Inactivation of dried cells and biofilms of Listeria monocytogenes by exposure to blue light at different wavelengths and the influence of surface materials.

Antimicrobial blue light (aBL) in the 400-470 nm wavelength range has been reported to kill multiple bacteria. This study assessed its potential for mitigating an important foodborne pathogen, Listeria monocytogenes (Lm), focusing on surface decontamination. Three wavelengths were tested, with gallic acid as a photosensitizing agent (Ps), against dried cells obtained from bacterial suspensions, and biofilms on stainless-steel (SS) coupons. Following aBL exposure, standard microbiological analysis of inoculated coupons was conducted to measure viability. Statistical analysis of variance was performed. Confocal laser scanning microscopy was used to observe the biofilm structures. Within 16 h of exposure at 405 nm, viable Lm dried cells and biofilms were reduced by approx. 3 log CFU/cm2 with doses of 2,672 J/cm2. Application of Ps resulted in an additional 1 log CFU/cm2 at 668 J/cm2, but its effect was not consistent. The highest dose (960 J/cm2) at 420 nm reduced viable counts on the biofilms by 1.9 log CFU/cm2. At 460 nm, after 800 J/cm2, biofilm counts were reduced by 1.6 log CFU/cm2. The effect of material composition on Lm viability was also investigated. Irradiation at 405 nm (668 J/cm2) of cells dried on polystyrene resulted in one of the largest viability reductions (4.0 log CFU/cm2), followed by high-density polyethylene (3.5 log CFU/cm2). Increasing the dose to 4,008 J/cm2 from 405 nm (24 h), improved its efficacy only on SS and polyvinyl chloride. Biofilm micrographs displayed a decrease in biofilm biomass due to the removal of biofilm portions from the surface and a shift from live to dead cells suggesting damage to biofilm cell membranes. These results suggest that aBL is a potential intervention to treat Lm contamination on typical material surfaces used in food production.IMPORTANCECurrent cleaning and sanitation programs are often not capable of controlling pathogen biofilms on equipment surfaces, which transmit the bacteria to ready-to-eat foods. The presence of native plant microbiota and organic matter can protect pathogenic bacteria by reducing the efficacy of sanitizers as well as promoting biofilm formation. Post-operation washing and sanitizing of produce contact surfaces might not be adequate in eliminating the presence of pathogens and commensal bacteria. The use of a dynamic and harmless light technology during downtime and close of operation could serve as a useful tool in preventing biofilm formation and persistence. Antimicrobial blue light (aBL) technology has been explored for hospital disinfection with very promising results, but its application to control foodborne pathogens remains relatively limited. The use of aBL could be a complementary strategy to inactivate surfaces in restaurant or supermarket deli settings.

SARS-CoV-2 remains infectious for at least a month on artificially-contaminated frozen berries.

The potential transmission of SARS-CoV-2 via food has been controversial since the beginning of the COVID-19 pandemic. To investigate these concerns, reliable detection methods and data on virus die-off rates in various foods are needed. Here, an FDA-standard method for the detection of enteric viruses' RNA from soft fruits was modified for the recovery of infectious SARS-CoV-2. Then, the survival of SARS-CoV-2 on berries was investigated as well as the effectiveness of washing virus-contaminated berries with water. The modified method did not significantly reduced log infectivity titers of recovered viruses, but berries did. The detection limit of the method for infectious SARS-CoV-2 was ∼2.97 log TCID50/g of berries. On SARS-CoV-2-inoculated berries that were stored at 4 °C for 7 days, significant reductions in SARS-CoV-2 infectivity were observed over time. In contrast, on frozen berries, infectious SARS-CoV-2 was recovered for 28 days without significant reductions. Washing SARS-CoV-2-inoculated berries with water removed >90% of infectious viruses within 10 min; however, infectious viruses were detected in wash water. Therefore, on fresh berries infectious viruses are markedly inactivated over time and can be largely removed by washing with water. However, the prolonged survival of SARS-CoV-2 on frozen berries suggests that the virus can potentially spread through frozen fruits.

Salmonella enterica and Escherichia coli in Wheat Flour: Detection and Serotyping by a Quasimetagenomic Approach Assisted by Magnetic Capture, Multiple-Displacement Amplification, and Real-Time Sequencing.

