Leakage Of Cellular Materials Research Articles

Susceptibility of dairy associated molds towards microbial metabolites with focus on the response to diacetyl

Food spoilage may cause food waste, economic losses and even brand image deterioration. In fermented dairy products, spoilage by molds is frequently occurring and the molds are commonly being introduced at the consumer stage after opening. Diverse molds may be involved with the main genera being Penicillium, Mucor, and Cladosporium. Fungal growth may be inhibited by bioprotective cultures by competition and/or production of inhibitive compounds. Several compounds produced by a range of lactic acid bacteria have been reported as having an antifungal activity against certain fungi. Susceptibility tests of a panel of dairy associated molds towards twelve of these pure compounds were performed using microtiter plate well method. Diacetyl was the most potent antifungal among the compounds followed by octanoic acid. Large variations were seen within the Penicillium strains tested with P. commune being more sensitive than P. roquefortii, and two Mucor strains generally exhibiting high tolerance. The growth of two susceptible Penicillium strains in yoghurt serum with pH 4.6 with increasing amounts of diacetyl added were followed. Susceptibility varied but at 64 μg/mL, both strains were completely inhibited on yoghurt regardless of temperature. At higher pH (5.6) in malt extract agar (MEA), growth was faster but inhibition was still observed. Upon exposure to diacetyl, the level of cytoplasmic reactive oxygen species (ROS) increased and impairment of membrane integrity and leakage of cellular materials were observed. These findings provide new knowledge of the activity of diacetyl against molds.

Mode of action of plectasin-derived peptides against gas gangrene-associated Clostridium perfringens type A.

NZ2114 and MP1102 are novel plectasin-derived peptides with potent activity against Gram-positive bacteria. The antibacterial characteristics and mechanism of NZ2114 and MP1102 against gas gangrene-associated Clostridium perfringens were studied for the first time. The minimal inhibitory concentration and minimal bactericidal concentration of NZ2114 and MP1102 against resistant C. perfringens type A strain CVCC 46 were 0.91 μM. Based on the fractional inhibitory concentration index (FICI) result, an additive or synergic effect was observed between NZ2114 (FICI = 0.5~0.75) or MP1102 (FICI = 0.375~1.0) and antibiotics. The flow cytometry, scanning and transmission electron microscopy analysis showed that both NZ2114 and MP1102 induced obviously membrane damage, such as the leakage of cellular materials, partial disappearance of the cell membrane and membrane peeling, as well as retracting cytoplasm and ghost cell. The gel retardation and circular dichroism (CD) detection showed that NZ2114 and MP1102 could bind to C. perfringens genomic DNA and change the DNA conformation. Moreover, NZ2114 also interfered with the double helix and unwind the genomic DNA. The cell cycle analysis showed that C. perfringens CVCC 46 cells exposed to NZ2114 and MP1102 were arrested at the phase I. These data indicated that both NZ2114 and MP1102 have potential as new antimicrobial agents for gas gangrene infection resulting from resistant C. perfringens.

Anti-biofilm activity of the metabolites of Streptomyces chrestomyceticus strain ADP4 against Candida albicans

Candida albicans is a commensal but a significant opportunistic pathogen. It forms biofilms, which protect them against anti-Candida compounds. Therefore, an agent capable of disrupting the Candida biofilms will be useful in the treatment of such infections. The metabolites of Streptomyces chrestomyceticus strain ADP4 displayed strong anti-Candida activity, hence were investigated further for their ability to inhibit biofilm. Strong inhibition of biofilms produced by several reference strains of C. albicans was observed with BIC90 values ranging from 4 to 8μg/mL. The anti-biofilm activity of ADP4 metabolites appeared to involve membrane disruption and leakage of cellular materials. Also, it effectively inhibited Candida cells from adhering to polystyrene surface and inhibited their conversion to the hyphal state, thereby preventing further development of the biofilm by the adherent cells. This is the first such report on the metabolites produced by any strain of S.chrestomyceticus.

Effect of Thermal and Radio Frequency Electric Fields Treatments on Escherichia coli bacteria in Apple Juice

The need for a nonthermal intervention technology that can achieve microbial safety without altering nutritional quality of liquid foods led to the development of the radio frequency electric fields (RFEF) process. However, insight into the mechanism of bacterial inactivation by this technology is limited. In this study, we investigated membrane damage of Escherichia coli bacterial (7.8 log CFU/ml) and leakage of intracellular membrane materials in RFEF treated apple juice at 25 kV/cm and operated at 25°C, 55°C and 75°C for 3.4 milliseconds at a flow rate of 540 ml/min. Damage to cell membrane was detected with Transmission Electron Microscopy (TEM) and leakage of cellular materials was determined with ATP luminometer (20 D) and electrostatic and hydrophobic interaction chromatography used to characterize changes in bacterial cell surfaces. RFEF treatment caused a significant decrease in bacterial cell surface hydrophobicity and loss of relative negative ions compared to heat treatment alone at 55°C and 75°C. Leakage of cellular materials into the media indicated cell damage and TEM observation showed altered intracellular membrane structure in RFEF treated E. coli cells. The results of this study suggest that the mechanism of inactivation of RFEF is by disruption of the bacterial cell surface hydrophobicity and loss of relative negative ions which led to injury and leakage of cellular materials and death.

