Evaluation of dairy and food plant sanitizers against Salmonella typhimurium and Listeria monocytogenes

Six commonly used dairy and food plant sanitizers were evaluated against Salmonella typhimurium and Listeria monocytogenes. Of these six, two were acid anionic sanitizers, one contained a quaternary ammonium compound, one was based on active iodine, and two contained active chlorine. Of the last two, one contained hypochlorite and the other contained active chlorine in organic form. The chlorine-based sanitizers were effective at 100 ppm of available chlorine against both these organisms. The sanitizer containing iodine was effective at 12.5 and 25 ppm titratable iodine against L. monocytogenes and S. typhimurium, respectively. The acid anionic sanitizers were effective at 200 ppm of active agent against both the bacteria, and the quaternary ammonium-based sanitizer was effective at 100 and 200 ppm of active compound against L. monocytogenes and S. typhimurium, respectively. The sanitizer containing iodine at 12.5 and 25 ppm of titratable iodine showed activity equivalent to 50 and 200 ppm of available chlorine, respectively, against L. monocytogenes and 100 and 200 ppm of available chlorine, respectively, against S. typhimurium.     

Minimum inhibitory concentrations of antimicrobials against micro-organisms related to citrus juice

Minimum inhibitory concentrations (MIC) of various cleaner or sanitizer compounds were determined by a Spiral Gradient Endpoint agar diffusion method against four micro-organisms associated with spoilage of orange juice (Lactobacillus plantarum,Leuconostoc mesenteroides,Gluconobacter oxydans, andSaccharomyces cerevisiae) and against eight unidentified micro-organisms isolated from citrus fruit surfaces (two yeasts and six bacteria). Survivor curve testing was conducted on the four known micro-organisms. Compounds that were effective at the lowest concentrations were chlorine dioxide, the iodophor and quaternary ammonium compound. Dimethyldicarbonate, hypochlorite, peracetic acid and a phosphoric acid anionic compound also possessed substantial antimicrobial activity. Citric and lactic acids andd-limonene were less effective as antimicrobial compounds.     

Impedimetric determination of activity of disinfectants and detergents on Listeria: preliminary study

The efficiency of several disinfectants or detergents against three strains of Listeria monocytogenes, one strain of Listeria innocua and two strains of Streptococcus group D was tested in water as well as in the presence of milk, whey and salt by an impedimetric method using a Bactometer M120. Certain synergistic effects between active agents and matrix could be observed. Differences in sensitivities were noticed amongst the tested strains. Products containing iodine, peroxide or quaternary ammonium as active agents were shown to be efficient, even at relatively low concentrations.     

Efficacy of sanitizing treatments against Penicillium expansum inoculated on six varieties of apples

The effectiveness of several wash treatments was evaluated against spores of Penicillium expansum inoculated on six varieties of apples (Red Delicious, Golden Supreme, Empire, Macintosh, Fuji, and Gala). The wash treatments were water, acidified water (pH 6.5), acidified sodium hypochlorite (pH 6.5), nonacidified sodium hypochlorite (pH 8.8, 9.3, and 9.7; 50, 100, and 200 ppm, respectively), and peracetic acid (50 and 80 ppm). Spores of P. expansum were dried on the surface of the apples for 2 h before exposure to the different sanitizer solutions. Each apple was submerged in 100 ml of each treatment solution for 30 s, and the number of spores remaining were recovered and enumerated. The efficacy of chlorine solutions was enhanced by decreasing the pH to 6.5 (up to 5-log reduction, depending on apple variety). Peracetic acid solutions (50 and 80 ppm) resulted in a reduction of less than 2 log spores per g and had the same efficacy (P < or = 0.05) as nonacidified chlorine solutions (50, 100, and 200 ppm). Control water solutions produced a reduction of 1.34 log spores per g. Chlorine solutions at pH 6.5 resulted in the largest reduction of P. expansum spores for all apple varieties tested.     

Effectiveness of sanitation with quaternary ammonium compound or chlorine on stainless steel and other domestic food-preparation surfaces.

