Phage cocktail administration to reduce Salmonella load in broilers.

Salmonella is a serious foodborne pathogen that can cause gastrointestinal disease through the consumption of contaminated foods; including poultry meat. Salmonella is commonly present in the intestinal tract of poultry and farm environments, posing a potential risk of contamination during the processing of poultry meat. This study was a continuation in evaluating the effects of our previously developed phage cocktail targeting Salmonella at large-scale trials in commercial broiler farms, in which this cocktail considerably lowered Salmonella colonization in the gut of broilers. The phage cocktail given to broilers showed resistance to temperatures of up to 65 °C (> 60% survivability), pH ranging from 2 to 12 (> 96% survivability), 0.5 to 15% (w/v) NaCl (> 98% survivability), chlorine up to 0.5% (v/v) (53% survivability), and chlorine neutralizer (100% survivability). In the animal challenge study, phage treatments, designed as "prevention" and "exclusion" programs, could control Salmonella on day 20 and 32 of the experiment, respectively; as indicated by the absence of Salmonella detection in cloacal swabs from broilers (0% prevalence). In the commercial-scale trial I, Salmonella was not detected in the phage-treated group from cloacal swabs, boot cover swabs, and bedding material samples after 16 days (0% prevalence) of phage administration. In the commercial-scale trial II, phage treatment extended the Salmonella control period in broilers during a 40-day growout period. In summary, a phage cocktail demonstrated high efficiency in controlling various serovars of Salmonella historically linked to contamination on these broiler farms. Phage cocktail application offers an effective, alternative to enhance food safety within the poultry value chain, protecting consumers and as well as the economic sustainability of the poultry sector.

Control of Salmonella in Chicken Meat by a Phage Cocktail in Combination with Propionic Acid and Modified Atmosphere Packaging.

Salmonella contamination in poultry meat is an important food safety issue as this pathogen can lead to serious illness and economic losses worldwide. In poultry meat processing, a variety of strong bacteriostatic agents has been introduced for controlling Salmonella including bacteriophages (phages), organic acids, and modified atmosphere packaging (MAP). In our study, two selected phages including vB_SenM_P7 and vB_SenP_P32 were used in combination with propionic acid (PA) and MAP for controlling Salmonella of multiple serovars on chicken meat under storage at 4 °C. The two phages showed strong lytic activity against over 72 serovars of Salmonella tested (25.0 to 80.6%). Phages, vB_SenM_P7 and vB_SenP_P32 showed 40% and 60% survival rates, respectively, after the exposure to temperatures up to 70 °C. Both phages remained active, with nearly 100% survival at a wide range of pH (2 to 12) and 15% NaCl (w/v). The available chlorine up to 0.3% (v/v) led to a phage survival rate of 80-100%. A combination of Salmonella phage cocktail and 0.5% PA could reduce Salmonella counts in vitro by 4 log CFU/mL on day 3 whereas a phage cocktail and 0.25% PA showed a 4-log reduction on day 5 during storage at 4 °C. For the phage treatment alone, a 0.3-log reduction of Salmonella was observed on day 1 of storage at 4 °C. In the chicken meat model, treatment by a phage cocktail and PA at both concentrations in MAP conditions resulted in a complete reduction of Salmonella cells (4-5 log unit/g) on day 2 of storage whereas each single treatment under MAP conditions showed a complete cell reduction on day 4. For the meat sensory evaluation, chicken meat treated with a phage cocktail-PA (0.5%) in MAP condition showed the highest preference scores, suggesting highly acceptability and satisfactory. These findings suggest that a combined treatment using a phage cocktail and PA in MAP conditions effectively control Salmonella in poultry meat during storage at low temperature to improve the quality and safety of food.

Effectiveness of the Organic Acid-Based Antimicrobial Agent to Prevent Bacterial Contamination in Fish Meal.

