of antiulcer agents against Helicobacter pylori (formerly called Campylobacter pylori) strains were determined by an agar dilution method. Among the compounds tested, two benzimidazole proton pump inhibitors, lansoprazole (AG-1749) and omeprazole, were found to have significant activities against this organism. The activity of lansoprazole was comparable to that of bismuth citrate, with MICs ranging from 3.13 to 12.5 micrograms/ml, and fourfold more potent than that of omeprazole. A major metabolite and two acid-converted rearrangement products of lansoprazole also exhibited good activities comparable or superior to that of the parent compound. Exposure to lansoprazole of H. pylori growing in a liquid medium led to an extensive loss of viability without a reduction in culture turbidity and produced an aberrant bacterial morphology characterized by the irregular constriction of cells and the collapse of cell surface structures. The antibacterial activity of lansoprazole and its related compounds was selective against H. pylori; common aerobic and anaerobic bacteria and Campylobacter jejuni were not inhibited by 100 micrograms/ml.

2-Pyridinylmethylsulfinylbenzimidazoles↗

PubMed1998

In vitro activities of co-amoxiclav at concentrations achieved in human serum against the resistant subpopulation of heteroresistant Staphylococcus aureus: a controlled study with vancomycin.

The effects of concentrations that simulated those in human serum after a single intravenous dose of amoxicillin (2 g), amoxicillin-clavulanic acid (2,000 and 200 mg, respectively), or vancomycin (500 mg), on the viability and beta-lactamase activity of two isogenic (beta-lactamase and non-beta-lactamase producer) heteroresistant Staphylococcus aureus strains were studied in an in vitro pharmacodynamic model. A reduction of > or = 97% of the initial inoculum was obtained with vancomycin and amoxicillin-clavulanic acid against both strains, with respect to the total bacterial population and the oxacillin-resistant subpopulation. The same pattern was observed with amoxicillin and the beta-lactamase-negative strain. beta-Lactamase activity in the beta-lactamase-positive strain changed over time parallel to viability, decreasing with amoxicillin-clavulanic acid or vancomycin and increasing in the amoxicillin and control groups. Clavulanic acid concentrations achievable in serum that changed over time allowed amoxicillin to act against the beta-lactamase-producing methicillin-resistant S. aureus to a similar extent as vancomycin.

Amoxicillin↗

PubMed1992

Simple method for measuring the antibiotic concentration required to kill adherent bacteria.

A simple method was developed for measuring the antibiotic susceptibility of bacteria adherent to plastic surfaces. Staphylococcus aureus cells adhered to the bottom of a 96-well plastic tissue culture plate were incubated in serially diluted antibiotic solutions. After a 24-hour-incubation the solutions were removed, and fresh medium without antibiotics was added to each well. The viability of the cells was judged by their growth after a further 24-hour-incubation. The antibiotic concentration required to kill adherent bacteria was far higher than that required for planktonic cells, although we used bactericidal drugs; beta-lactam, quinolone, and aminoglycoside antibiotics. The tolerance demonstrated by adherent cells is likely to play a role in the difficulties encountered in the chemotherapy of biofilm infections.

Anti-Bacterial Agents↗

PubMed1998

The effect of grain storage conditions on the viability of Fusarium and deoxynivalenol production in infested malting barley.

A continuing outbreak of Fusarium head blight occurred on barley in the upper Midwest from 1993 to 1995. This resulted in barley with levels of the mycotoxin deoxynivalenol (DON) reaching levels of concern for maltsters and brewers. This study evaluated the effect of 7 months of storage under different conditions (ambient outdoor temperature from October to April), -20 degrees C, 4 degrees C, 24 degrees C with quiescent air, and 24 degrees C with forced air) on the viability of Fusarium and Alternaria infesting stored grain. Additionally, the ability of Fusarium to produce DON after storage and during malting was evaluated. Initial levels of infestation of barley by Fusarium and Alternaria were 85 and 75%, respectively. All storage condition reduced the viability of both molds slightly and significantly for Fusarium. Forced air ventilation at 24 degrees C was the type of storage most effective in reducing the viability of Fusarium, dropping the percentage of infected kernels to 66%. DON levels did not change after 7 months with respect to storage conditions. However, DON levels were lower in malt produced from barley stored at 24 degrees C with or without aeration. On-farm storage of infected barley at elevated temperatures may provide a means to reduce the level of DON in finished malts.

