Hypochlorous Acid Research by Microbe - Results Type: Bacteria Microbe: Bacillus

This study discussed the effects of different bacterial concentrations and centrifugations on the antimicrobial efficacy of electrolyzed oxidizing (EO) water on Bacillus subtilis and Escherichia coli O157:H7. Overnight grown bacterial cultures were centrifuged 1 to 3 times and bacterial concentrations were adjusted to approximately 9 (high), 7 (medium), or 5 (low) log10 CFU/mL. Antimicrobial efficacy of acidic EO water (AEW) and neutral pH EO water (NEW) containing 0.2530 mg/L available chlorine was determined. In order to ascertain the effects of AEW and NEW on targeted pathogens, cellular properties at bio-molecular levels were also studied. The results showed that the susceptibility of both pathogens decreased significantly with increasing bacterial concentrations. AEW with 10, 0.25 and 0.25 mg/L and NEW with 30, 0.5 and 0.25 mg/L available chlorine were needed for high, medium and low bacterial concentrations, respectively to non-detectable levels by direct plating for E. coli O157:H7. B. subtilis was found more resistant to both EO water treatments and only 4.1 and 3.8 log reductions were achieved for AEW and NEW containing 30 mg/L available chlorine. On the other hand, it was observed that as centrifugation time increased, both bacteria became significantly more sensitive to EO water treatments. When centrifugation period increased from 1 to 3 times, additional 2.67 and 3.38 log E. coli O157:H7 reductions were observed for AEW and NEW treatments, respectively. A similar trend was observed for B. subtilis. DNA and protein leakage increased when pathogens were treated by AEW and NEW with increasing available chlorine concentration, but decreased DNA and protein leakage were observed with increased centrifugation times. These results indicate that initial bacterial concentration and the centrifugation time are two important factors and should be carefully considered in chlorine-based antimicrobial efficacy testing.

Slightly acid electrolyzed water (SAcEW) and ultrasound (US) treatment have emerged as an environmental-friendly antimicrobial agent. However, SAcEW treatment alone shows low antimicrobial efficiency. Therefore, the aim of this study was to develop a hurdle approach that combined SAcEW and US to improve the antimicrobial effect against Bacillus cereus as well as inhibition of the growth on potato. US treatment under different conditions of dip times, acoustic energy densities (AED) and temperatures were conducted to obtain the optimal condition. Our findings demonstrate that 3 min of US with 400 W/L of AED at 40 C treatment (US 40 C) significantly (p 0.05) reduced B. cereus population by 2.3 0.1 log CFU/g with minimal change in the color of potato. In addition, 3 min of SAcEW (pH, 5.35.5 ORP, 958981 mV ACC, 2830 mg/L) simultaneous with US40 C treatment (SAcEW US40 C) an approximately 3.0 log CFU/g reduction in B. cereus. Furthermore, SAcEW US40 C treatment efficiently extended lag time of B. cereus by 0.210.5 hrs, reduced that of specific growth rate by 0.010.23 log CFU/h during storage at different temperatures from 5 to 35 C. Therefore, this combined hurdle technology is capable of improving microbial safety of potato during storage and distribution.

Spores of some Bacillus species are responsible for food spoilage and foodborne disease. These spores are highly resistant to various interventions and cooking processes. In this study, the sporicidal efficacy of acidic electrolyzed oxidizing (EO) water (AEW) and slightly acidic EO water (SAEW) with available chlorine concentration (ACC) of 40, 60, 80, 100, and 120 mg/L and treatment time for 1, 2, 3, 4, 5, and 6 min were tested on Bacillus subtilis and Bacillus cereus spores in suspension and on carrier with or without organics. The reduction of spore significantly increased with increasing ACC and treatment time (P < 0.05). Nondetectable level of B. cereus spore in suspension occurred within 2 min after exposure to both EO waters containing 120 mg/L ACC, while only SAEW at 120 mg/L and 2 min treatment achieved >6 log reductions of B. subtilis spore. Both types of EO water with ACC of 60 mg/L and 6 min treatment achieved a reduction of B. subtilis and B. cereus spores to nondetectable level. EO water with ACC of 80 mg/L and treatment time of 3 min on carrier test without organics addition resulted in reductions of B. subtilis spore to nondetectable level. But, addition of 0.3 organics on carrier decreased the inactivation effect of EO water. This study indicated that EO water was highly effective in inactivation of B. subtilis and B. cereus spores in suspension or on carrier, and therefore, rendered it as a promising disinfectant to be applied in food industry.

