Bacteriocins are antimicrobial protein produced by certain Gram positive and negative bacteria as a defense mechanism against closely related bacteria competing for the same nutrient or in the same niche. The competition for the same nutrient is supported by the fact that bacteriocins have narrow range of effect and only likely to be effective against closely related bacteria for the same scares resources hence a bacteriocin produced by a Gram positive bacteria will be active against a Gram positive pathogens and a bacteriocin produced by a Gram negative bacteria will be active against Gram negative pathogens. This is due to the difference in cell wall composition, they are either bacteriocidal or bacteriostatic Bacteriocins have been used for thousands of years for food preservation unknowingly to man, they are considered advantageous not only to the producing bacteria, but it's now been used by the food industry as a tool to control both spoilage and pathogenic bacteria in food, in a natural manner which is acceptable to the consumer. With a lot of research been carried out on bacteriocins produced by Gram positive bacteria, antagonist to Gram positive food borne pathogens, little is known about bacteriocins produced by Gram negative bacteria which would be active against Gram negative food borne pathogens that predominate in produce. The objective of my research therefore is to screen for antimicrobial antagonist to Gram negative food borne pathogens (Escherichia coli O157:H7 and Salmonella Enteritidis) from produce, to determine an appropriate screening method, to carry out a preliminary characterization of antagonist discovered and also to determine antimicrobial spectrum of antagonist found. Lettuce was screened for antimicrobial antagonist against Gram negative pathogen (Escherichia coli O157:H7 and Salmonella Enteritidis) which were used as indicator strains With over 5000 colonies screen, 1 colony (Serratia plymuthica) was discovered to be antagonistic against these indicator strain. Further screening of cell free extract using the spot test method showed that extract from Serratia plymuthica grown alone in TSBYE showed antagonist activity against indicator strain with a little clearing on the spot of extract dropped. But extract of a co-culture of Serratia plymuthica and either Escherichia coli O157:H7 or Salmonella Enteritidis showed a more obvious clearing around spotted zone, which further indicates antagonism against indicator strains. Preliminary heat test indicates antagonist compound to be heat stable at 60oC for 30mins, 100oC for 30minutes and 60mins and 121oC for 20minites, and antagonist compound possessed antagonist activity against other strains of Escherichia coli when tested.

For years the issue of food waste has been recognized and quantified; however, food safety issues often go unrecognized as a source of food waste. One objective of this research is to estimate quantities and monetary value of fruits and vegetables implicated in food safety recalls, and thus wasted. Using publicly available data we identified all recalls involving vegetable or fruit commodities contaminated with Listeria monocytogenes, pathogenic E. coli, or Salmonella during 2015-2018. When quantities were provided, monetary value of recalled product was calculated using USDA ERS 2016 average retail prices. Although data limitations only allowed analysis of 17% of the recalls that met the criteria of this study, we estimated an annual loss of 38 million pounds and $61 million in revenue. Overall this shows that food safety issues can result in food waste, therefore mitigation strategies are needed. There are many ways that produce can become contaminated, however contaminated soils are a potential source of produce contamination and treatments to mitigate this risk while maintaining soil health is lacking. Current biofumigation methods that use glucosinolate hydrolysis products in mustard seed meal to control plant pathogens could also be effective against foodborne pathogens in soil. The purpose of this research is to determine the fate of E. coli O157:H7, Salmonella, and L. monocytogenes in soil treated with Brassica spp seed meal and plant material. Seed meals were successful in reducing pathogen concentrations in soil, significant reductions (p < 0.05) of E. coli O157:H7, L. monocytogenes, and Salmonella were observed in soil over 72 hours with the addition of 1.0 and 1.5 g of mustard seed meal. Increasing the seed meal concentration did not significantly (p > 0.05) increase the observed log reduction for L. monocytogenes or Salmonella, reductions ranged from 5.6 – 5.9 log CFU/g. However, for E. coli O157:H7 seed meal concentration was significant (p < 0.05). A 5.7 log CFU/g decrease was observed when 1.5 g of seed meal was used which was larger than 3.5 log CFU/g reduction observed with 1.0 g. Findings suggest that biofumigation with mustard seed meal could potentially be used to reduce E. coli O157:H7, L. monocytogenes, and Salmonella in contaminated soil. However, the use of plant material was not as successful as the use of the processed seed meals. In soil or in the absence of soil Brassica spp. plant material at 10% 15%, and 75% significantly increased E. coli O157:H7, L. monocytogenes, and Salmonella concentrations (p < 0.05). The results of these studies support literature indicating Brassica spp. processed plant products, like seed meals or extracts may be a more effective strategy in reducing human pathogen concentrations in contaminated agricultural soils. While the process of Biofumigation using Brassica spp. cover crops has been successful in eliminating plant pests from agricultural soils, due to its low isothiocyanate release efficiency and reactivity in soil organic matter, it may not be sufficient as a soil decontamination method against human pathogens.

