Chicken Juice Research Articles

Predictive modeling of Salmonella spp. growth behavior in cooked and raw chicken samples: Real-time PCR quantification approach and model assessment in different handling scenarios.

The increasing prevalence of Salmonella contamination in poultry meat emphasizes the importance of suitable predictive microbiological models for estimating Salmonella growth behavior. This study was conducted to evaluate the potential of chicken juice as a model system to predict the behavior of Salmonella spp. in cooked and raw chicken products and to assess its ability to predict cross-contamination scenarios. A cocktail of four Salmonella serovars was inoculated into chicken juice, sliced chicken, ground chicken, and chicken patties, with subsequent incubation at 10, 15, 20, and 25°C for 39h. The number of Salmonella spp. in each sample was determined using real-time polymerase chain reaction. Growth curves were fitted into the primary Baranyi and Roberts model to obtain growth parameters. Interactions between temperature and growth parameters were described using the secondary Ratkowsky's square root model. The predictive results generated by the chicken juice model were compared with those obtained from other chicken meat models. Furthermore, the parameters of the chicken juice model were used to predict Salmonella spp. numbers in six worst-case cross-contamination scenarios. Performance of the chicken juice model was evaluated using the acceptable prediction zone from -1.0 (fail-safe) to 0.5 (fail-dangerous) log. Chicken juice model accurately predicted all observed data points within the acceptable range, with the distribution of residuals being wider near the fail-safe zone (75%) than near the fail-dangerous zone (25%). This study offers valuable insights into a novel approach for modeling Salmonella growth in chicken meat products, with implications for food safety through the development of strategic interventions. PRACTICAL APPLICATION: The findings of this study have important implications in the food industry, as chicken juice could be a useful tool for predicting Salmonella behavior in different chicken products and thus reducing the risk of foodborne illnesses through the development of strategic interventions. However, it is important to recognize that some modifications to the chicken juice model will be necessary to accurately mimic all real-life conditions, as multiple factors particularly those related to food processing can vary between different products.

Transcriptome Analysis of Campylobacter jejuni and Campylobacter coli during Cold Stress.

Campylobacter spp. are known to cause campylobacteriosis, a bacterial disease that remains a public health threat. Campylobacter spp. are prevalent in retail meat and liver products, and the prolonged survival of Campylobacter in the low temperatures needed for storage is a challenge for food safety. In this study, RNA-seq was used for the analysis of the C. coli HC2-48 (Cc48) and C. jejuni OD2-67 (Cj67) transcriptomes at 4 °C in a nutrient-rich medium (chicken juice, CJ) and Mueller-Hinton broth (MHB) for 0 h, 0.5 h, 24 h and 48 h. Differentially expressed genes (DEGs) involved in flagellar assembly were highly impacted by low temperatures (4 °C) in C. coli HC2-48, whereas genes related to the ribosome and ribonucleoprotein complex were modulated for C. jejuni OD2-67 at 4 °C. Most of the DEGs in cells grown at 4 °C in the two medium formulations were not significantly expressed at different incubation times. Although more DEGs were observed in CJ as compared to MHB in both Campylobacter strains, the absence of common genes expressed at all incubation times indicates that the food matrix environment is not the sole determinant of differential expression in Campylobacter spp. at low temperatures.

Interference with Bacterial Conjugation and Natural Alternatives to Antibiotics: Bridging a Gap.