Responses of Redroot Pigweed (Amaranthus retroflexus) and Witchgrass (Panicum capillare) Seeds to Anesthetics

Substances having anesthetic-like properties were examined for stimulation of redroot pigweed and witchgrass seed germination. Chemicals included the n-alcohols through C-5, 2-propanol, benzyl alcohol, diethyl ether, chloroform, and 2-bromo-2-chloro-1,1,1-trifluoroethanol. Redroot pigweed seeds required the far-red absorbing form of phytochrome (Pfr) for an anesthetic substance to increase germination, but in witchgrass seeds the active anesthetic substances stimulated germination without added Pfr. The correlation coefficient comparing relative activity (RA) versus the membrane/buffer partition coefficient (M/B) of the active substances was very poor (−0.37) for pigweed seeds but was −0.91 for witchgrass. Leakage of cellular materials, including electrolytes, substances absorbing at 280 nm, and amino acids from seeds treated with either active or inactive anesthetics, was not particularly indicative of effects on germination in both species. Application of increased pressure (0.965 MPa) during anesthetic treatment, which in animals prevents anesthesia, was found to act oppositely and increase seed response to ethanol in redroot pigweed seeds, but in witchgrass seeds pressure suppressed the stimulatory action of ethanol. In both species, pressure appeared to have an action of its own, but for this action to be expressed the presence of a stimulatory anesthetic was also required. The results do not appear to support a good relationship of anesthetic activity with seed membrane lipophilic components in redroot pigweed seeds, but the relationship seems likely for witchgrass seeds.

Leakage of Cellular Materials and Damage to the Cellular Surface of Escherichia coli by Glycine and/or Sodium Chloride

The combined antimicrobial effect of glycine and sodium chloride against Escherichia coli IFO3301 was examined in terms of membrane damage. The cellular materials, protein, phospholipid, lipopolysaccharide and 260nm-absorbing materials, leaked from the cells treated with glycine proportionally to the glycine concentration. The leakage was suppressed by the addition of sodium chloride. On the other hand, the largest leakage was observed when the cells grown in 2% sodium chloride-containing medium were treated with glycine. Consequently, the antimicrobial activity of glycine and sodium chloride was not enhanced in the case of combined treatment, but that of glycine was enhanced by the change of the bacterial surface caused by prior sodium chloride treatment during cell growth.The observations by electron microscopy also supported the above results. The membrane of the cells treated with sodium chloride was intact, although a remarkable plasmolysis was observed. On treatment of the cells grown in sodium chloride-containing medium with glycine, extensive blebbing of the membrane and extracellular vesicles was found. The results reflect the injury to the membrane.

Effect of ethylene on the permeability of excised cantaloupe fruit tissue.

C(hanges in tisstue permeability are associated with the matutration and ripening of fruits. Blackman and Parija (3) suggested that permeability changes may cause the climacteric rise in respiration which is a characteristic of ripening in most fruits. Sacher (13) showed that leakage of solutes increased progressively with the climacteric rise in ripening banana and avocado fruits, and he noted that these results were consistent with the hvpothesis that alteration in membrane properties may catuse the climacteric. Ben-Yehoshua (2) reported that leakage of cellular materials from excised disks of avocado fruits began before the onset of the climacteric and continued to increase during the rise in respiration. He agreed with Sacher that changes in membrane permeability may bear a causal relationship to the climacteric. Baur and Workman (1) noted that changes in permeability and the onset of the climacteric apparently occulrred simultaneously in banana fruits. Bturg et al. (5) concluded that solute efflux from banana tissue was simply a reflection of sugar content, and that permeability did not change with ripening. However, using solute uptake to estimate free space, Sacher (14) has shown conclusively that permeability does increase with ripening of banana frtuits and, indeed, that changes in permeability precede the onset of the climacteric rise. There is evidence that an increase in ethylene prodtuction also precedes the climacteric rise in respiration. This has been shown in bananas (4) and in Honey Dew melons (12). Recalculation of the data of Lyons et al. (7) indicates that an increase in internal ethylene concentration probably precedes the onset of the climacteric in cantaloupe fruits, also.