Surface disinfection of wheat kernels using gas phase hydroxyl-radical processes: Effect on germination characteristics, microbial load, and functional properties.

Wheat kernels harbor a diverse microflora that can negatively affect the suitability of the grains for further processing. To reduce surface microflora, a kernel disinfection method is required that does not affect grain functionality. Three different versions of gas phase hydroxyl-radical processes were compared with the common method for grain disinfection, that is, a bleach treatment. The gas phase hydroxyl-radicals are generated by the UV-C mediated degradation of hydrogen peroxide and/or ozone in a near water-free process. It was found that treating kernels with a bleach solution could reduce total aerobic count (TAC) and fungal count to below the level of enumeration. In comparison, the gas phase hydroxyl-radical treatment, that is, H2 O2 -UV-ozone treatment, could support a 1.3 log count reduction (LCR) in TAC and a 1.1 LCR in fungal count. The microbial load reduction for the wholemeal samples was less pronounced as endophytic microorganisms were less affected by all treatments, hinting at a limited penetration depth of the treatments. Despite reducing the microbial load on the kernel surface through the bleach and H2 O2 -UV-ozone treatments, none of these treatments resulted in a reduced microbial count on grains that underwent sprouting after the treatments. No negative effect on germination power or development of the seedling was observed for any of the treatments. The gluten aggregation behavior and xylanase activity of the wholemeal also remained unchanged after the gas phase hydroxyl-radical treatments. Our findings suggest that UV-H2 O2 -ozone treatment shows promise for dry-kernel disinfection, but further optimization of the processing parameters is required.

Inactivation of Salmonella Enteritidis on Hatchery and Table Eggs Using a Gas-phase Hydroxyl-Radical Process.

Eggs represent a significant vehicle for Salmonella Enteritidis with the pathogen being transferred to chicks in the hatchery, or to consumers via table eggs. In the following, the efficacy of a gas-phase hydroxyl-radical process for decontaminating hatchery and table eggs was evaluated. Recovery of Salmonella was maximized through holding eggs in tryptic soy broth containing 20% w/v glycerol for 1 h prior to plating. By using this technique, it was possible to recover 63% of the theoretical Salmonella inoculated onto eggs. The continuous hydroxyl-radical reactor consisted of a bank of UV-C lamps (254 nm) that generated hydroxyl-radicals from the degradation of hydrogen peroxide (H2O2) mist and ozone gas. The optimal treatment was defined as that which supports a 5 log CFU/egg reduction of Salmonella without negatively affecting egg quality or leaving H2O2 residues. A process of 2% v/v H2O2 delivered at 30 mL/min with a UV-C dose of 19 mJ/cm2 and ozone (20 ppm) with a total treatment time of 10s was selected. The egg quality metrics (Haugh value, yolk index, albumin pH, yolk pH) did not negatively differ over a 35-day shelf-life at 4 or 25℃ compared to washed eggs or nontreated controls. The cuticle layer of eggs remained intact following hydroxyl-radical treatment. Fertilized eggs (n = 61) treated with the hydroxyl-radicals exhibited the same hatchery rate (75%) as nontreated controls (71-79%) with no defects (unhealed navels or red hocks) being observed. The same hydroxyl-radical treatment could be applied to table eggs to support >5 log CFU/egg reduction of Salmonella and was compatible with egg washing regimes practiced in industry. In comparison, the egg washing process based on sodium hydroxide and chlorine supported a 2.76 ± 0.38 log CFU/egg reduction of Salmonella. The hydroxyl-radical treatment represents a preventative control step to reduce the carriage of Salmonella on hatchery and table eggs.

Antibiotic resistance mechanism and diagnosis of common foodborne pathogens based on genotypic and phenotypic biomarkers.

The emergence of antibiotic-resistant bacteria due to the overuse or inappropriate use of antibiotics has become a significant public health concern. The agri-food chain, which serves as a vital link between the environment, food, and human, contributes to the large-scale dissemination of antibiotic resistance, posing a concern to both food safety and human health. Identification and evaluation of antibiotic resistance of foodborne bacteria is a crucial priority to avoid antibiotic abuse and ensure food safety. However, the conventional approach for detecting antibiotic resistance heavily relies on culture-based methods, which are laborious and time-consuming. Therefore, there is an urgent need to develop accurate and rapid tools for diagnosing antibiotic resistance in foodborne pathogens. This review aims to provide an overview of the mechanisms of antibiotic resistance at both phenotypic and genetic levels, with a focus on identifying potential biomarkers for diagnosing antibiotic resistance in foodborne pathogens. Furthermore, an overview of advances in the strategies based on the potential biomarkers (antibiotic resistance genes, antibiotic resistance-associated mutations, antibiotic resistance phenotypes) for antibiotic resistance analysis of foodborne pathogens is systematically exhibited. This work aims to provide guidance for the advancement of efficient and accurate diagnostic techniques for antibiotic resistance analysis in the food industry.

