Battling Salmonella enteritidis infections: integrating proteomics and in vivo assessment of Galla Chinensis tannic acid.

Salmonella infections pose a significant threat to animal and human health. Phytochemicals present a potential alternative treatment. Galla chinensis tannic acid (GCTA), a hydrolyzable polyphenolic compound, inhibits bacterial growth and demonstrates potential as an alternative or supplement to antibiotics to prevent Salmonella infections. However, little is known about the antimicrobial mechanism of GCTA against Salmonella. Here, we revealed 456 differentially expressed proteins upon GCTA treatment, impacting pathways related to DNA replication, repair, genomic stability, cell wall biogenesis, and lipid metabolism using TMT-labeled proteomic analysis. TEM analysis suggested altered bacterial morphology and structure post-treatment. A Salmonella-infected-mouse model indicated that GCTA administration improved inflammatory markers, alleviated intestinal histopathological alterations, and reduced Salmonella enterica serovar Enteritidis (S. Enteritidis) colonization in the liver and spleen of Salmonella-infected mice. The LD50 of GCTA was 4100 mg/kg with an oral single dose, vastly exceeding the therapeutic dose. Thus, GCTA exhibited antibacterial and anti-infective activity against S. Enteritidis. Our results provided insight into the molecular mechanisms of these antibacterial effects, and highlights the potential of GCTA as an alternative to antibiotics.

Assessment of environmental safety and protective efficacy of O-antigen deficient DIVA capable Salmonella Enteritidis against chicken salmonellosis.

In this study, we incorporated deletion of the O-antigen ligase gene to an attenuated Salmonella Enteritidis (SE) strain, JOL919 (SE PS; Δlon ΔcpxR), using the Lambda-Red recombination method and evaluated the safety and immunological aspects of the novel genotype, JOL2381 (SE VS: Δlon, ΔcpxR, ΔrfaL). Assessment of fecal shedding and organ persistence following administration via oral and IM routes revealed that the SE VS was safer than its parent strain, SE PS. Immunological assays confirmed that immunization via the oral route with SE PS was superior to the SE VS. However, chickens immunized with SE PS and SE VS strains via the IM route showed higher humoral and cell-mediated immune responses. Compared to PBS control, the IM route of immunization with SE VS resulted in a higher IgY antibody titer and expansion of CD4+ and CD8+ T-cell populations, which resulted in the clearance of Salmonella from the liver and splenic tissues. Furthermore, deletion of the O-antigen ligase gene caused lower production of LPS-specific antibodies in the host, promoting DIVA functionality and making it a plausible candidate for field utilization. Due to significant protection, high attenuation, and environmental safety concerns, the present SE VS strain is an ideal choice to prevent chicken salmonellosis and ensure public health.

Genetic and phenotypic assessment of the antimicrobial activity of three potential probiotic lactobacilli against human enteropathogenic bacteria.

Introduction: Lactobacilli are avid producers of antimicrobial compounds responsible for their adaptation and survival in microbe-rich matrices. The bactericidal or bacteriostatic ability of lactic acid bacteria (LAB) can be exploited for the identification of novel antimicrobial compounds to be incorporated in functional foodstuffs or pharmaceutical supplements. In this study, the antimicrobial and antibiofilm properties of Lactiplantibacillus pentosus L33, Lactiplantibacillus plantarum L125 and Lacticaseibacillus paracasei SP5, previously isolated form fermented products, were examined, against clinical isolates of Staphylococcus aureus, Salmonella enterica subsp. enterica serovar Enteritidis and Escherichia coli. Methods: The ability of viable cells to inhibit pathogen colonization on HT-29 cell monolayers, as well as their co-aggregation capacity, were examined utilizing the competitive exclusion assay. The antimicrobial activity of cell-free culture supernatants (CFCS) was determined against planktonic cells and biofilms, using microbiological assays, confocal microscopy, and gene expression analysis of biofilm formation-related genes. Furthermore, in vitro analysis was supplemented with in silico prediction of bacteriocin clusters and of other loci involved in antimicrobial activity. Results: The three lactobacilli were able to limit the viability of planktonic cells of S. aureus and E. coli in suspension. Greater inhibition of biofilm formation was recorded after co-incubation of S. enterica with the CFCS of Lc. paracasei SP5. Predictions based on sequence revealed the ability of strains to produce single or two-peptide Class II bacteriocins, presenting sequence and structural conservation with functional bacteriocins. Discussion: The efficiency of the potentially probiotic bacteria to elicit antimicrobial effects presented a strain- and pathogen-specific pattern. Future studies, utilizing multi-omic approaches, will focus on the structural and functional characterization of molecules involved in the recorded phenotypes.

Assessment of stn, sipB and sopB Virulence Genes in Various Salmonella Serovars.

