Mu Ye

Control of Listeria monocytogenes on ham steaks by antimicrobials incorporated into chitosan-coated plastic films

Contamination of ready-to-eat (RTE) meat products such as ham steaks with Listeria monocytogenes has been a concern for the meat processing industry. The objective of this study was to evaluate the antilisterial efficacy of chitosan-coated plastic films alone or incorporating five generally recognized as safe (GRAS) antimicrobials. Effect of chitosan-coated plastic film on the growth of L. monocytogenes was first investigated in an aqueous system of culture medium broth and chitosan-coated films were able to inhibit the growth of L. monocytogenes in a concentration-dependent manner. However, chitosan-coated plastic films were not able to control the growth of L. monocytogenes on ham steaks. Therefore, five GRAS antimicrobials were subsequently incorporated into chitosan-coated plastic films to enhance their antilisterial effectiveness. Ham steaks were surface-inoculated with a five-strain cocktail of L. monocytogenes and then packaged in chitosan-coated plastic films containing 500 IU/cm(2) of nisin, 0.01 g/cm(2) of sodium lactate (SL), 0.0025 g/cm(2) of sodium diacetate, 0.003 g/cm(2) of potassium sorbate (PB), or 0.001 g/cm(2) of sodium benzoate (SB). The samples were stored at room temperature (ca. 20 degrees C) for 10 days. Incorporating antimicrobials into chitosan-coated plastic films slowed down or inhibited the growth of L. monocytogenes. The chitosan-coated plastic film containing SL was the most effective antimicrobial film and its efficacy against L. monocytogenes on ham steaks was evaluated during 12-week storage at 4 degrees C. The film showed excellent long-term antilisterial effect with the counts of L. monocytogenes being slightly lower than the initial inoculum. Chitosan-coated plastic films containing 0.001 g/cm(2) of SL have a potential to be used on ham steaks to control L. monocytogenes.

Effectiveness of chitosan-coated plastic films incorporating antimicrobials in inhibition of Listeria monocytogenes on cold-smoked salmon

The objective of this study was to evaluate the efficacy of chitosan-coated plastic films incorporating five Generally Recognized as Safe (GRAS) antimicrobials (nisin, sodium lactate (SL), sodium diacetate (SD), potassium sorbate (PS) and sodium benzoate (SB)) against Listeria monocytogenes on cold-smoked salmon. Salmon samples were surface-inoculated with a five-strain cocktail of L. monocytogenes and packaged in chitosan-coated plastic films containing 500 IU/cm(2) of nisin, 9 mg/cm(2) of SL, 0.5 mg/cm(2) of SD, 0.6 mg/cm(2) of PS, or 0.2 mg/cm(2) of SB, and stored at room temperature (ca. 20 degrees C) for 10 days. The film incorporating SL was the most effective, completely inhibiting the growth of L. monocytogenes during 10 days of storage. L. monocytogenes in samples packaged in the other four antimicrobial films grew, but the increase in counts was lower than the control. The antilisterial efficacy of films containing lower concentrations of SL (2.3 mg/cm(2) and 4.5 mg/cm(2)) and binary combinations SL, PS, SD, SB and nisin were subsequently evaluated. Among all the treatments, chitosan-coated plastic films with 4.5 mg/cm(2) SL, 4.5 mg/cm(2) SL-0.6 mg/cm(2) PS and 2.3 mg/cm(2) SL-500 IU/cm(2) nisin were the most effective. These three most effective antimicrobial films were then tested at refrigerated temperature. They completely inhibited the growth of L. monocytogenes on smoked salmon for at least 6 weeks. Chitosan-coated plastic films containing 4.5 mg/cm(2) SL can potentially assist the smoked-salmon processing industry in their efforts to control L. monocytogenes.

Potential antimicrobials to control Listeria monocytogenes in vacuum-packaged cold-smoked salmon pâté and fillets

