Combined treatment of pulsed light and nisin-organic acid based antimicrobial wash for inactivation of Escherichia coli O157:H7 in Romaine lettuce, reduction of microbial loads, and retention of quality.

Microbial safety of fresh produce continues to be a major concern. Novel antimicrobial methods are needed to minimize the risk of contamination. This study investigated the antimicrobial efficacy of pulsed light (PL), a novel nisin-organic acid based antimicrobial wash (AW) and the synergy thereof in inactivating E. coli O157:H7 on Romaine lettuce. Treatment effects on background microbiota and produce quality during storage at 4 °C for 7 days was also investigated. A bacterial cocktail containing three outbreak strains of E. coli O157:H7 was used as inoculum. Lettuce leaves were spot inoculated on the surface before treating with PL (1-60 s), AW (2 min) or combinations of PL with AW. PL treatment for 10 s, equivalent to fluence dose of 10.5 J/cm2, was optimal and resulted in 2.3 log CFU/g reduction of E. coli O157:H7, while a 2 min AW treatment, provided a comparable pathogen reduction of 2.2 log CFU/g. Two possible treatment sequences of PL and AW combinations were investigated. For PL-AW combination, inoculated lettuce leaves were initially exposed to optimum PL dose followed by 2 min AW treatment, whereas for AW-PL combination, inoculated lettuce were subjected to 2 min AW treatment prior to 10 s PL treatment. Both combination treatments (PL-AW and AW-PL) resulted in synergistic inactivation as E. coli cells were not detectable after treatment, indicating >5 log pathogen reductions. Combination treatments significantly (P < 0.05) reduced spoilage microbial populations on Romaine lettuce and also hindered their growth in storage for 7 days. The firmness and visual quality appearance of lettuce were not significantly (P > 0.05) influenced due to combination treatments. Overall, the results reveal that PL and AW combination treatments can be implemented as a novel approach to enhance microbial safety, quality and shelf life of Romaine lettuce.

Nisin-Based Organic Acid Inactivation of Salmonella on Grape Tomatoes: Efficacy of Treatment with Bioluminescence ATP Assay.

The antimicrobial activity of a new nisin-based organic acid sanitizer (NOAS), developed in our laboratory, was tested against viable aerobic mesophilic bacteria and Salmonella populations inoculated on produce surfaces. The activity of NOAS was compared with 200 ppm of chlorinated wash water and a bioluminescence ATP technique to determine the efficacy of treatments compared with plate count methods. The activity of the 10% final concentration of NOAS against viable populations of 109 CFU/mL Salmonella in phosphate-buffered saline (PBS), sterile deionized distilled water, and buffered peptone water was tested in vitro and on grape tomatoes inoculated with Salmonella at 2.5 log CFU/g. A similar batch of inoculated tomatoes were treated with 200 ppm of total available chlorinated water. All treatments for inactivation of viable Salmonella in vitro was tested up to 30 min and 5 min for the attached populations on tomatoes. Inactivation of viable Salmonella at 109 log CFU/mL by 10% the NOAS solution averaged >107 log CFU/mL in PBS, sterile deionized distilled water, and buffered peptone water. Similarly, Salmonella bacteria inactivated on tomato surfaces by the NOAS solution was significantly (P < 0.05) greater than numbers on chlorinated washed tomatoes, and surviving bacterial populations on NOAS and chlorine-treated tomatoes were <1 and 4 CFU/g, respectively. A significant linear correlation coefficient (r2 = 0.99) between the bioluminescence ATP assay and aerobic plate counts of inoculated and untreated grape tomatoes were recorded but not with NOAS and chlorine-treated tomatoes, as bacterial populations were less than the minimum baseline for determination. Also, the results indicated that the NOAS solution is a better alternative antimicrobial wash solution than 200 ppm of chlorinated water.

Effects of pulsed light and sanitizer wash combination on inactivation of Escherichia coli O157:H7, microbial loads and apparent quality of spinach leaves.

