Characterization of highly pathogenic avian influenza virus in retail dairy products in the US.

In March 2024, clade 2.3.4.4b H5N1 highly pathogenic avian influenza virus (HPAIV) was detected in dairy cattle in the US, and it was discovered that the virus could be detected in raw milk. Although affected cow's milk is diverted from human consumption and current pasteurization requirements are expected to reduce or eliminate infectious HPAIV from the milk supply, a study was conducted to characterize whether the virus could be detected by quantitative real-time RT-PCR (qrRT-PCR) in pasteurized retail dairy products and, if detected, to determine whether the virus was viable. From 18 April to 22 April 2024, a total of 297 samples of Grade A pasteurized retail milk products (23 product types) were collected from 17 US states that represented products from 132 processors in 38 states. Viral RNA was detected in 60 samples (20.2%), with qrRT-PCR-based quantity estimates (non-infectious) of up to 5.4log1050% egg infectious doses per mL, with a mean and median of 3.0log10/mL and 2.9log10/mL, respectively. Samples that were positive for type A influenza by qrRT-PCR were confirmed to be clade 2.3.4.4 H5 HPAIV by qrRT-PCR. No infectious virus was detected in any of the qrRT-PCR-positive samples in embryonating chicken eggs. Further studies are needed to monitor the milk supply, but these results provide evidence that the infectious virus did not enter the US pasteurized milk supply before control measures for HPAIV were implemented in dairy cattle.IMPORTANCEHighly pathogenic avian influenza virus (HPAIV) infections in US dairy cattle were first confirmed in March 2024. Because the virus could be detected in raw milk, a study was conducted to determine whether it had entered the retail food supply. Pasteurized dairy products were collected from 17 states in April 2024. Viral RNA was detected in one in five samples, but infectious virus was not detected. This provides a snapshot of HPAIV in milk products early in the event and reinforces that with current safety measures, infectious viruses in milk are unlikely to enter the food supply.

Bacterial adhesion inhibition by microalgal EPSs from Cylindrotheca closterium and Tetraselmis suecica biofilms.

In the food industry, successful bacterial pathogen colonization and persistence begin with their adhesion to a surface, followed by the spatial development of mature biofilm of public health concerns. Compromising bacterial settlement with natural inhibitors is a promising alternative to conventional anti-fouling treatments typically based on chemical biocides that contribute to the growing burden of antimicrobial resistance. In this study, three extracellular polymeric substance (EPS) fractions extracted from microalgae biofilms of Cylindrotheca closterium (fraction C) and Tetraselmis suecica (fraction Ta rich in insoluble scale structure and fraction Tb rich in soluble EPS) were screened for their anti-adhesive properties, against eight human food-borne pathogens belonging to Escherichia coli, Staphylococcus aureus, Salmonella enterica subsp. enterica, and Listeria monocytogenes species. The results showed that the fraction Ta was the most effective inducing statistically significant reduction for three strains of E. coli, S. aureus, and L. monocytogenes. Overall, EPSs coating on polystyrene surfaces of the different fractions increased the hydrophilic character of the support. Differences in bacterial adhesion on the different coated surfaces could be explained by several dissimilarities in the structural and physicochemical EPS compositions, according to HPLC and ATR-FTIR analysis. Interestingly, while fractions Ta and Tb were extracted from the same microalgal culture, distinct adhesion patterns were observed, highlighting the importance of the extraction process. Overall, the findings showed that EPS extracted from microalgal photosynthetic biofilms can exhibit anti-adhesive effects against food-borne pathogens and could help develop sustainable and non-toxic anti-adhesive surfaces for the food industry. KEY POINTS: •EPSs from a biofilm-based culture of C. closterium/T. suecica were characterized. •Microalgal EPS extracted from T. suecica biofilms showed bacterial anti-adhesive effects. •The anti-adhesive effect is strain-specific and affects both Gram - and Gram + bacteria.

UHPLC-MS/MS and GC-MS Metabolic Profiling of a Medicinal Flourensia Fiebrigii Chemotype.

