Pour-plate Method Research Articles

Survival and transfer of microorganisms from kitchen sponges to surfaces of stainless steel and polyethylene

Contaminated sponges might lead to cross-contamination in kitchens since they can transfer microorganisms to surfaces where microorganisms can survive for hours or days and contaminate food. The main objective of this study was to evaluate the transfer and the survival of bacteria from kitchen sponges to surfaces of AISI 316 stainless steel and polyethylene. Twenty-four sponges were collected from industrial kitchens in the state of Rio Grande do Sul and aseptically split into two equal parts. One part was subjected to enumeration of heterotrophic microorganisms, faecal coliforms, coagulase-positive Staphylococcus and search detection of Salmonella enterica. The other part was rubbed on surfaces of AISI 316 stainless steel (12 sponges) or polyethylene (12 sponges). The transfer and survival of microorganisms was quantified by swab collection and pour-plate method using plate count agar. All sponges were contaminated by heterotrophic microorganisms (average of 6.8 log CFU/sponge) and 83.3% with faecal coliforms (average of 5 log CFU/sponge). None of the sponges were contaminated by S. enterica and/or coagulase-positive Staphylococcus. The average transfer of microorganisms varied between 3.3 and 5.5 log CFU/cm2 for stainless steel and from 3.5 to 5.6 log CFU/cm2 for polyethylene. Although the survival rate decreased over time, more than 1 log CFU/cm2 of heterotrophic microorganisms survived after 24 hours on both surfaces. The sponges used in food services were significantly contaminated and could transfer large amounts of microorganisms to surfaces of AISI 316 stainless steel and polyethylene.

Antagonistic effects of Saccharomyces cerevisiae on the growth of Aspergillus flavus and Aspergillus parasiticus at varying temperatures

A pour-plate dilution method was employed to assess the antagonistic effects of Saccharomyces cerevisiae on the growth of both Aspergillus flavus and Aspergillus parasiticus at 22°C, 25°C, and 32°C. Dilutions of S. cerevisiae were pour plated, and upon solidification, the plates were inoculated with a lawn of either A. flavus or A. parasiticus. Results of this experiment suggest that higher concentrations of S. cerevisiae are effective in inhibiting the growth of both Aspergillus species investigated at all temperatures, but its effects are most pronounced at 22°C. Broader implications of this study include the possibility that S. cerevisiae may be used as a biocontrol agent to protect agricultural products commonly consumed by humans from the toxigenic metabolites produced by A. flavus and A. parasiticus.

Microbial Analysis of Soyabean Milk Sold In Makurdi Metropolis

A survey of the production and consumption rate of soymilk within Makurdi metropolis and its microbial quality was carried out. Samples were collected from women and children who hawked the product in Northbank, Wurukum, Highlevel and Wadata areas of Makurdi. Data on soymilk production and consumption was collected using questionnaire method. Pour plate method and relevant biochemical tests were used for isolation, identification and characterisation of microbes. Proximate analysis was carried out to determine the nutrient content of milk samples. Producer educational status showed that 54.55% of the producers had primary school qualification while 27.27% had secondary school certificates; 18.18% were graduates. The consumption rate showed that 61.79% were daily consumers, 31.07% occasional consumers and 7.14% made up nonconsumers. Proximate chemical analysis showed that the samples had high moisture content ranging from 68.50 – 91.49% with the ash content being 0.30 – 0.90%. The fibre content ranged from 0.10% 0.35% and the lipid content, 1.56% 17.10%. The protein content ranged from 2.86 to 7.76% while the carbohydrate content ranged from 3.09 – 13.93%. The microbial load of soymilk ranged from 6.9 × 10 7 – 7.6 × 10 7 c.f.u./ml for North bank, 4.1 × 10 7 – 5.6 × 10 7 c.f.u./ml for Wurukum, 3.0 × 10 7 – 4.7 × 10 7 c.f.u./ml for High level and 6.0 × 10 7 – 8.5 × 10 7 c.f.u./ml for Wadata respectively. All samples were contaminated with members of the enterobacteriaceae, including E. coli, Klebsiella spp, Salmonella typhi, Streptococcus faecalis, and Staphylococcus aureus. Soymilk sold in the sampled areas were highly contaminated with faecal microbes, indicating poor hygiene of handlers and sanitary quality of processing water.