The aim of this study was to investigate the in-vitro antimicrobial activity of usage and normal concentrations of electrolyzed water in hospital. In our study, the effects of different concentrations of electrolyzed water on two gram positive, four gram negative standard strains and clinical isolates of four gram negative, two gram positive, one spore-forming bacillus and Myroides spp strains that lead to hospital infections were researched. The effects of different concentrations and different contact times of Envirolyte electrolyzed water on cited strains were researched through method of qualitative suspension tests. Petri dishes fo bacteria have been incubated at 37 C 48 hours. Bactericidal disinfectant was interpreted to be effective at the end of the period due to the lack of growth. Solutions to which disinfectant were not added were prepared with an eye to control reproduction and controlcultures were made by using neutralizing agents. 1/1, 1/2, and 1/10 concentrations of Envirolyte electrolyzed water were found to be effective on the bacteria that lead to hospital infections used during all test times. As a conclusion, based upon the results we acquired, it was observed that Envirolyte electrolyzed water of 100 concentration would be convenient to be used for disinfection when diluted to a usage concentration of 1/10.

Application of slightly acidic electrolyzed water (SAEW) in combination with ultrasound for decontamination of kashk was investigated. SAEW had a pH of 5.3-5.5, an oxidation reduction potential of 545-600 mV, and an available chlorine concentration of 20-22 mg/L. Kashk is a dairy product with a unique aroma and a high nutritive value produced in Iran. A 2/1 SAEW/kashk ratio showed 1.42, 1.13, 1.24, and 1.37 log CFU/mL microbial reductions in Staphylococcus aureus, Bacillus cereus, Escherichia coli, and Aspergillus fumigatus, respectively, at room temperature. A combination of SAEW treatment with ultrasound (SAEWultrasound) resulted in 1.87, 1.67, 1.71, and 1.91 log CFU/mL reductions in S. aureus, B. cereus, E. coli, and A. fumigatus, respectively. The developed hurdle approach can be a useful tool for sanitization of kashk and similar products. Application of SAEWultrasound in dairy microbial decontamination is first reported herein.

The combined effects of ultrasonication and slight acidic electrolyzed water were investigated to improve the microbial safety of brown rice against Bacillus cereus infection and to evaluate the growth kinetics of these bacteria during storage of untreated and treated rice at various temperatures (5, 10, 15, 20, 25, 30, and 35C). The results indicate that this combination treatment was bactericidal against B. cereus, resulting in an approximately 3.29-log reduction. Although B. cereus can be efficiently reduced by treatment, temperature abduring storage can allow B. cereus to recover and grow. A primary growth model (Baranyi and Roberts equation) was fitted to the raw growth data from untreated (control) and treated samples to estimate growth rate, lag time, and maximum population density, with a low standard error of the residuals (0.140) and high adjusted coefficient of determination (>0.990). The growth curves obtained from the Baranyi and Roberts model indicated that B. cereus grew more slowly on treated brown rice than on untreated brown rice. Secondary models predicting the square root of the maximum growth rate and the natural logarithm of the lag time as a function of temperature were satisfactory (bias factor = 0.993 to 1.013 accuracy factor = 1.290 to 1.352 standard error of prediction = 18.828 to 36.615%). Inactivation results and the model developed and validated in this study provided reliable and valuable growth kinetics information for quantitative microbiological risk assessment studies of B. cereus on brown rice.