The comparative metabolic profiling and their biological properties of eight extracts obtained from diverse parts (leaves, flowers, roots) of the medicinal plant Flourensia fiebrigii S.F. Blake, a chemotype growing in highland areas (2750 m a.s.l.) of northwest Argentina, were investigated. The extracts were analysed by GC-MS and UHPLC-MS/MS. GC-MS analysis revealed the presence of encecalin (relative content: 24.86 %) in ethereal flower extract (EF) and this benzopyran (5.93 %) together sitosterol (11.35 %) in the bioactive ethereal leaf exudate (ELE). By UHPLC-MS/MS the main compounds identified in both samples were: limocitrin, (22.31 %), (2Z)-4,6-dihydroxy-2-[(4-hydroxy-3,5-dimethoxyphenyl)methylidene]-1-benzofuran-3-one (21.31 %), isobavachin (14.47 %), naringenin (13.50 %), and sternbin, (12.49 %). Phytocomplexes derived from aerial parts exhibited significant activity against biofilm production of Pseudomonas aeruginosa and Staphylococcus aureus, reaching inhibitions of 74.7-99.9 % with ELE (50 µg/mL). Notably, the extracts did not affect nutraceutical and environmental bacteria, suggesting a selective activity. ELE also showed the highest reactive species scavenging ability. This study provides valuable insights into the potential applications of this chemotype.

Exploring the potential of 1,8-cineole from cardamom oil against food-borne pathogens: Antibacterial mechanisms and its application in meat preservation.

Food-borne pathogenic bacteria are a major public health concern globally. Traditional control methods using antibiotics have limitations, leading to the exploration of alternative strategies. Essential oils such as cardamom possess antimicrobial properties and have shown efficacy against food-borne pathogenic bacteria. The utilization of essential oils and their bioactive constituents in food preservation is a viable strategy to prolong the shelf-life of food products while ensuring their quality and safety. To the best of our knowledge, there are no studies that have utilized 1,8-cineole (the main active constituent of cardamom essential oil) as a preservative in meat, so this study might be the first to utilize 1,8-cineole as an antibacterial agent in meat preservation. The application of 1,8-cineole had a significant suppressive impact on the growth rate of Listeria monocytogenes, Staphylococcus aureus, Escherichia coli, and Salmonella Typhimurium in meat samples stored for 7 days at 4 °C. Additionally, the surface color of the meat samples was not negatively impacted by the application of 1,8-cineole. The minimum inhibitory concentration was 12.5-25 mg/ml, and the minimum bactericidal concentration was 25-50.0 mg/ml. The bacterial cell membrane may be the target of cardamom, causing leakage of intracellular proteins, ATP, and DNA. The obtained data in this study may pave a new avenue for using 1,8-cineole as a new perspective for dealing with this problem of food-borne pathogens and food preservation, such as meat.

Green synthesis of zinc oxide nanoparticles using novel bacterium strain (Bacillus subtilis NH1-8) and their in vitro antibacterial and antibiofilm activities against Salmonellatyphimurium.

Zinc oxide nanoparticles (ZnO NPs) are used in a range of applications, including food packaging, preservation, and storage. In the current investigation, extracellular green synthesis of ZnO NPs through an simple, eco-friendly, and rapid approach using a novel bacterial strain (Bacillus subtilis NH1-8) was studied. To assess the morphological, optical, and structural properties of ZnO NPs, transmission electron microscopy (TEM), energy-dispersive X-ray (EDX), scanning electron microscopy (SEM), fourier transform infrared (FTIR) spectroscopy, ultraviolet-visible (UV-vis) spectroscopy, and X-ray diffraction (XRD) techniques were carried out. In addition, disk diffusion, minimum bactericidal concentration (MBC), and minimum inhibitory concentration (MIC) methods were performed to determine the antibacterial activity of ZnO NPs. The average size of biosynthesized ZnO NPs was 39 nm, exhibiting semi-spherical, which was confirmed by TEM analyses. The UV-vis spectroscopy exhibited the absorption peak at 200-800nm. The ZnO NPs have shown effective antimicrobial and antibiofilm activities against S. typhimurium. Thus, biosynthesized ZnO NPs could be exploited as a breakthrough technology in the surface coating of food containers and cans to minimize contamination by S. typhimurium.

Diversity and epidemiological profile of Pseudomonas aeruginosa from drinking water in Brazil genotyped using multi-locus sequence typing.