Salmonella is a zoonotic bacterium that is considered to be one of the most common causes of foodborne infections worldwide. Bearing in mind the genes involved in its virulence, identifying these genes can enable experts to better understand bacterial pathogenicity, which could subsequently help develop more efficient means to control and prevent infections. This study aimed to analyze stn, sipB, and sopB genes in various Salmonella serovars. To carry out this study, 103 Salmonella serovars were extracted from livestock, poultry, and humans from existing samples at the Department of Microbiology of the Razi Serum and Vaccine Research Institute in Karaj, Iran. These samples were cultured in selection and differential media, and their serovars were identified using specific antibodies based on Kaufman-White Tables. Utilizing PCR and specific primers, stn, sopB, and sipB genes were detected among these serovars. In this investigation, the most common human serovars were Salmonella paratyphi A, Salmonella paratyphi B, and Salmonella enteritidis; the most common serovars among livestock consisted of Salmonella dublin and Salmonella typhimurium and the most common Salmonella serovars among poultry consisted of Salmonella infantis and Salmonella enteritidis. The results of PCR on stn, sipB, and sopB genes demonstrated segments with 617bp, 875 bp, and 220 bp on agar gel, respectively. Based on the obtained findings, stn, sipB, and sopB genes were detected in 96.11%, 99.02%, and 98.05% of Salmonella serovars, respectively. Considering the fact that the aforementioned genes play significant roles in bacterial virulence, they can be used to develop diagnostic ELISA kits and recombinant vaccines.

Assessment of foodborne transmission of Salmonella enteritidis in hens and eggs in Bangladesh.

OBJECTIVES: Salmonella is considered one of the leading causes of foodborne illnesses worldwide. Information about the transmission of pathogens to poultry and poultry products is necessary to implement control measures for reducing both human exposure and economic loss. The aim of this study was to analyze and evaluate the transmission characteristics of Salmonella enteritidis to laying-type hen flocks and their laid eggs. MATERIALS AND METHODS: For this purpose, 15 pairs of laying hens were used in which each pair consisted of one inoculated and one contact exposed hen. The eggs and cloacal swabs from these hens were subsequently analyzed. RESULTS: Of the 15 in-contact hens tested, 60% were found to be positive for S. enteritidis within 61 days postinoculation, of which 26.7% transmission occurred within the first 31 days postinoculation. Among the collected laid eggs tested, S. enteritidis was detected on 58% eggshells and 5.33% eggs internal contents. We also observed a 33.33% reduction in egg production from S. enteritidis-infected hens. In a cross-contamination study, we demonstrated that an experimentally inoculated container can act as a potential source of Salmonella spp. CONCLUSIONS: Our results will help establish effective monitoring programs to reduce the transmission of Salmonella spp. in poultry and poultry products.

Genome-Based Assessment of Antimicrobial Resistance and Virulence Potential of Isolates of Non-Pullorum/Gallinarum Salmonella Serovars Recovered from Dead Poultry in China.

Paratyphoid avian salmonellosis is considered one of the leading causes of poultry death, resulting in significant economic losses to poultry industries worldwide. In China, especially in Shandong province, the leading producer of poultry products, several recurrent outbreaks of avian salmonellosis have been reported during the last decade where the precise causal agent remains unknown. Moreover, the establishment of earlier and more accurate recognition of pathogens is a key factor to prevent the further dissemination of resistant and/or hypervirulent clones. Here, we aim to use whole-genome sequencing combined with in silico toolkits to provide the genomic features of the antimicrobial resistance and virulence potential of 105 regionally representative non-Pullorum/Gallinarum Salmonella isolates recovered from dead poultry between 2008 and 2019 in Shandong, China. Additionally, phenotypic susceptibility to a panel of 15 antibiotics representing 11 classes was assessed by the broth microdilution method. In this study, we identified eight serovars and nine multilocus sequence typing (MLST) types, with Salmonella enterica serovar Enteritidis sequence type 11 (ST11) being the most prevalent (84/105; 80%). Based on their phenotypic antimicrobial resistance, 77.14% of the isolates were defined as multidrug resistant (≥3 antimicrobial classes), with the detection of one S. Enteritidis isolate that was resistant to the 11 classes. The highest rates of resistance were observed against nalidixic acid (97.14%) and ciprofloxacin (91.43%), followed by ampicillin (71.43%), streptomycin (64.77%), and tetracycline (60%). Genomic characterization revealed the presence of 41 resistance genes, with an alarmingly high prevalence of blaTEM-1B (60%), in addition to genomic mutations affecting the DNA gyrase (gyrA) and DNA topoisomerase IV (parC) genes, conferring resistance to quinolones. The prediction of plasmid replicons detected 14 types, with a dominance of IncFIB(S)_1 and IncFII(S)_1 (87.62% for both), while the IncX1 plasmid type was considered the key carrier of antimicrobial resistance determinants. Moreover, we report the detection of critical virulence genes, including cdtB, rck, sodCI, pef, and spv, in addition to the typical determinants for Salmonella pathogenicity island 1 (SPI-1) and SPI-2. Furthermore, phylogenomic analysis revealed the detection of three intra-farm and five inter-farm transmission events. Overall, the detection of Salmonella isolates presenting high antimicrobial resistance and harboring different critical virulence genes is of major concern, which requires the urgent implementation of effective strategies to mitigate non-Pullorum/Gallinarum avian salmonellosis. IMPORTANCE Avian salmonellosis is one of the leading global causes of poultry death, resulting in substantial economic losses in China (constituting 9% of overall financial losses). In Shandong province, a top poultry producer (30% of the overall production in China, with 15% being exported to the world), extensive outbreaks of avian salmonellosis have been reported in the past decade where the causal agents or exact types remain rarely addressed. From approximately 2008 to 2019, over 2,000 Salmonella strains were isolated and identified from dead poultry during routine surveillance of 95 poultry farms covering all 17 cities in Shandong. Approximately 1,500 isolates were confirmed to be of non-Pullorum/Gallinarum Salmonella serovars. There is an urgent need to understand the mechanisms behind the implication of zoonotic Salmonella serovars in systemic infections of poultry. Here, we analyzed populations of clinically relevant isolates of non-Pullorum/Gallinarum Salmonella causing chicken death in China by a whole-genome sequencing approach and determined that antimicrobial-resistant Salmonella Enteritidis remained the major cause in the past decades.