Horizontal gene transfer (HGT) in food matrices has been investigated under conditions that favor gene exchange. However, the major challenge lies in determining the specific conditions pertaining to the adapted microbial pairs associated with the food matrix. HGT is primarily responsible for enhancing the microbial repertoire for the evolution and spread of antimicrobial resistance and is a major target for controlling pathogens of public health concern in food ecosystems. In this study, we investigated Salmonella Heidelberg (SH) and Escherichia coli (EC) regarding gene exchange under conditions mimicking the industrial environment, with the coproducts whey (SL) and chicken juice (CJ). The S. Heidelberg strain was characterized by antibiotic susceptibility standards and PCR to detect the blaTEM gene. A concentration of 0.39 mg/mL was determined to evaluate the anti-conjugation activity of nanostructured lipid nanocarriers (NLCs) of essential oils to mitigate β-lactam resistance gene transfer. The results showed that the addition of these coproducts promoted an increase of more than 3.5 (whey) and 2.5 (chicken juice) orders of magnitude in the conjugation process (p < 0.01), and NLCs of sage essential oil significantly reduced the conjugation frequency (CF) by 74.90, 90.6, and 124.4 times when compared to the transfers in the absence of coproducts and the presence of SL and CJ, respectively. For NLCs from olibanum essential oil, the decrease was 4.46-fold for conjugations without inhibitors and 3.12- and 11.3-fold in the presence of SL and CJ. NLCs associated with sage and olibanum essential oils effectively control the transfer of antibiotic resistance genes and are a promising alternative for use at industrial levels.

Development of a 3-plex droplet digital PCR for identification and absolute quantification of Salmonella and its two important serovars in various food samples

Salmonella is one of the most common foodborne pathogens that cause diarrhea in human, in which Enteritidis and Typhimurium are the top serovars frequently isolated from foodstuffs. A novel 3-plex droplet digital PCR (ddPCR) was successfully developed in this study for the simultaneous identification and absolute quantification of Salmonella and its two important serovars (Enteritidis and Typhimurium). The specificity of all the primers and probes was validated by testing 24 strains of Salmonella and 10 strains of other bacteria. Meanwhile, the anti-interference ability of this method was also evaluated using high proportion of non-target strains or various bacterial combinations. The detection limit of this 3-plex ddPCR was as low as 5 fg/μL for gDNA and 101 CFU/mL for pure cultures, respectively. Moreover, compared with quantitative PCR (qPCR), the detection limit of this ddPCR was much lower when it was applied to the detection of artificially contaminated food samples, which was 101 CFU/mL for lettuce, and 102 CFU/mL for milk and chicken juice samples, respectively. Additionally, a total of 100 retailed food samples were tested by this validated 3-plex ddPCR, which was also compared with traditional cultivation method. Notably, 19 of 20 samples showed consistent positive results by both methods, while the rest with the lowest concentration of Salmonella cells was detectable only by ddPCR. These results demonstrated that the 3-plex ddPCR established in this study presented highly specific, sensitive, and absolute quantitative capability, which had the potential to identify Salmonella, and to differentiate the two important serovars Enteritidis and Typhimurium in a wide variety of food samples.

Propionate and Butyrate Inhibit Biofilm Formation of Salmonella Typhimurium Grown in Laboratory Media and Food Models.

Salmonella is among the most frequently isolated foodborne pathogens, and biofilm formed by Salmonella poses a potential threat to food safety. Short-chain fatty acids (SCFAs), especially propionate and butyrate, have been demonstrated to exhibit a beneficial effect on promoting intestinal health and regulating the host immune system, but their anti-biofilm property has not been well studied. This study aims to investigate the effects of propionate or butyrate on the biofilm formation and certain virulence traits of Salmonella. We investigated the effect of propionate or butyrate on the biofilm formation of Salmonella enterica serovar Typhimurium (S. Typhimurium) SL1344 grown in LB broth or food models (milk or chicken juice) by crystal violet staining methods. Biofilm formation was significantly reduced in LB broth and food models and the reduction was visualized using a scanning electron microscope (SEM). Biofilm metabolic activity was attenuated in the presence of propionate or butyrate. Meanwhile, both SCFAs decreased AI-2 quorum sensing based on reporter strain assay. Butyrate, not propionate, could effectively reduce bacterial motility. Bacterial adhesion to and invasion of Caco-2 cells were also significantly inhibited in the presence of both SCFAs. Finally, two SCFAs downregulated virulence genes related to biofilm formation and invasion through real-time polymerase chain reaction (RT-PCR). These findings demonstrate the potential application of SCFAs in the mitigation of Salmonella biofilm in food systems, but future research mimicking food environments encountered during the food chain is necessitated.

