Volatilization and disinfection efficacy of gaseous hypochlorous acid from an air washer-type humidifier in a large space.

An air washer-type humidifier has two useful functions: humidification, and air purification, and it applies to large indoor spaces. In this study, the efficacy of an air washer-type humidifier fed with 24 L of weakly acidic electrolyzed water（WAEW) at pH 5.0 and 30 mg/L in disinfecting attached bacteria and airborne microorganisms was studied in a 480 m3 indoor space. The humidifier was operated at a shower volume of 9.0 L/min of WAEW and at an air flow rate of 29 m3/min. Volatilization of gaseous hypochlorous acid（HOCl(g)) proceeded according to first-order kinetics during the 60 min of operation. Fresh WAEW was supplied to the humidifier every 60 min, and the HOCl(g) concentration in the indoor space was maintained within the range of 25-52 ppb for at least 180 min of operation. The number of viable bacterial cells on wet agar plates placed on the floor at a distance of 5-20 m away from the humidifier decreased by 2.0-3.0 log after 30 min of operation, and no viable cells were detected after 60 min of operation. A logarithmic reduction of more than 2.7 was achieved within 15 min against bacteria-attached plates placed at a 1.5 m-height position where the outlet airflow from the humidifier was directly exposed. This indicates that the disinfection efficacy of HOCl(g) volatilized from the humidifier depends on the rate of outlet airflow reaching the bacteria-attached plates. The number of viable airborne microorganisms decreased by approximately 54% after 180 min of operation. This study demonstrated that an air-washer-type humidifier can spread HOCl(g) evenly throughout a large indoor space and is effective in disinfecting attached bacteria and airborne microorganisms.

Correlation between disinfection efficacy and cumulative amount of free chlorine reaching various positions during ultrasonic fogging with hypochlorite solution.

When a hypochlorite solution is ultrasonically fogged in a room, free chlorine, i.e., HOCl and OCl-, reaches various positions in two forms: fine fog droplets and gaseous hypochlorous acid（HOCl(g)）. In this study, the cumulative amount of free chlorine reaching various positions on the floor away from the fogger was measured in a 90-m3 room, using a sulfamate-carrying glass-fiber filter indicator. The fine droplets were blown out from the fogger into the spaces at different discharge port angles of 30 - 90°. Free chlorine was successfully trapped by sulfamate, forming monochlorosulfamate, which was stably retained on the indicator. The cumulative amount of free chlorine（ ng/indicator） increased with fogging time at each position and depended on the blow angle and distance from the fogger. Minor differences in the HOCl(g) concentration near the floor at all positions were observed. The disinfection efficacy of the fogging treatment against Staphylococcus aureus on wet surfaces was relatively higher at positions near the fogger and lower at positions far from the fogger. At each discharge port angle, a strong correlation between the logarithmic reduction in relative viable cells and the cumulative amount of free chlorine reaching S. aureus plates was observed. The slopes of the regression lines of correlation diagrams as a function of the cumulative amount of free chlorine were between -0.0362 and -0.0413 ng-1. This study demonstrated that the cumulative amount of free chlorine measured using the filter indicator could reflect the sum of the free chlorine of both fine droplets and HOCl(g), and that the disinfection efficiency depended on the cumulative amount of free chlorine reaching different areas.

Inhalation of gaseous hypochlorous acid and its effect on human respiratory epithelial cells in laboratory model systems.

