Inhibition of Helicobacter pylori by garlic extract (Allium sativum)

Abstract The antibacterial effect of aqueous garlic extract (AGE) was investigated against Helicobacter pylori . Sixteen clinical isolates and three reference strains of H. pylori were studied. Two different varieties of garlic were used. The concentration of AGE required to inhibit the bacterial growth was between 2–5 mg ml⁻¹. The concentration, for both AGE types, to inhibit 90% (MIC₉₀) of isolates was 5 mg ml⁻¹. The minimum bactericidal concentration (MBC) was usually equal to, or two-fold higher than, minimum inhibitory concentration (MIC). Heat treatment of extracts reduced the inhibitory or bactericidal activity against H. pylori ; the boiled garlic extract showed a loss of efficacy from two-to four-fold the values of MIC and the MBC obtained with fresh AGR. The antibacterial activity of garlic was also studied after combination with a proton pump-inhibitor (omeprazole) in a ratio of 250:1. A synergistic effect was found in 47% of strains studied; an antagonistic effect was not observed.     

Effect of various gas atmospheres on the growth and survival of Campylobacter jejuni on beef

Pieces of fresh beef were inoculated with three strains of Campylobacter jejuni . The meat was then allocated to three treatments: (a) vacuum packaged, (b) packaged in an atmosphere of 20% CO₂+ 80% N₂, and (c) packaged into sterile Petri dishes in anaerobic cultivation boxes, which were filled with a gas mixture of 5% O₂+ 10% CO₂+ 85% N₂. The packaging material in the first two treatments was PA 80/PE 100–PE 100/PA 80/PE 100. The survival of Campylobacter cells was followed at 37°C, 20°C and 4°C for 48 h, 4 days and 25 days, respectively. At 37°C the counts of two Campylobacter strains increased in each package treatment for 48 h. At 20°C and at 4°C the counts of the same two strains decreased by 1 to 2 log units and 0.5 to 1 log unit, respectively, during storage. The survival of the two strains was about the same in all package treatments. The third strain was the most sensitive of the strains studied. At 37°C its numbers increased only in the optimal gas atmosphere; at 20°C the strain was not detectable after 24 to 48 h storage and at 4°C after 4 days storage. The aerobic plate counts were determined for all samples at the same time as Campylobacter counts. The high indigenous bacterial numbers of the meat samples did not appear to have a great effect on the survival or growth of campylobacters.     

Antibacterial effect of protamine assayed by impedimetry

Impedimetric measurements were used to assay the antibacterial effect of protamine. A good linear correlation between the impedance detection time and the initial cell counts was obtained ( r = 0.99, n = 2). As basic peptides may cause clumping of cells, this correlation curve was used when estimating the cell number after protamine treatment, rather than colony counts.
Protamine from salmon killed growing Gram-positive bacteria and significantly inhibited growth of Gram-negative bacteria in Tryptone Soy Broth (TSB) at 25°C. In general Gram-positive bacteria were more sensitive to protamine than Gram-negative bacteria; the minimum inhibitory concentrations (MIC) determined for Gram-positive strains varied from 20 to 1000 μ ml^-1 and for Gram-negative strains from 500 μ ml^-1 to more than 4000 μ ml^-1.
The effect of protamine on non-growing Listeria monocytogenes Scott A suspended in buffer was not lethal as was the effect on growing cells; however, protamine (50–500 μg ml^-1) killed the Gram-negative fish spoilage bacteria Shewanella putrefaciens when the live cells were suspended in buffer.     

Antibiotic Susceptibility of Helicobacter pylori Clinical Isolates: Comparative Evaluation of Disk-Diffusion and E-Test Methods

Antimicrobial susceptibility of 25 Helicobacter pylori strains isolated from patients with acid peptic diseases were tested for in vitro sensitivity to commonly used antibiotics using disk-diffusion and E-test, methods. All strains tested were susceptible to tetracycline by E-test, with the minimum inhibitory concentration (MIC) values being <0.125 μg/ml for all strains except for 6 (<0.023 μg/ml). However 1 strain was resistant by disk-diffusion method. One strain was resistant to clarithromycin both by disk diffusion and E-test (MIC <48 μg/ml), and 1 strain was resistant only by disk diffusion. Only one strain was resistant to amoxicillin by disk diffusion and E-test (MIC >256 μg/ml). For ciprofloxacin, three strains were resistant by disk diffusion and two by E-test (MIC <32 μg/ml). Sixteen strains were resistant to metronidazole by disk diffusion and E-test (MIC ≥ 8 μg/ml), and 1 was resistant only by E-test (MIC <48 μg/ml). Overall, 64% of the strains were resistant to metronidazole. The MIC for metronidazole was also tested by agar-dilution method, and metronidazole resistant strains had an, MIC >8 μg/ml. The disk-diffusion method showed excellent correlation with E-test results; there was 100% agreement for amoxicillin a other antibiotics showed 90% to 95% accuracy. Disk diffusion is cheaper than E-test (approximately 2.6 cents vs. US$2.60), is easy to perform, and is a reliable method for testing H. pylori susceptibility to antimicrobial agents in the clinical microbiology laboratory.     