Animal feed production is an important step of the food animal production chain in a farm-to-table model. The contamination of raw ingredients with foodborne pathogens in feed production remains as an important safety issue where pathogens may spread into food animals to cause illnesses in humans when affected food animals are consumed. In the present study, we aimed to examine the quality and microbial contamination of fish meal and to investigate the effectiveness of the organic acid-based antimicrobial agent SALTEC 514TM against Salmonella to prevent bacterial contamination in fish meal. Fish meal samples (n = 4) collected from feed mills at different locations were analyzed for protein and total volatile basic nitrogen (TVBN) content to assess their nutritional value and freshness, and its microbiological quality. The protein and TVBN content ranged from 53.2 ± 3.1 to 67.5 ± 2.3 g/100 g and 73.8 ± 4.5 to 100.4 ± 11.2 mg/100 g meal, respectively. Total plate count of the fish meal samples ranged from 2.0 ± 0.3 to 4.5 ± 0.5 log units, whereas suspected foodborne bacteria, Escherichia coli and Salmonella, were not detected in all samples. Fish meal samples were artificially contaminated (day 0) and re-challenged (day 30 and 90) with Salmonella Enteritidis (3 log CFU/g) to test for the effectiveness of SALTEC 514TM, an organic acid-based antimicrobial formulation, in preventing Salmonella contamination and recontamination during storage. SALTEC 514TM, when applied at three different doses, was found to reduce the number of Salmonella in monitored samples after one day of storage. A low dose of 0.5 kg/ton SALTEC 514TM prevented Salmonella recontamination from occurring in fish meal samples stored for 37 days. In medium (1.0 kg/ton) and high doses (3.0 kg/ton), applications of SALTEC 514TM prevented the Salmonella recontamination for a maximum storage duration of 97 days. The application of SALTEC 514TM in fish meal and/or other feed ingredients may prove to be a safe alternative to reduce the microbial load, especially of foodborne-related microorganisms, to contribute to feed and food safety.

Oral Administration of a Phage Cocktail to Reduce Salmonella Colonization in Broiler Gastrointestinal Tract-A Pilot Study.

Salmonella contamination in poultry meat products can lead to serious foodborne illness and economic loss from product recalls. It is crucial to control Salmonella contamination in poultry from farm to fork. Bacteriophages (phages) are viruses of bacteria that offer several advantages, especially their specificity to target bacteria. In our study, three Salmonella phages (vB_SenS_KP001, vB_SenS_KP005, and vB_SenS_WP110) recovered from a broiler farm and wastewater treatment stations showed high lysis ability ranging from 85.7 to 96.4% on over 56 serovars of Salmonella derived from several sources, including livestock and a broiler farm environment. A three-phage cocktail reduced S. Enteritidis and S. Typhimurium, in vitro by 3.9 ± 0.0 and 3.9 ± 0.2 log units at a multiplicity of infection (MOI) of 103 and 3.8 ± 0.4 and 4.1 ± 0.2 log units at MOI of 104 after 6 h post-phage treatment. A developed phage cocktail did not cause phage resistance in Salmonella during phage treatments for three passages. Phages could survive under simulated chicken gastrointestinal conditions in the presence of gastric acid for 2 h (100.0 ± 0.0% survivability), bile salt for 1 h (98.1 ± 1.0% survivability), and intestinal fluid for 4 h (100 ± 0.0% survivability). Each phage was in the phage cocktail at a concentration of up to 9.0 log PFU/mL. These did not cause any cytotoxicity to human fibroblast cells or Caco-2 cells as indicated by the percent of cell viability, which remained nearly 100% as compared with the control during 72 h of co-culture. The phage cocktail was given to broilers raised in commercial conditions at a 9 log PFU/dose for five doses, while naturally occurring Salmonella cells colonized in the gastrointestinal tract of broilers were significantly reduced as suggested by a considerably lower Salmonella prevalence from over 70 to 0% prevalence after four days of phage treatment. Our findings suggest that a phage cocktail is an effective biocontrol agent to reduce Salmonella present in the guts of broilers, which can be applied to improve food safety in broiler production.

Broad lytic spectrum of novel Salmonella phages on ciprofloxacin-resistant Salmonella contaminated in the broiler production chain.