Carbon sequestration potential of Scenedesmus species (Microalgae) under the fresh water ecosystem

The study was conducted to investigate the carbon sequestration potential of Scenedesmus species (a microalgae species) under the fresh water ecosystem. Biofixation of CO2 by microalgae mass cultures represents an advanced, climate friendly biological process that enables the direct utilization of fossil CO2 streams produced from concentrated sources, such as power plants. The study was done in Nagpur region, the sample of Scenedesmus species (a microalgae species) were collected from fresh water ponds scattered over in Nagpur area. Collected samples were isolated for Scenedesmus species with Streak plate and Pour plate method by using serial dilution, purification was done by antibiotic treatment. The growth of Scenedesmus species was tested under different concentration of CO2, heavy metal treatment and different pH values and also chlorophyll contents were estimated over the various concentrations of CO2. Results revealed that the highest growth reported at 36% of CO2 treatment, 7.4 to 8.0 pH values and growth inhibited under high concentration of heavy metals. Chlorophyll synthesis was reported highest at 36% of CO2 treatment. With the results it concluded that the Scenedesmus species having good carbon sequestration potential as it can tolerate up to 48% CO 2 concentration in the medium, and can give excellent biomass accumulation under the 36% CO2 contents in the medium. This Scenedesmus species can be exploited as good source of bio-fixation of environmental CO2.

Dual-functional composite with anticoagulant and antibacterial properties based on heparinized silk fibroin and chitosan

Heparinized biomaterials exhibit great anticoagulant properties. However, they promote proliferation of Staphylococcus aureus (S. aureus) and therefore cause infection within the bloodstream upon implantation in vivo. In the present study, an interesting dual-functional composite with anticoagulant and antibacterial properties based on heparinized silk fibroin and chitosan was synthesized. First, heparin was grafted onto the silk fibroin by covalent immobilization with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS). All data gathered from Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and elemental analysis (EA) indicated that the heparin was successfully immobilized onto the silk fibroin. The dual-functional composite of heparinized silk fibroin and chitosan was then fabricated by a blending method. The anticoagulant activity of the heparinized materials was evaluated using the prothrombin time (PT), activated partial thromboplastin time (APTT) and thrombin time (TT). The results showed that both heparinized silk fibroin and the composite material exhibited better hemocompatibility in comparison with single silk fibroin or chitosan. The antibacterial property of the materials was investigated by the pour-plate method. Results further suggested that the composite antibacterial property with respect to S. aureus was significantly enhanced. The dual-functionality of the composite material may supply a potential choice in blood contact devices.

Microorganisms Associated with the Deterioration of Fresh Leafy Indian Spinach in Storage

This work investigated microorganisms associated with the deterioration of fresh leafy Indian spinach. Microbial counts involve the use of pour plate method (PPM). The deterioration, marked by loss of green colour to mushiness of the spinach leaves was caused by a total of four (4) bacterial species and five (5) fungal species. The bacteria isolates were Bacillus subtilis, Serratia marcescens, Lactobacillus sp, and Proteus mirabilis. The fungal isolates were Aspergillus niger, Aspergillus flavus, Mucor mucedo, Fusarium oxysporum and Penicillium sp. The spinach had high moisture content ranging from 90.50% in the white variety (Basella alba) and 90.00% in the purple variety (Basella rubra). The presence of these microorganisms in the leafy spinach indicated microbial contamination, hence spoilage and loss of quality.