A rapid method for the determination of bacterial fatty acid composition

Heat treatment of spores of non-proteolytic strains of Clostridium botulinum at 75–90°C, and enumeration of survivors on a nutrient medium containing lysozyme gave biphasic survival curves. A majority of spores were inactivated rapidly by heating, and the apparent heat-resistance of these spores was similar to that observed by enumeration on medium without lysozyme. A minority of spores showed much greater heat-resistance, due to the fact that the spore coat was permeable to lysozyme, which diffused into the spore from the medium and replaced the heat-inactivated germination system. The proportion of heated spores permeable to lysozyme was between 0.2 and 1.4% for spores of strains 17B (type B) and Beluga (type E), but was about 20% for spores of strain Foster B96 (type E). After treatment of heated spores with alkaline thioglycolate, all were permeable to lysozyme. D-values for heated spores that were permeable to lysozyme (naturally and after treatment with thioglycolate) were: for strain 17B, D₈₅°C, 100 min; D₉₀°C, 18.7 min; D₉₅°C, 4.4 min; for strain Beluga, D₈₅°C, 46 min; D₉₀°C, 11.8 min; D₉₅°C, 2.8 min. The z-values for these spores of strains 17B and Beluga were 7.6°C and 8.3°C.     

Inactivation of oenological lactic acid bacteria (Lactobacillus hilgardii and Pediococcus pentosaceus) by wine phenolic compounds

Aims:  To investigate the inactivation properties of different classes of phenolic compounds present in wine against two wine isolates of Lactobacillus hilgardii and Pediococcus pentosaceus , and to explore their inactivation mechanism.
Methods and Results:  After a first screening of the inactivation potency of 21 phenolic compounds (hydroxybenzoic and hydroxycinnamic acids, phenolic alcohols, stilbenes, flavan-3-ols and flavonols) at specific concentrations, the survival parameters (MIC and MBC) of the most active compounds were determined. For the L. hilgardii strain, the flavonols morin and kaempferol showed the strongest inactivation (MIC values of one and 5 mg l⁻¹, and MBC values of 7·5 and 50 mg l⁻¹, respectively). For the P. pentosaceus strain, flavonols also showed the strongest inactivation effects, with MIC values between one and 10 mg l⁻¹ and MBC values between 7·5 and 300 mg l⁻¹. Observations by epifluorescence and scanning electron microscopy revealed that the phenolics damaged the cell membrane and promoted the subsequent release of the cytoplasm material into the medium.
Conclusions:  The antibacterial activity of wine phenolics against L. hilgardii and P. pentosaceus was dependent on the phenolic compound tested, and led not only to bacteria inactivation, but also to the cell death.
Significance and Impact of the Study:  New information about the inactivation properties of wine lactic acid bacteria by phenolic compounds is presented. It opens up a new area of study for selecting/obtaining wine phenolic preparations with potential applications as a natural alternative to SO₂ in winemaking.     

Physiological differences between strains of Tribolium castaneum selected for resistance to hypoxia and hypercarbia, and the unselected strain

Abstract. The metabolic rates, as expressed by oxygen (O₂) consumption, carbon dioxide (CO₂) production, and losses in wet and dry weights, were examined for adults of three strains of the red flour beetle Tribolium castaneum (Herbst), during exposure to two modified atmospheres (MAs). Exposure of a strain selected for resistance over twenty-one generations to an atmosphere of 65% CO₂, 20% O₂ and the balance nitrogen (N₂), termed a high carbon dioxide concentration atmosphere (HCC) and exposure of an unselected strain to HCC, showed considerable levels of aerobic metabolism during exposure. For the unselected strain water loss and mobilization of energy reserves were rapid and mortality was followed by rapid desiccation. For the HCC-resistant strain water balance was maintained and energy reserves were utilized more slowly over a prolonged period. Exposure of a strain selected for resistance over twenty-one generations to a low oxygen concentration atmosphere (LOC) of 0.5% O₂ in N₂, and an unselected strain to LOC, revealed that even at 0.5% O₂, metabolism was largely aerobic in both strains. Maintenance of water balance was not a major factor causing mortality of either strain during exposure to LOC. In air, metabolic rates of both the resistant strains were lower than that of the unselected strain.     