Hydroxyl-radical activated water for inactivation of Escherichia coli O157:H7, Salmonella and Listeria monocytogenes on germinating mung beans.

The following reports on the generation of hydroxyl-radical activated water prepared by passing a hydrogen peroxide solution containing Fe(III) catalyst through a UV-C reactor. The activated water was subsequently evaluated for antimicrobial activity against Escherichia coli O157:H7 in suspension or when inoculated onto mung beans. Hydroxyl-radical generation was assessed through the oxidation of methylene blue when reacted with activated water prepared from solutions of different pH (4-10), UV-C dose (32-128 mJ/cm2), hydrogen peroxide (0-1000 mg/L) and Fe(III) concentration (0-100 mg/L). Methylene blue oxidation was associated with high concentrations of each reactant with a positive correlation with Fe(III) concentration. Inactivation curves of E. coli O157:H7 in activated water were diphasic with an initial slow rate that increased after 15 min contact time. In contrast to the methylene blue assay, the antimicrobial action of activated water was associated with high hydrogen peroxide (500 mg/mL) and low Fe(III) catalyst (1 mg/L) with no significant interaction with UV-C dose. Evidence would suggest that the mode-of-inactivation was through a radical propagation reaction that is rate-limited by the reduction of Fe (III) to Fe (II). Here, the initial activation process via UV-C illumination results in photo-reduction of Fe(III) and propagates the formation of hydroxyl-radicals. Fe(III) to Fe(II) cycling continues with oxidation of cell structures that ultimately leads to loss of viability due to accumulation of cellular damage. When activated water was used to soak mung beans inoculated with E. coli O157:H7 a 1 log reduction was obtained with a 19% increase in germinated beans and 8.5% higher sprout yield relative to controls. The oxidation reduction potential decreased from 477 mV to 288 mV and pH increased from 3.97 to 5.47, over the 24 h mung bean soak period. The reduction of Salmonella and Listeria monocytogenes on mung beans soaked in activated water was <1 log CFU/g with all three pathogens growing back over the sprouting period. From the results it can be concluded that activated water can enhance the germination of mung beans along with sprout yield but has limited capacity when applied alone as a seed disinfection method.

Pre-oxidation of spent lettuce wash water by continuous Advanced Oxidation Process to reduce chlorine demand and cross-contamination of pathogens during post-harvest washing.

A continuous Photo-Fenton Advanced-Oxidation-Process (AOP) for reducing the chlorine-demand of spent lettuce wash water was developed based on the generation of hydroxyl-radicals from the UV-C degradation of hydrogen peroxide in the presence of ferric-catalyst. It was found that an interaction between UV-C and hydrogen peroxide or ferric-catalyst concentration was associated with high hydroxyl-radical generation as determined from the oxidation of methylene blue. The optimal AOP treatment was identified as 320 mJ/cm2 UV-C dose, 9.6 mg/L H2O2, and 9 mg/L ferric-catalyst. When the treatment was applied to simulated lettuce spent wash water (6.6 g romaine lettuce per liter of distilled water containing 100 mg bentonite; pH 6.9) the chlorine demand was reduced from 150 ppm to 130 ppm. The chlorination of AOP treated water did not result in a greater log reduction of pathogens (Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella) on lettuce but did reduce cross-contamination between batches during washing. The chlorinated byproducts formed in AOP treated water exhibited higher antimicrobial activity compared to untreated controls. Although the treatment was successful in reducing cross-contamination of lettuce batches the cytotoxicity of disinfection byproducts requires to be assessed.

Vapor-Phase Hydroxyl or Chlorine Radical Treatment for Inactivating Listeria monocytogenes on Mushrooms (Agaricus bisporus) without Negatively Affecting Quality or Shelf Life.