In Vitro Effect of photodynamic therapy on Aggregatibacter actinomycetemcomitans and Streptococcus sanguinis

New periodontal disease treatments are needed to prevent infection progression. Photodynamic therapy (PDT) is one of the greatest pledges for this purpose. It involves the use of light of specific wavelength to activate a nontoxic photosensitizing agent in the presence of oxygen for eradication of target cells, and can be used for photokilling of microorganisms. This study evaluated in vitro the photodynamic effect of 0.01% toluidine blue-O (TBO) in combination with an AlGaInP diode laser light source on Aggregatibacter actinomycetemcomitans (A.a.) and Streptococcus sanguinis (S.s.). Suspensions (2 mL) containing A.a. and S.s. at 1.5 x 108 CFU/mL concentration were prepared and divided into 3 groups: Control group (no treatment), Dye group (inoculum and TBO for 5 min) and Dye/Laser group (inoculum, TBO for 5 min and laser for 3 min). Next, a dilution for subsequent subculture in 20 mL of Trypic Soy Agar (A.a) and Brucella Agar (S.s.) in Petri dishes (Pourplate Method) was done. Incubation of A.a. in microaerophilia and S.s. in aerobiosis at 35oC for 48 h was performed for subsequent visual counting of CFU/mL. Data were analyzed by one-way ANOVA and Tukey's HSD test at 5% significance level. For both strains, the control group showed a significantly higher (p<0.05) bacterial growth (1.5 x 108 CFU/mL), while the Dye group presented no significant reduction (p>0.05) in the CFU counts. The Dye/Laser group presented a significant decrease in the CFU counts (p<0.05) compared with the Control group (61.53% for A.a. and 84.32% for S.s.). It may be concluded that PDT was effective in reducing the numbers of A.a. and S.s. in vitro.

Bacterial species associated with soils contaminated with used petroleum products in Keffi town, Nigeria

This investigation was carried out to determine the bacterial flora of soils contaminated with used oil (petroleum products) in Keffi town. Pour plate method involving the use of serial dilutions was employed for the isolation of the bacteria. Soil samples from 10 different mechanic workshops in Keffi town were obtained and plated out on plate count agar, eosin methylene blue agar, brilliant green agar, desoxycholate citrate agar and mannitol salt agar to isolate the bacterial species from the soils contaminated with petroleum products. The bacterial species isolated were Pseudomonas sp., Streptococcus sp., Escherichia coli, Staphylococcus sp.,Klebsiella sp., Bacillus sp., Mycobacterium sp., Enterobacter aerogenes,Salmonella sp., and Micrococcus sp. The hydrocarbon substrates (petroleum products) used were petrol, kerosene, diesel and engine oil. Pseudomonas sp.,Streptococcus sp., and Bacillus sp., were found to utilize all the four petroleum products as their sole source of carbon and energy. Staphylococcus sp., andMicrococcus sp., utilized petrol, kerosene and diesel, while Klebsiella sp., andMycobacterium sp., utilized only petrol and diesel. Salmonella sp., E. aerogenes andE. coli did not utilize any of the test substrates (Petrol, kerosene, diesel and engine oil). The results of this study revealed that Pseudomonas sp., Streptococcus sp., andBacillus sp., are the most versatile species of bacteria that could utilized petroleum products in the soil environment of Keffi. The investigation demonstrates thatPseudomonas sp., Streptococcus sp. and Bacillus sp. could be harnessed for use in bioremediation of land polluted with petroleum and petroleum products. Key words: Bacterial species, contaminated soils, used petroleum products, Keffi, Nigeria.

Comparison of a modified peptone water transport medium with two commercially available transport media for the recovery of aerobes from swab specimens

A laboratory modified peptone water medium was evaluated alongside Stuart and Amies media for their relative suitability as transport media for aerobic bacteria isolated from wound specimens obtained from Central Hospital, Benin City, Nigeria. The survival rates of isolates from the three media were assessed quantitatively for a period of seven successive days, using the pour-plate method for making viable counts. The percentage of isolation was in the following decreasing order, Pseudomonas aeruginosa (36.4%), Klebsiella aerogenes (27.3%), Escherichia coli (22.7%), Staphylococcus aureus (18.2%) and Proteus sp (13.6%). The survival rate of aerobic bacterial isolates was enhanced in the modified peptone water transported medium (77.3%) compared with that obtained in Stuart medium (22.7%) and Amies medium 54.5%). Statistical analysis using the students t-test at 90% confidence limit showed significant difference (p < 0.10) when modified peptone water was compared with both Stuart and Amies transport media.