Food safety is a new area for novel applications of metagenomics analysis, which not only can detect and subtype foodborne pathogens in a single workflow but may also produce additional information with in-depth analysis capabilities. In this study, we applied a quasimetagenomic approach by combining short-term enrichment, immunomagnetic separation (IMS), multiple-displacement amplification (MDA), and nanopore sequencing real-time analysis for simultaneous detection of Salmonella and Escherichia coli in wheat flour. Tryptic soy broth was selected for the 12-h enrichment of samples at 42°C. Enrichments were subjected to IMS using beads capable of capturing both Salmonella and E. coli MDA was performed on harvested beads, and amplified DNA fragments were subjected to DNA library preparation for sequencing. Sequencing was performed on a portable device with real-time basecalling adaptability, and resulting sequences were subjected to two parallel pipelines for further analysis. After 1 h of sequencing, the quasimetagenomic approach could detect all targets inoculated at approximately 1 CFU/g flour to the species level. Discriminatory power was determined by simultaneous detection of dual inoculums of Salmonella and E. coli, absence of detection in control samples, and consistency in microbial flora composition of the same flour samples over several rounds of experiments. The total turnaround time for detection was approximately 20 h. Longer sequencing for up to 15 h enabled serotyping for many of the samples with more than 99% genome coverage, which could be subjected to other appropriate genetic analysis pipelines in less than a total of 36 h.IMPORTANCE Enterohemorrhagic Escherichia coli (EHEC) and Salmonella are of serious concern in low-moisture foods, including wheat flour and its related products, causing illnesses, outbreaks, and recalls. The development of advanced detection methods based on molecular principles of analysis is essential to incorporate into interventions intended to reduce the risk from these pathogens. In this work, a quasimetagenomic method based on real-time sequencing analysis and assisted by magnetic capture and DNA amplification was developed. This protocol is capable of detecting multiple Salmonella and/or E. coli organisms in the sample within less than a day, and it can also generate sufficient whole-genome sequences of the target organisms suitable for subsequent bioinformatics analysis. Multiplex detection and identification were accomplished in less than 20 h and additional whole-genome analyses of different nature were attained within 36 h, in contrast to the several days required in previous sequencing pipelines.

Evaluation of single and combined antimicrobial treatments to inhibit Salmonella in queso fresco.

Hispanic style soft non-fermented cheeses, such as queso fresco (QF) have been linked to outbreaks and recalls. Salmonella is one of the main causes of these incidents. Due to lack of ripening or post-processing antimicrobial treatments, incorporating GRAS antimicrobials to production process may be a suitable approach to minimize microbial risk in QF. The aim of this study was to evaluate the efficiency of nisin (N), caprylic acid (CA) and trans-cinnamaldehyde (CN) as single or combined treatments to reduce Salmonella populations in QF during storage. Batches of QF were inoculated after curding with approx. 4 Log CFU/g of 5-strain cocktails of Salmonella and stored at 8 °C for 20 days. The final Salmonella counts in control samples ranged from 6.96 to 7.14 Log CFU/g. Application of CN at 0.6 g/kg inhibited Salmonella growth during storage, resulting in at least 3 Log CFU/g difference with the untreated controls (p < 0.05). Addition of N (0.5 g/kg) and CA (0.4 g/kg) with CN (0.3 and 0.6 g/kg) further enhanced the antimicrobial activity resulting in complete suppression of growth and even caused a 1 Log CFU/g reduction by the end of the experimental period compared to initial counts. Samples treated with the combined treatment (N, CA, CN) were evaluated in a consumer panel (n = 112). Participants preferred the control and commercial QF to the treated samples. However, treated samples with 0.3 g/kg CN were still within the acceptable range of neutral to like slightly. Results obtained, revealed that combined treatment of N, CA and CN can provide a solution to reduce the count of Salmonella in QF, whether in process or during storage.

Assessment of Pediococcus acidilactici ATCC 8042 as potential Salmonella surrogate for thermal treatments of toasted oats cereal and peanut butter.