Inhibition of methanogenesis by several heavy metals using pure cultures

The effect of different concentrations of nickel, copper and zinc on methanogenesis using pure cultures of Methanobacterium formicicum, Methanobrevibacter arboriphilicus, Methanosarcina thermophila and Methanospirillum hungatei over time (1, 15 and 30 d) was evaluated. methanobacterium formicicum showed the highest resistance to all the metals tested, while Methanospirillum hungatei was the most sensitive strain. All strains were sensitive to copper and zinc (10–250 mg 1^-1, but were much more resistant to nickel (200–1200 mg 1^-1). An adaptation process of the methanogenic pure culture with the toxicants was observed over time, which indicates that the inhibitory effects of heavy metals may be reverted in optimal anaerobic conditions.     

Minimum inhibitory concentrations of intestinal Escherichia coli from broiler chickens after oral administration of apramycin

This study describes the influence of apramycin on minimum inhibitory concentrations (MIC) of intestinal Escherichia coli in young broiler chickens, after oral administration of the antibiotic at a dosage equivalent to a prophylactic course of treatment for 10 d. The bacteria were isolated from cloacal swabs and caecal contents. MICs were determined by agar dilution procedures. MIC of apramycin for the investigated strains ranged from 1 μg ml^-1 to 16 μg ml^-1. Strains obtained from undosed birds mainly had MIC values of 1 μg ml^-1. MIC values of 8 μg ml^-1 or more were recorded only among isolates obtained from chickens which had received apramycin. Administration of apramycin resulted in a slight but statistically significant increase in the average MIC. Statistically higher average MICs were recorded among isolates from cloacal swabs 10 d after withdrawal until the end of the experiment. For strains from caecal contents, this was demonstrated only on one sampling occasion, 15 d after withdrawal.     

Some factors affecting airborne survival of Pseudomonas fluorescens indoors

The effect of relative humidity (RH) on the airborne survival of a Pseudomonas fluorescens strain was studied at 20, 40, 60 and 80% RH indoors. The aero-stable spore of Bacillus subtilis subsp. niger , used as a tracer of physical losses was compared with a light scattering particle counter, as there were doubts about the reliability of the spore as tracer. The Rion counter was validated before use and found to give a good estimate of relative physical losses providing spray suspensions contained between 10⁷ and 10⁹ cfu ml⁻¹ cells and that the humidity was not more than 80% RH. Pseudomonas fluorescens , strain P⁺ S⁺, suspended in distilled water survived best at mid humidities and least at 80% RH. When suspended in 1% glycerol there was an apparent 'increase' in viability after an initial rapid reduction. This was thought to be due to delay in equilibration of glycerol in the cell membrane after concentration on dehydration. The cells were thought to be unstable and sensitive to the stress of rehydration before equilibration occurred. The findings are discussed in relation to Cox's theories of outer membrane damage on aerosolization.     

Col V plasmids and the resistance of Escherichia coli to divalent copper ions

Free organisms of both plasmid-free and plasmid-carrying strains of Escherichia coli were killed by incubation in water containing low levels of cupric ions. Sensitivity was temperature-dependent with killing being more marked at 20° or 25°C than at 10° or 15°C. In contrast to the effects of other inhibitors from natural waters (which affect free Col V⁺organisms more than Col^-ones), free Col^-and Col V⁺organisms were equally sensitive to kill by Cu²⁺. Attachment to glass beads essentially abolished sensitivity to cupric ions with full survival after exposure to 15 μ g/ml. This applied to both p⁺and p^-strains but attachment would have more effect on the survival of p⁺organisms in natural waters because some plasmids markedly enhance attachment.     

The mode of antistaphylococcal action of Eleutherine americana

The anti-methicillin-resistant Staphylococcus aureus (anti-MRSA) activity and the possible mechanism of action of a crude extract from red bulbs of Eleutherine americana Merr. were investigated. The crude ethanolic extract from E. americana produced minimum inhibitory concentrations (MICs) of 62.5–1000 and 250 μg mL⁻¹ against MRSA isolates and the reference strains, respectively. Treatment of S. aureus ATCC 27664 with a crude extract at 2MIC reduced the inoculum size by 5 log at 24 h compared with the control. The combined effect of the extract and 7.5% NaCl on the enterotoxin-producing ATCC strain resulted in no detection of organisms within 24 h compared with the control. The release of cell materials after extract treatment was determined by measuring OD_260 nm, the treatment resulted in cytoplasmic leakage. Determination of OD_620 nm showed that the extract did not cause gross cell wall damage. However, observation of S. aureus cells under an electron microscope after treatment with 2MIC and 4MIC of the crude extract revealed that the extract caused damage to membrane morphology. A knowledge of the mechanism of action of the E. americana extract may offer useful hints in the search for novel antibacterial substance.     