Antimicrobial activity of Thymus zygis essential oil against Listeria monocytogenes and its application as food preservative

Thymus zygis is an aromatic plant used in folk medicine. This work aimed to evaluate the anti-Listeria monocytogenes activity of T. zygis essential oil (EO), whose thymol is its major compound. Furthermore, the attenuation of this bacterium's virulence, namely by the inhibition of biofilm formation, motility and invasion of human cells, and the possible application of the EO in food were evaluated. The T. zygis EO showed antibacterial activity against L. monocytogenes with a minimum inhibitory concentration (MIC) of 0.05%, while showing a bactericidal effect. The EO significantly reduced the biofilm formation (inhibition from 16.85 to 89.86%) and motility (halos between 6.66 and 10.98 mm, compared to controls 13.12 to 17.22 mm), and not inducing cross-resistance to antibiotics, such as ampicillin, cefotaxime, erythromycin, gentamicin, tetracycline, and vancomycin. L. monocytogenes counts (initial inoculum of ~106 CFU/mL) were lowered by the use of 2× MIC of T. zygis EO in the chicken juice (1.53 log CFU/mL) and lettuce model (to below the detection limit) after two days of storage. The use of EO (0.2% (v/v)) for sanitizing fresh vegetables, reduce L. monocytogenes and natural microbiota for values below the detection limit of the method for iceberg lettuce after an immersion of 5 min. For the spinach, L. monocytogenes was reduced in 4.35 log CFU/mL and the natural microbiota was diminished in a range of 4.47 to 5.94 log CFU/mL, when compared with the washing with water. Overall, the T. zygis EO has demonstrated a promising antimicrobial activity and these findings point to the potential of the EO as a natural food preservative or sanitizer for controlling L. monocytogenes in food products.

Biofilm Formation of Listeria monocytogenes and Pseudomonas aeruginosa in a Simulated Chicken Processing Environment.

This study aims to investigate the mono- and dual-species biofilm formation of Listeria monocytogenes and Pseudomonas aeruginosa incubated in different culture mediums, inoculum ratios, and incubation time. The planktonic cell population and motility were examined to understand the correlation with biofilm formation. The results showed that chicken juice significantly inhibited the biofilm formation of L. monocytogenes (p < 0.05). Pseudomonas aeruginosa was the dominant bacteria in the dual-species biofilm formation in the trypticase soy broth medium. The dynamic changes in biofilm formation were not consistent with the different culture conditions. The growth of planktonic L. monocytogenes and P. aeruginosa in the suspension was inconsistent with their growth in the biofilms. There was no significant correlation between motility and biofilm formation of L. monocytogenes and P. aeruginosa. Moreover, scanning electron microscopy (SEM) results revealed that the biofilm structure of L. monocytogenes was loose. At the same time, P. aeruginosa formed a relatively dense network in mono-species biofilms in an initial adhesion stage (24 h). SEM results also showed that P. aeruginosa was dominant in the dual-species biofilms. Overall, these results could provide a theoretical reference for preventing and controlling the biofilm formation of L. monocytogenes and P. aeruginosa in the food processing environment in the future.

Antimicrobial and Antibiofilm Effect of ε-Polylysine against Salmonella Enteritidis, Listeria monocytogenes, and Escherichia coli in Tryptic Soy Broth and Chicken Juice.