The purpose of this study was to investigate the efficacy of pulsed light (PL), a new formula of sanitizer (HEN) consisting of hydrogen peroxide, EDTA and Nisin, as well as synergy of PL and HEN sanitizer (PL-HEN) wash in inactivating E. coli O157:H7 on spinach. The treatment effect on microbial loads and apparent quality during 13 days storage at 4 °C was also determined. A bacterial cocktail containing three strains of E. coli O157:H7 was used as inoculum based on their association with produce-related outbreaks. Spinach leaves were spot inoculated on surface before treating with PL (1-63J/cm2), HEN sanitizer wash (2 min) or their combinations. PL inactivation was influenced significantly at low doses. Treatment dose of 15.75 J/cm2, equivalent to 15 s intense PL treatment, was found optimal above which adverse quality effect was evident. The optimal PL dose resulted 2.7 log CFU/g reduction of E. coli O157:H7 while a rapid 2 min wash in sanitizer formulation HEN, provided comparatively low, 1.8 log CFU/g, reduction of the pathogen. Two different sequences of PL and HEN treatment combinations were tested. In PL-HEN treatment, inoculated leaves were first treated at optimal PL dose (15.75 J/cm2) followed by 2 min immersion in HEN whereas in HEN-PL treatment, leaves were first washed in HEN before PL exposure. HEN-PL treatment indicated a compound inactivation activity (4.6 logs reduction) while PL-HEN treatment indicated a strong synergistic inactivation as E. coli cells were not detectable after treatment indicating >5 log reduction. The PL-HEN treatment not only significantly reduced spoilage microbial populations on spinach but also slowed their growth during storage. Furthermore, the visual and firmness quality of spinach were not significantly affected by the PL-HEN treatment. Overall, our results demonstrate that integrated PL-HEN technology can be used to enhance microbial safety of spinach.

Inactivation of Salmonella in grape tomato stem scars by organic acid wash and chitosan-allyl isothiocyanate coating.

The objective of this study was to evaluate inactivation of inoculated Salmonella enterica on grape tomato stem scars exploiting integrated treatment of organic acid wash (AW) followed by chitosan-allyl isothiocyanate (CT-AIT) coating. The treatment effect on microbial loads and fruit quality during 21days storage at 10°C was also determined. A bacterial cocktail containing three serotypes of Salmonella enterica was used for this study based on their association with produce-related outbreaks. Tomatoes were spot inoculated on stem scars and then immersed in an organic acid solution (700ml) containing 0.5% (v/v) each of acetic (AA) and formic acid (FA) to wash under mild agitation for 1min at ambient temperature (22°C) followed by 1min dipping in a coating solution containing 6mlAIT/g CT. AW in 0.5% organic acid (AA+FA) for 1min reduced Salmonella population by 2.7logCFU/g from an initial load of 7.8logCFU/g, while additional coating treatment of AW tomatoes reduced the pathogens on stem scars to undetectable levels (<0.7logCFU/g), achieving, in combination, a >7logCFU/g reduction for the pathogen. Although the populations of Salmonella in the controls (approx. 7.8logCFU/g stem scar) did not change significantly during 21days of storage at 10°C, the populations were reduced to undetectable level in the integrated (AW plus CT-AIT) treated stem scars on day 1 and no regrowth was observed during storage. The treatment significantly (p<0.05) reduced background bacterial loads to approx. 1.3logCFU/g and the population remained unchanged through day 21 at 10°C. The treatment also completely inactivated mold and yeast on day 1 with no growth reoccurrence. These results indicate that the integrated treatment can provide a safe and effective intervention strategy for grape tomatoes.

Effect of Hydrogen Peroxide in Combination with Minimal Thermal Treatment for Reducing Bacterial Populations on Cantaloupe Rind Surfaces and Transfer to Fresh-Cut Pieces.