Colony growth of Philippine isolates of Blastocystis hominis in simplified, soft agar medium

The agar-cloning technique of Blastocystis hominis has been observed in both solid and semisolid agar using Iscove's modified Dulbecco's medium. In this study, Philippine isolates of B. hominis were grown by pour-plate method in semisolid agar using Locke's solution. Inoculated plates contained 0.7% agar, 10% horse serum, and 0.1% sodium thioglycollate. Plates were incubated at 37 degrees C in a microaerophilic jar for 7-10 days. Biconvex disk-shaped colonies were seen abound at the bottom half of the medium. Colonies growing at the agar-glass interface were flat and consisted of thin layers of cells. From these colonies, large amoeboid cells were frequently seen on the periphery, whereas smaller cells were concentrated at the core. Analysis of the SSU rDNA genetically established the identity of the clones to be B. hominis. This is the first report on agar cloning of Blastocystis in a compound medium.

Viability and virulence of selectedbeauveria brongniartii formulations againstHolotrichia serrata

Selected formulations of the entomopathogenic fungusBeauveria brongniartii were evaluated for conidial viability by media plating method and bioassay against eggs and larvae of its target hostHolotrichia serrata F. (Coleoptera: Scarabaeidae). When inoculated on rose Bengal agar medium, talc formulation (9 months) produced the highest colony forming units (CFUs) among all the formulations giving formulation strength of 2.4 - 4.2 x 106 CFUs/g in pour-plate method and 1.2 - 6.0 x 106 CFUs/g in spread-plate method. Lignite (4 months) produced colonies by both methods giving variable formulation strength at different dilutions. The detected strength (CFUs/g) was far lower than the label strength (spores/g) for these formulations. Formulations with vermicast, lignite (both 16 months) and dry yeast (7 months) as carrier materials did not produce colonies. In bioassay tests withH. serrata eggs, formulations produced lower infection rates (0-12%) than pure conidial suspension (28%). In third instar grubs too, formulations produced lower mortality rates (0-40%) than pure conidial suspensions (100%). The study indicated suitability of talc and lignite as carrier material and a shelf life of less then 12 months.

Flow-cytometric study of vital cellular functions in Escherichia coli during solar disinfection (SODIS)

The effectiveness of solar disinfection (SODIS), a low-cost household water treatment method for developing countries, was investigated with flow cytometry and viability stains for the enteric bacterium Escherichia coli. A better understanding of the process of injury or death of E. coli during SODIS could be gained by investigating six different cellular functions, namely: efflux pump activity (Syto 9 plus ethidium bromide), membrane potential [bis-(1,3-dibutylbarbituric acid)trimethine oxonol; DiBAC4(3)], membrane integrity (LIVE/DEAD BacLight), glucose uptake activity (2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-d-glucose; 2-NBDG), total ATP concentration (BacTiter-Glo) and culturability (pour-plate method). These variables were measured in E. coli K-12 MG1655 cells that were exposed to either sunlight or artificial UVA light. The inactivation pattern of cellular functions was very similar for both light sources. A UVA light dose (fluence) of <500 kJ m(-2) was enough to lower the proton motive force, such that efflux pump activity and ATP synthesis decreased significantly. The loss of membrane potential, glucose uptake activity and culturability of >80 % of the cells was observed at a fluence of approximately 1500 kJ m(-2), and the cytoplasmic membrane of bacterial cells became permeable at a fluence of >2500 kJ m(-2). Culturable counts of stressed bacteria after anaerobic incubation on sodium pyruvate-supplemented tryptic soy agar closely correlated with the loss of membrane potential. The results strongly suggest that cells exposed to >1500 kJ m(-2) solar UVA (corresponding to 530 W m(-2) global sunlight intensity for 6 h) were no longer able to repair the damage and recover. Our study confirms the lethal effect of SODIS with cultivation-independent methods and gives a detailed picture of the 'agony' of E. coli when it is stressed with sunlight.