In the wake of recent outbreaks associated with Listeria monocytogenes in ready-to-eat foods and an increasing desire for minimally processed foods, there has been a burgeoning interest in the use of natural antimicrobials by the food industry to control this pathogen. The minimum inhibitory concentrations (MICs) of nisin and salts of organic acids (sodium lactate (SL), sodium diacetate (SD), sodium benzoate (SB), and potassium sorbate (PS)) against twelve strains of L. monocytogenes in a TSBYE broth medium at 35 degrees C were determined. The MICs were strain-dependent and fell in the range of 0.00048-0.00190% for nisin, 4.60-5.60% for SL, 0.11-0.22% for SD, 0.25-0.50% for SB and 0.38-0.75% for PS, respectively. The two most antimicrobial-resistant strains were used as a cocktail in the following experiments to represent a worst case scenario. The five antimicrobials alone and in binary combinations were screened for their efficacy against the two-strain cocktail in TSBYE at sub-MIC and sub-legal levels at 35 degrees C. Seven effective antimicrobial treatments were then selected and evaluated for their long-term antilisterial effectiveness in cold-smoked salmon pâté and fillets during refrigerated storage (4 degrees C) of 3 and 6 weeks, respectively. The two most effective antimicrobial formulations for smoked salmon pâté, 0.25% SD and 2.4% SL/0.125% SD, were able to inhibit the growth of L. monocytogenes during the 3 weeks of storage. Surface application of 2.4% SL/0.125% SD was the most effective treatment for smoked salmon fillets which inhibited the growth of L. monocytogenes for 4 weeks. These antimicrobial treatments could be used by the smoked salmon industry in the U.S. and Europe in their efforts to control L. monocytogenes as they are effective against even the most antimicrobial-resistant strains tested in this study.

Bioactive alginate coatings to control Listeria monocytogenes on cold-smoked salmon slices and fillets

The relatively high incidence of Listeria monocytogenes in cold smoked salmon (CSS) is of concern as CSS is a ready-to-eat product. No post-processing measures are currently available to control this pathogen in CSS. The objective of this study was to develop an effective antimicrobial edible coating containing organic salts to control the growth of L. monocytogenes in CSS slices and fillets. An in-house made formulation consisting of sodium lactate (SL, 0-2.4%) and sodium diacetate (SD, 0-0.25%) as well as 2.5% OptiForm (a commercial formulation of SL and SD) were incorporated into five edible coatings: alginate, kappa-carrageenan, pectin, gelatin or starch. The coatings were applied onto the surface of CSS slices inoculated with L. monocytogenes to an inoculum level of 500 CFU/cm(2) ( approximately 3 log CFU/g) and stored at room temperature (22 degrees C) for 6 days. Alginate coating was found to be the most effective carrier for the various antimicrobial treatments in inhibiting the growth of L. monocytogenes. In the second phase of the study, CSS slices and fillets inoculated with the pathogen at a level of 500 CFU/cm(2) were coated with alginate incorporating the in-house made and the commercial (OptiForm) SL/SD based formulations and stored for 30 days at 4 degrees C. When cold-smoked salmon slices and fillets were stored at 4 degrees C, alginate coatings supplemented with 2.4%SL/0.25%SD and the commercial product OptiForm significantly delayed the growth of L. monocytogenes during the 30-day storage with final counts reaching 4.1 and 3.3 log CFU/g (slices) and 4.4 and 3.8 log CFU/g (fillets), respectively, while the counts in their untreated counterparts were significantly higher (P<0.05) reaching 7.3 and 6.8 log CFU/g for slices and fillets, respectively. Therefore, this study demonstrates the effectiveness of using an alginate-based coating containing lactate and diacetate to control the growth of L. monocytogenes to enhance the microbiological safety of filleted and sliced smoked salmon.

Pressure inactivation of Tulane virus, a candidate surrogate for human norovirus and its potential application in food industry

Human norovirus (HuNoV) is the leading causative agent for foodborne disease. Currently, studies of HuNoV usually rely on surrogates such as murine norovirus (MNV) due to the lack of a suitable cell culture system and a small animal model for HuNoV. Tulane virus (TV), a monkey calicivirus, is a cultivable enteric calicivirus that not only recognizes the same receptors as HuNoV, but is also genetically closely related to HuNoV. In this study, we determined the pH stability of TV and MNV-1, as well as the effect of high hydrostatic pressure (HHP) on inactivating both viruses in aqueous media, blueberries and oysters. We demonstrated that both TV and MNV-1 were very stable under an acidic environment. They were more resistant to pressure at an acidic environment than at neutral pH. Pressure treatment of 600 MPa for 2 min at different temperatures (4, 21 and 35 °C) barely caused any reduction of TV, as well as MNV-1, on un-wetted (dry) blueberries. However, both TV and MNV-1 on blueberries were successfully inactivated by a pressure of ≤400 MPa when blueberries were immersed in phosphate-buffered saline during HHP. Pressure inactivation of both TV and MNV-1 in blueberries and oysters increased as sample temperature decreased in the order of 4>21>35 °C. TV was more sensitive to pressure than MNV-1 for the three matrices tested, culture media, blueberries and oysters. This study provides important information on the use of TV as a surrogate for HuNoV study. Results obtained from this study lay a foundation for designing effective HHP treatments for inactivation of HuNoV in high-risk foods such as berries and oysters.