ABSTRACT: Processes based on generating vapor-phase hydroxyl radicals or chlorine radicals were developed for inactivating Listeria monocytogenes on mushrooms without negatively affecting quality. Antimicrobial radicals were generated from the UV-C degradation of hydrogen peroxide or hypochlorite and ozone gas. Response surface modeling was used to identify the interaction among the operating parameters for the hydroxyl radical process: UV-C254nm intensity, hydrogen peroxide concentration, and ozone delivered. There was an inverse relationship between hydrogen peroxide concentration and UV-C intensity in terms of the log reduction of L. monocytogenes. The independent parameters for the chlorine radical process were hypochlorite concentration, pH, and UV-C intensity. From predictive models, the optimal hydroxyl radical treatment was found to be 5% (v/v) H2O2, 2.86 mW/cm2 UV-C intensity (total UV-C dose 144 mJ/cm2), and 16.5 mg of ozone. The optimal parameters for the chlorine radical process were 10 ppm of hypochlorite (pH 3.0), 11.0 mg of ozone, and 4.60 mW/cm2 UV-C intensity. When inoculated mushrooms were treated with the optimal hydroxyl radical and chlorine radical processes, the reduction of L. monocytogenes was found to be 2.42 ± 0.42 and 2.61 ± 0.30 log CFU, respectively, without any negative effects on mushroom quality (weight loss and Browning index during 14 days of storage at 4°C). These reductions were significantly greater than those from application of the individual elements of the radical processes and those in the control process, which used a 90-s dip in 1% (v/v) hydrogen peroxide. The study has demonstrated that hydroxyl radical and chlorine radical vapor-phase treatments are equally effective at inactivating L. monocytogenes on mushrooms and can be considered as a preventative control step.

Effects of Vaccination Against Coccidiosis on Gut Microbiota and Immunity in Broiler Fed Bacitracin and Berry Pomace.

Feeding practices have been found to influence gut microbiota which play a major role in immunity of poultry. In the present study, changes in cecal microbiota and humoral responses resulting in the 55 ppm bacitracin (BACI), 1% each of cranberry (CP1) and wild blueberry (BP1) pomace alone or in combination (CP+BP) feeding in broiler Cobb 500 vaccinated or not against coccidiosis were investigated. In the non-vaccinated group, no significant treatment effects were observed on performance parameters. Vaccination significantly affected bird's performance parameters particularly during the growing phase from 10 to 20 days of age. In general, the prevalence of coccidiosis and necrotic enteritis (NE) was reduced by vaccination (P < 0.05). BACI-treated birds showed low intestinal lesion scores, and both CP1 and BP1 feed supplementations reduced Eimeria acervulina and Clostridium perfringens incidences similar to BACI. Vaccination induced change in serum enzymes, minerals, and lipid levels in 21-day old birds while, levels of triglyceride (TRIG) and non-esterified fatty acids (NEFA) were higher (P < 0.05) in CP1 treated non-vaccinated group than in the control. The levels of NEFA were lower in BACI- and CP1-fed birds than in the control in non-vaccinated day 28 old birds. The highest levels of all estimated three immunoglobulins (IgY, IgM, and IgA) were found in the vaccinated birds. Metagenomics analysis of the cecal bacterial community in 21-day old birds showed the presence of Firmicutes (90%), Proteobacteria (5%), Actinobacteria (2%), and Bacteroidetes (2%). In the vaccinated group, an effect of BACI was noted on Proteobacteria (P = 0.03). Vaccination and/or dietary treatments influenced the population of Lactobacillaceae, Enterobacteriaceae, Clostridiaceae, and Streptococcaceae which were among the most abundant families. Overall, this study revealed that besides their beneficial effects on performance, alike bacitracin, berry pomaces in poultry feed have profound impacts on the chicken cecal microbiota and blood metabolites that could be influenced by vaccination against coccidiosis.

Decontamination of N95 and surgical masks using a treatment based on a continuous gas phase-Advanced Oxidation Process.