Super-oxidized water is one of the broad spectrum disinfectants, which was introduced recently. There are many researches to find reliable chemicals which are effective, inexpensive, easy to obtain and use, and effective for disinfection of microorganisms leading hospital infections. Antimicrobial activity of super-oxidized water is promising. The aim of this study was to investigate the in-vitro antimicrobial activity of different concentrations of Medilox super-oxidized water that is approved by the Food and Drug Administration (FDA) as high level disinfectant. Material and methods In this study, super-oxidized water obtained from Medilox Soosan E & C, Korea device, which had been already installed in our hospital, was used. Antimicrobial activities of different concentrations of super-oxidized water (1/1, 1/2, 1/5, 1/10, 1/20, 1/50, 1/100) at different exposure times (1, 2, 5, 10, 30 min) against six ATCC strains, eight antibiotic resistant bacteria, yeasts and molds were evaluated using qualitative suspension test. Dey-Engley Neutralizing Broth Sigma-Aldrich, USA was used as neutralizing agent. Results Medilox was found to be effective against all standard strains (Acinetobacter baumannii 19606, Escherichia coli 25922, Enterococcus faecalis 29212, Klebsiella pneumoniae 254988, Pseudomonas aeruginosa 27853, Staphylococcus aureus 29213), all clinical isolates (Acinetobacter baumannii, Escherichia coli, vancomycin-resistant Enterococcus faecium, Klebsiella pneumoniae, Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, Bacillus subtilis, Myroides spp.), and all yeastsat 1/1 dilution in 1 minute. It was found to be effective on Aspergillus flavus at 1/1 dilution in 2 minutes and on certain molds in 5 minutes. Conclusion Medilox super-oxidized water is a broad spectrum, on-site producible disinfectant, which is effective on bacteria and fungi and can be used for the control of nosocomial infection.

The efficacy of slightly acidic electrolyzed water (SAEW) for reducing microbial contamination on fresh-cut cilantro was investigated in this study. The impacts of SAEW on the microbes of cilantro samples inoculated with two kinds of bacteria (Escherichia coli O78 and Bacillus subtilis 1.1849) were evaluated in comparison with NaClO solution and acidic electrolyzed water (AEW). Dipping with AEW, SAEW and NaClO solutions for 5 min resulted in a reduction in populations of E. coli O78 from 6.38 to 4.93, 3.89 and 4.88 log10 cfu/g and in populations of B. subtilis from 6.52 to 5.02, 4.98, 4.63 log10 cfu/g, respectively, The similar results were found that the populations on cilantro inoculated the mixture of two microbes of E. coli O78 treated with AEW, SAEW and NaClO solutions decreased to 4.15, 3.99, 5.10 log10 cfu/g, respectively, and the populations of B. subtili on cilantro decreased to 5.08,4.97,4.82 log10 cfu/g, respectively. The efficacies of SAEW wash in reducing natural micro flora on fresh-cut cilantro were studied. The results showed SAEW had strong disinfection ability to reduce the microbe population of fresh-cut cilantro and could be an alternative of AEW and NaClO solutions.

A study was carried out on the disinfection efficiency of electrolyzed oxidizing water (EOW) on spores of Bacillus subtilis var. niger. The results showed a remarkable fungicidal rate of 100% after 20 min duration of 191 mg/L active available chlorine (ACC). The disinfection effect was improved with increased ACC or prolonged disinfection time, while organic interferents exerted a strong concentration-dependent inhibition against the disinfection. The disinfection mechanism was also investigated at bio-molecular level. EOW decreased dehydrogenase activity, intensified membrane permeability, elevated suspension conductivity, and caused leakage of intracellular K+, proteins, and DNA, indicating a damage of cell walls and membranes. Effects of EOW on microbiological ultra-structures were also verified by transmission electronic microscopy (TEM) images, showing that EOW destroyed protective barriers of the microbe and imposed some damages upon the nucleus area.

In this study we investigated the effects of low concentration electrolyzed water (LcEW) and several other sanitizers (strong acid electrolyzed water (SAEW), aqueous ozone (AO), 1% citric acid (CA) and sodium hypochlorite solution (NaOCl)) on the inactivation of natural microflora (total aerobic bacteria counts (TBC) and yeasts and moulds (YM)) and foodborne pathogens (Escherichia coli O157:H7, Listeria monocytogenes, Salmonella Typhimurium and Bacillus cereus) on oyster mushroom. The effects of temperature and treatment time on the antimicrobial activity of LcEW to reduce the populations of foodborne pathogens were also determined. LcEW showed the strongest bactericidal efficacy among all the sanitizers on TBC, YM and pathogens by reductions of 1.35, 1.08 and 1.90 2.16 log CFU/g after 3 min treatment at room temperature (23 2 C), respectively. There was no significant difference between the antimicrobial effects of LcEW and SAEW (P > 0.05). Among those sanitizers, their relative influence of inactivation was LcEW > NaOCl > CA > AO.

