[Show abstract][Hide abstract] ABSTRACT: The in vitro antimicrobial effect of cinnamaldehyde and Sporan in combination with acetic acid against Escherichia coli O157:H7 and Salmonella was investigated in Luria-Bertani broth (7 log cfu/mL) containing cinnamaldehyde or Sporan (800 and 1,000 ppm) alone or in combination with 200 ppm acetic acid, and incubated at 37C for up to 6 h. Surviving populations of test pathogens were determined by spiral plating on selective media. E. coli O157:H7 and Salmonella were undetectable after 1 h in the presence of 800 ppm cinnamaldehyde. A 1,000 ppm Sporan significantly reduced Salmonella and E. coli O157:H7 populations by 1.83 and 3.02 log cfu/mL within 2 and 4 h, respectively. Scanning and transmission electron microscopy of oil-treated bacterial cells revealed cell structural damage and leakage of cellular content. Cinnamaldehyde was highly effective against both E. coli O157:H7 and Salmonella whereas the effect of Sporan was dependent on its concentration, exposure time and pathogen.Practical ApplicationsConsumers' preference for fewer chemicals in food has led to research on potential use of natural antimicrobials in food for controlling spoilage and pathogenic microorganisms. The antimicrobial effect of essential oils has been studied earlier. This is the first report on the antimicrobial activity of Sporan, and the combined effect of acetic acid. The results of the present study indicate that the antimicrobial activity of cinnamaldehyde and Sporan against enteric pathogens could be useful in food processing and preservation.
Journal of Food Processing and Preservation 06/2014; 38(3). · 0.45 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Contamination of fresh produce could represent a public health concern because no terminal kill step is applied during harvest or at the processing facility to kill pathogens. In addition, once contaminated, pathogens may internalize into produce and be protected from disinfectants during the postharvest processing step. The objective of the current study was to determine the potential internalization of Escherichia coli O157:H7 into spinach roots and subsequent transfer to the edible parts. Because curli are involved in biofilm formation, we investigated whether their presence influence the internalization of E. coli O157:H7 into spinach. Further, the effect of the spinach cultivar on E. coli O157:H7 internalization was evaluated. Spinach plants were grown in contaminated soil as well as hydroponically to prevent mechanical wounding of the roots and inadvertent transfer of pathogens from the contamination source to the non-exposed plant surfaces. Results showed that E. coli O157:H7 could internalize into hydroponically grown intact spinach plants through the root system and move to the stem and leaf level. The incidence of internalization was significantly higher in hydroponically grown plants when roots were exposed to 7log CFU/mL compared to those exposed to 5log CFU/mL. The effect of cultivar on E. coli O157:H7 internalization was not significant (P>0.05) for the analyzed spinach varieties, internalization incidences showing almost equal distribution between Space and Waitiki, 49.06% and 50.94% respectively. Wounding of the root system in hydroponically grown spinach increased the incidence of E. coli O157:H7 internalization and translocation to the edible portions of the plant. Experimental contamination of the plants grown in soil resulted in a greater number of internalization events then in those grown hydroponically, suggesting that E. coli O157:H7 internalization is dependent on root damage, which is more likely to occur when plants are grown in soil. Curli expression by E. coli O157:H7 had no significant effect on its root uptake by spinach plants.
International journal of food microbiology 12/2013; 173C:48-53. · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Spinach plants were irrigated biweekly with water containing 2.1 log CFU Salmonella/100 ml water (the maximum Escherichia coli MPN recommended by the Leafy Greens Marketing Agreement; LGMA), or 4.1 CFU Salmonella/100 ml water to determine Salmonella persistence on spinach leaves. Green Fluorescent protein expressing Salmonella were undetectable by most-probable number (MPN) at 24 h and 7 days following each irrigation event. This study indicates that Salmonella are unlikely to persist on spinach leaves when irrigation water is contaminated at a level below the LGMA standards. In a parallel study, persistence of Salmonella isolated from poultry or produce was compared following biweekly irrigation of spinach plants with water containing 6 log CFU Salmonella/100 ml. Produce Salmonella isolates formed greater biofilms on polystyrene, polycarbonate and stainless steel surfaces and persisted at significantly higher numbers on spinach leaves than those Salmonella from poultry origin during 35 days study. Poultry Salmonella isolates were undetectable (<1 log CFU/g) on spinach plants 7 days following each irrigation event when assayed by direct plating. This study indicates that Salmonella persistence on spinach leaves is affected by the source of contamination and the biofilm forming ability of the strain.