Inactivation of Escherichia coli O157:H7 and Salmonella spp. in strawberry puree by high hydrostatic pressure with/without subsequent frozen storage

The objectives of this study were to investigate the survival of Escherichia coli O157:H7 and Salmonella spp. in frozen strawberry puree and to assess the application of high pressure processing (HPP) to decontaminate strawberry puree from both pathogens. Fresh strawberry puree was inoculated with high (~6 log CFU/g) and low (~3 log CFU/g) levels of E. coli O157:H7 or Salmonella spp. and stored at -18°C for 12 weeks. Both pathogens were able to persist for at least 4weeks and samples with high inoculums were still positive for both pathogens after 12 weeks. Pressure treatment of 450 MPa for 2 min at 21°C was able to eliminate both pathogens in strawberry puree. Frozen storage at -18°C after pressure treatment substantially enhanced the inactivation of both pathogens and 4-8 days of frozen storage was able to reduce the pressure level needed for elimination of both pathogens to 250-300 MPa. Natural yeasts and molds in strawberry puree were effectively reduced by pressure of 300 MPa for 2 min at 21°C. No adverse impacts on physical properties such as color, soluble solids content, pH and viscosity of strawberry puree was found for pressure-treated samples. Therefore, the treatment of 300 MPa for 2 min at 21°C followed by 4 days frozen storage at -18°C was recommended for the minimal processing of strawberry puree with great retention of fresh-like sensory properties. HPP could be a promising alternative to traditional thermal processing for berry purees.

Potential application of high hydrostatic pressure to eliminate Escherichia coli O157:H7 on alfalfa sprouted seeds

Sprouts eaten raw are increasingly being perceived as hazardous foods as they have been implicated in Escherichia coli O157:H7 outbreaks where the seeds were found to be the likely source of contamination. The objective of our study was to evaluate the potential of using high hydrostatic pressure (HHP) technology for alfalfa seed decontamination. Alfalfa seeds inoculated with a cocktail of five strains of E. coli O157:H7 were subjected to pressures of 500 and 600 MPa for 2 min at 20 degrees C in a dry or wet (immersed in water) state. Immersing seeds in water during pressurization considerably enhanced inactivation of E. coli O157:H7 achieving reductions of 3.5 log and 5.7 log at 500 and 600 MPa, respectively. When dry seeds were pressurized, both pressure levels reduced the counts by <0.7 log. To test the efficacy of HHP to completely decontaminate seeds whilst meeting the FDA requirement of 5 log reductions, seeds inoculated with a ~5 log CFU/g of E. coli O157:H7 were pressure-treated at 600 and 650 MPa at 20 degrees C for holding times of 2 to 20 min. A >5 log reduction in the population was achieved when 600 MPa was applied for durations of > or =6 min although survivors were still detected by enrichment. When the pressure was stepped up to 650 MPa, the threshold time required to achieve complete elimination was 15 min. Un-inoculated seeds pressure-treated at 650 MPa for 15 min at 20 degrees C successfully sprouted achieving a germination rate identical to untreated seeds after eight days of sprouting. These results therefore demonstrate the promising application of HHP on alfalfa seeds to eliminate the risk of E. coli O157:H7 infections associated with consumption of raw alfalfa sprouts.

Effectiveness and stability of plastic films coated with nisin for inhibition of Listeria monocytogenes.