Pseudomonas aeruginosa is a Gram-negative bacillus associated with waterborne diseases. The objective of this study was to determine whether particular P. aeruginosa sequence types (STs) were associated with drinking water contamination in Brazil. This was achieved by searching the Pseudomonas PubMLST database, which contains the records for 8358 strains collected between 1938 and 2023. The majority (97.2%) had the complete 7-loci multilocus sequence typing profile and were assigned to 3486 STs. After eBURST (an algorithm used to infer patterns of evolutionary descent among clusters), 1219 groups with single-locus variant and 575 groups with double-locus variant were formed. Brazil was the South American country with the most isolates (n = 219, 58.24%), and the Simpson's index was 0.9392. Of the 219 Brazilian isolates, eight were isolated in water and identified as STs 252, 1417, 1605, 2502, 2620, 3078, and 3312. ST252, 1417, and 3078 have already been isolated from clinical cases worldwide. Furthermore, ST1605 and 2620, after the eBURST, they were grouped in the same clonal complex as STs involved in human infections. In conclusion, P. aeruginosa STs involved in human infections were found in bottled drinking water commercialized in Brazil, revealing that these types of drinking waters can be a vehicle of contamination.

NtrC Increases Fitness of Salmonella enterica Serovar Typhimurium under Low and Fluctuating Nutrient Conditions.

Enteric pathogens cycle between nutrient-rich host and nutrient-poor external environment. These pathogens compete for nutrients while cycling between host and external environment, and often experience starvation. In this context, we have studied the role of a global regulator (NtrC) of Salmonella Typhimurium. The ntrC knockout mutation caused extended lag phase (8 h) and slow growth in the minimal medium. In lag phase, the wild-type cells showed ~60-fold more expression of ntrC gene. Gene expression studies and biochemical assays showed that the extended lag phase and slow growth is due to slow metabolism, instead of nitrogen transport. Further, we observed that ntrC knockout mutation led extended lag phase and slow growth, made ΔntrC mutant unable to compete with wild-type S. Typhimurium in both static and fluctuating nutrient condition. In addition to this, ΔntrC knockout mutant was unable to survive long-term nitrogen starvation (150 days). The nutrient recycling assays and gene expression studies revealed that ntrC gene is essential for rapid recycling of nutrients from the dead cells. Moreover, in the absence of ntrC gene, magnesium limits the nutrient recycling efficiency of S. Typhimurium. Therefore, the ntrC gene, which is often studied with respect to nitrogen scavenging in a low nitrogen growing condition, is required even in the adequate supply of nitrogen to maintain optimal growth and fast exit from the lag phase. Hence, we conclude that, the ntrC expression is essential for competitive fitness of S. Typhimurium under the low and fluctuating nutrient condition. IMPORTANCE S. Typhimurium, both in host and external environment, faces enormous competition from other microorganisms. The competition may take place either in static or in fluctuating nutrient conditions. Thus, how S. Typhimurium survives under such overlapping stress conditions remained unclear. Therefore, using S. Typhimurium as model organism we report that a global regulator NtrC, found in enteric bacteria like Escherichia coli and Salmonella, activates the set of genes and operons involved in rapid adaptation and efficient nutrient recycling/scavenging. These properties enable cells to compete with other microbes under the characteristic feast-or-famine lifestyle of S. Typhimurium. Therefore, this work helps us to understand the starvation physiology of the enteric bacterial pathogen S. Typhimurium.

Inactivation of Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes on alfalfa seeds by the combination treatment of vacuumed hydrogen peroxide vapour and vacuumed dry heat.

We evaluated the combined effects of vacuumed hydrogen peroxide vapour (VHPV) and vacuum-sealed dry heat (vacuum heat, VH) to inactivate food-borne pathogens (Salmonella Typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes) on alfalfa seeds. Alfalfa seeds inoculated with food-borne pathogens were sequentially treated initially with 1·0 ml of 0 or 30% VHPV for 1 min and later with dry heat (DH) or VH for 2 h, and the rate of seed germination was evaluated. The combination treatment decreased the populations of three food-borne pathogens below the limit of detection (1·0 log CFU per gram) on alfalfa seeds without decreasing germinability. The sequential treatment using VHPV and VH greatly reduced the total treatment time needed to inactivate pathogens on alfalfa seeds by more than 5 log CFU per gram. These results demonstrate that a combination of VHPV and VH has potentially employed as a new method for pasteurization of alfalfa seeds without affecting their germinability.