The relative ability of various materials used for domestic and/or food-service sinks and countertops to be sanitized was determined. Both smooth (unused) and abraded surfaces were tested by exposure to 200 mg of quaternary ammonium compound per liter or 200 mg of sodium hypochlorite per liter. Surface materials tested included mechanically polished (type 304, #4 finish) and electropolished stainless steel, polycarbonate, and mineral resin. Surfaces were prepared for testing by allowing attachment of a Staphylococcus aureus culture for 4 h to achieve an initial attached population of 10(4) to 10(5) CFU/cm2. The test procedure involved immersion of the surface in sanitizer solution followed by wiping with a sanitizer-saturated cloth. Residual staphylococci were detected by overlaying agar directly on the treated surface. Results indicated that the stainless steels and the smooth polycarbonate, which had 0.5 log CFU/cm2 or fewer of residual staphylococci, were more readily sanitized by quaternary ammonium compound than were either the mineral resin surfaces, which had nearly 2.0 log CFU/cm2 of residual staphylococci, or the abraded polycarbonate which had nearly 1.0 log CFU/cm2 of residual staphylococci. Chlorine was most effective on the mechanically polished stainless steel, the unabraded electropolished stainless steel, and the polycarbonate surfaces, reducing cell populations to less than 1.0 log CFU/cm2. Chlorine was less effective on abraded electropolished stainless steel and mineral resin surfaces, where populations remained greater than 1.0 log CFU/cm2. Sanitation with quaternary ammonium compound or chlorine reduced S. aureus populations more than 1,000-fold on all surfaces except unabraded mineral resin.     

Direct microscopic observation of viability of Campylobacter jejuni on chicken skin treated with selected chemical sanitizing agents

The objective of this research was to determine the effect of chlorine, acidified sodium chlorite, and peracetic acid treatments on viable Campylobacter jejuni located at various depths within follicles or folds of chicken skin. Chicken skin was inoculated with C. jejuni transformed with P(c)gfp plasmid (GFP-Campylobacter), which also codes for kanamycin resistance. Effectiveness of sanitizer treatments was determined by plate count. C. jejuni were also observed on chicken skin by confocal scanning laser microscopy, whereby viable and nonviable cells were differentiated by their ability to take up staining with 5-cyano-2,3-ditolyl tetrazolium chloride. Sodium hypochlorite, peracetic acid, and acidified sodium chlorite were each applied at 40 or 100 ppm for 2 or 15 min. Each sanitizer resulted in approximately a 1-log decrease (CFU) when used at 100 ppm for 15 min and no significant decrease when used at 40 ppm for 2 min. Numbers of viable cells observed on the skin by direct microscopic count were similar to numbers obtained by plate count. Although viable counts decreased with sanitizer treatments, the total number of Campylobacter cells (live plus dead) attached to the skin remained unchanged. After each chemical treatment, viable C. jejuni were observed at depths of 0 to 10, 11 to 20, and 21 to 30 microm in folds or follicles of chicken skin. Most of the C. jejuni that survived treatment were located at 0 to 10 microm depth, which is where most of the viable cells were located before treatment. The inability of chemical sanitizers to effectively eliminate C. jejuni on chicken skin does not appear to be a result of protection by location in feather follicles or other depressions in the skin.     

Mitigation of Alicyclobacillus spp. spores on food contact surfaces with aqueous chlorine dioxide and hypochlorite

The prevalence of Alicyclobacillus spp. and other spore-forming spoilage organisms in food handling and processing environments presents a sanitation challenge to manufacturers of products such as juices and beverages. The objectives of this study were to determine the efficacy of chlorine dioxide and sodium hypochlorite in killing Alicyclobacillus spores in situ and to evaluate the efficacy of various chlorine dioxide and hypochlorite sanitizing regimes on Alicyclobacillus spp. spores on stainless steel, wood, and rubber conveyor material. Five or two log CFU/ml spore concentrations were left in aqueous solution or inoculated onto stainless steel, rubber, or wood coupons and challenged with sanitizer for varied time intervals. After treatment, the coupons were placed in sterile sample bags, massaged with neutralizing buffer, and enumerated on Ali agar. Surfaces were also examined before and after treatment by scanning electron microscopy to confirm destruction or removal of the spores. For both five and two log CFU/ml spore concentrations, treatments of 50 and 100 ppm of chlorine dioxide and 1000 and 2000 ppm of hypochlorite, respectively, were the most effective. Of the range of chlorine dioxide concentrations and contact time regimes evaluated for all surfaces, the most effective concentration/time regime applied was 100 ppm for 10 min. Reductions ranged from 0 to 4.5 log CFU/coupon. Chlorine dioxide was least effective when applied to wood. Hypochlorite was not efficient at eliminating Alicyclobacillus spores from any of the food contact surfaces at any time and concentration combinations tested. Chlorine dioxide is an alternative treatment to kill spores of Alicyclobacillus spp. in the processing environment.     