[Show abstract][Hide abstract] ABSTRACT: Similar to phytopathogens, human bacterial pathogens have been shown to colonize the plant phylloplane. In addition to environmental factors, such as temperature, UV, relative humidity, etc., the plant cultivar and, specifically, the leaf blade morphological characteristics may affect the persistence of enteropathogens on leafy greens. This study was conducted to evaluate the effect of cultivar-dependent leaf topography and the role of strain phenotypic characteristics on Escherichia coli O157:H7 persistence on organic spinach. Spinach cultivars Emilia, Lazio, Space, and Waitiki were experimentally inoculated with the foodborne E. coli O157:H7 isolate EDL933 and its isogenic mutants deficient in cellulose, curli, or both curli and cellulose production. Leaves of 6-week-old plants were inoculated with 6.5 log CFU per leaf in a biosafety level 2 growth chamber. At 0, 1, 7, and 14 days, E. coli O157:H7 populations were determined by plating on selective medium and verified by laser scanning confocal microscopy. Leaf morphology (blade roughness and stoma density) was evaluated by low-temperature and variable-pressure scanning electron microscopy. E. coli O157:H7 persistence on spinach was significantly affected by cultivar and strain phenotypic characteristics, specifically, the expression of curli. Leaf blade roughness and stoma density influenced the persistence of E. coli O157:H7 on spinach. Cultivar Waitiki, which had the greatest leaf roughness, supported significantly higher E. coli O157:H7 populations than the other cultivars. These two morphological characteristics of spinach cultivars should be taken into consideration in developing intervention strategies to enhance the microbial safety of leafy greens.
Journal of food protection 11/2013; 76(11):1829-37. · 1.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There is generally no kill-step when preparing salad vegetables, so there is a greater risk for foodborne illness from contaminated vegetables. Some essential oils have antimicrobial activities and could provide a natural way to reduce pathogens on fresh produce. The objective of this study was to investigate the antimicrobial activity of cinnamon oil wash against Salmonella enterica serotype Newport on organic leafy greens. Organic romaine and iceberg lettuce, and organic baby and mature spinach were inoculated with Salmonella Newport and then dip treated in a phosphate buffered saline (PBS) control and 3 different concentrations (0.1, 0.3, and 0.5% v/v) of cinnamon oil. The treatment time varied at either 1 or 2min, and storage temperature varied at either 4 or 8°C. Samples were collected at days 0, 1, and 3. For romaine and iceberg lettuce, S. Newport was not recovered on day 3 for 2min 0.3% and 0.5% cinnamon oil treatments. For mature spinach, S. Newport was not recovered by day 3 for the 2min 0.3% and 0.5% 4°C treatments. For baby spinach, there was no recovery of S. Newport by day 1 for all 0.5% treatments. Overall, the cinnamon oil treatments were concentration and time dependent with higher concentrations and longer treatment times providing the greatest reduction in S. Newport population on leafy greens. In addition, the treatments had a residual effect with the greatest reduction generally seen on the last day of sampling. Storage temperature did not have a significant effect on the reduction of S. Newport. Based on the results of this study, cinnamon oil has the potential to be used as a treatment option for washing organic baby and mature spinach, and iceberg and romaine lettuces.
