Ifigenia Geornaras

Overview of current meat hygiene and safety risks and summary of recent studies on biofilms, and control of Escherichia coli O157:H7 in nonintact, and Listeria monocytogenes in ready-to-eat, meat products

As meat consumption is increasing around the world, so do concerns and challenges to meat hygiene and safety. These concerns are mostly of a biological nature and include bacterial pathogens, such as Escherichia coli O157:H7, Salmonella and Campylobacter in raw meat and poultry, and Listeria monocytogenes in ready-to-eat processed products, while viral pathogens are of major concern at foodservice. A major goal of scientists, industry, public health and regulatory authorities is to control pathogenic microorganisms and improve meat product hygiene and safety within a country and internationally. This paper is not a comprehensive or critical review of the scientific literature on the broad area of meat hygiene and safety, but it provides an overview of major current meat hygiene and safety issues, and then a summary of studies on biofilm formation by pathogens, control of E. coli O157:H7 in nonintact meat products, and control of L. monocytogenes in ready-to-eat meat products, conducted at the Center for Meat Safety & Quality and Food Safety Cluster of Colorado State University in recent years.

Control of Listeria monocytogenes on Frankfurters with Antimicrobials in the Formulation and by Dipping in Organic Acid Solutions

The antilisterial activity of sodium lactate (SL) and sodium diacetate (SD) was evaluated in a frankfurter formulation and in combination with a dipping treatment into solutions of lactic acid or acetic acid after processing and inoculation. Pork frankfurters were formulated with 1.8% SL or 0.25% SD or combinations of 1.8% SL with 0.25 or 0.125% SD. After processing, frankfurters were inoculated (2 to 3 log CFU/cm2) with a 10-strain composite of Listeria monocytogenes and left undipped or were dipped (2 min) in 2.5% solutions of lactic acid or acetic acid (23 +/- 2 degrees C) before vacuum packaging and storage at 10 degrees C for 40 days. Total microbial populations and L. monocytogenes, lactic acid bacteria, and yeasts and molds were enumerated during storage. Sensory evaluations also were carried out on frankfurters treated and/or formulated with effective antimicrobials. The combination of 1.8% SL with 0.25% SD provided complete inhibition of L. monocytogenes growth throughout storage. Dipping in lactic acid or acetic acid reduced initial populations by 0.7 to 2.1 log CFU/cm2, but during storage (12 to 20 days), populations on dipped samples without antimicrobials in the formulation reached 5.5 to 7.9 log CFU/cm2. For samples containing single antimicrobials and dipped in lactic acid or acetic acid, L. monocytogenes growth was completely inhibited or reduced over 12 and 28 days, respectively, whereas final populations were lower (P < 0.05) than those in undipped samples of the same formulations. Bactericidal effects during storage (reductions of 0.6 to 1.0 log CFU/ cm2 over 28 to 40 days) were observed in frankfurters containing combinations of SL and SD that were dipped in organic acid solutions. Inclusion of antimicrobials in the formulation and/or dipping the product into organic acid solutions did not affect (P > 0.05) the flavor and overall acceptability of products compared with controls. The results of this study may be valuable to meat processors as they seek approaches for meeting new regulatory requirements in the United States.

Characterization and determination of origin of lactic acid bacteria from a sorghum-based fermented weaning food by analysis of soluble proteins and amplified fragment length polymorphism fingerprinting.

ABSTRACT The group that includes the lactic acid bacteria is one of the most diverse groups of bacteria known, and these organisms have been characterized extensively by using different techniques. In this study, 180 lactic acid bacterial strains isolated from sorghum powder (44 strains) and from corresponding fermented (93 strains) and cooked fermented (43 strains) porridge samples that were prepared in 15 households were characterized by using biochemical and physiological methods, as well as by analyzing the electrophoretic profiles of total soluble proteins. A total of 58 of the 180 strains wereLactobacillus plantarum strains, 47 were Leuconostoc mesenteroides strains, 25 were Lactobacillus sake-Lactobacillus curvatus strains, 17 were Pediococcus pentosaceus strains, 13 were Pediococcus acidilacticistrains, and 7 were Lactococcus lactis strains. L. plantarum and L. mesenteroides strains were the dominant strains during the fermentation process and were recovered from 87 and 73% of the households, respectively. The potential origins of these groups of lactic acid bacteria were assessed by amplified fragment length polymorphism fingerprint analysis.