ε-Polylysine (ε-PL) is a safe food additive that is used in the food industry globally. This study evaluated the antimicrobial and antibiofilm activity of antibacterial peptides (ε-PL) against food poisoning pathogens detected in chicken (Salmonella Enteritidis, Listeria monocytogenes, and Escherichia coli). The results showed that minimum inhibitory concentrations (MICs) ranged between 0.031–1.0 mg/mL, although most bacterial groups (75%) showed MICs of 1.0 mg/mL. The reduction in the cell viability of pathogens due to ε-PL depended on the time and concentration, and 1/2 × MIC of ε-PL killed 99.99% of pathogens after 10 h of incubation. To confirm biofilm inhibition and degradation effects, crystal violet assay and confocal laser scanning microscopy (CLSM) were used. The biofilm formation rates of four bacterial groups (Salmonella, Listeria, E. coli, and multi-species bacteria) were 10.36%, 9.10%, 17.44%, and 21.37% at 1/2 × MIC of ε-PL, respectively. Additionally, when observed under a CLSM, ε-PL was found to induce biofilm destruction and bacterial cytotoxicity. These results demonstrated that ε-PL has the potential to be used as an antibiotic and antibiofilm material for chicken meat processing.

Molecular Characterization and Survive Abilities of Salmonella Heidelberg Strains of Poultry Origin in Brazil.

The aim of the study was to evaluate the genotypic and phenotypic characteristics of 20 strains of S. Heidelberg (SH) isolated from broilers produced in southern Brazil. The similarity and presence of genetic determinants linked to virulence, antimicrobial resistance, biofilm formation, and in silico-predicted metabolic interactions revealed this serovar as a threat to public health. The presence of the ompC, invA, sodC, avrA, lpfA, and agfA genes was detected in 100% of the strains and the luxS gene in 70% of them. None of the strains carries the blaSHV, mcr-1, qnrA, qnrB, and qnrS genes. All strains showed a multidrug-resistant profile to at least three non-β-lactam drugs, which include colistin, sulfamethoxazole, and tetracycline. Resistance to penicillin, ceftriaxone (90%), meropenem (25%), and cefoxitin (25%) were associated with the presence of blaCTX–M and blaCMY–2 genes. Biofilm formation reached a mature stage at 25 and 37°C, especially with chicken juice (CJ) addition. The sodium hypochlorite 1% was the least efficient in controlling the sessile cells. Genomic analysis of two strains identified more than 100 virulence genes and the presence of resistance to 24 classes of antibiotics correlated to phenotypic tests. Protein-protein interaction (PPI) prediction shows two metabolic pathways correlation with biofilm formation. Virulence, resistance, and biofilm determinants must be constant monitoring in SH, due to the possibility of occurring infections extremely difficult to cure and due risk of the maintenance of the bacterium in production environments.

Antibiotic Resistance in the Alternative Lifestyles of Campylobacter jejuni.

Campylobacter jejuni is the main pathogen identified in cases of foodborne gastroenteritis worldwide. Its importance in poultry production and public health is highlighted due to the growing antimicrobial resistance. Our study comparatively investigated the effect of five different classes of antimicrobials on the planktonic and biofilm forms of 35 strains of C. jejuni with high phylogenetic distinction in 30 of them. In the planktonic form, the existence of susceptible strains to colistin (7/35 – 20%) and resistance to meropenem (3/35 – 8.6%) represent a novelty in strains evaluated in Brazil. In biofilms formed with the addition of chicken juice, the number of resistant strains was significantly higher for colistin, erythromycin and meropenem (100%), but the susceptibility to tetracycline was shown as a control strategy for specific cases. High concentrations (1,060 ± 172.1mg/L) of antibiotics were necessary to control the biofilm structure in susceptible strains in the planktonic form, which is consistent with the high biomass produced in these strains. Stainless steel and polyurethane were the most (BFI=2.1) and least (BFI=1.6) favorable surfaces for the production of biomass treated with antimicrobials. It is concluded that the antimicrobial action was detected for all tested drugs in planktonic form. In sessile forms, the biomass production was intensified, except for tetracycline, which showed an antibiofilm effect.