Drought tolerance by lentil rhizobia (Rhizobium leguminosarum) from arid and semiarid areas of Pakistan

Ten strains of lentil rhizobia (Rhizobium leguminosarum ) were evaluated for drought tolerance by exposing them to soil moisture potentials of −0·03, −1·0 and −1·5 MPa. Water availability, rhizobial strain and time of exposure to drought had a significant ( P ≤ 0·001) effect on the number of surviving rhizobia g⁻¹ of soil. Highest cell counts were observed at −0·03 MPa, followed by soil maintained at −1·0 and −1·5 MPa. Five strains originating from saline areas showed significantly ( P ≤ 0·05) better survival under low water potential after 35 days. Two strains exhibited greatest survival under low water potential and produced viable cell counts of more than 10⁷ rhizobia g⁻¹ of soil. These strains could probably be used successfully as inoculants for lentil production in arid and semi-arid environments.     

The interaction between diepoxybutane and ultraviolet light in yeast II. Experiments with UV-sensitive and resistant strains

Earlier experiments in which the effects of diepoxybutane (DEB) pretreatment on UV inactivation were studied in wild-type yeast, have been extended to include DEB pretreatment effects in the strain uvs- which is sensitive to both UV and DEB. The effects of post-UV treatment with DEB have also been studied in both strains.

DEB pretreatment produces a similar result in uvs- as the effect described earlier for wild-type in spite of the necessary reduction in doses of both agents. Post-treatment with DEB fails to affect photoreactivation in both resistant and sensitive strains but in the resistant strain the shoulder on the UV curve is obliterated. There is no such effect in the UV-sensitive material.     

Effect of protective solutes on leakage from and survival of immobilized Lactobacillus subjected to drying, storage and rehydration

When lactic acid bacteria are used industrially as fermentation starters it is important to obtain stable and highly viable bacterial cultures. Six strains of Lactobacillus encapsulated in Ca-alginate gel beads were investigated to determine whether dehydration, storage and rehydration may inflict injury. A negative relationship between leakage of lactate dehydrogenase and survival rates was found. Mesophilic lactobacilli showed only negligible leakage compared with thermophilic strains when dehydrated at 30 °C to a level of 0·11 g H₂0 (g dry wt)⁻¹. The choice of an appropriate suspending medium to be introduced before drying was therefore very important for thermophilic lactobacilli in order to increase the survival rates during dehydration, storage and rehydration. The osmoregulatory solutes tested were adonitol, betaine, glycerol and reconstituted non-fat milk solids (NFMS). Less injury was inflected during dehydration for Lactobacillus helveticus with adonitol, glycerol and NFMS. Survival rates for the strains subjected to immobilization, dehydration, storage and rehydration varied with the strain and the protective solute when fluidized-bed drying was used at 5 °C to a level as high as 0·34 g H₂0 (g dry wt)⁻¹. Non-fat milk solids gave the best protection for thermophilic lactobacilli, while adonitol and NFMS were best for mesophilic lactobacilli.     

In vitro susceptibility of Clostridium perfringens isolated from farm animals to growth-enhancing antibiotics

Minimal inhibitory concentration (MIC) determinations were carried out with seven growth-enhancing antibiotics against 95 Clostridium perfringens field isolates obtained during 1991 and 1992 from poultry, pigs and calves. All were resistant to 64 μg ml⁻¹ of the bambermycin antibiotic, flavomycin (flavophospholipol) and susceptible to avoparcin (MIC₉₀ 0.25 μg ml⁻¹), avilamycin (MIC₉₀ 0.5 μg ml⁻¹) and salinomycin (MIC₉₀≤ 0.12 μg ml⁻¹). Acquired resistance against bacitracin was detected in some isolates from poultry and bovines and resistance to tylosin and virginiamycin in some strains from all species investigated. Overall, the prevalence of resistance was comparable to the low levels recorded in 1979 in Cl. perfringens isolates from the same animal host species.     