A gas-phase Advanced Oxidation Process (gAOP) was evaluated for decontaminating N95 and surgical masks. The continuous process was based on the generation of hydroxyl-radicals via the UV-C (254 nm) photo-degradation of hydrogen peroxide and ozone. The decontamination efficacy of the gAOP was dependent on the orientation of the N95 mask passing through the gAOP unit with those positioned horizontally enabling greater exposure to hydroxyl-radicals compared to when arranged vertically. The lethality of gAOP was independent of the applied hydrogen peroxide concentration (2-6% v/v) but was significantly (P<0.05) higher when H2O2 was introduced into the unit at 40 ml/min compared to 20 ml/min. A suitable treatment for N95 masks was identified as 3% v/v hydrogen peroxide delivered into the gAOP reactor at 40 ml/min with continuous introduction of ozone gas and a UV-C dose of 113 mJ/cm2 (30 s processing time). The treatment supported >6 log CFU decrease in Geobacillus stearothermophilus endospores, > 8 log reduction of human coronavirus 229E, and no detection of Escherichia coli K12 on the interior and exterior of masks. There was no negative effect on the N95 mask fitting or particulate efficacy after 20 passes through the gAOP system. No visual changes or hydrogen peroxide residues were detected (<1 ppm) in gAOP treated masks. The optimized gAOP treatment could also support >6 log CFU reduction of endospores inoculated on the interior or exterior of surgical masks. G. stearothermophilus Apex spore strips could be applied as a biological indicator to verify the performance of gAOP treatment. Also, a chemical indicator based on the oxidative polymerization of pyrrole was found suitable for reporting the generation of hydroxyl-radicals. In conclusion, gAOP is a verifiable treatment that can be applied to decontaminate N95 and surgical masks without any negative effects on functionality.

Organic cranberry pomace and its ethanolic extractives as feed supplement in broiler: impacts on serum Ig titers, liver and bursal immunity.

With the pressure to reduce antibiotics use in poultry production, cost-effective alternative products need to be developed to enhance the bird's immunity. The present study evaluated the efficacy of cranberry fruit by-products to modulate immunity in broiler chickens. Broiler Cobb 500 chicks were fed a control basal diet, basal diet supplemented with bacitracin (BACI, 55 ppm), cranberry pomace at 1% and 2% (CP2), or cranberry pomace ethanolic extract at 150 and 300 ppm (COH300) for 30 d. Blood sera were analyzed at days 21 and 28 of age for Ig levels by ELISA. The innate and adaptive immune-related gene expression levels in the liver and bursa of Fabricius were investigated at 21 d of age by quantitative polymerase chain reaction arrays. At day 21, the highest IgY level was found in the blood serum of the CP2-fed birds. In the liver, 13 of the 22 differentially expressed genes were downregulated across all treatments compared with the control. Expression of genes belonging to innate immunity such as caspase 1 apoptosis-related cysteine peptidase, chemokine receptor 5, interferon gamma, myeloid differentiation primary response gene 88, and Toll-like receptor 3 were significantly downregulated mainly in BACI- and COH300-fed birds. In the bursa, 5 of 9 genes associated with the innate immunity were differentially expressed. The expression of anti-inflammatory IL-10 gene was upregulated in all treatment groups in bursa compared with the control. The expression of transferrin gene was significantly upregulated in livers of birds fed COH300 and in bursa of birds fed BACI, indicating feeding practices and organ-dependant modulation of this gene in broiler. Overall results of this study showed that cranberry product feed supplementation modulated the innate immune and suppressed proinflammatory cytokines in broilers, providing a platform for future investigations to develop berry products in poultry feeding.

Hazard assessment using an in-silico toxicity assessment of the transformation products of boscalid, pyraclostrobin, fenbuconazole and glyphosate generated by exposure to an advanced oxidative process.