Background and Aim: Ciprofloxacin (CIP) is recommended for salmonellosis treatment as the drug of choice; however, overuse of this drug can cause drug resistance issues and failure to treat diseases. Phage therapy is an alternative approach for combatting CIP-resistant infection. This study aimed to estimate the prevalence of CIP-resistant Salmonella isolated from the broiler production chain and evaluated the lytic ability of novel Salmonella phages isolated from water samples. Materials and Methods: Samples were obtained from the broiler production chain and used for Salmonella isolation. serovar and CIP resistance of each isolate were characterized through latex agglutination and agar disk diffusion test, respectively. Water samples from different sources were acquired for phage isolation. The lytic activity of novel-isolated phages was also examined. Results: In this study, 51 Salmonella isolates were recovered from the broiler production chain (two commercial farms, one free-range farm, two slaughterhouses, and three stalls from the wet market). Kentucky was the major serovar characterized (16), followed by Typhimurium (9), Agona (5), Corvalis (5), Schwarzengrund (5), Singapore (3), Weltevreden (3), Mbandaka (2), Give (2), and Albany (1). The serovars that exhibited CIP resistance were 14/16 isolates of serovar Kentucky (87.5%) and one isolate of serovar Give (50%), whereas eight other serovars were susceptible to this drug. Overall, the prevalence of CIP-resistant Salmonella recovered from the sources included in this study was 29.4%. This study identified 11 Salmonella phages isolated from wastewater samples derived from broiler farms, wastewater treatment stations, and natural reservoirs. Our phages showed the total percentage of lysis ability ranging from 33.3% to 93.3% against CIP-resistant isolates. However, only one bacterial isolate, namely 210SL, recovered from the food contact surface of a wet market stall and was resistant to all phages. Conclusion: Diverse serovars of Salmonella were recovered in the broiler production chain in this study, while the isolates presenting CIP-resistant Salmonella were as high as 29.4%. Overall, Salmonella phages showed high lysis ability against these CIP-resistant Salmonella isolates, suggesting the potential application of phage-based treatments or biocontrol in the broiler production chain.

Genomic Analysis of Prophages Recovered from Listeria monocytogenes Lysogens Found in Seafood and Seafood-Related Environment.

A prophage is a phage-related sequence that is integrated into a bacterial chromosome. Prophages play an important role in bacterial evolution, survival, and persistence. To understand the impact of Listeria prophages on their host genome organizations, this work sequenced two L. monocytogenes strains (134LM and 036LM), previously identified as lysogens by mitomycin C induction. Draft genomes were generated with assembly sizes of 2,953,877 bp and 3,000,399 bp. One intact prophage (39,532 bp) was inserted into the comK gene of the 134LM genome. Two intact prophages (48,684 bp and 39,488 bp) were inserted in tRNA-Lys and elongation-factor genes of the 036LM genome. The findings confirmed the presence of three corresponding induced phages previously obtained by mitomycin C induction. Comparative genomic analysis of three prophages obtained in the newly sequenced lysogens with 61 prophages found in L. monocytogenes genomes, available in public databases, identified six major clusters using whole genome-based phylogenetic analysis. The results of the comparative genomic analysis of the prophage sequences provides knowledge about the diversity of Listeria prophages and their distribution among Listeria genomes in diverse environments, including different sources or geographical regions. In addition, the prophage sequences and their insertion sites contribute to the genomic diversity of L. monocytogenes genomes. These data of prophage sequences, prophage insertion sites, and prophage sequence comparisons, together with ANIb confirmation, could be useful for L. monocytogenes classification by prophages. One potential development could be refinement of prophage typing tools for monitoring or surveillance of L. monocytogenes contamination and transmission.

Isolation and Characterization of Potential Salmonella Phages Targeting Multidrug-Resistant and Major Serovars of Salmonella Derived From Broiler Production Chain in Thailand.

Salmonella is a major foodborne pathogen that causes foodborne disease in humans through consumption of contaminated foods, especially those of animal origin. Multiple Salmonella strains are antibiotic-resistant due to the common use of antibiotics in farm animals, including broiler farms. In this study, an alternative strategy using phage-based treatment was evaluated against Salmonella isolated from the broiler production. The prevalence of Salmonella spp. showed up to 46.2 and 44.4% in bedding samples from the broiler farms located in eastern and southern Thailand, respectively. Overall, 21 samples (36.2%) were positive for Salmonella and eight serovars were recovered from cloacal swabs, bedding materials (rice husk), and boot swabs collected from five farms. Up to 20 Salmonella phages were isolated from seven water samples from wastewater treatment ponds, a river, and a natural reservoir in Songkhla province. Isolated phages were investigated, as well as their lysis ability on eight target Salmonella serovars derived from broiler farms, five foodborne outbreak-related serovars, and 10 multidrug-resistant (MDR) serovars. All phages showed a strong lytic ability against five serovars of Salmonella derived from broiler farms including Kentucky, Saintpaul, Schwarzengrund, Corvalis, and Typhimurium; three foodborne outbreak serovars including Enteritidis, Typhimurium, and Virchow; and eight MDR serovars including Agona, Albany, Give, Kentucky, Typhimurium, Schwarzengrund, Singapore, and Weltevreden. Three phages with the highest lysis potential including vB_SenS_WP109, vB_SenS_WP110, and vB_SenP_WP128 were selected for a phage cocktail preparation. Overall, a phage cocktail could reduce Salmonella counts by 2.2-2.8 log units at 6 h of treatment. Moreover, Salmonella did not develop a resistant pattern after being treated with a phage cocktail. Findings here suggest that a phage cocktail is an effective biocontrol to combat Salmonella derived from broiler production chain, other serovars linked to foodborne outbreaks, and MDR serovars.