Surface structure and biochemical characteristics of bacteria and produce play a major role in how and where bacteria attach, complicating decontamination treatments. Whole cantaloupe rind surfaces were inoculated with Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes at 10(7) CFU/ml. Average population size of Salmonella, Escherichia coli O157:H7, and L. monocytogenes recovered after surface inoculation was 4.8 ± 0.12, 5.1 ± 0.14, and 3.6 ± 0.13 log CFU/cm(2), respectively. Inoculated melons were stored at 5 and 22°C for 7 days before washing treatment interventions. Intervention treatments used were (i) water (H2O) at 22°C, (ii) H2O at 80°C, (iii) 3% hydrogen peroxide (H2O2) at 22°C, and (iv) a combination of 3% H2O2 and H2O at 80°C for 300 s. The strength of pathogen attachment (SR value) at days 0, 3, and 7 of storage was determined, and then the efficacy of the intervention treatments to detach, kill, and reduce transfer of bacteria to fresh-cut pieces during fresh-cut preparation was investigated. Populations of E. coli O157:H7 attached to the rind surface at significantly higher levels (P < 0.05) than Salmonella and L. monocytogenes, but Salmonella exhibited the strongest attachment (SR value) at all days tested. Washing with 3% H2O2 alone led to significant reduction (P < 0.05) of bacteria and caused some changes in bacterial cell morphology. A combination treatment with H2O and 3% H2O2 at 8°C led to an average 4-log reduction of bacterial pathogens, and no bacterial pathogens were detected in fresh-cut pieces prepared from this combination treatment, including enriched fresh-cut samples. The results of this study indicate that the microbial safety of fresh-cut pieces from treated cantaloupes was improved at day 6 of storage at 5°C and day 3 of storage at 10°C.

Identification and Quantification of Volatile Chemical Spoilage Indexes Associated with Bacterial Growth Dynamics in Aerobically Stored Chicken.

Volatile organic compounds (VOCs) as chemical spoilage indexes (CSIs) of raw chicken breast stored aerobically at 4, 10, and 21 °C were identified and quantified using solid phase microextraction (SPME) combined with gas chromatography-mass spectrometry (GC-MS). The growth dynamics of total viable count (TVC), psychrotrophs, Pseudomonas spp., lactic acid bacteria (LAB), Brochothrix thermosphacta and H2 S producing bacteria were characterized based on maximum growth rates (µmax ), maximal microbial concentration (Nmax ) and at the moment of microbial shelf life (Svalues ), calculated from Gompertz-fitted growth curves. Pseudomonas spp. was predominant species, while B. thermosphacta was characterized by the highest µmax . The microbiological and sensory shelf lives were estimated based on TVC, Pseudomonas spp., and B. thermosphacta counts and sensory evaluation, respectively. Among 27 VOCs identified by GC-MS in spoiled chicken samples, ethanol (EtOH), 1-butanol-3-methyl (1But-3M), and acetic acid (C2 ) achieved the highest Pearson's correlation coefficients of 0.66, 0.61, and 0.59, respectively, with TVC, regardless of storage temperature. Partial least squares (PLS) regression revealed that the synthesis of 1But-3M and C2 was most likely induced by the metabolic activity of B. thermosphacta and LAB, while EtOH was attributed to Pseudomonas spp. The increase in concentration of selected volatile spoilage markers (EtOH, 1But-3M, and C2 ) in the headspace over spoiled chicken breast was found to be statistically significant (P < 0.05) with TVC growth. These findings highlight the possibility of analyzing the combination of 3 selected spoilage markers: EtOH, 1But-3M, and C2 as rapid evaluation for poultry quality testing using SPME-GC-MS.

Microbial safety and overall quality of cantaloupe fresh-cut pieces prepared from whole fruit after wet steam treatment.

Fresh-cut cantaloupes have been associated with outbreaks of Salmonellosis. Minimally processed fresh-cut fruits have a limited shelf life because of deterioration caused by spoilage microflora and physiological processes. The objectives of this study were to use a wet steam process to 1) reduce indigenous spoilage microflora and inoculated populations of Salmonella, Escherichia coli O157:H7 and Listeria monocytogenes on the surface of cantaloupes, and 2) reduce the populations counts in cantaloupe fresh-cut pieces after rind removal and cutting. The average inocula of Salmonella, E. coli O157:H7 and Listeria monocytogenes was 10(7)CFU/ml and the populations recovered on the cantaloupe rind surfaces after inoculation averaged 4.5, 4.8 and 4.1logCFU/cm(2), respectively. Whole cantaloupes were treated with a wet steam processing unit for 180s, and the treated melons were stored at 5°C for 29days. Bacterial populations in fresh-cut pieces prepared from treated and control samples stored at 5 and 10°C for up to 12days were determined and changes in color (CIE L*, a*, and b*) due to treatments were measured during storage. Presence and growth of aerobic mesophilic bacteria and Salmonella, E. coli O157:H7 and L. monocytogenes were determined in fresh-cut cantaloupe samples. There were no visual signs of physical damage on all treated cantaloupe surfaces immediately after treatments and during storage. All fresh-cut pieces from treated cantaloupes rind surfaces were negative for bacterial pathogens even after an enrichment process. Steam treatment significantly (p<0.05) changed the color of the fresh-cut pieces. Minimal wet steam treatment of cantaloupes rind surfaces designated for fresh-cut preparation will enhance the microbial safety of fresh-cut pieces, by reducing total bacterial populations. This process holds the potential to significantly reduce the incidence of foodborne illness associated with fresh-cut fruits.