Salmonella enterica Serovar Minnesota Biofilms, Susceptibility to Biocides, and Molecular Characterization.

The presence of virulence genes, phylogenetic relationships, biofilm formation index (BFI), and ultrastructure in S. Minnesota at different temperatures (4, 25, and 36 °C) were analyzed. In addition, the ability of biocidal agents (chlorhexidine1%, sodium hypochlorite 1%, and peracetic acid 0.8%) to inhibit biofilms formed by 20 strains isolated from broiler slaughter plants from two Brazilian companies in 2009, 2010, and 2014 was determined. The presence of specific genes was evaluated by PCR and phylogeny between strains by pulsed-field gel electrophoresis. The BFI was determined using tryptone soy broth with 5% of chicken juice, and its structure was observed by scanning electron microscopy. The presence of specific genes indicated that S. Minnesota has the potential to cause disease in humans, adapting to adverse conditions. Temperatures of 25 and 36 °C favored biofilm formation, although at 4 °C, there was still biomass that could contaminate the final product. Tolerance to all biocides was identified in 12/20 (60%), representing a real risk of adaptation mechanisms development, especially regarding to resistance to sodium hypochlorite. Phylogenetic analysis indicated cross-contamination and spread among companies, which was probably related to biofilms formation. Results show the necessity of attention to this serovar considering its resistance to sodium hypochlorite, including the need for rigorous control, adopting low temperatures to prevent biofilms formation in the poultry industry.

Meat juice contributes to the stability of ethanol adaptation in Salmonella enterica serovar Enteritidis

Abstract Stability assessment of observed tolerance phenotypes is integral in understanding stress adaptation in food-borne pathogens. Therefore, the current work was carried out to determine whether ethanol adaptation induced by exposure to 5 per cent ethanol for 60 min is a stable phenomenon in Salmonella enterica serovar Enteritidis. The capacity of Salmonella Enteritidis (S. Enteritidis) to maintain the acquired ethanol adaptation in the absence of sublethal ethanol stress was investigated at 37 °C, 25 °C or 4 °C in Luria–Bertani broth and two types of meat juice. It was found that ethanol adaptation was completely reversed within 40 min at 37 °C or within 60 min at 25 °C, but was stable at 4 °C for at least 48 h in the broth assay. Ethanol adaptation was retained in chicken juice during 60-min incubation at 25 °C or 48-h incubation at 4 °C. Moreover, exposure to pork juice stored at either 25 °C or 4 °C significantly (P&amp;lt;0.05) increased the ethanol tolerance of ethanol-adapted cells. Collectively, these findings suggest that ethanol adaptation stability in S. Enteritidis under cold conditions and in meat juices should be taken into account when conducting a comprehensive risk analysis during food processing.

Determination of creatine, creatinine, free amino acid and heterocyclic aromatic amine contents of plain beef and chicken juices.

In the present study, creatine, creatinine, free amino acid and heterocyclic aromatic amine (HAA) contents of plain beef and chicken juices were investigated. For this aim, the boned and boneless beef and chicken juices were obtained by boiling for 3h at 100°C without any additives in the laboratory. Creatine and creatinine content of the beef juices varied between 1.33-3.16mgg-1 and 1.29-1.42mgg-1, respectively, while creatine and creatinine content of the chicken juices varied between 0.98-1.63mgg-1 and 0.89-1.30mgg-1, respectively. The total free amino acid content in the beef juices and the chicken juices ranged between 14.61-20.65mg 100gdm-1 and 19.66-57.88mg 100gdm-1, respectively. None of the heterocyclic aromatic amines analyzed in the present study could be detected in the beef juices and the chicken juices. Therefore, the boned and boneless beef and chicken juice can be considered as safe from the standpoint of these HAAs and are suggested to use in the preparation of meals.