International journal of food microbiology 06/2013; 166(1):193-199. · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The objective of this study was to evaluate the effectiveness of oregano oil on four organic leafy greens (Iceberg and Romaine lettuces and mature and baby spinaches) inoculated with Salmonella Newport as a function of treatment exposure times as well as storage temperatures. Leaf samples were washed, dip inoculated with S. Newport (6-log CFU/ml) and dried. Oregano oil was prepared at 0.1, 0.3, and 0.5% concentrations in sterile phosphate buffered saline (PBS). Inoculated leaves were immersed in the treatment solution for 1 or 2 min, and individually incubated at 4 or 8 °C. Samples were taken at day 0, 1, and 3 for enumeration of survivors. The results showed that oregano oil was effective against S. Newport at all concentrations. S. Newport showed reductions from the PBS control of 0.7-4.8 log CFU/g (Romaine lettuce), 0.8-4.8 log CFU/g (Iceberg lettuce), 0.8-4.9 log CFU/g (mature spinach), and 0.5-4.7 log CFU/g (baby spinach), respectively. The antibacterial activity also increased with exposure time. Leaf samples treated for 2 min generally showed greater reductions (by 1.4-3.2 log CFU/g), than those samples treated for 1 min; however, there was minimal difference in antimicrobial activity among samples stored under refrigeration and abuse temperatures. This study demonstrates the potential of oregano oil to inactivate S. Newport on organic leafy greens.
[Show abstract][Hide abstract] ABSTRACT: Abstract Foodborne outbreaks associated with the consumption of fresh produce have increased. In an effort to identify natural antimicrobial agents as fresh produce-wash, the effect of essential oils in reducing enteric pathogens on iceberg and romaine lettuce was investigated. Lettuce pieces were inoculated with a five-strain cocktail of Escherichia coli O157:H7 or Salmonella enterica (5 log CFU/g) and then immersed in a treatment solution containing 5 ppm free chlorine, cinnamaldehyde, or Sporan(®) (800 and 1000 ppm) alone or in combination with 200 ppm acetic acid (20%) for 1 min. Treated leaves were spin-dried and stored at 4°C. Samples were taken to determine the surviving populations of E. coli O157:H7, Salmonella, total coliforms, mesophilic and psychrotrophic bacteria, and yeasts and molds during the 14-day storage period. The effect of treatments on lettuce color and texture was also determined. Cinnamaldehyde-Tween (800 ppm, 800T) reduced E. coli O157:H7 by 2.89 log CFU/g (p<0.05) on iceberg lettuce at day 0; Sporan(®)-acetic acid (1000SV) reduced E. coli O157:H7 and Salmonella on iceberg and romaine lettuce by 2.68 and 1.56 log CFU/g (p<0.05), respectively, at day 0. The effect of essential oils was comparable to that of 5 ppm free chlorine in reducing E. coli O157:H7 and Salmonella populations on iceberg and romaine lettuce throughout the storage time. The natural microbiota on treated lettuce leaves increased during the storage time, but remained similar (p>0.05) to those treated with chlorine and control (water). The texture and the color of iceberg and romaine lettuce treated with essential oils were not different from the control lettuce after 14 days of storage. This study demonstrates the potential of Sporan(®) and cinnamaldehyde as effective lettuce washes that do not affect lettuce color and texture.
Foodborne Pathogens and Disease 12/2012; · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The effect of hydrodynamic pressure (HDP) treatment on the inactivation of Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes attached to beef surface was evaluated. Irradiated beef cubes were inoculated with these individual pathogens (ca. 4 log CFU.g−1) and treated with HDP. Surviving populations of E. coli O157:H7, S. Typhimurium, and L. monocytogenes were determined by spiral-plating on Sorbitol MacConkey agar (SMAC), XLT4 agar, and modified Oxford (MOX) agar, respectively. The HDP treatment significantly reduced E. coli O157:H7 (0.75 log CFU.g−1), S. Typhimurium (1.09 log CFU.g−1), and L. monocytogenes (0.55 log CFU.g−1) from beef surfaces. HDP-induced injury as determined by plating on non-selective media revealed that ca. 0.5 log CFU.g−1 of E. coli O157:H7 and S. Typhimurium cells were injured. Most cell injury occurred at the cell membrane level as observed by transmission electron microscopy (TEM).