The control of Salmonella in low water activity foods poses a challenge for the food industry because of its thermal resistance. The use of surrogate bacteria in a food plant is considered a critical component to validate processing steps. The objective of this study was to evaluate the use of Pediococcus acidilactici ATCC 8042, a generally recognized as safe bacterium (GRAS), as potential surrogate for Salmonella in commercial toasted oats cereal (TOC) and peanut butter. P. acidilactici was compared to a five-serovar cocktail of Salmonella and Enterococcus faecium NRRL-B2354, separately. Cultures were inoculated into TOC and thermal kinetic parameters (δ, ß) were determined at 80, 85, 90, and 95 °C using the Weibull model. In peanut butter, δ and ß parameters were obtained at 63, 68, 73, and 77 °C. In TOC, the δ values (initial decimal reduction time) of P. acidilactici were 63 and 7 min at 80 and 95 °C, respectively, and at all four temperatures they were not significantly different from δ values of E. faecium. The δ value of Salmonella at 80 °C (139 min) was two-fold greater than the other two bacteria's values (p < 0.05). In peanut butter, δ values of P. acidilactici ranged from 31 min at 63 °C to 2.6 min at 77 °C, and at all temperatures they were not significantly different from E. faecium's δ values. In peanut butter, all Salmonella cocktail's δ values were significantly smaller than P. acidilactici's with values of 2 min at 63 °C and 0.4 min at 77 °C. These results indicated that P. acidilactici was as heat tolerant as E. faecium in these food matrices. However, the thermal inactivation kinetic parameters suggested that P. acidilactici can only be considered a Salmonella surrogate in TOC at temperatures above 85 °C. Because of its greater thermal tolerance in peanut butter, P. acidilactici may be used as Salmonella surrogate if an additional safety factor is recommended.

Long-Term Survival and Thermal Death Kinetics of Enterohemorrhagic Escherichia coli Serogroups O26, O103, O111, and O157 in Wheat Flour.

Wheat flour has been associated with outbreaks of enterohemorrhagic Escherichia coli (EHEC), but little is known on EHEC's survival during storage and thermal processing. The objective of this study was to determine long-term viability and thermal inactivation kinetics of EHEC serogroups O26, O103, O111, and O157. Wheat flour samples were inoculated with a cocktail of five strains of a single serogroup and stored at 23 and 35°C. Inoculated samples were heated at 55, 60, 65, and 70°C. Viability was determined by plate counting. Decimal reduction time (D) and first decimal reduction time (δ) values were calculated with log-linear and Weibull models, respectively. At 23°C, EHEC counts declined gradually for 84 days and samples tested positive from 84 to 280 days. The thermal resistance (D and δ) values ranged from 7.5 to 8.2 and 3.1 to 5.3 days, respectively, but there were no significant differences among serogroups (P ≤ 0.05). At 35°C, no EHEC was quantifiable by day 7 and no positive samples were detected after 49 days. Heating at 55 and 65°C resulted in δ-value ranges of 15.6 to 39.7 min and 3.0 to 3.9 min, respectively, with no significant difference among serogroups either. Z values were 12.6, 6.7, 10.2, and 13.4°C for O26, O103, O111, and O157, respectively. Thermal death kinetics of EHEC in flour were better described using the Weibull model. Survival and inactivation rates of four serogroups were remarkably similar. These findings indicated that all EHEC serovars tested remained viable for at least 9 months at room temperature and survived for up to 60 min at 70°C in wheat flour.IMPORTANCE Enterohemorrhagic Escherichia coli (EHEC) and Salmonella have recently caused several gastroenteritis outbreaks and recalls of wheat flour. Because EHEC can cause illness with very low doses and there is very scarce information regarding their ability to survive storage and heating in flour, the present study was undertaken to assess the long-term survival of EHEC serogroups O26, O103, O111, and O157 in flour. These findings are relevant, as we report that EHEC can survive for more than 9 months in wheat flour during storage. In addition, results obtained suggest that thermal inactivation at 65°C for 30 min or 2 months of storage at 35°C may be feasible strategies to mitigate the risk of most EHEC serovars in wheat flour.

iTRAQ-Based Global Proteomic Analysis of Salmonella enterica Serovar Typhimurium in Response to Desiccation, Low Water Activity, and Thermal Treatment.