Xanthan Gum: An Economical Partial Substitute for Agar in Microbial Culture Media

Xanthan gum, microbial desiccation-resistant polysaccharide prepared commercially by aerobic submerged fermentation from Xanthomonas campestris, has been successfully used alone and in combination with agar for microbial culture media. As illustrative examples, eight bacteria and eight fungi were grown on media solidified with either agar (A, 1.5%), xanthan gum (X, 1%), or combinations of both (0.9% X + 0.1% A, 0.8% X + 0.2% A, 0.7% X + 0.3% A, 0.6% X + 0.4% A). All fungi and bacteria exhibited normal growth and differentiation in all these treatments. Rather, growth of most of the fungi was better on xanthan (alone) and xanthan + agar media than agar medium. As the media gelled with xanthan gum alone flow, it was not possible to incubate Petri plates in inverted position. Moreover, because of the softness, streaking of bacteria was difficult on such media. However, these problems could be overcome by partially replacing xanthan gum with 0.3% agar. Bacterial enumeration studies carried out for Serratia sp. and Pseudomonas sp. by serial dilution and pour-plate method on agar (1.5%), 0.7%/0.6% X + 0.3%/0.4% A yielded similar counts. Selective media, succinate medium for Pseudomonas sp., and MacConkey broth medium for Escherichia coli gelled with 0.7%/0.6% X + 0.3%/0.4% A did not support growth of other bacteria when inoculated along with the above-mentioned bacteria. Likewise, differential medium, CRMA (Congo red mannitol agar) gelled with xanthan-agar combination could differentiate between Agrobacterium tumefaciens and Rhizobium sp.

Susceptibility of Propionibacterium acnes and Staphylococcus epidermidis to killing by MPO-halide system products. Implication for taurine bromamine as a new candidate for topical therapy in treating acne vulgaris

Taurine chloramine (TauCl) and taurine bromamine (TauBr) are the main haloamines produced by activated neutrophils. TauCl exerts both anti-inflammatory and microbicidal activities. Clinical studies showed that TauCl may be useful as an antimicrobial agent in the local treatment of infections. Much less is known about TauBr. Circumstantial evidence suggests that Propionibacterium acnes (PA) has a role in the inflammation of acne. Available topical therapies include antimicrobial agents which reduce total PA numbers and anti-inflammatory agents which suppress activity of the cells present in acne inflammatory lesions. In this study the bactericidal activities of TauBr and TauCl against PA and Staphylococcus epidermidis (SE), as a control strain, were investigated. Moreover, the influence of these haloamines on the generation of reactive oxygen species (ROS) by activated neutrophils was also tested. TauBr and TauCl were prepared by reaction of taurine with HOBr and HOCl, respectively. The reaction was monitored by UV absorption spectra. The bactericidal activities of TauBr and TauCl were determined by the pourplate method. The generation of ROS by neutrophils was determined by luminol chemiluminescence assay. In our experimental set-up, TauBr showed stronger antibacterial activity than TauCl. Interestingly, PA was significantly more susceptible to TauBr than SE was. Moreover, TauBr at non-cytotoxic concentrations significantly reduced ROS generation by neutrophils. Since PA is considered to be an etiological agent in acne and ROS are closely correlated with the pathogenesis of inflammatory skin diseases, the reported data suggest that TauBr may be a good candidate for the topical therapy for acne vulgaris.

Comparison of the Compact Dry TC Method with the Standard Pour Plate Method (AOAC Official Method 966.23) for Determining Aerobic Colony Counts in Food Samples: Performance-Tested MethodSM

Compact Dry TC qualifies as a rapid method kit for determining aerobic colony counts in foods. The plates are presterilized and contain culture medium and a cold-soluble gelling agent. The medium is rehydrated by inoculating 1 mL diluted sample into the center of the self-diffusible medium and allowing the solution to diffuse by capillary action. The plates can then be incubated and the colonies counted without any additional steps. The Compact Dry TC method was validated with 5 different raw meats. The performance tests were conducted at 35 degrees and 30 degrees C. In all required performance studies, no apparent differences were observed between the Compact Dry TC method and the Standard Pour Plate method (AOAC Official Method 966.23) for the detection level of aerobic microorganisms. For the accuracy claim (n = 60), a correlation factor of r2(35) = 0.9977 (35 degrees C) and r2(30) = 0.9932 (30 degrees C) could be assigned, as stated in the application for "Performance Tested Method." Quality consistency and storage robustness studies, showed no significant variations in plate count results with different production lots or plates of diverse storage age.