[Show abstract][Hide abstract] ABSTRACT: The efficacy of cinnamaldehyde and Sporan for reducing Escherichia coli O157:H7 and Salmonella on spinach leaves was investigated. Spinach leaves were inoculated with a five-strain cocktail of Salmonella or E. coli O157:H7, air dried for ca. 30 min, and then immersed in a treatment solution containing 5 ppm of free chlorine, cinnamaldehyde, or Sporan (800 and 1,000 ppm) alone or in combination with 200 ppm of acetic acid (20%) for 1 min or with water (control). After spin drying, treated leaves were analyzed periodically during 14 days of storage at 4°C for Salmonella, E. coli O157:H7, total coliforms, mesophilic and psychrotrophic bacteria, and yeasts and molds. Treatment effects on color and texture of leaves also were determined. Sporan alone (1,000S), Sporan plus acetic acid (1,000SV), and cinnamaldehyde-Tween (800T) reduced E. coli O157:H7 by more than 3 log CFU/g (P < 0.05), and 1,000SV treatment reduced Salmonella by 2.5 log CFU/g on day 0. E. coli O157:H7 and Salmonella populations on treated spinach leaves declined during storage at 4°C. The 1,000SV treatment was superior to chlorine and other treatments for reducing E. coli O157:H7 during storage. Saprophytic microbiota on spinach leaves increased during storage at 4°C but remained lower on leaves treated with Sporan (800S) and Sporan plus acetic acid (1,000SV) than on control leaves. The color and texture of Sporan-treated leaves were not significantly different from those of the control leaves after 14 days. Sporan plus acetic acid (1,000SV) reduced E. coli O157:H7 and Salmonella on baby spinach leaves without adverse effects on leaf color and texture.
Journal of food protection 03/2012; 75(3):488-96. · 1.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Shiga-toxigenic Escherichia coli O157:H7 outbreaks have been linked to consumption of fresh produce. It is generally recognized that bacterial attachment to vegetal matrices constitutes the first step in contamination of fresh produce. Cellular appendages, such as curli fibers, and cellulose, a constituent of extracellular matrix, have been suggested to be involved in E. coli attachment and persistence in fresh produce. A comparative evaluation was conducted on the ability of Shiga toxin-producing E. coli O157:H7 strains EDL933 and 86-24, linked to two independent foodborne disease outbreaks in humans, and their mutants deficient in curli and/or cellulose expression to colonize and to firmly attach to spinach leaf. Inoculated spinach leaves were incubated at 22°C, and at 0, 24, and 48 h after incubation loosely and strongly attached E. coli O157:H7 populations were determined. Curli-expressing E. coli O157:H7 strains developed stronger association with leaf surface, whereas curli-deficient mutants attached to spinach at significantly (p<0.01) lower numbers. Attachment of cellulose-impaired mutants to spinach leaves was not significantly different from that of curliated strains. The relative attachment strength of E. coli O157:H7 to spinach increased with incubation time for the curli-expressing strains. Laser scanning confocal microscopy (LSCM) analysis of inoculated leaves revealed that curli-expressing E. coli O157:H7 were surrounded by extracellular structures strongly immunostained with anti-curli antibodies. Production of cellulose was not required to develop strong attachment to spinach leaf. These results indicate that curli fibers are essential for strong attachment of E. coli O157:H7 to spinach whereas cellulose is dispensable.