In this study, the changes in the global proteome of Salmonella in response to desiccation and thermal treatment were investigated by using an iTRAQ multiplex technique. A Salmonella enterica serovar Typhimurium strain was dried, equilibrated at high (1.0) and low (0.11) water activity (aw), and thermally treated at 75°C. The proteomes were characterized after every treatment. The proteomes of the different treatments differed in the expression of 175 proteins. On the basis of their proteomic expression profiles, the samples were clustered into two major groups, namely, "dry" samples and "moist" samples. The groups had different levels of proteins involved in DNA synthesis and transcription and in metabolic reactions, indicating that cells under either of the aw conditions need to strictly control energy metabolism, the rate of replication, and protein synthesis. The proteins with higher expression levels in moist samples were flagellar proteins (FlgEFGH), membrane proteins, and export systems (SecF, SecD, the Bam complex), as well as stress response proteins, suggesting that rehydration can trigger stress responses in moist cells. Dry samples had higher levels of ribosomal proteins, indicating that ribosomal proteins might be important for additional regulation of the cellular response, even when the synthesis of proteins is slowed down. At both aws, no differences in protein expression were observed between the thermally treated samples and the nonheated cells. In conclusion, our study indicates that the preadaptation to a dry condition was linked to increased thermal tolerance, while reversion from a dry state to a moist state induced a significant change in protein expression, possibly linked to the observed loss of thermal tolerance.IMPORTANCESalmonella enterica is able to survive in dry environments for very long periods. While it is well known that the initial exposure to desiccation is fundamental to trigger thermal tolerance in this organism, the specific physiological and molecular processes involved in this cross-protection phenomenon have not been fully characterized. Several studies have focused on the low-aw transcriptome of this pathogen when inoculated in different food matrices or on abiotic surfaces, but proteomic analyses have not been reported in the literature. Our study investigated the changes in proteomic expression in Salmonella enterica serovar Typhimurium during desiccation, exposure to low aw, and thermal treatment. A better knowledge of the systems involved in the response to desiccation and thermal tolerance, as well as a better understanding of their interplay, is fundamental to identify the most effective combination of interventions to prevent Salmonella's contamination of foods.

Antimicrobial treatments to control Listeria monocytogenes in queso fresco.

Queso fresco, is a Hispanic non-fermented cheese highly susceptible to contamination with L. monocytogenes. This research was aimed to determine the effect of GRAS antimicrobial ingredients to control L. monocytogenes. Antimicrobials included caprylic acid (CA), Nisaplin® (N, 2.5% nisin), a mixture of sodium lactate and sodium diacetate (SL/SD), Lactococcus lactis sbp. lactis DPC 3147, monolaurin, and lactic acid (LA). Batches of queso fresco curds were inoculated with 104 CFU/g and stored at 4 °C for three weeks. During storage the count of L. monocytogenes reached 7 to 8 Log CFU/g in control samples. Most individual antimicrobial treatments resulted in less than 1 Log CFU/g reductions in final counts, with the exception of N (0.5 g/kg) and CA (2.9 g/kg) that caused more than 3 and 5 Log CFU/g differences with controls, respectively. Mixtures of ingredients were more effective in inhibiting L. monocytogenes growth, and treatments with N and CA consistently delivered 6 Log CFU/g less counts than controls. Supplementation of 12 g/kg LA to treatments with SL/SD (3%/0.22%) caused differences of more than 4 Log CFU/g in final Listeria populations. Samples treated with the binary mixtures of N and CA (0.5 and 0.7 g/kg, respectively) were evaluated in a consumer panel (n = 67). Panelists slightly preferred control and commercial over treated samples, but all samples were in average rated between "slightly liking" and "moderately liking." These experiments indicated that combined use of antimicrobial ingredients may be an effective way to control the population of Listeria monocytogenes in queso fresco.

Factors influencing the Salmonella internalization into seedpods and whole plants of Arachis hypogaea (L.).

Here we investigated whether Salmonella enterica serovar Typhimurium ATCC 14028 was capable of internalizing in peanut seedpods and plants when exposed to inoculated soil and the edaphic factors that influenced uptake. Intact dry Virginia (DV) and fresh green Virginia (GV) seedpods were exposed to soil containing 6.5 Log (CFU/g) Salmonella under different soil moisture conditions. Internalization of S. Typhimurium into peanut plants germinated in inoculated soil was also examined with and without Bradyrhizobium (Arachis) sp.NC92. Salmonella counts recovered from GV seedpods were on average of 2.0 Log (CFU/pod) less than those recovered from DV seedpods. The internalization in DV pods was only observed at soil water content of 15% or greater in a loamy sand soil. S. Typhimurium was detected inside peanut plant tissues during most testing times. Cells were recovered from stem samples (3.5 Log CFU/g) at greater levels than it was observed for root (2.6 Log CFU/g) and leaf (1.7 Log CFU/g) samples. Overall, recovery of Salmonella from stem, root, and leaf samples were lower when B. NC92 was inoculated on seeds before sowing, but this trend was not significant. Our observations suggest possible routes of contamination of Salmonella into peanut products from soil.