Whole-Genome Sequencing Analysis of Salmonella Isolates from Poultry Farms, a Slaughterhouse, and Retail Stalls in Thailand.

The draft genome sequences of 21 Salmonella isolates obtained from poultry production chains in Hat Yai City, Songkhla Province, Thailand, are reported in this study. Our study revealed that there was Salmonella environmental contamination along poultry production chains and cross-contamination among poultry through inanimate surfaces in the environment.

Optimization of wall material for phage encapsulation via freeze-drying and antimicrobial efficacy of microencapsulated phage against Salmonella.

Microencapsulated phage as dry powder provides a protection to the phage particles from the harsh conditions while improving efficacy for controlling Salmonella. In this study, wall materials for phage encapsulation were optimized by altering the ratios of whey protein isolate (WPI) and trehalose prior to freeze-drying. Combination of WPI/trehalose at ratio of 3:1 (w/w) represented the optimal formulation with the highest encapsulation efficiency (91.9%). Fourier transform infrared spectroscopy analysis showed H-bonding in the mixture system and glass transition temperature presented at 63.43 °C. Encapsulated form showed the phage survivability of > 90% after 5 h of exposure to pH 1.5, 3.5, 5.5, 7.5 and 9.5. Phages in the non-encapsulated form could not survive at pH 1.5. In addition, microencapsulated phage showed high effectiveness in decreasing the numbers of S. Enteritidis and S. Typhimurium by approximately 1 log CFU/ml at 10 °C and 30 °C for both serovars. Phage powder newly developed in this study provides a convenient form for Salmonella control application and this form exhibits high stability over a wide range of temperatures and pH. This encapsulated phage thus can be used in various food applications without being interfered by physiological acidic or alkaline pH of foods or environments where phages are applied.

Shelf-life of refrigerated Asian sea bass slices treated with cold plasma as affected by gas composition in packaging.

Effect of in-bag dielectric barrier discharge cold plasma (IB-DBD-CP) on the keeping quality of Asian sea bass slices (ASBS) packaged under different gases during refrigerated storage at 4 °C was studied. ASBS without and with IB-DBD-CP treatment packaged under the gas combination of argon and oxygen (10:90) (gas A) or the mixtures of carbon dioxide, argon and oxygen (60:30:10) (gas B) and the control (kept in air) were monitored for quality changes up to 18 days. ASBS treated with IB-DBD-CP, regardless of gas composition, had lower microbial loads than those without treatment and the control (p < 0.05). The shelf-life of ASBS was prolonged to 9 and 12 days after being packaged under gas A and B, respectively without IB-DBD-CP treatment, while 6 days were recorded for the control. However, ASBS treated with IB-DBD-CP, packaged under gas A and B had the shelf-life of 12 and 15 days, respectively. Throughout the storage, trimethylamine content and total volatile nitrogen base content were lower in ASBS treated with IB-DBD-CP, particularly those packaged under gas B than that without treatment and the control (p < 0.05). Nevertheless, lipid oxidation as well as protein oxidation were higher in samples treated with IB-DBD-CP, regardless of gas composition used, in comparison with untreated counterpart. Therefore, IB-DBD-CP of ASBS packaged under high ratio of CO2 (60%) along with argon and oxygen was the potential method for augmenting the shelf-life of ASBS for >15 days at 4 °C.

Liposomal Encapsulated Ethanolic Coconut Husk Extract: Antioxidant and Antibacterial Properties.