During the disinfection of indoor spaces using gaseous hypochlorous acid (HOCl(g)), inhalation is the most common route of exposure for humans. In this study, an artificial human respiratory tract model was exposed to 12-140 ppb HOCl(g) at an aspiration flow rate of 800 mL/s for 15 h in a 1 m3 chamber. The respiratory tract model was equipped with 5th order bronchi and all gas-contact parts were made of silicone rubber with no other chlorine-consuming substances. The concentration of HOCl(g) reaching the lung pseudo-space was approximately 47.4% of the HOCl(g) concentrations in the chamber and was calculated to be very close to zero when the chamber concentration was less than 20.5 ppb. The disappearance of HOCl(g) during inhalation is likely due to the adsorption of HOCl(g) on the gas-contact silicone rubber surfaces. The cytotoxicity of HOCl(g) on respiratory epithelial cells was also examined using human air-liquid-interface airway tissue models. Human nasal epithelium and bronchiolar epithelium were exposed to 100 ppb and 500 ppb HOCl(g) for 8 h and 5 d, respectively. No significant effects of HOCl(g) on cell viability and ciliary activity were observed in any cell type, indicating that low concentrations of HOCl(g）, less than 500 ppb, had no cytotoxic effect.

Efficacy of sodium chlorite in inactivating Vibrio parahaemolyticus attached to polyethylene terephthalate surfaces.

The inactivation of Vibrio parahaemolyticus cells attached to a polyethylene terephthalate (PET) disc in a sodium chlorite (NaClO2) solution was kinetically studied in a weakly acidic pH range of 4.0 - 6.5. The logarithmic reduction in the survival ratio depended on the concentration-time product. All inactivation curves showed a linear reduction phase, and the reduction in viable cells was greater than 4-log. No significant desorption of attached cells was observed during the inactivation treatment. The first-order inactivation rate constant (k) increased by approximately 4.5-fold for every 1.0 unit fall in pH. At all pH values, the k values calculated for the attached cells were approximately half of those for the unattached cells. These findings indicate that a weakly acidic NaClO2 solution is effective in inactivating bacteria attached to hard surfaces.

Uses of gaseous hypochlorous acid for controlling microorganisms in indoor spaces.

Hypochlorous acid (HOCl) is an active species in the chlorination process. Hypochlorite salts that release hypochlorite ion (OCl-) have been used for more than 200 years as disinfecting, cleaning, deodorizing, and decolorizing agents in various technological fields. In the food industry, sodium hypochlorite is the most widely used among chlorine compounds. The antimicrobial activity of a dilute hypochlorite solution is attributed largely to HOCl because of its cell membrane permeability. OCl- exhibits an excellent cleaning action for organic soils on solid surfaces. HOCl has been used as an aqueous solution, and its objects to be treated are things. In hypochlorite solution, HOCl is volatile and easily volatilized by stirring, bubbling, atomizing, or forced-air vaporization. On the other hand, OCl- is non-volatile and stays in the solution. Recently, the scope of objects to be treated with hypochlorite solution has been expanded to indoor spaces, and the use of gaseous hypochlorous acid (HOCl（g) ) has been studied intensively. This review describes the mechanisms of actions of hypochlorous acid as liquid-based and gaseous disinfectants and provides the evidence for the safety and effectiveness of HOCl（g) for controlling microorganisms in indoor spaces.

Cobalt-Chromium Alloy Has Superior Antibacterial Effect Than Titanium Alloy: In Vitro and In Vivo Studies.

STUDY DESIGN: In vitro and in vivo laboratory studies. OBJECTIVE: This study aimed to compare bacterial survival on titanium alloy (Ti) and cobalt-chromium alloy (CC) using in vitro and in vivo experiments. SUMMARY OF BACKGROUND DATA: Spinal implants are frequently manufactured from Ti and CC. These foreign materials are thought to be susceptible to biofilm formation that contributes to the development of surgical site infections. Certain metals (i.e., silver, cobalt) are known to have antibacterial properties. METHODS: In the in vitro study, discs made of Ti or CC were incubated with one of two common bacteria: Staphylococcus aureus (S. aureus) and Propionibacterium acnes (P. acnes). After incubation, discs were assessed to determine the number of viable bacterial cells. In the in vivo study, the discs that were made of CC or Ti were implanted into the subcutaneous layer of BALB/c mice. After skin closure, a suspension including either S. aureus or P. acnes was directly inoculated on the implanted discs. The discs were retrieved and analyzed to determine the number of viable bacteria at 0.5, 1, and 3 days after inoculation. RESULTS: The number of viable S. aureus cultured from the CC discs was 0.9 ±â€Š0.2 × 103 CFU/disc, which was significantly lower than the cultured Ti discs (114.8 ±â€Š18.3 × 103 CFU/disc). Moreover, a significantly lower mean number of P. acnes were cultured with CC (1.9 ±â€Š1.2 × 103 CFU/disc) compared with the Ti (180.0 ±â€Š72.1 × 103 CFU/disc). The in vivo infection model testing against S. aureus or P. acnes showed a significantly lower number of viable S. aureus or P. acnes on CC discs than Ti discs. The result was seen at all measured time points. CONCLUSION: CC suppressed S. aureus and P. acnes proliferation compared with Ti in vitro and in an in vivo infection model.Level of Evidence: N/A.