Validation of good agricultural practices (GAP) on Minnesota vegetable farms.

The United States Food and Drug Administration and the Department of Agriculture jointly published the "Guide to Minimize Microbial Food Safety Hazards for Fresh Fruits and Vegetables," which is used as a basis for Good Agricultural Practices (GAP) audits. To understand barriers to incorporation of GAP by Minnesota vegetable farmers, a mail survey completed in 2008 was validated with visits to a subset of the farms. This was done to determine the extent to which actual practices matched perceived practices. Two hundred forty-six producers completed the mail survey, and 27 participated in the on-farm survey. Over 75% of the on-farm survey respondents produced vegetables on 10 acres or less and had 10 or fewer employees. Of 14 questions, excellent agreement between on-farm interviews and mail survey responses was observed on two questions, four questions had poor or slight agreement, and eight questions had no agreement. Ninety-two percent of respondents by mail said "they took measures to keep animals and pests out of packing and storage buildings." However, with the on-site visit only 45% met this requirement. Similarly, 81% of respondents by mail said "measures were taken to reduce the risk of wild and/or domestic animals entering into fruit and vegetable growing areas." With direct observation, 70% of farms actually had taken measures to keep animals out of the growing areas. Additional, on-farm assessments were done regarding employee hygiene, training, presence of animals, water sources, and composting practices. This validation study demonstrated the challenge of creating nonleading and concise questions that are not open to broad interpretation from the respondents. If mail surveys are used to assess GAP, they should include open-ended questions and ranking systems to better assess farm practices. To provide the most accurate survey data for educational purposes or GAP audits, on-farm visits are recommended.

Inhibitory effect of combinations of caprylic acid and nisin on Listeria monocytogenes in queso fresco.

Queso fresco (QF), a fresh Hispanic cheese has been linked to outbreaks and recalls caused by Listeria contamination. The use antimicrobial treatments may be a potential solution. The goal of this research was to test the addition of nisin (N), caprylic acid (CA) and trans-cinnamaldehyde (CN) as anti-listerial ingredients in QF. QF batches were inoculated with approx. 10(4) CFU/g of 5- or 6-strain mixtures of Listeria monocytogenes and treated with antimicrobials. Samples were stored at 4 °C for three weeks and Listeria counts were determined by plating on PALCAM agar. The impact on the QF's natural indicator microorganisms was also assessed during refrigerated storage. All N and CA combinations (≥0.4 g/kg each) were effective against L. monocytogenes and reduced the final counts by at least 3 log CFU/g after 20 days of storage compared to controls. The levels of most strain mixtures were reduced immediately after treatment and their numbers remained below 10(3) CFU/g during storage. CN (1.2 g/kg) was bacteriostatic against L. monocytogenes, but it did not reduce initial counts. The addition of CN to the combination of N and CA did not enhance their antimicrobial effect. Results indicated that combinations of N and CA could control L. monocytogenes in QF with little impact on the natural flora of the cheese, providing a solution to control post processing L. monocytogenes contamination of QF.

Concentration, detection and discrimination of Bacillus anthracis spores in orange juice using aptamer based surface enhanced Raman spectroscopy.

Herein, we developed an innovative label free platform for concentration, detection and discrimination of Bacillus anthracis spores. Based on aptamer capture and surface enhanced Raman scattering detection, we were able to detect 10(4) CFU spores spiked in orange juice and discriminate between spores B. anthracis and B. mycoides within 40 min.

Transcriptional and functional responses of Escherichia coli O157:H7 growing in the lettuce rhizoplane.