Characteristics of liposomal encapsulated ethanolic coconut husk extract (LE-ECHE) prepared using two levels of lipid phase (LP) containing soybean phosphatidylcholine/cholesterol mixture of 4:1 mol ratio (60 and 80 µmol/mL) and two ECHE concentrations (1% and 2%) were investigated. Poly-dispersity index, zeta-potential, and particle size of LE-ECHE samples were 0.22% to 0.28%, -70.4 to -53.63 mV, and 232 to 697.65 nm, respectively. Encapsulation efficiency of all samples was 75.25% to 90.11%. LE-ECHE prepared with LP content of 60 µmol/mL and 1% ECHE (LP60-EC1) was milky, whereas UN-EC1 (un-encapsulated ECHE) was brownish in color. ECHE retained its antioxidant activity even after entrapment in liposome, although higher activity was recorded for UN-EC1. Encapsulation of ECHE in liposome enhanced antibacterial properties of ECHE. Hence, LP60-EC1 showed promising potential as a delivery based system for lowering dark color, a drawback associated with ECHE as well as improving the antibacterial properties of ECHE. PRACTICAL APPLICATION: Ethanolic coconut husk extract (ECHE) contains polyphenols with diverse biological activities such as antimicrobial and antioxidant properties. However, there are limited applications of ECHE in food industries, mainly because of its distinctive dark brown color. A homogeneous and stable liposomal system was demonstrated to be an efficient delivery based system for ECHE. Remarkably, antimicrobial property of ECHE was enhanced with liposomal encapsulation, whereas antioxidant activities of ECHE were retained. Also, liposomal encapsulation was shown as the potential technique to mask the undesirable dark brown, a drawback associated with ECHE for wider application.

Evaluation of storage conditions and efficiency of a novel microencapsulated Salmonella phage cocktail for controlling S. enteritidis and S. typhimurium in-vitro and in fresh foods.

S. Enteritidis and S. Typhimurium are typically linked to foodborne outbreaks. Phages have continued to expand in various food applications. In this study, microencapsulation is applied for enhancing the stability and efficacy of phages as bio-control agent. Microencapsulated phage cocktail kept in aluminium laminated foil bag (LF) at 4 °C showed the highest survivability with a titer loss of 0.5 log PFU/g after 12 weeks of storage. Titer loss of phage cocktail lysate >4 log PFU/mL was observed after 12 weeks, at 4 °C. Color change of microencapsulated phage cocktail kept in LF at 4 °C did not show any significant difference during storage, and water activity (free water content) at 0.13 was found in these conditions. In-vitro study, S. Enteritidis and S. Typhimurium were decreased 1.79 and 3.63 log CFU/mL, respectively at 37 °C. Whereas, 0.43 and 0.76 log CFU/mL, respectively were observed at 10 °C. In foods, S. Enteritidis and S. Typhimurium were decreased 0.57 and 1.78 log CFU/cm2, respectively in meat. Whereas, 0.86 and 1.2 log CFU/g, respectively were observed in sprout. Foods with/without microencapsulated phage cocktail showed non-significant differences in liking scores after 2 days of storage. Overall, microencapsulated phage cocktail suggests another alternative for phage-based biocontrol with improved stability and efficacy for food application.

Dielectric Barrier Discharge High Voltage Cold Atmospheric Plasma: An Innovative Nonthermal Technology for Extending the Shelf-Life of Asian Sea Bass Slices.