Microbial community analysis in the gills of abalones suggested possible dominance of epsilonproteobacterium in Haliotis gigantea.

Gills are important organs for aquatic invertebrates because they harbor chemosynthetic bacteria, which fix inorganic carbon and/or nitrogen and provide their hosts with organic compounds. Nevertheless, in contrast to the intensive researches related to the gut microbiota, much is still needed to further understand the microbiota within the gills of invertebrates. Using abalones as a model, we investigated the community structure of microbes associated with the gills of these invertebrates using next-generation sequencing. Molecular identification of representative bacterial sequences was performed using cloning, nested PCR and fluorescence in situ hybridization (FISH) analysis with specific primers or probes. We examined three abalone species, namely Haliotis gigantea, H. discus and H. diversicolor using seawater and stones as controls. Microbiome analysis suggested that the gills of all three abalones had the unclassified Spirochaetaceae (one OTU, 15.7 ± 0.04%) and Mycoplasma sp. (one OTU, 9.1 ± 0.03%) as the core microbes. In most libraries from the gills of H. gigantea, however, a previously unknown epsilonproteobacterium species (one OTU) was considered as the dominant bacterium, which accounted for 62.2% of the relative abundance. The epsilonproteobacterium was only detected in the gills of H. diversicolor at 0.2% and not in H. discus suggesting that it may be unique to H. gigantea. Phylogenetic analysis performed using a near full-length 16S rRNA gene placed the uncultured epsilonproteobacterium species at the root of the family Helicobacteraceae. Interestingly, the uncultured epsilonproteobacterium was commonly detected from gill tissue rather than from the gut and foot tissues using a nested PCR assay with uncultured epsilonproteobacterium-specific primers. FISH analysis with the uncultured epsilonproteobacterium-specific probe revealed that probe-reactive cells in H. gigantea had a coccus-like morphology and formed microcolonies on gill tissue. This is the first report to show that epsilonproteobacterium has the potential to be a dominant species in the gills of the coastal gastropod, H. gigantea.

Diversity, enumeration, and isolation of Arcobacter spp. in the giant abalone, Haliotis gigantea.

Arcobacter have been frequently detected in and isolated from bivalves, but there is very little information on the genus Arcobacter in the abalone, an important fishery resource. This study aimed to investigate the genetic diversity and abundance of bacteria from the genus Arcobacter in the Japanese giant abalone, Haliotis gigantea, using molecular methods such as Arcobacter-specific clone libraries and fluorescence in situ hybridization (FISH). Furthermore, we attempted to isolate the Arcobacter species detected. Twelve genotypes of clones were obtained from Arcobacter-specific clone libraries. These sequences are not classified with any other known Arcobacter species including pathogenic Arcobacter spp., A. butzleri, A. skirrowii, and A. cryaerophilus, commonly isolated or detected from bivalves. From the FISH analysis, we observed that ARC94F-positive cells, presumed to be Arcobacter, accounted for 6.96 ± 0.72% of all EUB338-positive cells. In the culture method, three genotypes of Arcobacter were isolated from abalones. One genotype had a similarity of 99.2%-100.0% to the 16S rRNA gene of Arcobacter marinus, while the others showed only 93.3%-94.3% similarity to other Arcobacter species. These data indicate that abalones carry Arcobacter as a common bacterial genus which includes uncultured species.