Comparison of Campylobacter jejuni Slaughterhouse and Surface-Water Isolates Indicates Better Adaptation of Slaughterhouse Isolates to the Chicken Host Environment.

Campylobacter jejuni is an emerging food-borne pathogen that poses a high risk to human health. Knowledge of the strain source can contribute significantly to an understanding of this pathogen, and can lead to improved control measures in the food-processing industry. In this study, slaughterhouse and surface-water isolates of C. jejuni were characterized and compared in terms of their antimicrobial resistance profiles and adhesion to stainless steel and chicken skin. Resistance of C. jejuni biofilm cells to benzalkonium chloride and Satureja montana ethanolic extract was also tested. The data show that the slaughterhouse isolates are more resistant to ciprofloxacin, and adhere better to stainless steel at 42 °C, and at 37 °C in 50% chicken juice. Additionally, biofilm cells of the isolate with the greatest adhesion potential (C. jejuni S6) were harvested and tested for resistance to S. montana ethanolic extract, benzalkonium chloride, and erythromycin; and for efflux-pump activity, as compared to their planktonic cells. The biofilm cells showed increased resistance to both S. montana ethanolic extract and erythromycin, and increased efflux-pump activity. These data indicate adaptation of C. jejuni slaughterhouse isolates to the chicken host, as well as increased biofilm cell resistance due to increased efflux-pump activity.

Chlorous acid is a more potent antibacterial agent than sodium hypochlorite against Campylobacter

Foodborne disease caused by campylobacters is one of the major global problems for food safety. Infection source of Campylobacter to human is mainly through contaminated meat particularly chicken. Contamination of meat with Campylobacter usually occurs during processing at slaughterhouse and to prevent such contaminations, sodium hypochlorite is commonly used. However, it is well known that bactericidal activity of sodium hypochlorite becomes weak under organic matter rich conditions. In this study, we compared the strength of bactericidal activity of chlorous acid and sodium hypochlorite against Campylobacter jejuni and Campylobacter coli strains under organic matter rich conditions. Bactericidal activity against 5 representative C. jejuni and C. coli strains in chicken juice (an organic matter rich condition) showed that minimum concentration of chlorous acid required for complete killing of C. jejuni and C. coli cells is 200–400 ppm while that of sodium hypochlorite is 2,000 to 4,000 ppm. Similar results were obtained by using Bolton broth. Furthermore, it was observed that 400 ppm of chlorous acid but not 400 ppm of sodium hypochlorite is highly effective in killing of 25 different Campylobacter strains (12 C. jejuni and 13 C. coli strains) under the same conditions. To determine whether 400 ppm of chlorous acid treatment had killed bacterial cells or induced them into viable but non-culturable (VBNC) state, live and dead cell assay using DAPI and propidium iodide fluorescent dyes was done. Such assay clearly indicated that Campylobacter cells were indeed killed and not induced to VBNC state. Moreover, SDS-PAGE analysis of whole-cell lysates of campylobacters indicated distinct effects in protein profiles of chlorous acid but not sodium hypochlorite treated cells. The results strongly suggest that chlorous acid could efficiently kill C. jejuni and C. coli cells with much lower concentration than sodium hypochlorite and the bactericidal mechanisms of chlorous acid may be due to damages of bacterial proteins. Thus, chlorous acid could be a better disinfectant in chicken slaughtering and processing to kill campylobacters and prevent contamination.