This study focused on evaluating the antimicrobial activity of chlorine dioxide (ClO2) and different plant-based antimicrobials (carvacrol, thymol, and eugenol as bioactive compounds and muscadine extract- ME and blueberry extract- BBE as plant extracts) against selected foodborne pathogens under in vitro conditions and on produce (spinach and blueberries). In addition, bacterial microbiota associated with blueberries and blueberry farm environments from three different regions: Cundinamarca, Colombia; Mississippi, United States; and Guadalajara, Mexico, was determined. Under in vitro conditions, carvacrol and thymol were more effective (lower MICs and MBCs) than eugenol against Salmonella spp., Escherichia coli O157:H7 (MIC=MBC=0.2 mg/ml), and Listeria monocytogenes (MIC=MBC=0.4 mg/ml). Both plant extracts had the same MIC and MBC for Salmonella spp. while BBE had stronger bactericidal effect on Escherichia coli O157:H7 (MBC=150 mg/ml) and ME on L. monocytogenes (MBC=100 mg/ml). ClO2 had stronger bacteriostatic effect on L. monocytogenes (MIC=1 ppm) than on the Gram-negative bacteria (MIC=3 ppm). For the produce study, 300 mg/ml ME exerted the highest (P ≤ 0.05) E. coli O157:H7 reduction (4.5 log CFU/g at day 1) on spinach, and 3 ppm ClO2 + 300 mg/ml ME had the highest (P ≤ 0.05) L. monocytogenes reduction on both (4.5-5.6 log CFU/g). There was a similar (P>0.05) E. coli O157:H7 reduction on blueberries, regardless of antimicrobial treatment. For the microbiota study, Proteobacteria was the most abundant phylum in blueberries, soil, and water, with the exception of fruits from Mexico. Blueberries grown on the different regions shared two predominant genera: Heliorestis (10.5-47.4%) and Thiomonas (5.0-9.1%). Nonetheless, alpha and beta diversity revealed that blueberry microbiota structures were distinctive. PCoA plots revealed that within regions the microbial composition distribution was different (P ≤0.05) among fruits, soil, and water. Based on the results, ME combined with ClO2, could represent an antimicrobial alternative against foodborne pathogens for the produce industry. Furthermore, the study of the microbiota provided a good understanding on the bacterial community profile in blueberries and the blueberry farming environment across regions.

A bifidobacteria mix (nine strains) was evaluated for its effect on the growth and survival of Escherichia coli O157:H7 (ATCC 43890) in 11% NFDM and MRS broth under the optimum growth conditions for E. coli O157:H7 growth (37¡ãC, aerobic). Preliminary experiments were conducted to obtain the growth curves of the 9 strains of bifidobacteria and E. coli O157:H7 and confirm the inhibitory effect of acidity on E. coli O157:H7 (pH of media adjusted to 3.8). Acidapted E. coli O157:H7 showed no difference in resistance toward bifidobacteria (P>0.05) when compared to the non-acid adapted one. Escherichia coli O157:H7 did not survive in the supernatant of the bifodbacteria mix collected after incubation (37¡ãC) with aerobic shaking (8 h). However, the pathogen was able to grow after the pH of the supernatant was adjusted to 6.50 (pH of fresh MRS broth). Results suggest that a high content of bifidobacteria has a strong inhibitory effect on E. coli O157:H7, in part due to the low pH. However, products from bifidobacteria may also exert inhibitory effects.

No description available.

No description available.