Lettuce and spinach are increasingly implicated in foodborne illness outbreaks due to contamination by Escherichia coli O157:H7. While this bacterium has been shown to colonize and survive on lettuce leaf surfaces, little is known about its interaction with the roots of growing lettuce plants. In these studies, a microarray analyses, mutant construction and confocal microscopy were used to gain an understanding of structure and function of bacterial genes involved in the colonization and growth of E. coli O157:H7 on lettuce roots. After three days of interaction with lettuce roots, 94 and 109 E. coli O157:H7 genes were significantly up- and down-regulated at least 1.5 fold, respectively. While genes involved in biofilm modulation (ycfR and ybiM) were significantly up-regulated, 40 of 109 (37%) of genes involved in protein synthesis were significantly repressed. E. coli O157:H7 was 2 logs less efficient in lettuce root colonization than was E. coli K12. We also unambiguously showed that a ΔycfR mutant of E. coli O157:H7 was unable to attach to or colonize lettuce roots. Taken together these results indicate that bacterial genes involved in attachment and biofilm formation are likely important for contamination of lettuce plants with Shiga toxin-producing E. coli strains.

Impact of management practices and distillers' grains feeding on the prevalence of Escherichia coli O157 in feedlot cattle in Minnesota.

Escherichia coli O157 is a foodborne pathogen that can be transmitted by contaminated ground beef and is shed naturally in cattle feces. Recent reports indicated that feeding distillers' grains (DG) to cattle increased fecal shedding and prevalence of E. coli O157. In Minnesota, feeding DG with solubles (DGS) to livestock became widespread within the last 10 years, but there is no report about the prevalence of E. coli O157 in beef cattle in this state. This study was undertaken to survey the fecal prevalence of E. coli O157 in cattle fed diets containing DG and its association with environmental conditions and management practices. Fecal samples were collected from three feedlots during a 1-year period. All animals in those feedlots were fed different DGS levels. E. coli O157 presence was determined using a combination of enrichment, immunomagnetic separation, plating onto sorbitol MacConkey agar, and confirmation of isolates by immunoassay and multiplex virulence genes polymerase chain reaction analysis. Overall, E. coli O157 was confirmed in 9.7% of samples. Prevalence during summer was 30% and declined to less than 10% the rest of the year. In animals grouped by dietary DGS concentration, no significant difference in prevalence (12.0 and 5.5%) was detected between the low and the high average groups (less and more than 20%). Previous feeding of DGS before arriving to the feedlot also had no influence on fecal prevalence. The presence of several interacting variables, uncontrolled in a real-life feedlot environment, was the likely reason for our observation and suggested that at the levels studied, DGS had no effect on the STEC O157 prevalence in cattle populations.

Herd-level risk factors associated with fecal shedding of Shiga toxin-encoding bacteria on dairy farms in Minnesota, USA.

This study aimed to identify herd-level risk factors associated with fecal shedding of Shiga toxin-encoding bacteria (STB) on dairy cattle farms in Minnesota, USA. After adjustment for farm size, risk factors included: use of total mixed ration (TMR) for lactating dairy cows [odds ratio (OR) = 3.0; 95% confidence interval (CI): 1.8 to 5.1], no use of monensin for weaned calves (OR = 4.8, 95% CI: 2.5, 9.3), and no use of decoquinate for preweaned calves (OR = 2.2, 95% CI: 1.4, 3.6). Fecal shedding of STB was more common in small herds (< 100 cows, OR = 3.6, 95% CI: 2.1, 6.2) than in large herds (≥ 100 cows). Herd management factors related to cattle feeding practices were associated with fecal shedding of STB.

Facteurs de risque au niveau du troupeau associés à l'excrétion fécale des bactéries encodant la toxine de Shiga dans les fermes laitières du Minnesota, États-Unis. Cette étude avait pour but d'identifier les facteurs de risque au niveau du troupeau associés à l'excrétion fécale de bactéries encodant la shiga-toxine dans les fermes de bovins laitiers au Minnesota, États-Unis. Après un ajustement pour la taille de la ferme, les facteurs de risque incluaient : l'utilisation de la ration mixte totale (RMT) pour les vaches laitières en lactation [rapport de cotes (RC) = 3,0; intervalle de confiance (IC) de 95 % : de 1,8 à 5,1], pas d'utilisation de monensin pour les veaux sevrés (RC = 4,8, IC de 95 % : 2,5, 9,3) et pas d'utilisation de décoquinate pour les veaux présevrés (RC = 2,2, IC de 95 % : 1,4, 3,6). L'excrétion fécale de la bactérie encodant la shiga-toxine était plus commune dans les petits troupeaux (< 100 vaches, RC = 3,6, IC de 95 % : 2,1, 6,2) que dans les grands troupeaux (≥ 100 vaches). Des facteurs de gestion du troupeau se rapportant aux pratiques d'alimentation du bétail ont été associés à l'excrétion fécale de la bactérie encodant la shiga-toxine.(Traduit par Isabelle Vallières).