Physical and chemical changes in whey protein concentrate stored at elevated temperature and humidity.

In a case study, we monitored the physical properties of 2 batches of whey protein concentrate (WPC) under adverse storage conditions to provide information on shelf life in hot, humid areas. Whey protein concentrates with 34.9 g of protein/100g (WPC34) and 76.8 g of protein/100g (WPC80) were stored for up to 18 mo under ambient conditions and at elevated temperature and relative humidity. The samples became yellower with storage; those stored at 35 °C were removed from the study by 12 mo because of their unsatisfactory appearance. Decreases in lysine and increases in water activity, volatile compound formation, and powder caking values were observed in many specimens. Levels of aerobic mesophilic bacteria, coliforms, yeast, and mold were <3.85 log10 cfu/g in all samples. Relative humidity was not a factor in most samples. When stored in sealed bags, these samples of WPC34 and WPC80 had a shelf life of 9 mo at 35 °C but at least 18 mo at lower temperatures, which should extend the market for these products.

Volatile chemical spoilage indexes of raw Atlantic salmon (Salmo salar) stored under aerobic condition in relation to microbiological and sensory shelf lives.

The purpose of this investigation was to identify and quantify the volatile chemical spoilage indexes (CSIs) for raw Atlantic salmon (Salmo salar) fillets stored under aerobic storage conditions at 4, 10 and 21 °C in relation to microbial and sensory shelf lives. The volatile organic compounds (VOCs) were analyzed with SPME-GC-MS technique. Through multivariate chemometric method, hierarchical cluster analysis (HCA) and Pearson's correlations, the CSIs: trimethylamine (TMA), ethanol (EtOH), 3-methyl-1-butanol (3Met-1But), acetoin and acetic acid (C2) were selected from the group of 28 detected VOCs. At the moment of microbiological shelf life established at total viable count (TVC) of 7.0 log CFU/g, the CSIs achieved levels of 11.5, 38.3, 0.3, 24.0 and 90.7 µg/g of salmon for TMA, EtOH, 3M-1But, acetoin and C2, respectively. Pseudomonas spp. was found as major specific spoilage organism (SSOs), suitable for shelf life prediction using modified Gompertz model at the cut-off level of 6.5 log CFU/g. H2S producing bacteria and Brochothrix thermosphacta were considered as important spoilage microorganisms; however, they were not suitable for shelf life estimation. Partial least square (PLS) regression revealed possible associations between microorganisms and synthetized VOCs, showing correlations between Pseudomonas spp. and 3Met-1But and aldehydes synthesis, lactic acid bacteria were linked with EtOH, C2 and esters, and B. thermosphacta with acetoin formation.

Efficacy of Sanitizer Treatments on Survival and Growth Parameters of Escherichia coli O157:H7, Salmonella, and Listeria monocytogenes on Fresh-Cut Pieces of Cantaloupe during Storage.