Food residuals on the food‐contacting surfaces of stainless steel and polypropylene influence the efficacy of ultraviolet light in killing foodborne pathogens

AbstractThis study was conducted to examine effects of food residues on the survivals of pathogens on stainless steel (SSS) and polypropylene (PPP) after ultraviolet‐radiation (UVR) surface decontamination. Cultures of Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, Staphylococcus aureus, and Listeria monocytogenes were inoculated on coupons containing deionized water, tryptic soy broth, pork, chicken, cabbage, and milk, respectively. The surface coupons were incubated at 100% relative humidity (RH) and 25°C for 24 hr to produce their own biofilms. UVR (=254 nm) surface decontamination for 120 min resulted in bacterial reductions in the levels of ≥2.5 log10 cfu/coupon. Populations of S. Typhimurium and L. monocytogenes within biofilms declined on SSS harboring chicken juice after UVR for 120 min, showing by &lt;1.5 log10‐reductions. The presence of food residues on the food‐contacting surfaces would facilitate the strong adhesion of these organisms, indicating that bacteria enclosed in biofilms were more resistant to UVR sanitization.Practical applicationsIn this study, it seemed to indicate that the incidence of varying food residues such as deionized water, tryptic soy broth, pork, chicken, cabbage, and milk could affect bacterial adhesion–attachment to the food‐contacting surfaces significantly. At least, some food residues such as pork, chicken, and milk would be supposed to provide a favorable environment where major foodborne pathogens are able to grow and produce their biofilms strongly. Especially, it was determined that the gram‐negative bacteria such as Escherichia coli O157:H7 and Salmonella typhimurium showed the higher sensitivity on coupons against ultraviolet‐radiation surface decontamination.

Evaluation of a Hydrogel Matrix for Baiting Western Yellowjacket (Vespidae: Hymenoptera).

Baiting is an effective method to manage Vespula spp. yellowjacket (Hymenoptera: Vespidae) populations without having to locate and treat nests. Here, we assessed the utility of a commercially available polyacrylamide hydrogel as an alternative bait material for yellowjacket baiting. The experimental bait (hereafter referred to as 'hydrogel bait') consisted of diluted chicken juice (from canned chicken meat) and fipronil (0.025%, wt/wt) absorbed into granular polyacrylamide hydrogel particles. Three separate 24-h baiting trials were conducted at two different field sites with the western yellowjacket, Vespula pensylvanica (Saussare), as the target species. The monitoring data from pre- and posttreatment periods indicated that baiting with polyacrylamide hydrogel baits provided ≈74-96% reduction in the foraging activity of V. pensylvanica during its active season. In addition to their ability to absorb large quantities of aqueous bait containing phagostimulants and toxicants, the hydrogels' tactile resemblance to fresh meat upon hydration makes them a promising option as a non-meat material for delivering small amounts of insecticides to yellowjacket populations in a highly targeted manner.

Predictive Modeling for the Growth of Salmonella Enteritidis in Chicken Juice by Real-Time Polymerase Chain Reaction.

The goals of this study were to monitor the growth kinetics of Salmonella Enteritidis in chicken juice using real-time polymerase chain reaction (PCR) and to evaluate its efficacy by comparing the results with an experimental database. Salmonella Enteritidis was inoculated in chicken juice samples at an initial inoculum of 104 CFU/mL with inoculated samples incubated at six different temperatures (10, 15, 20, 25, 30, and 35°C). Sampling was carried out for 36 h to observe the growth of Salmonella Enteritidis. The total DNA was extracted from the samples, and the copy number of the Salmonella invasion gene (invA) was quantified by real-time PCR and converted to Salmonella Enteritidis cell concentration. Growth kinetics data were analyzed by the Baranyi and Roberts model to obtain growth parameters, whereas the Ratkowsky's square-root model was used to describe the effect of the interactions between growth parameters and temperature on the growth of Salmonella Enteritidis. The growth parameters of Salmonella Enteritidis obtained from an experiment conducted at a constant temperature were validated with growth data from chicken juice samples that were incubated under fluctuating temperature conditions between 5°C and 30°C for 30-min periods. A high correlation was observed between maximum growth rate (μmax) and storage temperature, indicating that the real-time PCR-monitoring method provides a precise estimation of Salmonella Enteritidis growth in food material with a microbial flora. Moreover, the μmax data reflected data from microbial responses viewer database and ComBase. The results of this study suggested that real-time PCR monitoring provides a precise estimation of Salmonella Enteritidis growth in food materials with a background microbial flora.