Agricultural pesticide use is ongoing and consumer concern regarding the safety of pesticide residues on produce has generated interest in techniques that can safely reduce residues post-harvest. Recently an advanced oxidative process has shown promise in substantial residue reduction on the surface of produce. Given the potential for oxidative transformation of pesticides to generate transformation products with greater toxicity than the parent residue, take for example the oxon products of the organophosphorus insecticides, it is important to consider what transformation products are generated by pesticide exposure to an oxidative process and their potential toxicity. In this study, previously published transformation products of boscalid, pyraclostrobin, fenbuconazole and glyphosate were identified after exposure to 3% hydrogen peroxide, UV-C irradiation or their combination in an advanced oxidative process on glass, their oral toxicity, carcinogenicity and developmental toxicity were identified in-silico and an initial tier hazard assessment was conducted. Of the 87 total structures that were searched for, 53 were detected by UPLC-QTOF-MS and identified by mass spectra: 15, 13, 22 and 3 structures for boscalid, pyraclostrobin, fenbuconazole and glyphosate respectively, including the parent residues. Oral toxicity of the transformation products of pyraclostrobin and glyphosate was similar to or lower than the parent residue. Several transformation products of boscalid and fenbuconazole were estimated to be significantly more orally toxic than their parent residues. While the majority of the transformation products of boscalid, pyraclostrobin and fenbuconazole were predicted to be carcinogenic there were 11 that were consistently identified to have carcinogenic potential by several assessments. 29 of the 53 molecules were predicted to be probable developmental toxicants. An initial tier hazard assessment was conducted for Cramer rules classification and mutagenicity using the threshold of toxicological concern approach and predicted rat oral LD50. Two exposure scenarios were considered, one highly protective considering each transformation product to be at the highest maximum residue limit (MRL) for the pesticide and whole produce consumption at the highest consumption rate from the USEPA Exposures Handbook, the other considering only apple consumption with the relevant MRL. As indicated by the hazard assessment, several transformation products of boscalid, pyraclostrobin and fenbuconazole should be strongly considered for further testing, either by quantifying their production or in-vivo and in-vitro toxicity tests due to their predicted toxicity and associated hazard.

The progress of type II persisters of Escherichia coli O157:H7 to a non-culturable state during prolonged exposure to antibiotic stress with revival being aided through acid-shock treatment and provision of methyl pyruvate.

Persisters are a form of dormancy in bacteria that provide temporary resistance to antibiotics. The following reports on the formation of Escherichia coli O157:H7 E318 type II persisters from a protracted (8 days) challenge with ampicillin. Escherichia coli O157:H7 followed a multiphasic die-off pattern with an initial rapid decline (Phase I) of susceptible cells that transitioned to a slower rate representing tolerant cells (Phase II). After 24 h post-antibiotic challenge, the E. coli O157:H7 levels remained relatively constant at 2 log CFU/mL (Phase III), but became non-culturable within 8-days (Phase IV). The revival of persisters in Phase III could be achieved by the removal of antibiotic stress, although those in Phase IV required an extended incubation period or application of acid-shock. The carbon utilization profile of persister cells was less diverse compared with non-persisters, with only methyl pyruvate being utilized from the range tested. Inclusion of methyl pyruvate in tryptic soy agar revived non-cultural persisters, presumably by stimulating metabolism. The results suggest that persisters could be subdivided into culturable or non-culturable cells, with the former representing a transition state to the latter. The study provided insights into how to revive cells from dormancy to aid enumeration and control.

Gut Microbiota, Blood Metabolites, and Spleen Immunity in Broiler Chickens Fed Berry Pomaces and Phenolic-Enriched Extractives.

This study evaluated the performance, gut microbiota, and blood metabolites in broiler chickens fed cranberry and blueberry products for 30 days. A total of 2,800 male day-old broiler Cobb-500 chicks were randomly distributed between 10 diets: control basal diet; basal diet with bacitracin (BACI); four basal diets with 1 and 2% of cranberry (CP1, CP2) and blueberry (BP1, BP2) pomaces; and four basal diets supplemented with ethanolic extracts of cranberry (COH150, COH300) or blueberry (BOH150, BOH300) pomaces. All groups were composed of seven replicates (40 birds per replicate). Cecal and cloacal samples were collected for bacterial counts and 16S rRNA gene sequencing. Blood samples and spleens were analyzed for blood metabolites and gene expressions, respectively. The supplementation of COH300 and BOH300 significantly increased the body weight (BW) during the starting and growing phases, respectively, while COH150 improved (P < 0.05) the overall cumulated feed efficiency (FE) compared to control. The lowest prevalence (P = 0.01) of necrotic enteritis was observed with CP1 and BP1 compared to BACI and control. Cranberry pomace significantly increased the quinic acid level in blood plasma compared to other treatments. At days 21 and 28 of age, the lowest (P < 0.05) levels of triglyceride and alanine aminotransferase were observed in cranberry pomace and blueberry product-fed birds, respectively suggesting that berry feeding influenced the lipid metabolism and serum enzyme levels. The highest relative abundance of Lactobacillaceae was found in ceca of birds fed CP2 (P < 0.05). In the cloaca, BOH300 significantly (P < 0.005) increased the abundances of Acidobacteria and Lactobacillaceae. Actinobacteria showed a significant (P < 0.05) negative correlation with feed intake (FI) and FE in COH300-treated birds, whereas Proteobacteria positively correlated with the BW but negatively correlated with FI and FE, during the growing phase. In the spleen, cranberry products did not induce the release of any pro-inflammatory cytokines but upregulated the expression of several genes (IL4, IL5, CSF2, and HMBS) involved in adaptive immune responses in broilers. This study demonstrated that feed supplementation with berry products could promote the intestinal health by modulating the dynamics of the gut microbiota while influencing the metabolism in broilers.