Foodborne Pathogens and Disease 02/2012; 9(2):160-7. · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Salmonella enterica is one of the most common bacterial pathogens implicated in foodborne outbreaks involving fresh produce in the last decade. In an effort to discover natural antimicrobials for use on fresh produce, the objective of the present study was to evaluate the effectiveness of different antimicrobial plant extract-concentrate formulations on four types of organic leafy greens inoculated with S. enterica serovar Newport. The leafy greens tested included organic romaine and iceberg lettuce, and organic adult and baby spinach. Each leaf sample was washed, dip inoculated with Salmonella Newport (10(6) CFU/ml), and dried. Apple and olive extract formulations were prepared at 1, 3, and 5% concentrations, and hibiscus concentrates were prepared at 10, 20, and 30%. Inoculated leaves were immersed in the treatment solution for 2 min and individually incubated at 4°C. After incubation, samples were taken on days 0, 1, and 3 for enumeration of survivors. Our results showed that the antimicrobial activity was both concentration and time dependent. Olive extract exhibited the greatest antimicrobial activity, resulting in 2- to 3-log CFU/g reductions for each concentration and type of leafy green by day 3. Apple extract showed 1- to 2-log CFU/g reductions by day 3 on various leafy greens. Hibiscus concentrate showed an overall reduction of 1 log CFU/g for all leafy greens. The maximum reduction by hydrogen peroxide (3%) was about 1 log CFU/g. The antimicrobial activity was also tested on the background microflora of organic leafy greens, and reductions ranged from 0 to 2.8 log. This study demonstrates the potential of natural plant extract formulations to inactivate Salmonella Newport on organic leafy greens.
Journal of food protection 10/2011; 74(10):1676-83. · 1.83 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Outbreaks associated with leafy greens have focused attention on the transfer of human pathogens to these commodities during harvest with commercial equipment. Attachment of Escherichia coli O157:H7 on new or rusty spinach harvester blades immersed in spinach extract or 10% tryptic soy broth (TSB) was investigated. Bacteriophages specific for E. coli O157:H7 were evaluated to kill cells attached to blade. A cocktail of five nalidixic acid-resistant E. coli O157:H7 isolates was transferred to 25 mL of spinach extract or 10% TSB. A piece of sterilized spinach harvester blade (2×1") was placed in above spinach extract or 10% TSB and incubated at room (22 °C) or dynamic (30 °C day, 20 °C night) temperatures. E. coli O157:H7 populations attached to blade during incubation in spinach extract or 10% TSB were determined. When inoculated at 1 log CFU/mL, E. coli O157:H7 attachment to blades after 24 and 48 h incubation at dynamic temperature (6.09 and 6.37 log CFU/mL) was significantly higher than when incubated at 22 °C (4.84 and 5.68 log CFU/mL), respectively. After 48 h incubation, two blades were sprayed on each side with a cocktail of E. coli O157-specific bacteriophages before scraping the blade, and subsequent plating on Sorbitol MacConkey media-nalidixic acid. Application of bacteriophages reduced E. coli O157:H7 populations by 4.5 log CFU on blades after 2 h of phage treatment. Our study demonstrates that E. coli O157:H7 can attach to and proliferate on spinach harvester blades under static and dynamic temperature conditions, and bacteriophages are able to reduce E. coli O157:H7 populations adhered to blades.