Efficacy of sanitizers to inactivate Escherichia coli O157:H7 on fresh-cut carrot shreds under simulated process water conditions

Chlorine is widely used as a sanitizer to maintain the microbial quality and safety of fresh-cut produce; however, chlorine treatment lacks efficacy on pathogen reduction, especially when the fresh-cut processing water contains heavy organic loads. A more efficacious sanitizer that can tolerate the commercial processing conditions is needed to maintain microbial safety of fresh-cut produce. This study evaluated the efficacy of Escherichia coli O157:H7 reduction on fresh-cut carrots using new and traditional sanitizers with tap water and fresh-cut processing water scenarios. Fresh-cut carrot shreds inoculated with E. coli O157:H7 were washed in sanitizer solutions including 200 ppm chlorine, citric acid-based sanitizer (Pro-San), 80 ppm peroxyacetic acid-based sanitizer (Tsunami 100), and 1,000 ppm acidified sodium chlorite (SANOVA) prepared in fresh tap water or simulated processing water with a chemical oxygen demand level of approximately 3,500 mg/liter. Samples were packaged and stored at 5 degrees C. Microbial analyses performed at days 0, 7, and 14 indicate that the organic load in the process water significantly affected the efficacy of chlorine on pathogen removal and was especially evident on samples tested during storage. Acidified sodium chlorite provided a strong pathogen reduction even under process water conditions with up to a 5.25-log reduction when compared with the no-wash control. E. coli O157:H7 was not recovered on acidified sodium chlorite-treated samples during the entire 14 days of storage, even following an enrichment step. These results suggest that acidified sodium chlorite holds considerable promise as an alternative sanitizer of fresh-cut produce.     

AN ASSESSMENT OF CHEMICAL SANITIZERS ON THE MICROBIOLOGICAL PROFILE OF AIR IN A MILK PROCESSING PLANT

The efficiency of chemical sanitizing agents to control the microbiota of milk processing areas at a dairy plant was evaluated. Diluted solutions of chlorhexidine digluconate at 1,000 and 2,000 mg.L^?1, pH = 5.2 and 5.3, respectively; peracetic acid, at 45 and 75 mg.L^?1, pH = 4.2 and 3.8; and a quaternary ammonium compound, pH = 9.2 and 9.3, respectively, were pulverized, at ambient temperature (20–25C) in the milk processing areas using spray equipment which produced a consistent fogging. For each 30 m² of processing area, 0.5 L of sanitizer solutions were applied at 9 Kgf.cm^?2 in 10 min. The sanitizer's efficiency was evaluated against mesophilic aerobic bacteria and yeast and molds. The microbial counts (APC) in the processing areas were detected by impaction technique, as proposed by APHA. Analysis were done before sanitizer application (T₀) and after 0.5, 12 and 24 h (T₁, T₂ and T₃), respectively. Activity of 2000 mg.L^?1 of chlorhexidine and 700 mg.L^?1 of quaternary ammonium against yeast and molds was observed. The T₀ number of microorganisms (1.5 and 1.3 log cfu.m^?3) decreased in 0.45 and 0.5 log cycle, respectively. Solutions containing 45 mg.L^?1 of peracetic acid were effective to control mesophilic aerobic microorganisms. The T₀ number of microorganisms (2.1 log cfu.m^?3) decreased in 0.55 log cycle. Residual effects against yeasts and molds were observed for 1000 mg.L^?1 of chlorhexidine digluconate and 75 mg.L^?1 of peracetic acid. Chemical sanitizer applications to control the microbiological quality of the air in milk processing areas is a barrier technology that helps in the production of foods with good microbiological, sensory and organoleptic characteristics.     