Impact of dielectric barrier discharge high-voltage cold atmospheric plasma (DBD-HVCAP) generated with the mixture of oxygen and argon (10:90) for various treatment times (2.5 to 10 min) on the qualities of Asian sea bass slices during 4 °C storage was investigated. Microbial load of slices treated with DBD-HVCAP were lower than the control. The efficacy of bacteria reduction by DBD-HVCAP was dependent on the treatment times (P < 0.05). Total viable bacteria count (TVBC) was more than 6.0 Log CFU/g at day 6 for the control kept in air. Slices treated with DBD-HVCAP for all treatment times used had TVBC lower than the limit at day 12. Total volatile nitrogen base content (TVNB) as well as trimethylamine (TMA) content in slices treated with DBD-HVCAP were lower than that of the control throughout the storage. TVNB as well as TMA contents were lower in HVCAP treated slices in a treatment time-dependent manner. Nevertheless, lipid oxidation in samples treated with DBD-HVCAP was higher than that of the control. Polyunsaturated fatty acids were decreased in slices treated with DBD-HVCAP for more than 5 min after 12 days of storage. Therefore DBD-HVCAP treatment for 5 min was demonstrated to be potential means for increasing the shelf-life of Asian sea bass slices with minimal negative effect on chemical and sensory properties, in which they could be stored longer than 12 days at 4 °C. PRACTICAL APPLICATION: Microbial inactivation capacity of dielectric barrier discharge high-voltage cold atmospheric plasma (DBD-HVCAP) has been documented with limited information on its application in extending the shelf-life of foods. DBD-HVCAP was demonstrated as an innovative technology for extending the shelf-life of Asian sea bass slices, which could be implemented in seafood industries for assuring safety and extending shelf-life of products. The shelf-life of the slices treated with DBD-HVCAP was extended to 12 days of storage at 4 °C as compared to the 6 days of the untreated counterpart.

Lysis Profiles of Salmonella Phages on Salmonella Isolates from Various Sources and Efficiency of a Phage Cocktail against S. Enteritidis and S. Typhimurium.

Salmonella enterica serovar Enteritidis and Salmonella enterica serovar Typhimurium are major foodborne pathogens of concern worldwide. Bacteriophage applications have gained more interest for biocontrol in foods. This study isolated 36 Salmonella phages from several animal farms in Thailand and tested them on 47 Salmonella strains from several sources, including farms, seafood processing plant and humans in Thailand and USA. Phages were classified into three major groups. The estimated phage genome size showed the range from 50 ± 2 to 200 ± 2 kb. An effective phage cocktail consisting of three phages was developed. Approximately 4 log CFU/mL of S. Enteritidis and S. Typhimurium could be reduced. These phages revealed a burst size of up to 97.7 on S. Enteritidis and 173.7 PFU/cell on S. Typhimurium. Our phage cocktail could decrease S. Enteritidis on chicken meat and sunflower sprouts by 0.66 log CFU/cm² and 1.27 log CFU/g, respectively. S. Typhimurium on chicken meat and sunflower sprouts were decreased by 1.73 log CFU/cm² and 1.17 log CFU/g, respectively. Overall, animal farms in Thailand provided high abundance and diversity of Salmonella phages with the lysis ability on Salmonella hosts from various environments and continents. A developed phage cocktail suggests a potential biocontrol against Salmonella in fresh foods.

Characterization of Listeria prophages in lysogenic isolates from foods and food processing environments.

Prophages are commonly found in Listeria genomes, potentially enhancing survival or fitness of Listeria spp. Currently, there is still limited information on the distribution of prophages among Listeria isolates of different allelic types and from various sources. In this study, by using mitomycin C induction, prophages were found in 23/144 isolates (16.0%), including 13 L. monocytogenes and 10 Listeria spp. isolates, resulting in 28 and 11 induced phages, respectively. These prophage-carrying isolates (lysogens) were obtained from foods and food-related environments presenting 3 common allelic types (ATs) of L. monocytogenes (lineage I, II and IV), 4 ATs of L. innocua and 1 AT of L. welshimeri. The likelihood of prophage-carrying isolates of L. monocytogenes was 14.4 (95% CI: 4.9-35.4), and 18.5 (95% CI: 4.8-50.2) for Listeria spp. The 39 induced phages were classified into 3 lysis groups by the host range test against 9 major serotypes of L. monocytogenes and 5 species of Listeria. Most phages were host-specific with higher ability to lyse L. monocytogenes serotype 4 than other serotypes. The genome size of phages ranged from 35±2 kb to 50±2 kb and belonged to two common phage families, Myoviridae and Siphoviridae. Restriction analysis classified 19 selected phages into 16 restriction profiles, suggesting highly diverse prophages with at least 16 types. This may contribute to the variation in the genomes of Listeria. Information obtained here provides basic knowledge for further study to understand the overall role of prophages in Listeria, including roles in survival or fitness in foods and food processing environments.

Antioxidant Properties and Antibacterial Effects of Eucalyptus camaldulensis Ethanolic Leaf Extract on Biofilm Formation, Motility, Hemolysin Production, and Cell Membrane of the Foodborne Pathogen Listeria monocytogenes.