Attachment and biofilm formation by Escherichia coli O157:H7 at different temperatures, on various food-contact surfaces encountered in beef processing.

Escherichia coli O157:H7 attached to beef-contact surfaces found in beef fabrication facilities may serve as a source of cross-contamination. This study evaluated E. coli O157:H7 attachment, survival and growth on food-contact surfaces under simulated beef processing conditions. Stainless steel and high-density polyethylene surfaces (2×5cm) were individually suspended into each of three substrates inoculated (6log CFU/ml or g) with E. coli O157:H7 (rifampicin-resistant, six-strain composite) and then incubated (168h) statically at 4 or 15°C. The three tested soiling substrates included sterile tryptic soy broth (TSB), unsterilized beef fat-lean tissue (1:1 [wt/wt]) homogenate (10% [wt/wt] with sterile distilled water) and unsterilized ground beef. Initial adherence/attachment of E. coli O157:H7 (0.9 to 2.9log CFU/cm(2)) on stainless steel and high-density polyethylene was not affected by the type of food-contact surface but was greater (p<0.05) through ground beef. Adherent and suspended E. coli O157:H7 counts increased during storage at 15°C (168h) by 2.2 to 5.4log CFU/cm(2) and 1.0 to 2.8log CFU/ml or g, respectively. At 4°C (168h), although pathogen levels decreased slightly in the substrates, numbers of adherent cells remained constant on coupons in ground beef (2.4 to 2.5log CFU/cm(2)) and increased on coupons in TSB and fat-lean tissue homogenate by 0.9 to 1.0and 1.7 to 2.0log CFU/cm(2), respectively, suggesting further cell attachment. The results of this study indicate that E. coli O157:H7 attachment to beef-contact surfaces was influenced by the type of soiling substrate and temperature. Notably, attachment occurred not only at a temperature representative of beef fabrication areas during non-production hours (15°C), but also during cold storage (4°C) temperatures, thus, rendering the design of more effective sanitation programs necessary.

Postprocessing antimicrobial treatments to control Listeria monocytogenes in commercial vacuum-packaged bologna and ham stored at 10°C

The antilisterial effect of chemical dipping solutions on commercial bologna and ham slices, inoculated (3 to 4 log CFU/ cm2) after processing, was evaluated during storage in vacuum packages at 10 degrees C. Samples were inoculated with a 10-strain composite of Listeria monocytogenes and subsequently immersed (25+/-2 degrees C) for 2 min in 2.5% acetic acid (AA), 2.5% lactic acid (LA), 5% potassium benzoate (PB), or 0.5% Nisaplin (commercial form of nisin, equivalent to 5,000 IU/ml of nisin) solutions, either singly or sequentially (Nisaplin plus AA, Nisaplin plus LA, or Nisaplin plus PB), and then vacuum packaged and stored at 10 degrees C for 48 days. In addition to microbiological analysis, sensory evaluations were performed on uninoculated samples treated with AA, LA, or PB. Initial reductions (day 0) of the pathogen, compared with the controls, on bologna and ham samples treated with AA, LA, or PB ranged from 0.4 to 0.7 log CFU/cm2. Higher (P < 0.05) initial reductions (2.4 to 2.9 log CFU/cm2) were obtained for samples treated with Nisaplin alone and when followed by AA, LA, or PB. L. monocytogenes populations on control bologna and ham samples increased from 3.4 log CFU/cm2 (day 0) to 7.4 and 7.8 log CFU/ cm2, respectively, in 8 days at 10 degrees C. Listericidal effects were observed for all treatments tested, except for Nisaplin applied on its own, during storage at 10 degrees C. The sequential treatment of Nisaplin plus LA reduced L. monocytogenes to undetectable levels in both products at the end of storage. The sequential treatments were also found to inhibit growth of spoilage microorganisms. Sensory evaluations indicated that dipping (2 min) of ham samples in AA (2.5%), LA (2.5%), or PB (5%) led to lower sensory scores. However, since results of this study indicated that these treatments caused extensive listericidal effects, there is possibly a potential to reduce the levels of chemicals applied and still achieve adequate antilisterial activity without major negative effects on product quality.