Efficacy of alkyl dimethyl benzyl ammonium chloride on suppression of Physalospora vaccinii in laboratory assays

Growth of Physalospora vaccinii on inoculated agar growth medium and cranberries treated with 0.1, 1, 10, 100, and 1,000 ppm of alkyl dimethyl benzyl ammonium chloride (ADBAC) was investigated in the laboratory. In vitro growth assays, the colony diameter, and mycelial dry weight of P. vaccinii was reduced at 1,000 ppm ADBAC. Mild or no reduction of fungal growth and mycelial dry weight was observed at concentrations less than 100 ppm when compared with the nonamended control. Growth of P. vaccinii on inoculated cranberries was inhibited by treatment with 10 and 100 ppm ADBAC. Complete inhibition of fungus growth was also achieved at 1,000 ppm ADBAC. Area under the disease progress curve values in wounded fruits were 75, 77, and 100% at 10, 100, and 1,000 ppm ADBAC, respectively, whereas area under the disease progress curve values in fruits immersed in ADBAC and pathogen were reduced 47 to 100% compared with the untreated fruits used as controls. No P. vaccinii or other fungi were detected on the control fruits inoculated with sterile distilled water. This is the first report on the use of ADBAC to control a field and storage rotting fungus, P. vaccinii. ADBAC is likely to be an important component to any integrated approach for reducing the risks associated with the presence of pathogenic microorganisms in or on foods.     

Effect of chemical sanitizers with and without ultrasonication on Listeria monocytogenes as a biofilm within polyvinyl chloride drain pipes

As part of a biofilm in a floor drain, Listeria monocytogenes is exceedingly difficult to eradicate with standard sanitizing protocols. The objective of these studies was to test the use of ultrasonication to break up biofilm architecture and allow chemical sanitizers to contact cells directly. L. monocytogenes biofilms were created in model polyvinyl chloride drain pipes. Chemical sanitizers (quaternary ammonium, peroxide, or chlorine) were applied to the drain pipes with and without a 30-s ultrasonication treatment. Controls using sterile water were included for comparison. L. monocytogenes cells were enumerated from the liquid in the drain and the inside wall surface of the pipe. All chemicals lowered numbers of planktonic cells from 6.6 log CFU/ml in the water control to < 100 CFU/ml. Attached cells were also affected by the chemical sanitizers. Approximately 6.0 log CFU/cm2 of the inner wall surface was detected in water control pipes, and ultrasonication did not lower these numbers. With or without ultrasonication, the peroxide-based sanitizer was effective for reducing the numbers of attached L. monocytogenes cells, resulting in approximately 2.0 log CFU/cm2. Both the chlorine- and quaternary ammonium-based sanitizers reduced the number of attached L. monocytogenes cells to a lesser degree, resulting in 4.2 to 4.4 log CFU/cm2. However, addition of ultrasonication improved the performance of both these sanitizers, causing a further reduction to 3.1 and 2.9 CFU/ cm2 for quaternary ammonium- and chlorine-based chemicals, respectively. These results indicate that a peroxide-based sanitizer alone can be very effective against biofilm L. monocytogenes in drain pipes, and the addition of ultrasonication can improve the effectiveness of chlorine or quaternary ammonium sanitizers.     

Salmonella sensitivity to sodium hypochlorite and citric acid in washing water of lettuce residues

Salmonella spp. is one of the main lettuce pathogens and should be inactivated during the disinfection of these vegetables before consumption. In minimally processed vegetable industries, residues of organic matter can prevent the inactivation of this pathogen by disinfectants. The objective of the present work was to evaluate the inactivation of Salmonella isolated from organic lettuce to sodium hypochlorite (25 and 50 ppm) and citric acid (0.5 and 1%) in washing water added with lettuce residues. To do so, a washing water with lettuce residues was elaborated, and Salmonella was added in the order of 10⁶ CFU/ml. Thereafter, each sanitizer was added separately to evaluate its effect on reducing Salmonella counts. After 1, 2, 3, 4, 5, 10, and 15 min of contact with the sanitizers, serial dilutions using neutralizer (0.5% sodium thiosulfate) were performed and each dilution was sown in Xylose-Lysine-Desoxycholate medium. Total aerobic mesophilic counts of wash water with lettuce residues before testing (without Salmonella) and after 15 min of exposure to each sanitizer (with Salmonella) were also performed. In addition, the free chlorine still present in the samples after the contact of sodium hypochlorite with lettuce residues for 15 min. The results demonstrated that 50 and 25 ppm sodium hypochlorite could reduce 6 log CFU/ml of Salmonella in 1 and 3 min of contact, respectively, while 0.5 and 1% citric acid was able to reduce 1.26 and 1.74 log CFU/ml respectively from the same microorganism within 15 min of contact. The total aerobic mesophilic counts of the wash water before being tested were, on average, 1.5 log CFU/ml. After addition of Salmonella, with 15 min of contact with the sanitizer, the results of total counts showed the same magnitude as the Salmonella counts. Organic matter may have reacted with the free chlorine present, reducing chlorine concentrations, since values of 30.4 ppm were observed when the initial concentration should be 50 and 17.1 ppm when the initial concentration should be 25 ppm. Based on the results, sodium hypochlorite demonstrated a greater microbial reduction capacity in wash water with lettuce residues, indicating that it is more appropriate to avoid cross-contamination between batches during sanitation of lettuce in washing tanks.     