For health reasons, people are consuming fresh-cut fruits with or without minimal processing and, thereby, exposing themselves to the risk of foodborne illness if such fruits are contaminated with bacterial pathogens. This study investigated survival and growth parameters of Escherichia coli O157:H7, Salmonella, Listeria monocytogenes, and aerobic mesophilic bacteria transferred from cantaloupe rind surfaces to fresh-cut pieces during fresh-cut preparation. All human bacterial pathogens inoculated on cantaloupe rind surfaces averaged ∼4.8 log CFU/cm(2), and the populations transferred to fresh-cut pieces before washing treatments ranged from 3 to 3.5 log CFU/g for all pathogens. A nisin-based sanitizer developed in our laboratory and chlorinated water at 1,000 mg/liter were evaluated for effectiveness in minimizing transfer of bacterial populations from cantaloupe rind surface to fresh-cut pieces. Inoculated and uninoculated cantaloupes were washed for 5 min before fresh-cut preparation and storage of fresh-cut pieces at 5 and 10°C for 15 days and at 22°C for 24 h. In fresh-cut pieces from cantaloupe washed with chlorinated water, only Salmonella was found (0.9 log CFU/g), whereas E. coli O157:H7 and L. monocytogenes were positive only by enrichment. The nisin-based sanitizer prevented transfer of human bacteria from melon rind surfaces to fresh-cut pieces, and the populations in fresh-cut pieces were below detection even by enrichment. Storage temperature affected survival and the growth rate for each type of bacteria on fresh-cut cantaloupe. Specific growth rates of E. coli O157:H7, Salmonella, and L. monocytogenes in fresh-cut pieces were similar, whereas the aerobic mesophilic bacteria grew 60 to 80 % faster and had shorter lag phases.

Effect of storage temperature on survival and recovery of thermal and extrusion injured Escherichia coli K-12 in whey protein concentrate and corn meal.

Previously, we reported inactivation of Escherichia coli populations in corn product (CP) and whey protein product (WPP) extruded at different temperatures. However, information on the effect of storage temperatures on injured bacterial populations was not addressed. In this study, the effect of storage temperatures on the survival and recovery of thermal death time (TDT) disks and extrusion injured E. coli populations in CP and WPP was investigated. CP and WPP inoculated with E. coli bacteria at 7.8 log(10) CFU/g were conveyed separately into the extruder with a series 6300 digital type T-35 twin screw volumetric feeder set at a speed of 600 rpm and extruded at 35°C, 55°C, 75°C, and 95°C, or thermally treated with TDT disks submerged into water bath set at 35°C, 55°C, 75°C, and 95°C for 120 s. Populations of surviving bacteria including injured cells in all treated samples were determined immediately and every day for 5 days, and up to 10 days for untreated samples during storage at 5°C, 10°C, and 23°C. TDT disks treatment at 35°C and 55°C did not cause significant changes in the population of the surviving bacteria including injured populations. Extrusion treatment at 35°C and 55°C led to significant (p<0.05) reduction of E. coli populations in WPP as opposed to CP. The injured populations among the surviving E. coli cells in CP and WPP extruded at all temperatures tested were inactivated during storage. Population of E. coli inactivated in samples extruded at 75°C was significantly (p<0.05) different than 55°C during storage. Percent injured population could not be determined in samples extruded at 95°C due to absence of colony forming units on the agar plates. The results of this study showed that further inactivation of the injured populations occurred during storage at 5°C for 5 days suggesting the need for immediate storage of 75°C extruded CP and WPP at 5°C for at least 24 h to enhance their microbial safety.

Effect of native microflora, waiting period, and storage temperature on Listeria monocytogenes serovars transferred from cantaloupe rind to fresh-cut pieces during preparation.

The most recent outbreak of listeriosis linked to consumption of fresh-cut cantaloupes indicates the need to investigate the behavior of Listeria monocytogenes in the presence of native microflora of cantaloupe pieces during storage. Whole cantaloupes were inoculated with L. monocytogenes (10(8)-CFU/ml suspension) for 10 min and air dried in a biosafety cabinet for 1 h and then treated (unwashed, water washed, and 2.5% hydrogen peroxide washed). Fresh-cut pieces (∼3 cm) prepared from these melons were left at 5 and 10°C for 72 h and room temperature (20°C) for 48 h. Some fresh-cut pieces were left at 20°C for 2 and 4 h and then refrigerated at 5°C. Microbial populations of fresh-cut pieces were determined by the plate count method or enrichment method immediately after preparation. Aerobic mesophilic bacteria, yeast and mold of whole melon, and inoculated populations of L. monocytogenes on cantaloupe rind surfaces averaged 6.4, 3.3, and 4.6 log CFU/cm(2), respectively. Only H(2)O(2) (2.5%) treatment reduced the aerobic mesophilic bacteria, yeast and mold, and L. monocytogenes populations to 3.8, 0.9, and 1.8 log CFU/cm(2), respectively. The populations of L. monocytogenes transferred from melon rinds to fresh-cut pieces were below detection but were present by enrichment. Increased storage temperatures enhanced the lag phases and growth of L. monocytogenes. The results of this study confirmed the need to store fresh-cut cantaloupes at 5°C immediately after preparation to enhance the microbial safety of the fruit.