Quantitative risk assessment for Listeria monocytogenes in selected categories of deli meats: impact of lactate and diacetate on listeriosis cases and deaths.

Foodborne disease associated with consumption of ready-to-eat foods contaminated with Listeria monocytogenes represents a considerable pubic health concern. In a risk assessment published in 2003, the U.S. Food and Drug Administration and the U.S. Food Safety and Inspection Service estimated that about 90% of human listeriosis cases in the United States are caused by consumption of contaminated deli meats. In this risk assessment, all deli meats were grouped into one of 23 categories of ready-to-eat foods, and only the postretail growth of L. monocytogenes was considered. To provide an improved risk assessment for L. monocytogenes in deli meats, we developed a revised risk assessment that (i) models risk for three subcategories of deli meats (i.e., ham, turkey, and roast beef) and (ii) models L. monocytogenes contamination and growth from production to consumption while considering subcategory-specific growth kinetics parameters (i.e., lag phase and exponential growth rate). This model also was used to assess how reformulation of the chosen deli meat subcategories with L. monocytogenes growth inhibitors (i.e., lactate and diacetate) would impact the number of human listeriosis cases. Use of product-specific growth parameters demonstrated how certain deli meat categories differ in the relative risk of causing listeriosis; products that support more rapid growth and have reduced lag phases (e.g., turkey) represent a higher risk. Although reformulation of deli meats with growth inhibitors was estimated to reduce by about 2.5- to 7.8-fold the number of human listeriosis cases linked to a given deli meat subcategory and thus would reduce the overall risk of human listeriosis, even with reformulation deli meats would still cause a considerable number of human listeriosis cases. A combination of strategies is thus needed to provide continued reduction of these cases. Risk assessment models such as that described here will be critical for evaluation of different control approaches and to help define the combinations of control strategies that will have the greatest impact on public health.

Effect of single or sequential hot water and lactic acid decontamination treatments on the survival and growth of Listeria monocytogenes and spoilage microflora during aerobic storage of fresh beef at 4, 10, and 25°C

The survival and growth of Listeria monocytogenes and spoilage microflora during storage of fresh beef subjected to different decontamination treatments was studied. Fresh beef inoculated with a five-strain mixture of L. monocytogenes (5.18 log CFU/cm2) was left untreated (control) or was immersed (30 s) in hot water (HW; 75 degrees C), 2% lactic acid (LA; 55 degrees C), hot water followed by lactic acid (HW-LA), or lactic acid followed by hot water (LA-HW) and then stored aerobically at 4, 10, and 25 degrees C for 25, 17, and 5 days, respectively. Initial populations of L. monocytogenes were reduced by 0.82 (HW), 1.43 (LA), 2.73 (HW-LA), and 2.68 (LA-HW) log CFU/cm2. During storage, the pathogen grew at higher rates in HW than in control samples at all storage temperatures. Acid decontamination treatments (LA. HW-LA, and LA-HW) resulted in a weaker inhibition of L. monocytogenes (P < 0.05) at 25 degrees C than at 4 and 10 degrees C. In general, the order of effectiveness of treatments was HW-LA > LA > LA-HW > HW > control at all storage temperatures tested. In untreated samples, the spoilage microflora was dominated by pseudomonads, while lactic acid bacteria, Enterobacteriaceae, and yeasts remained at lower concentrations during storage. Brochothrix thermosphacta was detected periodically in only a limited number of samples. Although decontamination with HW did not affect the above spoilage microbial profile, acid treatments shifted the predominant microflora in the direction of yeasts and gram-positive bacteria (lactic acid bacteria). Overall, the results of the present study indicate that decontamination with LA and combinations of LA and HW could limit growth of L. monocytogenes and inhibit pseudomonads, which are the main spoilage bacteria of fresh beef stored under aerobic conditions. However, to optimize the efficacy of such treatments, they must be applied in the appropriate sequence and followed by effective temperature control.