Fresh produce is increasingly associated with foodborne outbreaks. In order to develop effective intervention and measures to reduce microbial risks, it is essential to attain a better understand the mechanisms of attachment of foodborne pathogens to fruits and vegetables. Using lettuce as a model, the attachment of Escherichia coli O157:H7 to produce surfaces was studied. Strains expressing various extracellular proteins (curli, O157-antigen, and intimin) known to influence attachment of E. coli to intestinal cells were evaluated for their physicochemical properties and ability to adhere to cut edge and whole leaf lettuce. Escherichia coli O157:H7 strains included: 0018, 43894 and 43895 (curli producing and non-producing); 86-24 (WT), F-12 (O157-antigen negative), pRFBE (O-antigen replaced on plasmid); and 86-24, 86-24Ã eae10 (intimin negative). The eleven strains were surveyed for their hydrophobicity and cell charge using hydrophobic interaction chromatography (HIC) and electrostatic interaction chromatography (ESIC) techniques. Iceberg lettuce squares (2 x 2 cm) were inoculated with E. coli O157:H7 strains separately (7.0 log CFU/square) and dried in a laminar flow hood. Lettuce was sampled before (unrinsed) and after being rinsed twice with sterile de-ionized water (rinsed). Strips (2 mm wide) of each cut edge of the lettuce were aseptically removed. Cut-edge and whole-leaf samples were homogenized and spiral plated onto Luria-Bertani agar, supplemented with nalidixic acid (50ppm), to assess levels of bacteria remaining on the lettuce leaf after rinsing. The rinse steps were not effective in significantly removing bacteria from lettuce (p>0.05). Curli-producing and non-producing strains preferentially attached to cut edge versus the whole leaf portions of lettuce (p<0.05); however the 86-24 strains showed no preference for attachment. With the exception of 0018 curli-producing and non-producing strains, presence/absence of extracellular proteins surveyed did not influence attachment of E. coli O157:H7 to either cut edge or whole leaf lettuce. There was significantly greater attachment of the curli-producing 0018 strain over the curli non-producing 0018 strain to cut and whole lettuce surfaces (p<0.05). Production of curli and O-polysaccharide significantly increased (p<0.05) the cell's overall hydrophobicity of the cell; however this did not affect attachment (p<0.05). The overall cell charge of all strains was negative; however, charge did not affect attachment of E. coli O157:H7 to lettuce. The presence of extracellular appendages (curli, O157-antigen, intimin) as well as hydrophobicity and cell charge properties had no affect on attachment of the cell to lettuce. / Ph. D.

The antibacterial efficacy of hydrogen peroxide treatments in four fruit juices was determined. Preservative free apple cider, white grape, and purple grape juice were inoculated with ~ 6.4 log CFU/ml of a five strain, acid adapted, nalidixic acid resistant E. coli O157:H7 cocktail. Orange juice was inoculated with a comparable Salmonella spp. cocktail. In the first study, 0.017% and 0.012% H₂O₂ was added in combination with 0.1% and 0.3% of the dominant organic acid (OA) to 4°C and 25°C juices, with samples taken each day for 21 days. H₂O₂ was a significant factor in all juices (p < 0.05) except white grape (lack of data), and both 0.017% H₂O₂ treatments reduced counts in apple cider, orange juice, and white grape to undetectable numbers within 48 hrs as cultured on tryptone soy agar + 0.05% nalidixic acid (TSAN). Treatments in purple grape juice were less effective overall, and more dependent on OA concentration (p < 0.001) than H₂O₂. There were instances where bacterial survival in apple cider, purple grape, and orange juice continued for 21 days after treatment, and sometimes outlasted the control. These occurrences were dependent on temperature (25°C) and H₂O₂ (0.012%), but not on OA. However, OA concentration was a significant factor (p < 0.05) overall in apple cider and purple grape juice, but not in orange juice. In the second study, 0.015% and 0.03% H₂O₂ was added to 10, 25, and 40°C apple cider and orange juice inoculated with 6.4 log CFU/ml E. coli O157:H7 and Salmonella spp. respectively. Only 0.03% H₂O₂ was effective in reducing counts to undetectable numbers in both juices. However, both temperature and H₂O₂ were significant factors (p < 0.0001) in bacterial destruction, with 0.03% H₂O₂ at 40°C giving undetectable numbers at ≤ 3 and ≤ 6 hours in orange juice and apple cider respectively. It has been demonstrated that at ~ ≥ 0.017%, H₂O₂ can provide a 5 log reduction of these pathogens in fruit juice. Increasing temperature and organic acid concentration can improve its rate of effectiveness in certain juices. However, sensory concerns may negate its use in some products. / Master of Science