Antimicrobial and Antioxidant Potential of Scenedesmus obliquus Microalgae in the Context of Integral Biorefinery Concept.

Small-scale photobioreactors (PBRs) in the inoculum stage were designed with internal (red or green) and external white LED light as an initial step of a larger-scale installation aimed at fulfilling the integral biorefinery concept for maximum utilization of microalgal biomass in a multifunctional laboratory. The specific growth rate of Scenedesmus obliquus (Turpin) Kützing biomass for given cultural conditions was analyzed by using MAPLE software. For the determination of total polyphenols, flavonoids, chlorophyll "a" and "b", carotenoids and lipids, UHPLC-HRMS, ISO-20776/1, ISO-10993-5 and CUPRAC tests were carried out. Under red light growing, a higher content of polyphenols was found, while the green light favoured the flavonoid accumulation in the biomass. Chlorophylls, carotenoids and lipids were in the same order of magnitude in both samples. The dichloromethane extracts obtained from the biomass of each PBR synergistically potentiated at low concentrations (0.01-0.05 mg/mL) the antibacterial activity of penicillin, fluoroquinolones or oregano essential oil against the selected food-borne pathogens (Staphylococcus aureus, Escherichia coli and Salmonella typhimurium) without showing any in vitro cytotoxicity. Both extracts exhibited good cupric ion-reducing antioxidant capacity at concentrations above 0.042-0.08 mg/mL. The UHPLC-HRMS analysis revealed that both extracts contained long chain fatty acids and carotenoids thus explaining their antibacterial and antioxidant potential. The applied engineering approach showed a great potential to modify microalgae metabolism for the synthesis of target compounds by S. obliquus with capacity for the development of health-promoting nutraceuticals for poultry farming.

Reduced Antibacterial Drug Resistance and blaCTX-M ß-Lactamase Gene Carriage in Cattle-Associated Escherichia coli at Low Temperatures, at Sites Dominated by Older Animals, and on Pastureland: Implications for Surveillance.

Little is known about the drivers of critically important antibacterial resistance in species with zoonotic potential present on farms (e.g., CTX-M ß-lactamase-positive Escherichia coli). We collected samples monthly between January 2017 and December 2018 on 53 dairy farms in South West England, along with data for 610 variables concerning antibacterial usage, management practices, and meteorological factors. We detected E. coli resistant to amoxicillin, ciprofloxacin, streptomycin, and tetracycline in 2,754/4,145 (66%), 263/4,145 (6%), 1,475/4,145 (36%), and 2,874/4,145 (69%), respectively, of samples from fecally contaminated on-farm and near-farm sites. E. coli positive for blaCTX-M were detected in 224/4,145 (5.4%) of samples. Multilevel, multivariable logistic regression showed antibacterial dry cow therapeutic choice (including use of cefquinome or framycetin) to be associated with higher odds of blaCTX-M positivity. Low average monthly ambient temperature was associated with lower odds of blaCTX-ME. coli positivity in samples and with lower odds of finding E. coli resistant to each of the four test antibacterials. This was in addition to the effect of temperature on total E. coli density. Furthermore, samples collected close to calves had higher odds of having E. coli resistant to each antibacterial, as well as E. coli positive for blaCTX-M Samples collected on pastureland had lower odds of having E. coli resistant to amoxicillin or tetracycline, as well as lower odds of being positive for blaCTX-MIMPORTANCE Antibacterial resistance poses a significant threat to human and animal health and global food security. Surveillance for resistance on farms is important for many reasons, including tracking impacts of interventions aimed at reducing the prevalence of resistance. In this longitudinal survey of dairy farm antibacterial resistance, we showed that local temperature-as it changes over the course of a year-was associated with the prevalence of antibacterial-resistant E. coli We also showed that prevalence of resistant E. coli was lower on pastureland and higher in environments inhabited by young animals. These findings have profound implications for routine surveillance and for surveys carried out for research. They provide important evidence that sampling at a single time point and/or single location on a farm is unlikely to be adequate to accurately determine the status of the farm regarding the presence of samples containing resistant E. coli.

Rhizobacteria Impact Colonization of Listeria monocytogenes on Arabidopsis thaliana Roots.