Corrigendum: Gut Microbiota, Blood Metabolites, and Spleen Immunity in Broiler Chickens Fed Berry Pomaces and Phenolic-Enriched Extractives.

[This corrects the article DOI: 10.3389/fvets.2020.00150.].

Validation of a vapor-phase advanced oxidation process for inactivating Listeria monocytogenes, its surrogate Lactobacillus fructivorans, and spoilage molds associated with green or red table grapes.

A method based on vapor-phase advanced oxidation process (AOP) for decontaminating red or green grapes was validated for inactivating Listeria monocytogenes and spoilage molds. A Central Composite Design (CCD) and Response Surface Methodology (RSM) were applied to determine the contribution of UV-C (254 nm) dose, hydrogen peroxide, and ozone concentration on the lethality toward Aspergillus niger spores (biodensiometer) and changes to the grape quality (firmness and color over 14-day post-treatment storage at 4 °C). A high UV-C dose (>129 mJ/cm2 ) or >4.0 % v/v hydrogen peroxide induced-blistering and darkening of grapes at the end of the storage period. Yet, an optimized AOP treatment (with regards to preserving grape quality) was derived to be 1.3% v/v hydrogen peroxide (5 mL/10 berries) with 9-mg ozone gas and a UV-C dose of 123 mJ/cm2 (10 s at UV-C intensity of 12 mW/cm2 ). A predictive model was constructed and verified based on the log reduction of A. niger spores and changes in quality characteristics of red grapes. The optimal AOP treatment supported a 1.6-log CFU/g reduction of Aspergillus spores and decreased L. monocytogenes counts by 3.92 ± 0.17 and 4.77 ± 0.30 log CFU/g on green and red grapes, respectively, that were not significantly different to the surrogate, Lactobacillus fructivorans. There was no significant difference in the reduction of L. monocytogenes with grapes arranged in a single or double layer. Botrytis cinerea counts were reduced by 1.08 to 1.35 log CFU/g using the optimized AOP treatment with no change in grape color or firmness during storage. A sensory panel could not differentiate AOP-treated grapes from nontreated controls although 3 of 15 panelists did note subtle flavor notes. PRACTICAL APPLICATION: Postharvest washing of fresh produce has limited efficacy in removing foodborne pathogens and spoilage microbes. This is especially relevant to berries, such as grapes, that are susceptible to spoilage following washing. The vapor-phase AOP treatment provides a supplemental or alternative approach for produce decontamination. However, the operating parameters need to be optimized to ensure that decontamination of grapes is not at the expense of quality. In the current study, this was achieved by ensuring a balance between hydrogen peroxide, ozone, and UV-C dose that form the elements of an AOP treatment.

Inactivation of Salmonella and Listeria monocytogenes on dried fruit, pistachio nuts, cornflakes and chocolate crumb using a peracetic acid-ethanol based sanitizer or Advanced Oxidation Process.