Kinetics of the Inactivation of Vibrio parahaemolyticus in Weakly Acidic Sodium Chlorite Solution.

　The kinetics of the inactivation of Vibrio parahaemolyticus in sodium chlorite (NaClO2) solution was studied in the weakly acidic pH range of 4.0 to 6.5 and at various temperatures. The logarithmic reduction of the survival ratio depended on the concentration-time product, and all the inactivation curves showed a linear reduction phase. The first-order inactivation rate constant (k) increased by approximately twice for every 0.44 unit fall in pH. During the inactivation experiments, no formation of chlorine dioxide occurred. These data indicated that undissociated HClO2 was the active species governing the inactivation of V. parahaemolyticus. It was also shown that the use of weakly acidic NaClO2 solutions containing high concentrations of ionized ClO2- gave slower kinetics of the inactivation, whereas it could achieve the significant reduction of viable cells of more than 4-log. The k value showed an Arrhenius-type temperature dependence in the temperature range of 5 to 40℃. The apparent activation energy for the inactivation of V. parahaemolyticus was estimated to be 43.5 kJ/mol. The k value increased by approximately 1.8 times for every 10℃ rise in temperature.

Inactivation of Vibrio parahaemolyticus Unattached and Attached to a Solid Surface in pH-Controlled Sodium Hypochlorite Solutions.

　The inactivation of Vibrio parahaemolyticus cells that were unattached or attached to a polyethylene terephthalate (PET) disc in pH-controlled sodium hypochlorite (NaOCl) solutions was studied under turbulent conditions. No significant desorption of attached cells occurred at the free available chlorine (FAC) concentrations from 0.1 to 1.0 mg/l. The number of viable cells was estimated by microbial calorimetry. The logarithmic relative reduction of viable cells was proportional to the product of the FAC concentration and time. In the pH range of 5.6 to 9.3, the first-order inactivation rate constants for unattached and attached cells increased with decreasing solution pH. It was found that the rate constants for unattached cells were approximately 6 to 7 times higher than those for attached cells at all pH values examined. It was confirmed that attached cells were more resistant to NaOCl solutions than unattached cells even when accessibility of attached cells to HOCl/OCl- was enhanced under turbulent conditions.

The mode of action of sodium hypochlorite in the decolorization of azo dye orange II in aqueous solution.

The effect of sodium hypochlorite (NaOCl) on the decolorization of azo dye orange II (4-[(2-hydroxy-1-naphthalenyl) azo]-benzenesulfonic acid, monosodium salt) in aqueous solution was studied as a function of pH. The first-order rate constant of color reduction (k) was evaluated from the curve of color reduction in the initial stage of decolorization. The k values increased with increasing the pH from 5.4 to 9.3 and the NaOCl concentration from 2.82 × 10(-4) M to 1.13 × 10(-3) M. It was found that k depended on the concentration of OCl(-), but not on HOCl. On the other hand, k decreased markedly at pHs more than 10, depending on pH. This suggested that the reactivity of the orange II molecule with OCl(-) decreased probably due to the competitive action of the hydroxide ion under alkaline conditions. It could be concluded that controlling the solution pH and the OCl(-) concentration was indeed a key factor determining the rate of decolorization of orange II in aqueous solution.

Improvement of the cleanability of pseudo-sensitized stainless steel surfaces by solution treatment.