In spite of its relevance as a foodborne pathogen, we have limited knowledge about Listeria monocytogenes in the environment. L. monocytogenes outbreaks have been linked to fruits and vegetables; thus, a better understanding of the factors influencing its ability to colonize plants is important. We tested how environmental factors and other soil- and plant-associated bacteria influenced L. monocytogenes' ability to colonize plant roots using Arabidopsis thaliana seedlings in a hydroponic growth system. We determined that the successful root colonization of L. monocytogenes 10403S was modestly but significantly enhanced by the bacterium being pregrown at higher temperatures, and this effect was independent of the biofilm and virulence regulator PrfA. We tested 14 rhizosphere-derived bacteria for their impact on L. monocytogenes 10403S, identifying one that enhanced and 10 that inhibited the association of 10403S with plant roots. We also characterized the outcomes of these interactions under both coinoculation and invasion conditions. We characterized the physical requirements of five of these rhizobacteria to impact the association of L. monocytogenes 10403S with roots, visualizing one of these interactions by microscopy. Furthermore, we determined that two rhizobacteria (one an inhibitor, the other an enhancer of 10403S root association) were able to similarly impact 10 different L. monocytogenes strains, indicating that the effects of these rhizobacteria on L. monocytogenes are not strain specific. Taken together, our results advance our understanding of the parameters that affect L. monocytogenes plant root colonization, knowledge that may enable us to deter its association with and, thus, downstream contamination of, food crops. IMPORTANCE Listeria monocytogenes is ubiquitous in the environment, being found in or on soil, water, plants, and wildlife. However, little is known about the requirements for L. monocytogenes' existence in these settings. Recent L. monocytogenes outbreaks have been associated with contaminated produce; thus, we used a plant colonization model to investigate factors that alter L. monocytogenes' ability to colonize plant roots. We show that L. monocytogenes colonization of roots was enhanced when grown at higher temperatures prior to inoculation but did not require a known regulator of virulence and biofilm formation. Additionally, we identified several rhizobacteria that altered the ability of 11 different strains of L. monocytogenes to colonize plant roots. Understanding the factors that impact L. monocytogenes physiology and growth will be crucial for finding mechanisms (whether chemical or microbial) that enable its removal from plant surfaces to reduce L. monocytogenes contamination of produce and eliminate foodborne illness.

"One-step" characterization platform for pathogenic genetics of Staphylococcus aureus.

Staphylococcus aureus (S. aureus) is an important human pathogen causing a variety of life-threatening diseases. In recent years, the health problem caused by S. aureus contaminated food has become a global health problem. S. aureus can express various pathogenic factors, mainly used for adhesion, colonization, invasion and infection of the host. Therefore, rapid and accurate detection of virulence genes in S. aureus is necessary to prevent outbreaks caused by this pathogen. PCR is a useful tool for rapid detection of foodborne pathogens. The objective of this study was to detect the presence of major toxin genes in S. aureus, including sea, seb, sec, see, pvl and tsst, by using a PCR plate. Of the 13 strains tested, 12 (92.3%) were found to be positive for one or more toxin genes. This study realized the one-step detection of main toxin factors in S. aureus.

Antimicrobial Properties of Palladium and Platinum Nanoparticles: A New Tool for Combating Food-Borne Pathogens.

Although some metallic nanoparticles (NPs) are commonly used in the food processing plants as nanomaterials for food packaging, or as coatings on the food handling equipment, little is known about antimicrobial properties of palladium (PdNPs) and platinum (PtNPs) nanoparticles and their potential use in the food industry. In this study, common food-borne pathogens Salmonella enterica Infantis, Escherichia coli, Listeria monocytogenes and Staphylococcus aureus were tested. Both NPs reduced viable cells with the log10 CFU reduction of 0.3-2.4 (PdNPs) and 0.8-2.0 (PtNPs), average inhibitory rates of 55.2-99% for PdNPs and of 83.8-99% for PtNPs. However, both NPs seemed to be less effective for biofilm formation and its reduction. The most effective concentrations were evaluated to be 22.25-44.5 mg/L for PdNPs and 50.5-101 mg/L for PtNPs. Furthermore, the interactions of tested NPs with bacterial cell were visualized by transmission electron microscopy (TEM). TEM visualization confirmed that NPs entered bacteria and caused direct damage of the cell walls, which resulted in bacterial disruption. The in vitro cytotoxicity of individual NPs was determined in primary human renal tubular epithelial cells (HRTECs), human keratinocytes (HaCat), human dermal fibroblasts (HDFs), human epithelial kidney cells (HEK 293), and primary human coronary artery endothelial cells (HCAECs). Due to their antimicrobial properties on bacterial cells and no acute cytotoxicity, both types of NPs could potentially fight food-borne pathogens.