Foodborne Pathogens and Disease 04/2011; 8(4):541-6. · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Salmonella can survive in soil for months to years; consequently, soil can be a preharvest source of contamination of produce. Elimination of Salmonella with natural products and processes such as essential oils is important to prevent infection among consumers. Essential oils (distilled extract from plants) have been mainly evaluated in liquid medium and foods in which minimum inhibitory concentration is determined. However, there are no reports describing the impact of essential oils in soil, especially organic soil. We evaluated essential oils for controlling Salmonella enterica serovars in organic soil. Two essential oils (cinnamaldehyde and eugenol), two bio-pesticides (Ecotrol and Sporan), and an organic acid (20% acetic acid) at 0.5%, 1.0%, 1.5%, and 2.0% were mixed with organic sandy soil and inoculated with six different serovars of S. enterica separately. Soils were incubated at room temperature, and samples obtained at 1, 7, and 28 days were enumerated to determine survival. The bactericidal effect of cinnamaldehyde was evident at 0.5%, 1.0%, 1.5%, and 2.0% and during all times of incubation. Overall, Salmonella Negev was the most sensitive strain to oils resulting in significant reductions compared with other strains. Increases in oil concentration resulted in further reduction of Salmonella with all oils used in the study. Up to six log reductions in Salmonella serovars Typhimurium, Negev, and Newport were found after 1 day when cinnamaldehyde, Ecotrol, eugenol, Sporan, or acetic acid was used at 2% level. This study shows the potential use of essential oils to effectively reduce Salmonella populations in soil. The significant reduction of Salmonella could greatly reduce potential contamination of fresh organic produce inadvertently contaminated by soil.
Foodborne Pathogens and Disease 02/2011; 8(2):311-7. · 2.28 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: This study investigated the ability of five Salmonella enterica serovars to attach to and colonize intact and cut lettuce (Iceberg, Romaine) and cabbage surfaces. Biofilm formation and attachment of Salmonella serovars to intact and cut leaves were determined. Populations of loosely and strongly attached Salmonella were obtained to calculate the attachment strength (S(R)). Biofilm formation, as determined by microtiter plate assay, varied with strain and growth medium used. Salmonella Tennessee and S. Thompson produced stronger biofilms compared to S. Newport, S. Negev, and S. Braenderup. Biofilm formation was also stronger when Salmonella spp. were grown in diluted TSB (1:10). S. Tennessee, which produced strong biofilms, attached to produce surfaces at significantly higher numbers than the populations of S. Negev. Overall, S. Tennessee displayed more biofilm formation in vitro and attached more strongly to lettuce than other serovars. All Salmonella serovars attached rapidly on intact and cut produce surfaces. Salmonella spp. attached to Romaine lettuce at significantly higher numbers than those attached to Iceberg lettuce or cabbage. Salmonella attached preferentially to cut surface of all produce; however, the difference between Salmonella populations attached to intact and cut surfaces was similar (P>0.05). Salmonella attachment to both intact and cut produce surfaces increased with time. The overall attachment strength of Salmonella was significantly lower on cabbage (0.12) followed by Iceberg (0.23) and Romaine lettuce (0.34). Cabbage, intact or cut, did not support attachment of Salmonella as well as Romaine lettuce. Understanding the attachment mechanisms of Salmonella to produce may be useful in developing new intervention strategies to prevent produce outbreaks.
International journal of food microbiology 02/2010; 139(1-2):41-7. · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Soil can be a significant source of preharvest contamination of produce by pathogens. Demand for natural pesticides such as essential oils for organic farming continues to increase. We examined the antimicrobial activity of several essential oils against Escherichia coli O157:H7 in soil. Two essential oils (cinnamaldehyde and eugenol), two bio-pesticides (Ecotrol and Sporan) containing essential oils, and an organic acid (acetic acid) at 0.5%, 1.0%, 1.5% and 2.0%, were mixed with organic sandy soil and inoculated with five different strains of E. coli O157:H7. Soils were incubated at room temperature (22 °C) and samples obtained at 1, 7 and 28 days were enumerated to determine survival. E. coli O157:H7 populations in soil were reduced by up to 5 log cfu/g after 24 h incubation at room temperature with 2% cinnamanaldehyde, Ecotrol, Sporan or vinegar. Reduction in E. coli O157:H7 by eugenol was not significantly different from control. Overall, E. coli O157:H7 strain 4406 was the most sensitive of all the five strains tested and cinnamaldehyde was superior to other treatments in reducing E. coli O157:H7 in soil. In general, increases in essential oil concentrations corresponded to reduced survival of E. coli O157:H7 with all oils used in this study. The results suggest that oils can reduce potential contamination of fresh organic produce inadvertently contaminated by soil.