Influence of milk, chicken residues and oxygen levels on biofilm formation on stainless steel, gene expression and small RNAs in Salmonella enterica

Salmonella spp. are highly versatile foodborne pathogens as they can adapt and shift from the aerobic environment outside the host to the anaerobic environment inside the host through changes in gene expression. These changes can be mediated by small RNAs (sRNAs), a class of post-transcriptional regulators that can modulate diverse functions from biofilm formation to motility and virulence. In this study, biofilm formation on stainless steel, motility, morphotype, and transcription of biofilm- and virulence-related genes and sRNAs were evaluated in fourteen Salmonella enterica strains under aerobiosis, microaerobiosis and anaerobiosis conditions. In order to mimic actual food industry conditions and to compare with laboratory media, chicken exudate and milk were used as culture media. In all growth media tested, biofilm formation was significantly greater (P < 0.05) in aerobiosis than in microaerobiosis or anaerobiosis. The RDAR (red, dry, and rough) morphotype was only produced in aerobiosis, while motility was significantly higher in anaerobiosis (P < 0.05) than in microaerobiosis or aerobiosis. Whereas the gene csgD, which codifies the biofilm master regulator, was downregulated, the virulence genes hilA and invA were upregulated in microaerobiosis and anaerobiosis in all growth media. The transcription of sRNAs was highly influenced by both atmosphere and growth media. Positive regulators of biofilm formation arcZ, sroC and csrB were downregulated in microaerobiosis and anaerobiosis in comparison to aerobiosis. Curiously, the negative regulators of biofilm formation oxyS and rprA were also downregulated in microaerobiosis and anaerobiosis. However, the virulence promoter and negative regulator of biofilm formation dsrA was significantly upregulated in chicken juice. The results of this study indicate that oxygen levels have a considerable influence on biofilm formation and motility in Salmonella. Modification of the gene expression and transcription of sRNAs could be useful in understanding how Salmonella adapt to different conditions within the food chain. This information could then be taken into account in the development of new strategies with which to control the growth of this pathogen.

Effect of Pseudomonas aeruginosa on the sanitizer sensitivity of Salmonella Enteritidis biofilm cells in chicken juice

Salmonella Enteritidis may be encountered in poultry processing plants in the form of biofilms on food contact surfaces. Little is known about the fate of Salmonella in mixed-species biofilms with other bacteria commonly found in food processing environment. Thus, the objectives of this study were to determine biofilm formation and sanitizer resistance of two S. Enteritidis strains (rdar and bdar, cellulose expression positive and negative, respectively), in the absence and presence of Pseudomonas aeruginosa, under conditions encountered in poultry processing plants. S. Enteritidis in single-species and dual-species biofilms with P. aeruginosa at two mixed ratios (Salmonella: Pseudomonas cells at 1:1 and 0.01:1, respectively) were formed on stainless steel coupons in chicken juice for 6 days at 25 °C. On days 1 and 6 of the incubation period, biofilm cell populations were determined before and after chlorine treatment (50 ppm, 1 min). Results showed that population of S. Enteritidis in dual-species biofilms was significantly lower (P < 0.05) than that in single-species biofilms on day 6. On the other hand, S. Enteritidis in dual-species biofilms demonstrated greater chlorine resistance, with an average of 1 log CFU/cm2 lower reduction than that in single-species biofilms, except for the rdar strain which became more sensitive to chlorine treatment in dual-species biofilms after 6 days. Fluorescence microscopy images showed that more glycoconjugates were present in the dual-species biofilms, which might have contributed to the enhanced chlorine resistance of S. Enteritidis. Thus, this study indicates that although the presence of P. aeruginosa could negatively influence S. Enteritidis biofilm formation, it might protect Salmonella cells from sanitizer treatment in poultry processing environment.