Alternaria↗

PubMed2000

A new bioactive steroidal saponin, furcreastatin, from the plant Furcraea foetida.

Microbial and plant secondary metabolites were screened for compounds that are selectively cytotoxic to mutant p53-expressing mouse fibroblasts. As a result, furcreastatin, a novel steroidal saponin, was isolated from an EtOH extract of the leaves of Furcraea foetida. Furcreastatin consisted of hecogenin as the aglycone and a hexasaccharide containing D-galactose, L-rhamnose and four D-glucose residues. The structure was determined to be (3 beta,5 alpha,25R)- 3-hydroxyspirostan-12-one 3-O-[alpha-L-Rhap-(1-->4)-beta-D-Glcp-(1-->3)-¿beta-D-Glcp-(1-->3) -beta-D- Glcp-(1-->2)¿-beta-D-Glcp-(1-->4)-beta-D-Galp] by extensive NMR spectroscopic studies. Furcreastatin decreased the viability of mutant p53-over-expressing cells with an ED50 of 4.0 micrograms/mL, and decreased that of the parental cell-line with an ED50 of 9.6 micrograms/mL.

3T3 Cells↗

PubMed2006

In vitro antibacterial and cytotoxicity assay of multilayered polyelectrolyte-functionalized stainless steel.

Infection of implanted materials by bacteria constitutes one of the most serious complications following prosthetic and implant surgery. In the present study, a new strategy for confering stainless steel with antibacterial property via the alternate deposition of quaternized polyethylenimine (PEI) or quaternized polyethylenimine-silver complex and poly(acrylic acid) (PAA) was investigated. The success of the deposition of the polyelectrolyte multilayers (PEM) and its chemical nature was investigated by static water contact angle and X-ray photoelectron spectroscopy (XPS), respectively. The antibacterial activity was assessed using Escherichia coli (E. coli, a gram-negative bacterium) and Staphylococcus aureus (S. aureus, a gram-positive bacterium). The inhibition of E. coli and S aureus growth on the surface of functionalized films was clearly shown using the LIVE/DEAD Baclight bacterial viability kits and fluorescence microscopy. The cytotoxicity of the PEM to mammalian cells, evaluated by the MTT assay, was shown to be minimal and long-term antibacterial efficacy can be maintained. These results indicate new possibilities for the use of such easily built and functionalized architectures for the functionalization of surfaces of implanted medical devices.

Anti-Bacterial Agents↗

PubMed1993

General resistance to sterol biosynthesis inhibitors in Saccharomyces cerevisiae.

Screening for resistance to fenpropimorph was undertaken in order to isolate yeast mutants affected in the regulation of the ergosterol pathway. Among the mutants isolated, one bearing the recessive fen1-1 mutation was characterized by a 1.5-fold increase in the ergosterol level and a general resistance to sterol biosynthesis inhibitors. The fen1-1 mutation was linked to MAT locus on chromosome III. The measurement of enzyme activities involved in the ergosterol pathway revealed that isopentenyl diphosphate (IPP) isomerase activity was specifically increased 1.5-fold as compared to the wild type strain. However, overexpression of IPP isomerase in the wild type strain was not by itself sufficient to lead to sterol increase or resistance to sterol biosynthesis inhibitors, showing that IPP isomerase is not a limiting step in the pathway. The fen1-1 mutation permits viability in aerobiosis of yeast disrupted for sterol-14 reductase in absence of exogenous ergosterol supplementation, whereas the corresponding strain bearing the wild type FEN1 allele grows only in anaerobiosis. This result shows that ignosterol is able to efficiently replace ergosterol as bulk membrane component and that the fen1-1 mutation eliminates the specific ergosterol requirement in yeast.

Antifungal Agents↗

PubMed1992

Treatment of mycoplasma-contaminated continuous cell lines with mycoplasma removal agent (MRA).