Two decontamination methods were evaluated for inactivating a cocktail of Salmonella or Listeria monocytogenes inoculated onto model low moisture foods (LMFs; dried strawberry, dried apple, raisins, chocolate crumb, cornflakes, shell-on or deshelled pistachio nuts). One treatment was based on a peracetic acid-ethanol (PAA-ethanol) sanitizer combination with the other being an Advanced Oxidation Process (AOP) that simultaneously applied UV-C (254 nm), ozone and hydrogen peroxide. The low moisture food was spray inoculated then dried prior to treatment. With Salmonella it was found that a pre-incubation step in 1% w/v glycerol-tryptic soy broth for 1 h prior to plating, significantly increased recovery of the pathogen compared to TSB alone. However, no increased recovery of L. monocytogenes was observed using the TSB-glycerol pre-incubation step. No Salmonella was detected on cornflakes, chocolate crumb and strawberry using 1.25 parts per thousand (‰) PAA-ethanol. The inactivation of Salmonella on deshelled pistachio was significantly higher using 2.5‰ PAA-ethanol sanitizer compared to the AOP treatments tested. Only negligible reductions of Salmonella (<1 log cfu) were obtained with shell-on pistachio treated with PAA-ethanol sanitizer or AOP. Salmonella could be reduced on dried apple slices by >4 log CFU when 5.0‰ PAA-ethanol was applied. L. monocytogenes was more sensitive to PAA-ethanol compared to Salmonella and could be eliminated on all the LMFs apart from shell-on pistachio. An AOP treatment applied 10% v/v hydrogen peroxide, ozone and 54 mJ/cm2 UV-C could significantly reduce Salmonella on dried apple slices compared to when the individual elements (hydrogen peroxide, ozone or UV-C) were applied. Salmonella was also eliminated by AOP on the other LMFs (apart from shell-on pistachio) although the same level of inactivation was achieved by spraying with 10% v/v hydrogen peroxide alone. L. monocytogenes was sensitive to hydrogen peroxide and AOP being eliminated from all the LMFs. Although this may suggest that hydrogen peroxide spray was equivalent to AOP treatment it was noted that no residual H2O2 or changes in visual appearance was evident on samples treated with the latter process. The study has demonstrated that the two decontamination methods assessed can be applied to reduce Salmonella and L. monocytogenes on LMFs although efficacy is dependent on the pathogen and product type.

Transcriptional profiling of Salmonella enterica serovar Enteritidis exposed to ethanolic extract of organic cranberry pomace.

Non-typhoidal Salmonella enterica serovars continue to be an important food safety issue worldwide. Cranberry (Vaccinium macrocarpon Ait) fruits possess antimicrobial properties due to their various acids and phenolic compounds; however, the underlying mechanism of actions is poorly understood. We evaluated the effects of cranberry extracts on the growth rate of Salmonella enterica serovars Typhimurium, Enteritidis and Heidelberg and on the transcriptomic profile of Salmonella Enteritidis to gain insight into phenotypic and transcriptional changes induced by cranberry extracts on this pathogen. An ethanolic extract from cranberry pomaces (KCOH) and two of its sub-fractions, anthocyanins (CRFa20) and non-anthocyanin polyphenols (CRFp85), were used. The minimum inhibitory (MICs) and bactericidal (MBCs) concentrations of these fractions against tested pathogens were obtained using the broth micro-dilution method according to the Clinical Laboratory Standard Institute's guidelines. Transcriptional profiles of S. Enteritidis grown in cation-adjusted Mueller-Hinton broth supplemented with or without 2 or 4 mg/ml of KCOH were compared by RNASeq to reveal gene modulations serving as markers for biological activity. The MIC and MBC values of KCOH were 8 and 16 mg/mL, respectively, against all tested S. enterica isolates. The MIC value was 4 mg/mL for both CRFa20 and CRFp85 sub-fractions, and a reduced MBC value was obtained for CRFp85 (4 mg/ml). Treatment of S. Enteritidis with KCOH revealed a concentration-dependent transcriptional signature. Compared to the control, 2 mg/ml of KCOH exposure resulted in 89 differentially expressed genes (DEGs), of which 53 and 36 were downregulated and upregulated, respectively. The upregulated genes included those involved in citrate metabolism, enterobactin synthesis and transport, and virulence. Exposure to 4 mg/ml KCOH led to the modulated expression of 376 genes, of which 233 were downregulated and 143 upregulated, which is 4.2 times more DEGs than from exposure to 2 mg/ml KCOH. The downregulated genes were related to flagellar motility, Salmonella Pathogenicity Island-1 (SPI-1), cell wall/membrane biogenesis, and transcription. Moreover, genes involved in energy production and conversion, carbohydrate transport and metabolism, and coenzyme transport and metabolism were upregulated during exposure to 4 mg/ml KCOH. Overall, 57 genes were differentially expressed (48 downregulated and 9 upregulated) in response to both concentrations. Both concentrations of KCOH downregulated expression of hilA, which is a major SPI-1 transcriptional regulator. This study provides information on the response of Salmonella exposed to cranberry extracts, which could be used in the control of this important foodborne pathogen.