Plastic films were coated with a cellulose-based carrier solution containing nisin, a natural antimicrobial peptide with the potential to inhibit growth of food spoilage and pathogenic microorganisms such as Listeria monocytogenes. Five commercial plastic films with different chemical compositions and surface properties were compared in this study: low-density polyethylene, ethylene-vinyl acetate copolymer, and three types of ethylene-methacrylic acid copolymers: Surlyn 1601, Nucrel 0403, and Nucrel 0903. The films were coated with nisin at a concentration of 1000 IU/cm2. Nisin-coated films were stored at room temperature (21 degrees C) and at 4 degrees C and analyzed weekly for 12 weeks. Antimicrobial activity of the different nisin-coated films against a nisin indicator strain, Lactococcus lactis subsp. cremoris ATCC 14365, and against L. monocytogenes ATCC 19115 was assessed using an inhibition zone assay. Nisin incorporated into the films was recovered by a boiling and extraction procedure, and its activity was quantified using an agar well diffusion assay. Film type did not have any significant effect on the antimicrobial activity of the nisin-coated films (P < 0.05); all five film types had comparable inhibition zones on both assays. The films maintained stable activity for the duration of the study, both at room temperature and refrigeration. The results of this study demonstrate that commercially available packaging films can be coated with nisin and the resulting antimicrobial films can be conveniently stored at room temperature with no adverse effect on nisin activity.

Inactivation of Vibrio parahaemolyticus and Vibrio vulnificus in oysters by high-hydrostatic pressure and mild heat.

Several recent outbreaks associated with oysters have heightened safety concerns of raw shellfish consumptions, with the majority being attributed to Vibrio spp. The objective of this study was to determine the effect of high-hydrostatic pressure (HHP) followed by mild heating on the inactivation of Vibrio parahaemolyticus and Vibrio vulnificus in live oysters. Inoculated oysters were randomly subjected to: a) pressurization at 200-300 MPa for 2 min at 21 °C, b) mild heat treatment at 40, 45 or 50 °C for up to 20 min and c) pressure treatment of 200-300 MPa for 2 min at 21 °C followed by heat treatment at 40-50 °C. Counts of V. parahaemolyticus and V. vulnificus were then determined using the most probable number (MPN) method. Pressurization at 200-300 MPa for 2 min resulted in various degrees of inactivation, from 1.2 to >7 log MPN/g reductions. Heat treatment at 40 and 45 °C for 20 min only reduced V. parahaemolyticus and V. vulnificus by 0.7-2.5 log MPN/g while at 50 °C for 15 min achieved >7 log MPN/g reduction. HHP and mild heat had synergistic effects. Combinations such as HHP at 250 MPa for 2 min followed by heat treatment at 45 °C for 15 min and HHP at 200 MPa for 2 min followed by heat treatment at 50 °C for 5 min reduced both V. parahaemolyticus and V. vulnificus to non-detectable levels by the MPN method (<3 MPN/g). HHP at ≥275 MPa for 2 min followed by heat treatment at 45 °C for 20 min and HHP at ≥200 MPa for 2 min followed by heat treatment at 50 °C for 15 min completely eliminated both pathogens in oysters (negative enrichment results). This study demonstrated the efficiency of HHP followed by mild heat treatments on inactivation of V. parahaemolyticus and V. vulnificus and could help the industry to establish parameters for processing oysters.

Inactivation of human norovirus in contaminated oysters and clams by high hydrostatic pressure.

ABSTRACT Human norovirus (NoV) is the most frequent causative agent of food-borne disease associated with shellfish consumption. In this study, the effect of high hydrostatic pressure (HHP) on inactivation of NoV was determined. Genogroup I.1 (GI.1) or genogroup II.4 (GII.4) NoV was inoculated into oyster homogenates and treated at 300 to 600 MPa at 25, 6, and 1°C for 5 min. After HHP, samples were treated with RNase and viral particles were extracted with porcine gastric mucin (PGM)-conjugated magnetic beads (PGM-MBs). Viral RNA was then quantified by real-time reverse transcription (RT)-PCR. Since PGM contains histo-blood group-like antigens, which can act as receptors for NoV, deficiency for binding to PGM is an indication of loss of infectivity of NoV. After binding to PGM-MBs, RT-PCR-detectable NoV RNA in oysters was reduced by 0.4 to >4 log10 by HHP at 300 to 600 MPa. The GI.1 NoV was more resistant to HHP than the GII.4 NoV (P < 0.05). HHP at lower temperatures significantly enhanced the inactivation of NoV in oysters (P < 0.05). Pressure treatment was also conducted for clam homogenates. Treatment at 450 MPa at 1°C achieved a >4 log10 reduction of GI.1 NoV in both oyster and clam homogenates. It is therefore concluded that HHP could be applied as a potential intervention for inactivating NoV in raw shellfish. The method of pretreatment of samples with RNase, extraction of viral particles using PGM-MB binding, and quantification of viral RNA using RT-PCR can be explored as a practical means of distinguishing between infectious and noninfectious NoV.