Prevention of surface colonization and anti-biofilm effect of selected phytochemicals against Listeria innocua strain.

This work aimed to determine the ability of Listeria innocua (L.i.) to colonize eight materials found in food-processing and packaging settings and to evaluate the viability of the sessile cells. We also selected four commonly used phytochemicals (trans-cinnamaldehyde, eugenol, citronellol, and terpineol) to examine and compare their efficacies against L.i. on each surface. Biofilms were also deciphered in chamber slides using confocal laser scanning microscopy to learn more about how phytochemicals affect L.i. The materials tested were silicone rubber (Si), polyurethane (PU), polypropylene (PP), polytetrafluoroethylene (PTFE), stainless steel 316 L (SS), copper (Cu), polyethylene terephthalate (PET), and borosilicate glass (GL). L.i. colonized Si and SS abundantly, followed by PU, PP, Cu, PET, GL, and PTFE surfaces. The live/dead status ranged from 65/35% for Si to 20/80% for Cu, and the estimates of cells unable to grow on Cu were the highest, reaching even 43%. Cu was also characterized by the highest degree of hydrophobicity (ΔGTOT = -81.5 mJ/m2). Eventually, it was less prone to attachment, as we could not recover L.i. after treatments with control or phytochemical solutions. The PTFE surface demonstrated the least total cell densities and fewer live cells (31%) as compared to Si (65%) or SS (nearly 60%). It also scored high in hydrophobicity degree (ΔGTOT = -68.9 mJ/m2) and efficacy of phytochemical treatments (on average, biofilms were reduced by 2.1 log10 CFU/cm2). Thus, the hydrophobicity of surface materials plays a role in cell viability, biofilm formation, and then biofilm control and could be the prevailing parameter when designing preventive measures and interventions. As for phytochemical comparison, trans-cinnamaldehyde displayed greater efficacies, with the highest reductions seen on PET and Si (4.6 and 4.0 log10 CFU/cm2). The biofilms in chamber slides exposed to trans-cinnamaldehyde revealed the disrupted organization to a greater extent than other molecules. This may help establish better interventions via proper phytochemical selection for incorporation in environment-friendly disinfection approaches.

Transcriptional responses of Escherichia coli K-12 and O157:H7 associated with lettuce leaves.

An increasing number of outbreaks of gastroenteritis recently caused by Escherichia coli O157:H7 have been linked to the consumption of leafy green vegetables. Although it is known that E. coli survives and grows in the phyllosphere of lettuce plants, the molecular mechanisms by which this bacterium associates with plants are largely unknown. The goal of this study was to identify E. coli genes relevant to its interaction, survival, or attachment to lettuce leaf surfaces, comparing E. coli K-12, a model system, and E. coli O157:H7, a pathogen associated with a large number of outbreaks. Using microarrays, we found that upon interaction with intact leaves, 10.1% and 8.7% of the 3,798 shared genes were differentially expressed in K-12 and O157:H7, respectively, whereas 3.1% changed transcript levels in both. The largest group of genes downregulated consisted of those involved in energy metabolism, including tnaA (33-fold change), encoding a tryptophanase that converts tryptophan into indole. Genes involved in biofilm modulation (bhsA and ybiM) and curli production (csgA and csgB) were significantly upregulated in E. coli K-12 and O157:H7. Both csgA and bhsA (ycfR) mutants were impaired in the long-term colonization of the leaf surface, but only csgA mutants had diminished ability in short-term attachment experiments. Our data suggested that the interaction of E. coli K-12 and O157:H7 with undamaged lettuce leaves likely is initiated via attachment to the leaf surface using curli fibers, a downward shift in their metabolism, and the suppression of biofilm formation.

Thermal Death Kinetics of Three Representative Salmonella enterica Strains in Toasted Oats Cereal.