Genetic factors in Rhizobium affecting the symbiotic carbon costs of N2 fixation and host plant biomass production

The effect of genetic factors in Rhizobium on host plant biomass production and on the carbon costs of N₂ fixation in pea root nodules was studied. Nine strains of Rhizobium leguminosarum were constructed, each containing one of three symbiotic plasmids in combination with one of three different genomic backgrounds. The resulting strains were tested in symbiosis with plants of Pisum sativum using a flow-through apparatus in which nodule nitrogenase activity and respiration were measured simultaneously under steady state conditions. Nodules formed by strains containing the background of JI6015 had the lowest carbon costs of N₂ fixation (7.10–8.10 μmol C/μmol N₂), but shoot dry weight of those plants was also smaller than that of plants nodulated by strains with the background of B151 or JI8400. Nodules formed by these two strain types had carbon costs of N₂ fixation varying between 11.26 and 13.95 μmol C/μmol N₂. The effect of symbiotic plasmids on the carbon costs was relatively small. A time-course experiment demonstrated that nodules formed by a strain derived from JI6015 were delayed in the onset of nitrogenase activity and had a lower rate of activity compared to nodules induced by a strain with the background of B151. The relationship between nitrogenase activity, carbon costs of N₂ fixation and host plant biomass production is discussed.     

Chemotherapeutic sensitivity and viability of Nocardia asteroides and Nocardia brasiliensis in drugs

Procedures for testing drug sensitivity ofNocardia were standardized.Five strains each ofN. asteroides (NA) andN. brasiliensis (NB) isolated from pathogenic materials were tested for drug sensitivity in Sabouraud's glucose agar (SGA), Sabouraud's glucose broth (SGB) and SGB with addition of 10 % horse serum against sulphadiazine (S), penicillin (P), streptomycin (St), chloramphenicol (C), tetracycline (T) and 4-4-diamino diphenyl sulphone (DDS). The viability of strains at minimum inhibitory concentrations (MIC) of different drugs were also tested.Results showed that liquid medium is preferable to solid medium for drug testing as pellicle formation observed on the surface of liquid medium helps in accurate assessment of MIC. Serum addition did not appear to be necessary. Ten days incubation at 37 °C gives optimum results for determination of MIC. Taking the maximum drug concentration attainable in blood during therapy, the strains were classified as sensitive and resistant. All strains were resistant to DDS and P. One strain of NB was sensitive to St. One strain each of NA and NB was borderline sensitive to C. Four out of 5 strains in both species were sensitive to S. All strains were sensitive to T. The organisms remained viable in drugs, thus suggesting the limitations of antinocardial therapy.From Mycology Research Unit, Department of Dermatology, Seth G.S. Medical College and K.E.M. Hospital, Bombay-12.Hon. Professor Dermato-Venereology.Research Fellow.     

Sensitivity of Bacillus coagulans spores to combinations of high hydrostatic pressure, heat, acidity and nisin

We investigated the combined effects of pressure, temperature, pH, initial spore concentration and the presence of nisin on the survival of spores of Bacillus coagulans. Spores were more sensitive to pressure both at lower pH and at higher treatment temperatures. An additional 1.5-log₁₀ reduction in cfu ml^-1 was observed when pH was lowered from 7.0 to 4.0 during pressurization at 400 Mpa and 45°C. A 4-log₁₀ cfu ml^-1 reduction was observed when the temperature was increased from 25°C to 70°C during pressurization at 400 Mpa. The spores were sensitive to nisin at concentrations as low as 0.2 IU ml^-1. At least a 6-log₁₀ reduction was generally achieved with pressurization at 400 Mpa in pH 4.0 buffer at 70°C for 30 min when plated in nutrient agar containing 0.8 IU ml^-1 nisin.     

Chronic toxicity of ammonia to juvenile gilthead seabream Sparus aurata and related histopathological effects

Effect of 20 days exposure of juvenile gilthead seabream ( Sparus aurata ) to elevated levels of ammonia on growth and survival was examined in a continuous flow system. Suppressed growth and reduced survival were observed at concentrations of 8.2 and 13 mg l⁻¹ total ammonia-N (0.5 and 0.7 mg l⁻¹ un-ionized ammonia-N, respectively) and higher. The maximum acceptable toxic concentration (MATC) for growth was between 4.8 to 8.2 mg l⁻¹ total ammonia-N (0.3 and 0.5 mg l⁻¹ un-ionized ammonia-N, respectively). Fish exposed to high ammonia levels (13 mg l⁻¹ total ammonia-N, 0.7 mg l⁻¹ un-ionized ammonia-N) displayed clear signs of liver pathology. Existing evidence suggests that S. aurata is less sensitive to ammonia than other reported marine and freshwater fish.
Under certain conditions ammonia concentration in the intensive fish ponds in Eilat may exceed the no observed effect concentration for S. aurata .