Evaluation of the Antimicrobial Efficacy of N-Acetyl-l-Cysteine, Rhamnolipids, and Usnic Acid-Novel Approaches to Fight Food-Borne Pathogens.

In the food industry, the increasing antimicrobial resistance of food-borne pathogens to conventional sanitizers poses the risk of food contamination and a decrease in product quality and safety. Therefore, we explored alternative antimicrobials N-Acetyl-l-cysteine (NAC), rhamnolipids (RLs), and usnic acid (UA) as a novel approach to prevent biofilm formation and reduce existing biofilms formed by important food-borne pathogens (three strains of Salmonella enterica and two strains of Escherichia coli, Listeria monocytogenes, Staphylococcus aureus). Their effectiveness was evaluated by determining minimum inhibitory concentrations needed for inhibition of bacterial growth, biofilm formation, metabolic activity, and biofilm reduction. Transmission electron microscopy and confocal scanning laser microscopy followed by image analysis were used to visualize and quantify the impact of tested substances on both planktonic and biofilm-associated cells. The in vitro cytotoxicity of the substances was determined as a half-maximal inhibitory concentration in five different cell lines. The results indicate relatively low cytotoxic effects of NAC in comparison to RLs and UA. In addition, NAC inhibited bacterial growth for all strains, while RLs showed overall lower inhibition and UA inhibited only the growth of Gram-positive bacteria. Even though tested substances did not remove the biofilms, NAC represents a promising tool in biofilm prevention.

Effects of Green Tea Powder, Pomegranate Peel Powder, Epicatechin and Punicalagin Additives on Antimicrobial, Antioxidant Potential and Quality Properties of Raw Meatballs.

Alternative technologies, which have been developed in order to meet the consumers' demand for nourishing and healthy meat and meat products, are followed by the food industry. In the present study, it was determined, using the HPLC method, that green tea contains a high level of epicatechin (EP) under optimal conditions and that pomegranate peel contains a high level of punicalagin (PN). Green tea, pomegranate peel, EP and PN were added to meatballs at different concentrations in eight groups. The antioxidant capacities of extracts were measured. The antimicrobial activity was examined for 72 h using three different food pathogens. The highest level of antimicrobial activity was achieved in the 1% punicalagin group, whereas the minimum inhibition concentration (L. monocytogenes, S. typhimurium) was found to be 1.87 mg/mL. A statistically significant decrease was found in FFA, POV and TBARS levels of meatballs on different days of storage (p < 0.05). When compared to the control group, the bioactive compounds preserved the microbiological and chemical properties of meatballs during storage at +4 °C (14 days). It was concluded that the extracts with high EP and PN concentrations can be used as bio-preservative agents for meat and meat products.

Both Handwashing and an Alcohol-Based Hand Sanitizer Intervention Reduce Soil and Microbial Contamination on Farmworker Hands during Harvest, but Produce Type Matters.