Comparative Effects of Thermal, High Hydrostatic Pressure, and UV-C Processing on the Quality, Nutritional Attributes, and Inactivation of Escherichia coli, Salmonella, and Listeria Introduced into Tiger Nut Milk.

HIGHLIGHTS: Thermal and nonthermal methods can support a 5-log CFU reduction of model bacteria introduced into tiger nut milk. Thermal treatment of tiger nut milk results in significant loss of protein, antioxidants, and quality properties. HHP or UV-C treatment of tiger nut milk retains quality and nutritional characteristics. HHP or UV-C are suitable for the pasteurization of tiger nut milk.

Decontamination of Chia and Flax Seed Inoculated with Salmonella and Surrogate, Enterococcus faecium NRRL B-2354, Using a Peracetic Acid Sanitizing Solution: Antimicrobial Efficacy and Impact on Seed Functionality.

Raw chia and flax seeds are increasingly associated with Salmonella contamination. However, intervention technologies for these seeds that maintain them in a raw state, without causing clumping because of mucilage production upon moisture exposure, are limited. In this study, a commercial ethanol and paracetic acid sanitizing solution meeting these criteria was evaluated for efficacy against Salmonella and Enterococcus faecium NRRL B-2354, a known Salmonella surrogate for thermal intervention technologies. Samples (100 g each) of chia and flax seeds ( n = 5) were inoculated with either a cocktail of Salmonella Newport, Senftenberg, Oranienburg, Saintpaul, Typhimurium DT104, and Cubana or E. faecium NRRL B-2354. After overnight acclimatization, samples were treated with 4 mL of sanitizing solution per sample and then held at ambient temperature (20 to 25°C) for 1 h before bacterial enumeration. Separate 1-kg-treated batches were evaluated for germination ability (4 replicates of 100-g samples), as well as nutrient content and rancidity ( n = 3), compared with untreated control. Following the posttreatment holding time, these batches were dried back to original moisture content at 70°C to evaporate residual sanitizing solution, thereby stopping treatment. The sanitizing solution was found to be an effective intervention method for chia and flax seeds, reducing Salmonella to below the level of detection by more than 4 and more than 5 average log CFU/g, respectively. Germination was not significantly affected ( P ≥ 0.05) for chia seed. For both seeds, nutrition and rancidity were not significantly affected ( P ≥ 0.05). Furthermore, E. faecium NRRL B-2354 was found to be an appropriate Salmonella surrogate for treatment of chia and flax seeds with this sanitizing solution, showing comparable but higher resistance to treatment with the sanitizing solution than the Salmonella cocktail.

Droplet digital polymerase chain reaction (ddPCR) assays integrated with an internal control for quantification of bovine, porcine, chicken and turkey species in food and feed.

Food adulteration and feed contamination are significant issues in the food/feed industry, especially for meat products. Reliable techniques are needed to monitor these issues. Droplet Digital PCR (ddPCR) assays were developed and evaluated for detection and quantification of bovine, porcine, chicken and turkey DNA in food and feed samples. The ddPCR methods were designed based on mitochondrial DNA sequences and integrated with an artificial recombinant plasmid DNA to control variabilities in PCR procedures. The specificity of the ddPCR assays was confirmed by testing both target species and additional 18 non-target species. Linear regression established a detection range between 79 and 33200 copies of the target molecule from 0.26 to 176 pg of fresh animal tissue DNA with a coefficient of determination (R2) of 0.997-0.999. The quantification ranges of the methods for testing fortified heat-processed food and feed samples were 0.05-3.0% (wt/wt) for the bovine and turkey targets, and 0.01-1.0% (wt/wt) for pork and chicken targets. Our methods demonstrated acceptable repeatability and reproducibility for the analytical process for food and feed samples. Internal validation of the PCR process was monitored using a control chart for 74 consecutive ddPCR runs for quantifying bovine DNA. A matrix effect was observed while establishing calibration curves with the matrix type under testing, and the inclusion of an internal control in DNA extraction provides a useful means to overcome this effect. DNA degradation caused by heating, sonication or Taq I restriction enzyme digestion was found to reduce ddPCR readings by as much as 4.5 fold. The results illustrated the applicability of the methods to quantify meat species in food and feed samples without the need for a standard curve, and to potentially support enforcement activities for food authentication and feed control. Standard reference materials matching typical manufacturing processes are needed for future validation of ddPCR assays for absolute quantification of meat species.