Type 316L stainless steel particles were pseudo-sensitized by being heated at 700 degrees C for 100 h under a reduced pressure of 1.3 x 10(-3) Pa. Pseudo-sensitization treatment resulted in the formation of chromium-rich precipitates at the outermost surfaces of the stainless steel particles. The curve for the apparent surface charge density (sigma app) of the pseudo-sensitized particles showed a more basic character compared with the curve for the sigma app of the original particles. There was no significant difference between the amounts of bovine serum albumin (BSA) adsorbed on the original and the pseudo-sensitized particles. On the contrary, the efficiency of BSA removal from the pseudo-sensitized particles during batch-wise and continuous cleaning operations with 0.1M NaOH solution were significantly smaller than that for the original particles. It was indicated that the extremely lowered cleanability of the pseudo-sensitized particles was caused by a marked increase in the positive sigma app values after the pseudo-sensitization. Solution treatment of the pseudo-sensitized particles at 1,050 degrees C for 1 h under a reduced pressure of 1.3 x 10(-3) Pa could put again the chromium precipitates into a solid solution, thereby returning the surface chemical composition and the sigma app nearly to the original states. In addition, the solution-treated particles showed a good cleanability comparable to that of the original particles. These results demonstrated that solution treatment could improve the lowered cleanability of the pseudo-sensitized stainless steel surfaces.

Cleanability of titanium and stainless steel particles in relation to surface charge aspects.

The cleanability of titanium and 316L stainless steel particles was studied in terms of their apparent surface charge density (sigma(app)). Bovine serum albumin (BSA) was used as the model fouling agent. Curves for the sigma(app) of titanium and stainless steel particles showed the apparent points of zero charge (pzc(app)) of 4.6 and 8.5, respectively. Compared with the curve for the sigma(app) of stainless steel, that of titanium was characterized by small positive and large negative sigma(app) values. The isotherms for BSA adsorption and the saturation amount of BSA adsorbed on titanium and stainless steel depended largely on the intrinsic properties of BSA. In continuous cleaning in a plug-flow column fed by a 0.05M NaOH solution, BSA was found to be faster desorbed from titanium than from stainless steel, and smaller amounts of BSA remaining after 120-min cleaning were observed on titanium. Kinetic analysis showed that the two first-order desorption rate constants, reflecting the rate of BSA desorption in the initial and later stages of cleaning, for titanium were respectively 1.7-fold and 1.3-fold higher than those values for stainless steel. It could be suggested that the better cleanability of titanium was probably due to the small binding strength of BSA on slightly negatively-charged titanium surfaces and due to their large negative sigma(app) values under alkaline cleaning conditions.

Mechanisms of actions of sodium hypochlorite in cleaning and disinfection processes.

Sodium hypochlorite (NaOCl) is the most widely used disinfectant in the food industry despite the increasing availability of other disinfectants. Sodium hypochlorite fulfills many requirements as the ideal disinfectant and furthermore it has an excellent cleaning action. The effectiveness of sodium hypochlorite in the cleaning and disinfection processes depends on the concentration of available chlorine and the pH of the solution. Hypochlorous acid (HOCl) is a weak acid and dissociates to the hypochlorite ion (-OCl) and proton (H+) depending on the solution pH. It is generally believed that HOCl is the active species in the germicidal action, whereas the concentration of -OCl is a key factor determining the cleaning efficiency. This implies that the optimal pH region of the germicidal activity of sodium hypochlorite differs from that of its cleaning activity. This paper describes the theory and practice of the cleaning and disinfecting operations based on the use of sodium hypochlorite solution.

Role of a surfactant in caustic alkali cleaning of polyethylene terephthalate surfaces fouled with organic substances.

The role of polyoxyethylene lauryl ether (POELE), a nonionic surfactant, in removing bovine serum albumin (BSA) and catechin from polyethylene terephthalate (PET) particles was quantitatively studied. Relatively large fractions of adsorbed BSA and catechin were removed from PET particles under weak alkaline conditions below pH 9, whereas cleaning with high concentrations of NaOH was not so effective, especially on the removal of BSA. This was attributed to the lower susceptibility of PET particles to water-based alkali (OH-) cleaning because of the lower degree of the polarity of PET surfaces. The combined use of NaOH solutions of moderate pHs and POELE could successfully improve the efficiency of cleaning of PET particles without a significant dissolution of PET materials. There was a good correlation between the lowered surface tension (7) of the NaOH solution with POELE and the removal efficiency of BSA or catechin. No effect of POELE was observed on the NaOH cleaning of hydrophilic alumina (A12O3) particles fouled with BSA. It could be concluded that the role of POELE in NaOH cleaning of PET particles was to lower the gamma of the NaOH solution and hence to facilitate penetration of OH- ions into the PET-BSA and PET-catechin interfaces.