Thirty-nine continuous adherent or suspension cell lines were treated with a quinolone antibiotic, Mycoplasma Removal Agent (MRA), for the elimination of chronic mycoplasma contamination. In preliminary experiments MRA did not show any cytostatic or cytotoxic effects on mycoplasma-free cell cultures in concentrations up to ten-fold the concentration used for mycoplasma eradication. Twenty-eight cell lines (72%) were effectively cleansed of the mycoplasma contaminants by MRA treatment. The persistent removal of the mycoplasma infection was monitored by three mycoplasma detection assays. In seven cell lines (18%) the mycoplasmas were resistant to treatment with MRA. The resistant species was mainly M. arginini followed by M. orale and A. laidlawii; however, other cell lines harboring these species were cured. Four cell lines (10%) which prior to treatment presented with decreased viability and poor or no cell growth were lost during or shortly after the exposure to the antibiotic. If an antibiotic elimination is attempted it is imperative to closely examine the effectiveness of treatment and possible eukaryotic cytotoxicity. The treated mycoplasma-free cells may also no longer express the original features as a result of treatment or the absence of mycoplasma.

Cell Survival↗

PubMed2002

Cytometric monitoring of growth, sporogenesis and spore cell sorting in Paenibacillus polymyxa (formerly Bacillus polymyxa).

AIMS: Formation of bacterial endospores is a basic process in Gram-positive bacteria and has implications for health, industry and the environment. Flow cytometry offers a practical alternative for the rapid detection, enumeration and characterization of bacterial endospores. METHODS AND RESULTS: Paenibacillus polymyxa was chosen for this study because its spores cause sporangium deformation and have thick walls with a star-shaped section. Sporulating populations were analysed with a particle analyser and a flow cytometer after labelling with propidium iodide and Syto-13. Flow cytometric detection of single spores was confirmed by optical and scanning electron microscopy after cell sorting. Four cell sub-populations were cytometrically detected in P. polymyxa cultures grown in liquid sporulation medium. Two sub-populations consisted of vegetative cells differing in both morphology and viability; the other two sub-populations consisted of spores differing in their viability. CONCLUSIONS: This work has shown that flow cytometry is a simple and fast method (less than 15 minutes for sample preparation and analysis) for the study of the sporulation in P. polymyxa. The use of this technique allowed both detection and quantification of sporulation inside a culture, and distinguished cells that differed in viability despite being morphologically identical under microscopic observation. SIGNIFICANCE AND IMPACT OF THE STUDY: Flow cytometry has been proved to be a valuable tool for the analysis of sporulation in P. polymyxa cultures, with the unique capacity of distinguishing between endospores and vegetative cells, and between live and dead cells, in the same analysis. An important percentage of non-viable endospores has been found in aged cultures using this method.

Bacillus↗

PubMed2005

The mycotoxins citrinin and gliotoxin differentially affect production of the pro-inflammatory cytokines tumour necrosis factor-alpha and interleukin-6, and the anti-inflammatory cytokine interleukin-10.

BACKGROUND: Microbial growth is considered one of the major causes of indoor air problems. Moulds have been associated with asthma, allergy and a wide range of diffuse indoor air-related symptoms. However, mechanisms of the adverse health effects are not well understood. OBJECTIVE: We hypothesized that the mycotoxins citrinin and gliotoxin could cause an imbalance between the secretion of the pro-inflammatory cytokines TNF-alpha and IL-6 and the anti-inflammatory cytokine IL-10. METHODS: We investigated the influence of citrinin and gliotoxin on the human monocytic cell line Mono-Mac-6 (MM6) with and without lipopolysaccharide -stimulation. The levels of IL-10, IL-6 and TNF-alpha were analysed in cell culture supernatants by ELISA. Cell viability and cell apoptosis were measured by flow cytometry. RESULTS: The strongest inhibition of cytokine secretion was found for IL-10. IL-6 levels were found to decrease in a dose-dependent manner along with reduced cell viability. TNF-alpha levels increased with low gliotoxin exposure (less than 100 ng/mL), but decreased significantly at 375 ng/mL and higher along with increased cell apoptosis and reduced cell viability. TNF-alpha levels were not reduced by citrinin exposure. CONCLUSION: We observed a cytokine imbalance with a more pronounced reduction of IL-10 concentrations compared with those of TNF-alpha and IL-6. We suggest that low exposure doses of citrinin and gliotoxin (corresponding to less than 100 ng/mL gliotoxin and less than 10 mug/mL citrinin) may inhibit IL-10 and lead to increased risk of an inflammatory response with relative overproduction of TNF-alpha and IL-6. The findings and their clinical implications must be verified by human studies. However, we speculate that the observed biological effects may be of importance as they may partly explain the occurrence of diffuse general indoor air-related symptoms as well as the worsening of asthmatic inflammatory reactions experienced in mouldy environments.