Guar gum: a cheap substitute for agar in microbial culture media

To determine the possibility of using guar gum, a colloidal polysaccharide, as a cheap alternative to agar for gelling microbial culture media. As illustrative examples, 12 fungi and 11 bacteria were cultured on media solidified with either guar gum or agar. All fungi and bacteria exhibited normal growth and differentiation on the media gelled with guar gum. Microscopic examination of the fungi and bacteria grown on agar or guar gum gelled media did not reveal any structural differences. However, growth of most of the fungi was better on guar gum media than agar, and correspondingly, sporulation was also more advanced on the former. Bacterial enumeration studies carried out for Serratia sp. and Pseudomonas sp. by serial dilution and pour-plate method yielded similar counts on both agar and guar gum. Likewise, a selective medium, succinate medium used for growth of Pseudomonas sp. did not support growth of Bacillus sp. when inoculated along with Pseudomonas on both agar or guar gum supplemented medium. Guar gum, a galactomannan, which is 50 times cheaper than Difco-bacto agar, can be used as a gelling agent in place of agar in microbial culture media. As the media gelled with guar gum do not melt at temperature as high as 70 degrees C, these can be used for isolation and maintenance of thermophiles.

Is there a role of taurine bromamine in inflammation? Interactive effects with nitrite and hydrogen peroxide

The myeloperoxidase system of neutrophils generates chlorinating and brominating oxidants in vivo. The major haloamines of the system are taurine chloramine (TauCl) and taurine bromamine (TauBr). It has been demonstrated in vitro that TauCl exerts both antiinflammatory and anti-bacterial properties. Much less is known about TauBr. The present study was conducted to compare bactericidal and immunoregulatory capacity of TauBr with that of the major chlorinating oxidants: HOCl and TauCl. Moreover, the effect of nitrites and H(2)O(2) on TauBr activity was investigated. TauBr was prepared by reaction of HOBr with taurine. The reaction was monitored by UV absorption spectra. Bactericidal activity of TauBr, TauCl and HOCl was tested by incubation of E. coli with the compounds and determined by the pour-plate method. To test the anti-inflammatory activity the compounds were incubated with LPS and IFN-gamma stimulated murine peritoneal macrophages. The production of following mediators was measured: nitrites by Griess reaction; TNF-alpha, IL-6, IL-10, IL-12p40 using capture ELISA. In some experiments the compounds were incubated with either nitrites or H(2)O(2). In our experimental set-up TauBr and HOCl exerted strong bactericidal effects on E. coli (MBC = 110 microM and 8 microM, respectively), while TauCl (< 1000 microM) did not kill test bacteria. However, both, TauBr and TauCl, at noncytotoxic concentrations (< 300 microM) inhibited the cytokine and nitric oxide production by macrophages. H(2)O(2) completely abolished the biological activities of TauBr but not those of TauCl. Nitrites did not affect any activity of TauBr or TauCl while they diminished the HOCl(-) mediated bacterial killing. TauBr, despite very low concentration of Br(-) in body fluids, may support TauCl and HOCl in the regulation of inflammatory response and in killing of bacteria by neutrophils. However, TauBr activity in vivo will depend on the presence of H(2)O(2) and possible other mediators of inflammation which can compete with target molecules for TauBr.

Antibacterial activity of Aristolochia bracteata root extracts.

Aristolochia bracteata (Aristolochiaceae) is used in traditional medicines as a gastric stimulant and in the treatment of cancer, lung inflammation, and dysentery and snake bites. The present investigation was undertaken to evaluate the antibacterial activity of Aristolochia bracteata root extracts. Roots of A. bracteata were powdered and successively extracted with ethyl acetate, acetone, methanol, and water for 8 hours each using a Soxhlet extractor. Antibacterial activity of dried extracts was evaluated by the pour-plate method against a few Gram-positive and Gram-negative bacteria. All the crude extracts showed a broad spectrum of antibacterial activity. The ethyl acetate extract was found to be the most effective. This study shows the potential for replacement of synthetic preservatives by the use of natural extracts.