Comparison of the traditional three-tube most probable number method with the Petrifilm, SimPlate, BioSys optical, and Bactometer conductance methods for enumerating Escherichia coli from chicken carcasses and ground beef

A study was conducted to compare commonly used methods, such as Petrifilm and SimPlate, and the rapid microbiological methods BioSys optical and Bactometer conductance to the standard most probable number (MPN) procedure for enumerating Escherichia coli from poultry carcasses and ground beef. Broiler carcasses and ground beef were evaluated in each of three replicate trials. Five groups of carcasses or ground beef were sampled and analyzed using Petrifilm, SimPlate, BioSys optical, and Bactometer conductance measurements after temperature abuse at 37 degrees C for 0 (Petrifilm and SimPlate only), 2, 4, 6, or 8 h. The correlation coefficients for the regression lines comparing the standard E. coli MPN procedure to Petrifilm and SimPlate for chicken and ground beef, respectively, were as follows: 0.95, 0.94, 0.93, and 0.91. The correlation coefficients for the regression lines comparing the standard E. coli MPN procedure to BioSys optical and Bactometer conductance measurements for chicken and ground beef, respectively, were -0.91, -0.90, -0.93, and -0.96. Although Petrifilm and SimPlate performed well, E. coli could not be enumerated from 16.7 and 10% of samples, respectively, using these methods. The BioSys optical and Bactometer conductance methods performed very well when compared with Petrifilm and SimPlate. Using rapid methods (BioSys optical and Bactometer conductance), results were obtained in 1 to 11 h rather than the 48 h required to conduct Petrifilm or SimPlate or the 5 days required to conduct the MPN procedure. These methods may allow processors to test products and obtain results before shipping, avoiding the cost and loss of reputation associated with a recall or foodborne illness outbreak.     

Detergent-sterilant spraying of mobile milk tanks before cleaning

The work described was undertaken to assess the value of spraying mobile tanks with a detergent-sterilant immediately after emptying but before proper cleaning at the farm. In a laboratory experiment, mobile tanks were divided into sections, soiled and treated by spraying with various detergent-sterilants. After a suitable contact time, sections were sampled for microbiological analyses. An iodophor and a non-ionic quaternary ammonium compound proved most effective. The iodophor was used in a field trial involving 20 farms over a period of eight weeks. Milk and tank/lid surfaces were sampled weekly. All samples were subjected to microbiological analyses. Results showed that spraying with iodophor did not lead to improved hygienic status of tanks/lids or to an improvement in milk quality.     

Impedimetric method for estimating the residual activity of freeze-dried Lactobacillus delbrueckii ssp. bulgaricus

The residual activity of Lactobacillus delbrueckii ssp. bulgaricus cultures was analysed using pH and various impedimetric methods (impedance detection time (IDT), conductance and capacitance) to quantify the loss in activity following freeze-drying. The large variation recorded in IDT values for similar levels of activity suggests that IDT is not an adequate parameter for estimating the culture's fermentative activity. Comparison of the impedance signals generated revealed that capacitance yields values that are more reproducible than those of conductance, and also gives a better correlation with pH. Statistical analysis (p<0.05) indicated that there are no significant differences between the capacitance and the pH method when attempting to estimate residual activity.     

Adaptive and cross-adaptive responses of persistent and non-persistent Listeria monocytogenes strains to disinfectants