Consumer concerns toward chemical preservatives have resulted in increased search for healthy green alternative. In this study, the antioxidant activity and antibacterial effects of Eucalyptus camaldulensis ethanolic leaf extract against Listeria monocytogenes, a serious foodborne pathogen, was evaluated. Total phenolic and flavonoid contents of the extract were 11.10 mg garlic acid equivalent/mg extract and 15.05 mg quercetin equivalent/mg extract, respectively. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration of the extract was 64-128 µg/mL and 256-512 µg/mL, respectively. Time-kill assay revealed growth inhibitory effects after 4-h treatment of the bacteria with the extract. A reduction of ≈2-3 log colony-forming units per milliliter was observed against the tested food and environmental isolates after challenging the pathogens with the extract at MIC for 6 h. Sub-MICs of the extract significantly inhibited motility and listeriolysin O production up to 80%, with 60% inhibition of biofilm formation (p < 0.05). Antioxidant assay revealed free radical scavenging activity with 50% inhibitory concentration (IC50) of 57.07 µg/mL for 2,2-diphenyl-1-picrylhydrazyl and 29.01 µg/mL for ABTS [2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)] assay. Ferric reducing antioxidant power assay further showed a total antioxidant power equivalent to 92.93 µM ascorbic acid equivalent/mg extract. As the extract exhibited profound antilisterial activity and good radical scavenging ability, it might serve as a potential alternative source of biopreservative agent against L. monocytogenes.

Prospects for Food Fermentation in South-East Asia, Topics From the Tropical Fermentation and Biotechnology Network at the End of the AsiFood Erasmus+Project.

Fermentation has been used for centuries to produce food in South-East Asia and some foods of this region are famous in the whole world. However, in the twenty first century, issues like food safety and quality must be addressed in a world changing from local business to globalization. In Western countries, the answer to these questions has been made through hygienisation, generalization of the use of starters, specialization of agriculture and use of long-distance transportation. This may have resulted in a loss in the taste and typicity of the products, in an extensive use of antibiotics and other chemicals and eventually, in a loss in the confidence of consumers to the products. The challenges awaiting fermentation in South-East Asia are thus to improve safety and quality in a sustainable system producing tasty and typical fermented products and valorising by-products. At the end of the "AsiFood Erasmus+ project" (www.asifood.org), the goal of this paper is to present and discuss these challenges as addressed by the Tropical Fermentation Network, a group of researchers from universities, research centers and companies in Asia and Europe. This paper presents current actions and prospects on hygienic, environmental, sensorial and nutritional qualities of traditional fermented food including screening of functional bacteria and starters, food safety strategies, research for new antimicrobial compounds, development of more sustainable fermentations and valorisation of by-products. A specificity of this network is also the multidisciplinary approach dealing with microbiology, food, chemical, sensorial, and genetic analyses, biotechnology, food supply chain, consumers and ethnology.

Combined effect of bacteriophage and antibiotic on the inhibition of the development of antibiotic resistance in Salmonella typhimurium.

This study was designed to evaluate the combined effects of bacteriophage and antibiotic on the reduction of the development of antibiotic-resistance in Salmonella typhimurium LT2. The susceptibilities of S. typhimurium to ciprofloxacin and erythromycin were increased when treated with bacteriophages, showing more than 10% increase in clear zone sizes and greater than twofold decrease in minimum inhibitory concentration values. The growth of S. typhimurium was effectively inhibited by the combination of bacteriophage P22 and ciprofloxacin. The combination treatment effectively reduced the development of antibiotic resistance in S. typhimurium. The relative expression levels of efflux pump-related genes (acrA, acrB, and tolC) and outer membrane-related genes (ompC, ompD, and ompF) were decreased at all treatments. This study provides useful information for designing new antibiotic therapy to control antibiotic-resistant bacteria.

Genome Sequences of Listeria Phages Induced from Lysogenic Isolates of Listeria monocytogenes from Seafood and a Seafood Processing Environment in Thailand.

We report here the complete genome sequences of three Listeria phages (PSU-VKH-LP019, PSU-VKH-LP040, and PSU-VKH-LP041), which were newly induced from lysogenic isolates of Listeria monocytogenes from seafood and a seafood processing environment in Thailand. The three phages show circularly permuted double-stranded DNA genomes with sizes of 38.6, 39.6, and 48.3 kb.