The bactericidal effect of slightly acidic hypochlorous water (SAHW) on Salmonella Enteritidis, Escherichia coli, Staphylococcus aureus, Listeria monocytogenes, and Bacillus cereus, as well as some bacterial strains isolated from fresh lettuce was evaluated. Viable counts of all tested bacterial samples decreased immediately after treatment by SAHW. Most bacterial cells with the exception of B. cereus, and S. aureus were not culturable on TSA after treatment by 1 to 30 mg/L SAHW. Likewise, Pseudomonas sp., and Flavobacterium or Xanthomonas sp., Kurthia sp., Micrococcus sp., and Corynebacterium or Microbacterium sp. were not culturable on TSA after treatment by 30 mg/L SAHW. Viable counts of S. aureus, E. coli, Flavobacterium or Xanthomonas sp., and Pseudomonas sp. showed a 5 to 6 log cfu/mL reduction at day 0 and maintained a count of less than 1 log cfu/mL from day 1 to day 7 following treatment by 30 mg/L SAHW. Sodium hypochlorite (NaOCl, 0.5-1.0 mg/L) decreased the viable counts of S. Enteritidis to less than the lower limit of detection, 1 log cfu/mL, from day 1 to day 7 following treatment by 1 mg/L. NaOCl was not sufficient at 0.5-0.75 mg/L in reducing viable counts of S. Enteritidis because of a 2 to 5 log cfu/mL increase from day 2 to day 5 due to recovery from injury. Initial counts of S. Enteritidis after hydrogen

Background : Disinfection is essential for the prevention of hospital infection. Medilox, an super oxidized water generated by Medilox (SOO SAN EC CO., LTD. Yongin, Korea) was developed as a disinfectant in Korea. This is not costly and does not caany clinical problems and environmental pollution. We evaluated bactericidal activity ofMedilox against several clinical isolates and standard strains. Method : Clinical isolates and reference ATCC strains were exposed to Medilox, an super oxidized water (80 ppm ofHOCl) generated by Medilox (SOO SAN ECCO., LTD. Yongin, Korea) for the various periods (0.5, 1, 2, 5, and 10 minutes). After the exposure mixture of microorganisms and Medilox solution was inoculated into tryptic soy broth and onto tryptic soy agar, Sabouraud dextrose agar or Ogawa medium and cultured at 35C. Results: All strains of bacteria, yeasts, mycobacteria and vegetative form of Bacillus subtilis were killed within 30 seconds after an exposure to Medilox (80 ppm ofHOCl) under clean and dirty conditions. But, spore form of Bacillus subtilis was killed within 5 minutes. Conclusion: It may be recommended that Medilox can be used for the effective disinfectant for hospital environments and high-level disinfectant for hospital infection control.

The effects of acidic electrolyzed water (AcEW), alkaline electrolyzed water (AlEW), 100 ppm sodium hypochlorite (NaClO), and 1% citric acid (CA) alone, and combinations of AcEW with 1% CA (AcEW + CA) and AlEW with 1% CA (AlEW + CA) against Bacillus cereus vegetative cells and spores was evaluated as a function of temperature (25, 30, 40, 50, or 60 C) and dipping time (3 or 6 h). A 3-strain cocktail of Bacillus cereus cells or spores of approximately 107 CFU/g was inoculated in various cereal grains (brown rice, Job s tear rice, glutinous rice, and barley rice). B. cereus vegetative cells and spores were more rapidly inactivated at 40 C than at 25 C. Regardless of the dipping time, all treatments reduced the numbers of B. cereus vegetative cells and spore by more than 1 log CFU/g, except the deionized water (DIW), which showed approximately 0.7 log reduction. The reductions of B. cereus cells increased with increasing dipping temperature (25 to 60 C). B. cereus vegetative cells were much more sensitive to the combined treatments than spores. The effectiveness of the combined electrolyzed water (EW) and 1% CA was considerable in inhibiting B. cereus on cereal grains. The application of combined EW and CA for controlling B. cereus cells and spores on cereal grains has not been previously reported. Therefore, the synergistic effect of EW and CA may provide a valuable insight on reducing foodborne pathogens on fruits, vegetables, and cereal grains.