Persistent and non-persistent Listeria monocytogenes strains were tested for initial resistance and adaptive and cross-adaptive responses towards two quaternary ammonium compounds, alkyl-benzyl-dimethyl ammonium chloride and n-alkyldimethyl ethylbenzyl ammonium chloride, one tertiary alkylamine, 1,3-propanediamine-N-(3-aminopropyl)N-dodecyl, sodium hypochlorite and potassium persulphate. The initial resistance of two persistent and two non-persistent L. monocytogenes strains was observed to differ. Both types of strains adapted after a 2-h sublethal exposure to the quaternary ammonium compounds and the tertiary alkylamine, the highest increase in the minimum inhibitory concentration (MIC) being 3-fold. Progressively increasing disinfecting concentrations at 10 and 37 degrees C resulted in adaptation of L. monocytogenes to all disinfectants except potassium sulphate. The highest observed increase in MIC was over 15-fold, from 0.63 to 10 microg/ml of n-alkyldimethyl ethylbenzyl ammonium chloride. All strains reached approximately similar MICs. Stability of the increased resistance was tested by measuring MICs every seventh day for 28 days. The increased resistance to sodium hypochlorite disappeared in 1 week, but the quaternary ammonium compounds and the tertiary alkylamine showed increased resistance for 28 days. These results suggest that cellular changes due to adaptive responses continue to have an effect on the resistance some time after the exposure. All disinfectants were shown to cause cross-adaptation of L. monocytogenes, the highest increase in MIC being almost 8-fold. The only agent that L. monocytogenes could not be shown to cross-adapt to was potassium persulphate which did, however, cause cross-adaptation to the other disinfectants. The mechanism behind these adaptive responses seemed to be non-specific as cross-adaptation was observed not only between related but also unrelated disinfectants. These findings suggest that sustaining high disinfectant effectiveness may be unsuccessful by rotation, even when using agents with different mechanisms of action.     

Possible synergistic effect of nisin and propionic acid on the growth of the mycotoxigenic fungi Aspergillus parasiticus, Aspergillus ochraceus, and Fusarium moniliforme.

The effects of nisin and propionic acid (PA) on aflatoxin production and on mycelial growth and spore germination of the mycotoxigenic fungi Aspergillus parasiticus, A. ochraceus, and Fusarium moniliforme were investigated. The growth of A. ochraceus was completely inhibited on media containing PA with nisin in concentrations of 0.05% PA with 1,000 ppm nisin, and 0.1% PA with 500 or 1,000 ppm nisin. The growth of both F. moniliforme and A. parasiticus was completely inhibited by PA with nisin at a concentration of 0.1% PA with 1,000 ppm nisin. Nisin alone caused a significant increase in mycelial growth when applied to A. ochraceus at 500 or 1,000 ppm and when applied to A. parasiticus at 1,000 ppm. Spore germination of A. ochraceus was completely inhibited on media containing 0.1% PA with 500 or 1,000 ppm nisin. Spores of F. moniliforme failed to germinate in 0.05% PA with 500 or 1,000 ppm nisin, whereas spores of A. parasiticus did not germinate on media containing 0.1% PA with 1,000 ppm nisin. For all three fungi tested, the inhibitory effect on mycelial growth was found to be fungistatic rather than fungicidal. The combined treatment of PA with nisin produced better fungistatic activity than treatment involving either material alone. Nisin, applied alone, did not stimulate aflatoxin production (expressed by microg toxin/mg mycelium), but the combined treatment at certain concentrations was inhibitory to aflatoxin B1 or G1. The production of aflatoxin G1, but not of B1, was stimulated in 0.05% PA with 1,000 ppm nisin and on media containing 0.1% PA with 100 ppm nisin. Nisin is currently applied in foods to prevent spoilage induced by bacteria but not by mold. The results of the present study indicate that a combined treatment of nisin in small concentrations of PA might be useful in preventing mold damage in certain foods and stored grain.     

A comparison of different chemical sanitizers for inactivating Escherichia coli O157:H7 and Listeria monocytogenes in solution and on apples, lettuce, strawberries, and cantaloupe