Involvement of radical species in inactivation of Vibrio parahaemolyticus in saline solutions by direct-current electric treatment.

The effect of pulsed low-direct-current (DC) electric treatment on the viability of Vibrio parahaemolyticus in artificial seawater and 3.0% (w/v) NaCl solution was studied as a function of available chlorine (AC) concentration. The amount of AC generated during the DC electric treatment increased in proportion to the amount of passed DC. The survival fraction of V. parahaemolyticus cells decreased depending on AC concentration. When the generated AC components were completely reduced in the presence of sufficient sodium thiosulfate, no inactivation of V. parahaemolyticus in the NaCl solution was observed during the DC electric treatment. Based on the AC concentration, the inactivation efficacies of the DC electric treatment of the seawater and NaCl solution were approximately 4-fold and 30-fold that of the exogenous addition of sodium hypochlorite, respectively. Fluorometric analysis using 2-[6-(4'-hydroxy)phenoxy-3H-xanthen-3-on-9-yl]benzoic acid showed that the generation of highly reactive radical species such as hydroxyl radical in the seawater and NaCl solution occurred during the DC electric treatment. The amount of generated radical species depended on the amount of passed DC. It is concluded that pulsed low-DC electric treatment of saline solutions exerts superior inactivation efficacy against V. parahaemolyticus to sodium hypochlorite owing to the generation of radical species.

Cleanability of stainless steel surfaces as influenced by heat treatment.

The cleanability of heat-treated 316L stainless steel particles treated at various temperatures of 100 to 500 degrees C was studied in a plug-flow column fed by a 0.1M NaOH solution. Bovine Serum albumin (BSA) was used as the model fouling agent. Heat treatment resulted in the enrichment of iron in passive films on stainless steel particles depending on temperature. The degree of surface hydroxylation and the apparent surface charge density (sigma(app)) of stainless steel particles decreased markedly with increasing heating temperatures. The saturation amounts of BSA adsorbed (gamma(sat)) were larger on the particles heated at higher temperatures. No correlation was observed between the gamma(sat) and sigma(app) values. With increasing heating temperatures, the rate of BSA desorption from stainless steel particles decreased gradually in the initial and later stages of cleaning, resulting in larger amounts of BSA remaining on the particles at the end of 120-min of cleaning. The susceptibility to BSA adsorption and the cleanability were found to be correlated with the iron content of the passive films on stainless steel particles. It could be suggested that the decrease in the cleanability was probably due to the formation of iron-enriched passive films with lower degrees of surface hydroxylation of stainless steel particles due to heat treatment.

Kinetic study of desorption of two species of bovine serum albumin from alumina during alkali elution process.

Kinetics of desorption of irreversibly adsorbed bovine serum albumin (BSA) from alumina (Al2O3) particles at various surface coverages (theta) was studied in a plug-flow column by feeding 0.05 M NaOH solution. The desorption curve obtained by plotting the logarithm of the amount of residual BSA against elution time was analyzed by using an integrated model that describes two simultaneous first-order processes. At each of theta, except for an initial lag time, the curve of BSA desorption could be reduced to the sum of two independent and simultaneous first-order processes occurring at different rates. The desorption rate constant (k(f)) for a faster-desorbing BSA (BSA(f)) was influenced by theta, especially above 0.7. On the other hand, the rate constant (k(s)) for a slower-desorbing BSA (BSA(s)) was fairly constant irrespective of theta and approximately 25- to 100-fold lower than k(f). The amount of BSA(f) estimated was proportional to theta, whereas that of BSA(s) reached a plateau value corresponding to 12.8% of saturation amount at theta above 0.42.