Effects of storage temperature (1, 5, and 10 C) on growth of microbial populations (total aerobic bacteria, coliform bacteria, Bacillus cereus, and psychrotrophic bacteria) on acidic electrolyzed water (AcEW)-treated fresh-cut lettuce and cabbage were determined. A modified Gompertz function was used to describe the kinetics of microbial growth. Growth data were analyzed using regression analysis to generate best-fit modified Gompertz equations, which were subsequently used to calculate lag time, exponential growth rate, and generation time. The data indicated that the growth kinetics of each bacterium were dependent on storage temperature, except at 1 C storage. At 1 C storage, no increases were observed in bacterial populations. Treatment of vegetables with AcEW produced a decrease in initial microbial populations. However, subsequent growth rates were higher than on nontreated vegetables. The recovery time required by the reduced microbial population to reach the initial (treated with tap water [TW]) population was also determined in this study, with the recovery time of the microbial population at 10 C being <3 days. The benefits of reducing the initial microbial populations on fresh-cut vegetables were greatly affected by storage temperature. Results from this study could be used to predict microbial quality of fresh-cut lettuce and cabbage throughout their distribution.

Acidic electrolyzed water (acidic EW), which is prepared by the electrolysis of an aqueous NaCl solution, has recently become of great importance for disinfection in a variety of fields, including medicine, the food industry and agriculture. In a previous paper we showed that: 1) acidic EW is a mixture of hypocholorite ion, hypochlorous acid and chlorine, depending upon the pH; 2) hypochlorous acid is primarily responsible for disinfection in the case of Escherichia coli K12 and Bacillus subtilis PCI219, both in clean culture media. In practice, however, the use of acidic EW is in many cases severely hampered due to the presence of a variety of non-selective reducing agents. In view of the salient nature of acidic EW, it is therefore strongly urged to establish an optimum way to use acidic EW in a variety of systems. The present paper is the first report on our attempt along this line in order to characterize the nature of the chemical changes that the bactericidal activity of the acidic EW deteriorates in the presence of organic materials, which include amino acids and proteins.

To identify the primary component responsible in electrolyzed oxidizing (EO) water for inactivation, this study determined the concentrations of hypochlorous acid (HOCl) and hypochlorite ions (OCl-) and related those concentrations to the microbicidal activity of the water. The ultraviolet absorption spectra were used to determine the concentrations of HOCl and OCl- in EO water and the chemical equilibrium of these species with change in pH and amperage. EO water generated at higher amperage contained a higher chlorine concentration. The maximum concentration of HOCl was observed around pH 4 where the maximum log reduction (2.3 log10 CFU/ml) of Bacillus cereus F4431/73 vegetative cells also occurred. The high correlation (r = 0.95) between HOCl concentrations and bactericidal effectiveness of EO water supports HOCl s role as the primary inactivation agent. Caution should be taken with standard titrimetric methods for measurement of chlorine as they cannot differentiate the levels of HOCl present in EO water of varying pHs.

This study was undertaken to evaluate the efficacy of electrolyzed oxidizing (EO) and chemically modified water with properties similar to the EO water for inactivation of different types of foodborne pathogens (Escherichia coli O157:H7, Listeria monocytogenes and Bacillus cereus). A five-strain cocktail of each microorganism was exposed to deionized water (control), EO water and chemically modified water. To evaluate the effect of individual properties (pH, oxidation-reduction potential (ORP) and residual chlorine) of treatment solutions on microbial inactivation, iron was added to reduce ORP readings and neutralizing buffer was added to neutralize chlorine. Inactivation of E. coli O157:H7 occurred within 30 s after application of JAW EO water with 10 mg/l residual chlorine and chemically modified solutions containing 13 mg/l residual chlorine. Inactivation of Gram-positive and -negative microorganisms occurred within 10 s after application of ROX EO water with 56 mg/l residual chlorine and chemically modified solutions containing 60 mg/l residual chlorine. B. cereus was more resistant to the treatments than E. coli O157:H7 and L. monocytogenes and only 3 log10 reductions were achieved after 10 s of ROX EO water treatment. B. cereus spores were the most resistant pathogen. However, more than 3 log10 reductions were achieved with 120-s EO water treatment.