Ozone (3 ppm), chlorine dioxide (3 and 5 ppm), chlorinated trisodium phosphate (100- and 200-ppm chlorine), and peroxyacetic acid (80 ppm) were assessed for reduction of Escherichia coli O157:H7 and Listeria monocytogenes in an aqueous model system and on inoculated produce. Initially, sanitizer solutions were inoculated to contain approximately 10(6) CFU/ml of either pathogen, after which aliquots were removed at 15-s intervals over a period of 5 min and approximately plated to determine log reduction times. Produce was dip inoculated to contain approximately 10(6) E. coli O157:H7 or L. monocytogenes CFU/g, held overnight, submerged in each sanitizer solution for up to 5 min, and then examined for survivors. In the model system study, both pathogens decreased > 5 log following 2 to 5 min of exposure, with ozone being most effective (15 s), followed by chlorine dioxide (19 to 21 s), chlorinated trisodium phosphate (25 to 27 s), and peroxyacetic acid (70 to 75 s). On produce, ozone and chlorine dioxide (5 ppm) were most effective, reducing populations approximately 5.6 log, with chlorine dioxide (3 ppm) and chlorinated trisodium phosphate (200 ppm chlorine) resulting in maximum reductions of approximately 4.9 log. Peroxyacetic acid was the least effective sanitizer (approximately 4.4-log reductions). After treatment, produce samples were stored at 4 degrees C for 9 days and quantitatively examined for E. coli O157:H7, L. monocytogenes, mesophilic aerobic bacteria, yeasts, and molds. Populations of both pathogens remained relatively unchanged, whereas numbers of mesophilic bacteria increased 2 to 3 log during storage. Final mold and yeast populations were significantly higher than initial counts for chlorine dioxide- and ozone-treated produce. Using the nonextended triangle test, whole apples exposed to chlorinated trisodium phosphate (200 ppm chlorine) and shredded lettuce exposed to peroxyacetic acid were statistically different from the other treated samples.     

Inactivation of Listeria monocytogenes/Pseudomonas biofilms by peracid sanitizers.

The ability of peracetic acid and peroctanoic acid sanitizers to inactivate mixed-culture biofilms of a Pseudomonas sp. and Listeria monocytogenes on stainless steel was investigated. Types of biofilms tested included a 4-h attachment of the mixed-cell suspension and a 48-h biofilm of mixed culture formed in skim milk or tryptic soy broth. Biofilm-containing coupons were immersed in solutions of hypochlorite, peracetic acid, and peroctanoic acid either with or without organic challenge. Organic challenge consisted of either coating the biofilms with milk that were then allowed to dry, or adding milk to the sanitizing solution to achieve a 5% concentration. Surviving cells were enumerated by pouring differential agar directly on the treated surfaces. The peracid sanitizers were more effective than chlorine for inactivating biofilm in the presence of organic challenge. The 48-h mixed-culture biofilm grown in milk was reduced to less than 3 CFU/cm2 by 160 ppm of peracid sanitizer after 1 min of exposure. Peroctanoic acid was more effective than peracetic acid against biofilm cells under conditions of organic challenge. Pseudomonas and L. monocytogenes were inactivated to similar levels by the sanitizer treatments, even though Pseudomonas predominated in the initial biofilm population.     

Influence of extended acid stressing in fresh beef decontamination runoff fluids on sanitizer resistance of acid-adapted Escherichia coli O157:H7 in biofilms

This study evaluated resistance to sanitizing solutions of Escherichia coli O157:H7 cells forming biofilms on stainless steel coupons exposed to inoculated meat decontamination runoff fluids (washings). A previously acid-adapted culture of a rifampicin-resistant derivative of E. coli O157:H7 strain ATCC 43895 was inoculated in unsterilized or sterilized combined hot-water (85 degrees C) and cold-water (10 degrees C) (50/50 [vol/vol]) composite water (W) washings (pH 6.29 to 6.47) and in W washings mixed with 2% acetic acid (pH 4.60 to 4.71) or in 2% lactic acid W washings (pH 4.33 to 4.48) at a ratio of 1/99 (vol/vol). Stainless steel coupons (2 by 5 by 0.08 cm) were submerged in the inoculated washings and stored for up to 14 days at 15 degrees C. Survival of E. coli O157:H7 was determined after exposure (0 to 60 s for cells in suspension and 0 to 300 s for attached cells) to two commercial sanitizers (150 ppm peroxyacetic acid and 200 ppm quaternary ammonium compound) at 2, 7, and 14 days. E. coli O157:H7 attached more rapidly to coupons submerged in washings containing the natural flora than to those without. The attached cells were more resistant to the effects of the sanitizers than were the cells in suspension, and survival was highest in the presence of the natural flora. Attached cells in the presence of dilute acid washings were more sensitive to subsequent sanitizer treatments than were cells generated in the presence of W washings. Under the conditions of this study, cells of E. coli O157:H7 in W washings were more sensitive to acidic (peroxyacetic acid) than to alkaline (quaternary ammonium) sanitizers during storage. These results suggest that meat processing plants that apply no decontamination or that use only water washings of meat should consider using acidic sanitizers to enhance biofilm removal. Plants that apply both water and acidic washings may create a sublethal acid-stressing environment in the runoff fluids, sensitizing biofilm cells to subsequent sanitizing treatments.