Air Pollution, Indoor↗

PubMed2006

Assessment of DNA damage and repair in Mycobacterium terrae after exposure to UV irradiation.

AIM: Ultraviolet (UV) irradiation for drinking water treatment was examined for inactivation and subsequent dark and photo-repair of Mycobacterium terrae. METHODS AND RESULTS: UV sources tested were low pressure (monochromatic, 254 nm) and medium pressure (polychromatic UV output) Hg lamps. UV exposure resulted in inactivation, and was followed by dark or photo-repair experiments. Inactivation and repair were quantified utilizing a molecular-based endonuclease sensitive site (ESS) assay and conventional colony forming unit (CFU) viability assay. Mycobacterium terrae was more resistant to UV disinfection compared to many other bacteria, with approximately 2-log reduction at a UV fluence of 10 mJ cm(-2) ; similar to UV inactivation of M. tuberculosis. There was no difference in inactivation between monochromatic or polychromatic UV lamps. Mycobacterium terrae did not undergo detectable dark repair. Photo-repair resulted in recovery from inactivation by approximately 0.5-log in less than 30 min for both UV lamp systems. CONCLUSIONS: Mycobacterium terrae is able to photo-repair DNA damage within a short timeframe. The number of pyrimidine dimers induced by UV light were similar for Escherichia coli and M. terrae, however, this similarity did not hold true for viability results. SIGNIFICANCE AND IMPACT OF THE STUDY: There is no practical difference between UV sources for disinfection or prevention of DNA repair for M. terrae. The capability of M. terrae to photo-repair UV damage fairly quickly is important for wastewater treatment applications where disinfected effluent is exposed to sunlight. Finally, molecular based assay results should be evaluated with respect to differences in the nucleic acid content of the test micro-organism.

Colony Count, Microbial↗

PubMed1999

Exposure to antibiotics induces expression of the Mycobacterium tuberculosis sigF gene: implications for chemotherapy against mycobacterial persistors.

The sigF gene encodes an alternate sigma factor found in Mycobacterium tuberculosis and related pathogenic mycobacteria. Determination of conditions of sigF expression is an important step in understanding the conditional gene regulation which may govern such processes as virulence and dormancy in mycobacteria. We constructed an in-frame translational lacZ-kan fusion within the sigF gene to determine the conditions of sigF expression. This reporter construct was expressed from a multicopy plasmid in a strain of BCG harboring an integrated luciferase reporter gene under the control of the mycobacteriophage L5 gp71 promoter. Antibiotic exposure, in particular, ethambutol, rifampin, streptomycin, and cycloserine treatment, increased the level of SigF reporter specific expression in a dose-dependent fashion. The level of SigF reporter specific expression increased over 100-fold in late-stationary-phase growth compared to that in exponential growth. During the exponential phase, SigF specific expression could be induced by a number of other stresses. Anaerobic metabolism induced SigF by greater than 150-fold, particularly in the presence of metronidazole. Cold shock increased the level of SigF specific expression, while heat shock decreased it. Oxidative stress was also an important inducer of SigF specific expression; a greater induction was seen with cumene hydroperoxide than with hydrogen peroxide. Comparisons of bacterial viability as determined by the luciferase assay or by plating serial dilutions revealed that luciferase gp71-dependent activity was an unreliable predictor of the numbers of CFU during stationary-phase growth and anaerobic metabolism. The induction of sigF following antibiotic exposure suggests that this bacterial transcription factor may control genes which are important for mycobacterial persistence in the host during chemotherapy.

Anaerobiosis↗