Hand hygiene interventions are critical for reducing farmworker hand contamination and preventing the spread of produce-associated illness. Hand hygiene effectiveness may be produce-commodity specific, which could influence implementation strategies. This study's goal was to determine if produce commodity influences the ability of handwashing with soap and water or two-step alcohol-based hand sanitizer (ABHS) interventions to reduce soil and bacteria on farmworker hands. Farmworkers (n = 326) harvested produce (cantaloupe, jalapeño, and tomato) for 30 to 90 minutes before engaging in handwashing, two-step ABHS (jalapeño and cantaloupe), or no hand hygiene. Hands were rinsed to measure amounts of soil (absorbance at 600 nm) and indicator bacteria (coliforms, Enterococcus sp., generic Escherichia coli, and Bacteroidales universal [AllBac] and human-specific [BFD] 16S rRNA gene markers). Without hand hygiene, bacterial concentrations (0.88 to 5.1 log10 CFU/hand) on hands significantly differed by the produce commodity harvested. Moderate significant correlations (ρ = -0.41 to 0.56) between soil load and bacterial concentrations were observed. There were significant produce-commodity-specific differences in the ability of handwashing and two-step ABHS interventions to reduce soil (P < 0.0001), coliforms (P = 0.002), and Enterococcus sp. (P = 0.003), but not the Bacteroidales markers AllBac (P = 0.4) or BFD (P = 0.3). Contamination on hands of farmworkers who harvested cantaloupe was more difficult to remove. Overall, we found that a two-step ABHS intervention was similar to handwashing with soap and water at reducing bacteria on farmworker hands. In summary, produce commodity type should be considered when developing hand hygiene interventions on farms.IMPORTANCE This study demonstrated that the type of produce commodity handled influences the ability of handwashing with soap and water or a two-step alcohol-based hand sanitizer (ABHS) intervention to reduce soil and bacterial hand contamination. Handwashing with soap and water, as recommended by the FDA's Produce Safety Rule, when tested in three agricultural environments, does not always reduce bacterial loads. Consistent with past results, we found that the two-step ABHS method performed similarly to handwashing with soap and water but also does not always reduce bacterial loads in these contexts. Given the ease of use of the two-step ABHS method, which may increase compliance, the two-step ABHS method should be further evaluated and possibly considered for implementation in the agricultural environment. Taken together, these results provide important information on hand hygiene effectiveness in three agricultural contexts.

Inactivation of Escherichia coli, Salmonella enterica serovar Typhimurium, and Bacillus cereus in roasted grain powder by radio frequency heating.

AIMS: The objective of this study was to evaluate the antimicrobial effects of radio frequency (RF) heating and the combination treatment of RF heating with ultraviolet (UV) radiation against foodborne pathogens in roasted grain powder (RGP). METHODS AND RESULTS: Foodborne pathogens inoculated on RGP were subjected to RF heating or RF-UV combination treatments. After 120 s of RF heating, 4·68, 3·89 and 4·54 log reductions were observed for Escherichia coli, Salmonella Typhimurium and Bacillus cereus vegetative cells respectively. The combined RF-UV treatment showed synergistic effects of over 1 log unit compared to the sum of individual treatment for E. coli and S. Typhimurium, but not for B. cereus vegetative cells because of their high UV resistance. Germinated B. cereus cells were not significantly inactivated by RF heating (<1 log CFU per gram), and increased heat resistance compared to the vegetative cells was verified with mild heat treatment. The colour of RGP was not significantly affected by the RF or RF-UV treatments. CONCLUSIONS: Applying RF heating to grain-based food products has advantages for the inactivation of E. coli and S. Typhimurium in RGP. SIGNIFICANCE AND IMPACT OF THE STUDY: The results of the present study could be used as a basis for determining the treatment conditions for inactivating E. coli and other foodborne pathogens such as S. Typhimurium and B. cereus in RGP.

Heat and Chemical Treatments Affect the Viability, Morphology, and Physiology of Staphylococcus aureus and Its Subsequent Antibody Labeling for Flow Cytometric Analysis.

The effects of heat and chemical treatments on Staphylococcus aureus viability and physiology and their subsequent effects on antibody binding ability and cell morphology were measured. Treatments included lethal and sublethal heat; exposure to organic acids, salt, and sodium hydroxide; and freeze-thawing. Strain-related differences in viability were noted depending on treatment and were reflected in changes in physiology as monitored by flow cytometry (FCM) using three different staining protocols: SYTO 9/propidium iodide (PI), DiOC2(3), or calcein acetoxymethyl ester (calcein-AM)/PI. Treatments that resulted in significant losses in viability as measured by plate counting were reflected better by the first two staining combinations, as intracellular calcein-AM uptake may have been impaired by certain treatments. FCM analysis using labeling by commercial anti-S. aureus antibodies indicated that differences in cell physiology as a result of treatments influenced immunofluorescence detection. The ratio of the mean fluorescence intensities of stained cells to those of unstained cells [MFI/MFI(us)] varied with treatment, five of these treatments, including freeze-thaw, citric acid, oxalic acid, NaCl, and NaOH treatments, resulted in significantly lower fluorescence values compared to controls.IMPORTANCE FCM data indicated that cells conventionally considered to be dead and which would not give rise to CFU in a plate count assay, e.g., cells heated to 80°C, were labeled by antibody staining. This finding suggests that without the inclusion of a live/dead discriminating dye, these cells would be erroneously detected as viable within an FCM assay. Reductions in antibody staining due to physicochemical treatment were strain related, reflecting the complexity of the phenomenon under study and illustrating that substantial validation of any new antibody detection-based method, including physiological staining and cell sorting, should be undertaken. Researchers should be aware of physicochemical treatments causing false-negative results: in this study, freeze-thawing severely reduced antibody binding without affecting the viability of a substantial percentage of cells. Scanning electron microscopy carried out on treated cells revealed a range of morphological changes resulting from physicochemical treatments which may have hindered antibody binding.