Postprocess Control of Listeria monocytogenes on Commercial Frankfurters Formulated with and without Antimicrobials and Stored at 10°C

The antilisterial effect of postprocess antimicrobial treatments on commercially manufactured frankfurters formulated with and without a 1.5% potassium lactate–0.05% sodium diacetate combination was evaluated. Frankfurters were inoculated (ca. 3 to 4 log CFU/cm2) with 10-strain composite Listeria monocytogenes cultures originating from different sources. The inocula evaluated were cells grown planktonically in tryptic soy broth plus 0.6% yeast extract (30°C, 24 h) or in a smoked sausage homogenate (15°C, 7 days) and cells that had been removed from stainless steel coupons immersed in an inoculated smoked sausage homogenate (15°C, 7 days). Inoculated frankfurters were dipped (2 min, 25 ± 2°C) in acetic acid (AA; 2.5%), lactic acid (LA; 2.5%), potassium benzoate (PB; 5%), or Nisaplin (commercial form of nisin; 0.5%, equivalent to 5,000 IU/ml of nisin) solutions, or in Nisaplin followed by AA, LA, or PB, and were subsequently vacuum packaged and stored for 48 days at 10°C. In addition to microbiological anal...

Escherichia coli O157:H7 survival, biofilm formation and acid tolerance under simulated slaughter plant moist and dry conditions.

Microorganisms persisting in slaughter plant environments may develop acid resistance and be translocated to other environmental surfaces or products. The objective of this study was to evaluate the potential of Escherichia coli O157:H7 to form biofilms and maintain acid resistance, under different culture habituation scenarios, on stainless steel coupons (2 x 5 x 0.08 cm), in the presence of beef carcass decontamination runoff fluids (washings). Coupons were stored in test tubes with unsterilized water washings (WW; pH 6.94) or lactic acid washings (LAW; pH 4.98), which were inoculated with E. coli O157:H7 (10(3)-10(4)CFU/ml) and incubated at 15 (24 or 48 h) or 35 degrees C (7 or 24 h), simulating different habituation scenarios on sites of a slaughter plant, including sanitation and overnight drying, during consecutive operational shifts. Acid resistance (AR) of planktonic and detached E. coli O157:H7 cells was assessed in tryptic soy broth adjusted to pH 3.5 with lactic acid. The highest pre-drying attachment and AR of E. coli O157:H7 were observed after 24h at 35 degrees C and 48 h at 15 degrees C. Drying reduced (P<0.05) recovery of attached E. coli O157:H7 cells; however, exposure of dried coupons to uninoculated washings allowed recovery of attached E. coli O157:H7, which restored AR, especially under conditions that favored post-drying growth. Exposure of attached cells to 50 ppm PAA for 45 s before drying, as well as habituation in LAW, reduced the recovery and AR of E. coli O157:H7. Therefore, incomplete removal of biofilms may result in cells of increased AR, especially in sites within a slaughter plant, in which liquid meat wastes may remain for long periods of time.

Differences in survival among 13 Listeria monocytogenes strains in a dynamic model of the stomach and small intestine.

ABSTRACT Thirteen Listeria monocytogenes strains were compared for the ability to survive in a dynamic gastrointestinal model. Strains displayed various degrees of susceptibility to gastric acidity; however, strain-to-strain variations became evident mainly after 90 min of exposure (pH 2.0). Cell levels transferred to the intestine depended on initial populations, while reductions during intestinal exposure were relatively small for all strains.