Several reports have indicated that the thermal tolerance of Salmonella at low-water activity increases significantly, but information on the impact of diverse food matrices is still scarce. The goal of this research was to determine the kinetic parameters (decimal reduction time, D; time required for the first decimal reduction, δ) of thermal resistance of Salmonella in a previously cooked low water activity food. Commercial toasted oats cereal (TOC) was used as the food model, with or without sucrose (25%) addition. TOC samples were inoculated with 108 CFU/mL of a single strain of one of three Salmonella serovars (Agona, Tennessee, Typhimurium). TOC samples were ground and equilibrated to aw values of 0.11, 0.33 and 0.53, respectively. Ground TOC was heated at temperatures between 65 °C and 105 °C and viable counts were determined over time (depending on the temperature for up to 6 h). Death kinetic parameters were determined using linear and Weibull regression models. More than 70% of Weibull's adjusted regression coefficients (Radj2) and only 38% of the linear model's Radj2 had values greater than 0.8. For all serovars, both D and δ values increased consistently at a 0.11 aw compared to 0.33 and 0.53. At 0.33 aw, the δ values for Typhimurium, Tennessee and Agona were 0.55, 1.01 and 2.87, respectively, at 85 °C, but these values increased to 65, 105 and 64 min, respectively, at 0.11 aw. At 100 °C, δ values were 0.9, 5.5 and 2.3 min, respectively, at 0.11 aw. The addition of sucrose resulted in a consistent reduction of eight out of nine δ values determined at 0.11 aw at 85, 95 and 100 °C, but this trend was not consistent at 0.33 and 0.53 aw. The Z values (increase of temperature required to decrease δ-value one log) were determined with modified δ values for a fixed ß (a fitting parameter that describes the shape of the curve), and ranged between 8.9 °C and 13.4 °C; they were not influenced by aw, strain or sugar content. These findings indicated that in TOC, high thermal tolerance was consistent among serovars and thermal tolerance was inversely dependent on aw.

Detection of a foreign protein in milk using surface-enhanced Raman spectroscopy coupled with antibody-modified silver dendrites.

Herein we developed a rapid and simple method which used surface-enhanced Raman spectroscopy (SERS) coupled with antibody-modified silver dendrites to detect ovalbumin (OVA), the egg white protein, introduced into whole milk. OVA was first captured out of milk by use of antibody-modified silver dendrites and then directly measured on the silver dendrites by Raman spectroscopy. Results show that this method is capable of detecting OVA at 0.1 µg/mL in phosphate buffered saline (PBS) and 5 µg/mL in milk within 30 min based on the principal component analysis. This method has the potential for wide use in areas such as allergenic protein detection and bioterrorism agent detection in complex matrixes.

Reduction of Escherichia coli O157:H7 viability on leafy green vegetables by treatment with a bacteriophage mixture and trans-cinnamaldehyde.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 has been recognized as a major foodborne pathogen responsible for frequent gastroenteritis outbreaks. Phages and essential oils can be used as a natural antimicrobial method to reduce bacterial pathogens from the food supply. The objective of this study was to determine the effect of a bacteriophage cocktail, BEC8, alone and in combination with the essential oil trans-cinnameldehyde (TC) on the viability of a mixture of EHEC O157:H7 strains applied on whole baby romaine lettuce and baby spinach leaves. The EHEC O157:H7 strains used were Nal(R) mutants of EK27, ATCC 43895, and 472. Exponentially growing cells from tryptic soy (TS) broth cultures were spot inoculated on leaves and dried. EHEC cells were placed at low, medium, and high inoculum levels (10(4), 10(5), and 10(6) CFU/mL, respectively). Appropriate controls, BEC8 (approx. 10(6) PFU/leaf), and TC (0.5% v/v) were applied on treated leaves. The leaves were incubated at 4, 8, 23, and 37 °C in Petri dishes with moistened filter papers. EHEC survival was determined using standard plate count on nalidixic acid (50 µg/mL) Sorbitol MacConkey agar. No survivors were detected when both leaves were treated with BEC8 or TC individually at low inoculum levels after 24 h at 23 and 37 °C. When the EHEC inoculum size increased and/or incubation temperature decreased, the efficacy of BEC8 and TC decreased. However, when the two treatments were combined, no survivors were detected after 10 min at all temperatures and inoculum levels on both leafy greens. These results indicated that the BEC8/TC combination was highly effective against EHEC on both leafy greens. This combination could potentially be used as an antimicrobial to inactivate EHEC O157:H7 and reduce their incidence in the food chain.