Hydrophobic and electrostatic interaction chromatography for estimating changes in cell surface charge of Escherichia coli cells treated with pulsed electric fields.

Pulsed electric field (PEF) treatments, a nonthermal process, have been reported to injure and inactivate bacteria in liquid foods. However, the effect of this treatment on bacterial cell surface charge and hydrophobicity has not been investigated. Apple juice (pH 3.8) purchased from a wholesale distributor was inoculated with cocktail of Escherichia coli O157:H7 at 7.4 log CFU/mL, processed with a PEF at a field strength of 18.4 kV/cm and 32.2 kV/cm at 25°C, 35°C, and 45°C with a treatment time of 160 µs and a flow rate of 120 mL/min. Bacterial cell surface charge and hydrophobicity of untreated and PEF-treated E. coli O157:H7 were determined immediately and after storage at 5°C and 23°C using hydrophobic and electrostatic interaction chromatography. Similarly, the populations surviving the PEF treatments including injured cells were determined by plating 0.1 mL of the sample on sorbitol MacConkey agar and tryptic soy agar (TSA) plates. The surviving populations of E. coli cells after PEF treatment varied depending on field strength and treatment temperature used. Percent injury in the surviving populations was high immediately after PEF treatment and varied among treatment temperatures. Cell surface charge of E. coli bacteria before PEF treatment averaged 32.10±8.12. PEF treatments at 25°C, 35°C, and 45°C reduced the above surface charge to 26.34±1.24, 14.24±3.30, and 6.72±2.82, respectively. Similarly, the surface hydrophobicity of untreated E. coli cells at 0.194±0.034 was increased to an average of 0.268±0.022, 0.320±0.124, and 0.586±0.123 after PEF treatments at 25°C, 35°C, and 45°C, respectively. The results of this study indicate that PEF treatment affects the outer cell envelope of E. coli bacteria as evidenced by the changes in surface hydrophobicity and cell surface charge leading to injury and subsequent inactivation of the cells.

A combined treatment of UV-light and radio frequency electric field for the inactivation of Escherichia coli K-12 in apple juice.

Radio frequency electric fields (RFEF) and UV-light treatments have been reported to inactivate bacteria in liquid foods. However, information on the efficacy of bacterial inactivation by combined treatments of RFEF and UV-light technologies is limited. In this study, we investigated the relationship between cell injury and inactivation of Escherichia coli K-12 in apple juice treated with a combination of RFEF and UV-light. Apple juice purchased from a wholesale distributor was inoculated with E. coli K-12 at 7.8 log CFU/ml, processed with a laboratory scale RFEF unit at 20 kHz, 15 kV/cm for 170 micros at a flow rate of 540 ml/min followed by UV-light treatment (254 nm) for 12s at 25, 30 and 40 degrees C. Treated samples were analyzed for leakage of UV-substances as a function of membrane damage and were plated (0.1 ml) on Sorbitol MacConkey Agar (SMAC) and Trypticase Soy Agar (TSA) plates to determine the viability loss and percent injury. At 40 degrees C, UV-light treatment alone caused 5.8 log reduction of E. coli in apple juice while RFEF caused only 2.8 log reduction. A combination of the two processing treatments did not increase cell injury or leakage of intracellular bacterial UV-substances more than that from the UV-light treatment. Similarly, the viability loss determined was not significantly (P<0.05) different than UV-light treatment alone. However, the UV-substances determined in apple juice treated with RFEF was significantly (P>0.05) different than UV-light treated samples. The results of this study suggest that RFEF treatment causes more injury to the bacterial cells leading to more leakage of intracellular UV-substances than cells treated with UV-light alone. Also, the effect of the two processing treatment combination on bacterial inactivation was not additive.