Cinnamic acid (CA) is a naturally occurring organic acid that is found in some fruits and a number of spices. CA has antimicrobial activity against certain spoilage microorganisms and pathogenic bacteria. However, the acid is poorly soluble in water. Cyclodextrin molecules have a hydrophobic cavity that allows them to serve as a host for insoluble molecules in aqueous matrices. This study was conducted to determine if the aqueous solubility of cinnamic acid could be improved via complexation with α- or β-cyclodextrins, and if these complexes could be used to control bacterial pathogens in juices. Based upon phase solubility analysis, α-cyclodextrin was chosen as the host molecule for the remainder of this study. In complex with α-cyclodextrin, the solubility of cinnamic acid increased from approximately 400 mg/L to 3800 mg/L. Prepared cinnamic acid complexed with α-cyclodextrin was aseptically added (400 mg/L and 1000 mg/L) to orange juice inoculated with a Salmonella enterica (7 log CFU/mL) and apple cider inoculated with Escherichia coli O157:H7 (7 log CFU/mL). Cider and orange juice samples were extracted on day 0 and at 24 h intervals for seven days and spread plated onto Tryptic Soy Agar. Cinnamic acid was effective for reducing populations of both bacterial pathogens in juice. Populations of E. coli O157:H7 in the apple cider were significantly reduced after 7 days at 25.6 ± 0.42°C at concentrations of 400 mg/L (5-log CFU/mL reduction) and 1000 mg/L (6-log CFU/mL reduction) cyclodextrin-cinnamic acid. S. enterica counts were also reduced in orange juice at 4° C treated with 400 mg/L (2.7-log CFU/mL reduction) and 1000 mg/L (3.2-log CFU/mL reduction) complexed cinnamic acid. The much improved solubility of this compound provides food processors with greater flexibility in using cinnamic acid in their product formulations. / Master of Science

Master of Science / Food Science / Kelly J.K. Getty / James L. Marsden / Escherichia coli O157:H7 outbreaks have been linked to leafy green produce and bagged spinach. The objective of this study was to evaluate a Photohydroionization (PHI) panel (novel advanced oxidation technology) and varying concentrations of lactic acid washes for controlling E. coli O157:H7 on baby spinach. Leaves were dip inoculated with a five-strain cocktail of E. coli O157:H7 inoculum having a concentration between 5-6 log CFU/ml. Leaves were submerged in inoculum for 30 s and dried for 1 h. Non-inoculated and inoculated leaves were washed for 30 s in food grade lactic acid diluted to concentrations of 0.5, 1.0, or 2.0% and allowed to dry for 10 min. For PHI treatment, leaves were placed under the PHI panel and treated for 1, 2, or 5 min on both sides for total treatment times of 2, 4 or 10 min. Following treatments, leaves were either sealed in low-density polyethylene bags or enumerated. Samples were enumerated at 0, 3, 7, 10, and 14 days following inoculation. Ten gram samples were diluted with sterile peptone and stomached for one min, and then 0.1 ml was plated onto sorbitol MacConkey agar with cefixime and tellurite plates that were incubated at 37°C for 24 h. For lactic acid treatments, E. coli O157:H7 populations were different (P < 0.05) compared to the control. There was no difference (P > 0.05) due to sampling time so sampling times where pooled together for each lactic acid concentration of 0.5, 1.0, and 2.0% and resulted in 2.01, 2.78, and 3.67 log CFU/g reductions, respectively. Leaves treated with 1.0% and 2.0% lactic acid had color degradation and were organoleptically unacceptable by day 14. When leaves were treated with PHI for 1, 2, or 5 min per side, E. coli O157:H7 populations were reduced 1.6, 1.49, or 1.95 log CFU/g, respectively. Leaves treated with PHI were not different from one another, but were different (P < 0.05) from the positive control. No color change occurred in leaves treated with PHI. The PHI panel and lactic acid washes of 0.5% or higher are effective in reducing E. coli O157:H7 in baby spinach.