Bacteriological qualities of indoor and out-door drinking water in Kibera sub-location of Nairobi, Kenya.

To compare the bacteriological quality of out-house (tank or standpipe) water and in-house drinking water (storage containers) and determine the risk factors influencing it. A cross-sectional study. The study was carried out in Kibera slums located 7 km southwest from the Nairobi City centre. Water samples from twenty outside tanks/standpipes and sixty from in-house water storage containers. Pour plate method was used to enumerate total bacterial counts in water, while the multiple tube technique was used to determine faecal coliform (FC) and faecal streptococci (FS) numbers. A questionnaire and environmental observation were used to determine the risk factors influencing bacteriological quality of water. The mean total bacterial counts (TBC) for out-house water was 46.6 per 100 ml while that for in-house water was 818.2 per 100 ml. Faecal coliforms were isolated from 7 (35%) standpipes and 57 (95%) in-house storage containers. The mean faecal coliform count was 93 and 103.4 per 100 ml for out-house and in-house water, respectively. The counts were significantly higher in the latter. Faecal streptococci were isolated from 2 (10%) standpipes and 37 (61.7%) in-house storage containers. The mean faecal streptococci counts were 35 and 65 per 100 ml for out-house and in-house water sources, respectively. Escherichia coli was isolated in 2 (10%) of out-house water and 30 (50%) of in-house. Of these, four were enteropathogenic, serotype 011 from one out-house water source and serotypes 011, 011, 0112ac from in-house water sources. Bacteriological contamination of water at the source with a further deterioration between the collection points and homes was observed. A defective water delivery system and inadequate environmental sanitation were a potential source of contamination for out-house water. Scoops were a major source of contamination for stored water.

Antimicrobial and cytotoxic activity of hypochlorous acid: interactions with taurine and nitrite.

HOCl, a major bactericidal product of neutrophil MPO-halide system reacts with taurine to form taurine chloramine (TauCl), a less toxic anti-inflammatory mediator. Recently, it has been reported that HOCl may also react with nitrite (NO2-), a major end-product of nitric oxide (NO) metabolism, to form very active oxidant, nitryl chloride (NO2Cl). The present study was conducted to elucidate the effect of nitrite on bactericidal and some immunoregulatory properties of HOCl and TauCl. TauCl was prepared from NaOCl and taurine. The reaction was carried out at pH 5.0 and pH 7.4, in the presence or absence of nitrite. All reactions were monitored by UV absorption spectra. Bactericidal activity of HOCl and TauCl in the presence of nitrite was tested by incubation of E. coli with the compounds and determined by the pour-plate method. To test the effect of the compounds on activity of inflammatory cells, murine peritoneal neutrophils (PMN) and macrophages were used. The cells were activated in vitro with either LPS, IFN-gamma or zymosan and the production of following mediators was measured: reactive oxygen species using luminol-dependent chemiluminescence; nitric oxide by Griess reaction; TNF-alpha using capture ELISA. In addition, we tested the effect of HOCl and TauCl on activity of myeloperoxidase (MPO). At physiological pH nitrite reacts with HOCl but not with TauCl. This reaction was abolished in the presence of taurine. Nitrite prevented HOCl-mediated bacterial killing, inhibition of MPO activity, cellular cytotoxicity and inhibition of TNF-alpha production. Nitrite did not affect any activity of TauCl. We have shown that nitrite may react in vitro with HOCl but not with TauCl, to form new biologically active product(s). We did not confirm the hypothesis that a product of HOCl reaction with nitrite is more toxic than HOCl. To the contrary, we found that nitrite diminished bactericidal and immunoregulatory properties of HOCl. In vivo, nitrite will also compete with taurine for reaction with PMN-released HOCl. Nevertheless, due to high concentration of taurine in PMN cytosol, formation of TauCl will be a major regulatory mechanism of MPO-halide-system.