Growth parameters of Escherichia coli O157:H7, Salmonella spp., Listeria monocytogenes, and aerobic mesophilic bacteria of apple cider amended with nisin-EDTA.

The effect of nisin (0 or 300 IU/mL), ethylenediamine tetraacetic acid (EDTA, 20 mM), and nisin (300 IU)-EDTA (20 mM) on growth parameters, including lag period (LP) and generation time, of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella spp. in the presence or absence of aerobic mesophilic bacteria of apple cider during storage at 5 degrees C for up to 16 days or 23 degrees C for 16 h was investigated. The growth data were analyzed and fitted to the modified Gompertz model. The LP values for aerobic mesophilic bacteria of apple cider (control) and those amended with EDTA and nisin during storage at 5 degrees C were 1.61, 1.76, and 5.45 days, respectively. In apple cider stored at 23 degrees C for 16 h, the LP values for the same bacteria and treatment were 3.24, 3.56, and 5.85 h, respectively. The LP values for E. coli O157:H7 determined in the presence of aerobic mesophilic bacteria of apple cider stored at 23 degrees C for 16 h was 1.48 h, while populations for L. monocytogenes and Salmonella in the same cider declined. In sterile apple cider left at 23 degrees C for 16 h, the LP values for E. coli O157:H7, Salmonella, and L. monocytogenes averaged 2.74, 2.37, and 3.16 h, respectively. The generation time for these pathogens were 0.402, 0.260, and 0.187 log (CFU/mL)/h, respectively. Addition of nisin and EDTA combination caused a decline in lag phase duration and the populations for all pathogens tested, suggesting possible addition of this additive to freshly prepared apple cider to enhance its microbial safety and prevent costly recalls.

Membrane damage and viability loss of Escherichia coli K-12 and Salmonella enteritidis in liquid egg by thermal death time disk treatment.

Bacterial injury, including leakage of intracellular substance and viability loss, of Escherichia coli K-12 (ATCC 23716) and Salmonella Enteritidis (ATCC 13076) inoculated in liquid egg white and liquid whole egg was determined by thermal death time disk. E. coli K-12 and Salmonella Enteritidis were inoculated in liquid egg white and liquid whole egg to a final count of 7.8 log CFU/ml and were thermally treated with thermal death time disks at room temperature (23"C), 54, 56, 58, and 60 degrees C from 0 to 240 s. Sublethal injury, leakage of intracellular substances, and viability loss of E. coli K-12 and Salmonella Enteritidis was investigated by plating 0.1 ml on selective trypticase soy agar containing 3% NaCl, 5% NaCl, sorbitol MacConky agar, and xylose lysine sodium tetradecylsulfate and nonselective trypticase soy agar. No significant (P > 0.05) differences on percent injury or viability loss for E. coli K-12 and Salmonella populations were determined in all samples treated at 23 degrees C. Sublethal injury occurred in E. coli and Salmonella populations at 54 degrees C or above for 120 s. Viability losses for both bacteria averaged 5 log at 54 degrees C or above for 180 s, and the surviving populations were below detection (<10 CFU/ml). Thermal treatment at 40 degrees C and above led to membrane damage, leakage, and accumulation of intracellular ATP from 2 to 2.5 log fg/ml and UV-absorbing substances of 0.1 to 0.39 in the treated samples. These results indicate similar thermal injury/damage on both E. coli and Salmonella membranes as determined by the amount of inactivation, viability loss, and leakage of intracellular substances of bacteria.

Membrane damage and viability loss of Escherichia coli K-12 in apple juice treated with radio frequency electric field.