Assessment of the bioprotective potential of lactic acid bacteria against Listeria monocytogenes on vacuum-packed cold-smoked salmon stored at 8 °C.

Smoked salmon is a highly appreciated delicatessen product. Nevertheless, this ready-to-eat (RTE) product is considered at risk for Listeria monocytogenes, due to both the prevalence and growth potential of this bacteria on the product. Biopreservation may be considered a mild and natural effective strategy for minimizing this risk. In this study, we evaluated the following three potential bioprotective lactic acid bacterial strains against L. monocytogenes in three smoked salmon types with different physicochemical characteristics, primarily fat, moisture, phenol and acid acetic content: two bacteriocin-like producers that were isolated from smoked salmon and identified as Lactobacillus curvatus and Carnobacterium maltaromaticum and a recognized bioprotective bacteriocin producer from meat origin, Lactobacillus sakei CTC494. L. sakei CTC494 inhibited the growth of L. monocytogenes after 21 days of storage at 8 °C in all the products tested, whereas L. curvatus CTC1742 only limited the growth of the pathogen (<2 log increase). The effectiveness of C. maltaromaticum CTC1741 was dependent on the product type; this strain limited the growth of the pathogen in only one smoked salmon type. These results suggest that the meat-borne starter culture, L. sakei CTC494, may potentially be used as a bioprotective culture to improve the food safety of cold-smoked salmon.

Effect of modified atmosphere packaging on physicochemical and microbiological characteristics of Graviera Agraphon cheese during refrigerated storage.

The aim of this work was to examine the effect of modified atmosphere packaging on the physicochemical and microbiological changes of Graviera Agraphon cheese during refrigerated storage. Blocks of Graviera Agraphon cheese weighing around 200 g were packaged under natural (control) or modified atmosphere packaging (MAP) conditions (50% N2 - 50% CO2) and stored at 4 °C or 10 °C for up to 85 d. Prior to packaging, groups of cheese blocks were inoculated with one each of the following foodborne pathogens at around 104 log cfu/g: Listeria monocytogenes, Salmonella Typhimurium, Escherichia coli O157:H7 or Staphylococcus aureus, whilst further groups of cheese blocks were not inoculated. The protein, fat, moisture and salt contents as well as the pH of control and MAP cheese samples did not change significantly (P > 0.05) throughout 4 °C storage, while the pH values of control and MAP cheese samples were significantly (P < 0.05) reduced at 10 °C storage. At 10 °C storage, yeasts and molds, psychrotrophs and lactic acid bacteria (LAB) were significantly higher (P < 0.05) for the normal atmosphere than the MAP cheese samples after the 4th, 8th and 4th days, respectively. At 4 °C storage, the yeasts and molds or psychrotrophs were significantly higher (P < 0.05) than those of control after the 6th and 15th days, respectively at 4 °C storage. All foodborne pathogens showed a higher decrease (P < 0.05) at 10 °C than 4 °C storage. S. aureus proved more sensitive in inactivation in the MAP conditions than atmospheric conditions. L. monocytogenes and S. aureus presented a higher decrease than that of E. coli O157:H7 and S. Typhimurium. In conclusion, MAP proved efficient in retarding the growth of yeasts, molds, psychrotrophs and E. coli O157:H7, L. monocytogenes, S. Typhimurium and S. aureus in Graviera Agraphon cheese during refrigerated storage at 4 and 10 °C.