The need for a nonthermal intervention technology that can achieve microbial safety without altering nutritional quality of liquid foods led to the development of a radio frequency electric fields (RFEF) process. In order to understand the mechanism of inactivation of bacteria by RFEF, apple juice purchased from a wholesale distributor was inoculated with Escherichia coli K-12 at 7.8 log CFU/ml and then treated with RFEF. The inoculated apple juice was passed through an RFEF chamber operated at 20 kHz, 15 kV/cm for 170 micros at a flow rate of 540 ml/min. Treatment condition was periodically adjusted to achieve outlet temperatures of 40, 45, 50, 55, and 60 degrees C. Samples at each outlet temperature were plated (0.1 ml) and the number of CFU per milliliter determined on nonselective and selective agar media was used to calculate the viability loss. Bacterial inactivation and viability loss occurred at all temperatures tested with 55 degrees C treatment, leading to 4-log reductions. No significant effect was observed on bacterial population in control samples treated at 55 degrees C with a low-RFEF (0.15 kV/cm) field strength. These observations suggest that the 4-log reduction in samples treated at 15 kV/cm was entirely due to nonthermal effect. RFEF treatment resulted in membrane damage of the bacteria, leading to the efflux of intracellular ATP and UV-absorbing materials. Populations of injured bacteria recovered immediately (<30 min) from the treated apple juice averaged 0.43 log and were below detection after 1 h of RFEF treatment and determination using selective plates (tryptic soy agar containing 5% sodium chloride). The results of this study suggest that mechanism of inactivation of RFEF is by disruption of the bacterial surface structure leading to the damage and leakage of intracellular biological active compounds.

Effect of time before storage and storage temperature on survival of Salmonella inoculated on fresh-cut melons.

The effects of a waiting period at room temperature ( approximately 22 degrees C) before refrigerating fresh-cut watermelon, cantaloupe and honeydew pieces contaminated with Salmonella on survival of the inoculated pathogen were investigated. Whole cantaloupes, honeydew melons and watermelons were washed with water, and fresh-cut pieces from individual melons were prepared and inoculated with a five strain cocktail of Salmonella at 10(5)cfu/ml. Populations of aerobic mesophilic bacteria, yeast and mold and Pseudomonas spp. were higher for fresh-cut cantaloupe than for fresh-cut watermelon and honeydew immediately after preparation. Populations of Salmonella, aerobic mesophilic bacteria, yeast and mold and Pseudomonas ssp. in fresh-cut melons left at room temperature for up to 5h before refrigeration were significantly (P<0.05) higher than populations in fresh-cut melons stored at 5 degrees C immediately after preparation. Populations of Salmonella recovered in fresh-cut melon after inoculation with the cocktail of Salmonella strains averaged 2 log(10)cfu/g for all three types of melons. Populations in fresh-cut watermelon and honeydew pieces declined by 1 log when stored immediately at 5 degrees C for 12 days, while the populations in fresh-cut cantaloupe did not show significant (P>0.05) changes. Populations of Salmonella in fresh-cut melons stored immediately at 10 degrees C for 12 days increased significantly (P<0.05) from 2.0 to 3.0 log(10)cfu/g in watermelon, 1.9 to 3.0 log(10)cfu/g in honeydew and 2.0 to 3.6 log(10)cfu/g in cantaloupe pieces. Holding freshly prepared, contaminated fresh-cut melon pieces at 22 degrees C for 3h or more prior to refrigerated storage would increase the chances of Salmonella proliferation, especially if the fresh-cut melons were subsequently stored at an abusive temperature.

Effect of sanitizing treatments on removal of bacteria from cantaloupe surface, and re-contamination with Salmonella.

There are many reports of disease due to consumption of cantaloupes contaminated at the surface with enteric pathogens. Salmonella is among the most frequently reported cause of foodborne outbreaks of gastroenteritis in the United States. Research was undertaken to determine the effects of sanitizer and hot water treatments on microbial populations on cantaloupe surfaces and to determine whether prior decontamination of melons by sanitizer treatment affects vulnerability to recontamination by Salmonella. Cantaloupes were sanitized with 200 ppm chlorine or 2.5% hydrogen peroxide solution for 2 min, or hot water (96 degrees C) for 2 min and were held at 5 degrees C for 24 h. Hot water treatments reduced the microbial populations on cantaloupe surface by 4.9 log reduction while H2O2 or chlorine caused approximately 2.6 log unit reduction on cantaloupe surfaces. When sanitized or hot water treated whole cantaloupes were re-inoculated with Salmonella. Higher populations of Salmonella were recovered from sanitized cantaloupes than from the untreated controls; recovery was greater from hot water treated cantaloupes than from cantaloupes treated with chlorine or hydrogen peroxide. The results of this study clearly show that sanitized cantaloupes are susceptible to recontamination if exposed to a human bacterial pathogen during subsequent handling.