Global spread of Salmonella Enteritidis via centralized sourcing and international trade of poultry breeding stocks.

A pandemic of Salmonella enterica serotype Enteritidis emerged in the 1980s due to contaminated poultry products. How Salmonella Enteritidis rapidly swept through continents remains a historical puzzle as the pathogen continues to cause outbreaks and poultry supply becomes globalized. We hypothesize that international trade of infected breeding stocks causes global spread of the pathogen. By integrating over 30,000 Salmonella Enteritidis genomes from 98 countries during 1949-2020 and international trade of live poultry from the 1980s to the late 2010s, we present multifaceted evidence that converges on a high likelihood, global scale, and extended protraction of Salmonella Enteritidis dissemination via centralized sourcing and international trade of breeding stocks. We discovered recent, genetically near-identical isolates from domestically raised poultry in North and South America. We obtained phylodynamic characteristics of global Salmonella Enteritidis populations that lend spatiotemporal support for its dispersal from centralized origins during the pandemic. We identified concordant patterns of international trade of breeding stocks and quantitatively established a driving role of the trade in the geographic dispersal of Salmonella Enteritidis, suggesting that the centralized origins were infected breeding stocks. Here we demonstrate the value of integrative and hypothesis-driven data mining in unravelling otherwise difficult-to-probe pathogen dissemination from hidden origins.

Walnut (Juglans regia L.) Septum: Assessment of Bioactive Molecules and In Vitro Biological Effects.

Walnut (Juglans regia L.) septum represents an interesting bioactive compound source by-product. In our study, a rich phenolic walnut septum extract, previously selected, was further examined. The tocopherol content determined by liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed higher amounts of α-tocopherol compared to γ- and δ-tocopherols. Moreover, several biological activities were investigated. The in vitro inhibiting assessment against acetylcholinesterase, α-glucosidase, or lipase attested a real management potential in diabetes or obesity. The extract demonstrated very strong antimicrobial potential against Staphylococcus aureus, Pseudomonas aeruginosa and Salmonella enteritidis. It also revealed moderate (36.08%) and strong (43.27%) antimutagenic inhibitory effects against TA 98 and TA 100 strains. The cytotoxicity of the extract was assessed on cancerous (A549, T47D-KBluc, MCF-7) and normal (human gingival fibroblasts (HGF)) cell lines. Flow cytometry measurements confirmed the cytotoxicity of the extract in the cancerous cell lines. Additionally, the extract demonstrated antioxidant activity on all four cell types, as well as anti-inflammatory activity by lowering the inflammatory cytokines (interleukin-6 (IL-6), interleukin-8 (IL-8), interleukin-1 ß (IL-1ß)) evaluated in HGF cells. To the best of our knowledge, most of the cellular model analyses were performed for the first time in this matrix. The results prove that walnut septum may be a potential phytochemical source for pharmaceutical and food industry.

Assessment of the Risk of Salmonellosis Linked to the Consumption of Liquid Egg Products Made from Internally Contaminated Shell Eggs Initially Stored at 65°F (18°C) Compared with Eggs Stored at 45°F (7°C).

ABSTRACT: According to the U.S. Food and Drug Administration's (FDA's) rule on "Prevention of Salmonella Enteritidis in Shell Eggs during Production, Storage, and Transportation," shell eggs intended for human consumption are required to be held or transported at or below 45°F (7.2°C) ambient temperature beginning 36 h after time of lay. Meanwhile, eggs in hatcheries are typically stored at a temperature of 65°F (18.3°C). Although most of those eggs are directed to incubators for hatching, excess eggs have the potential to be diverted for human consumption as egg products through the "breaker" market if these eggs are refrigerated in accordance with FDA's requirement. Combining risk assessment models developed by the U.S. Department of Agriculture's Food Safety and Inspection Service for shell eggs and for egg products, we quantified and compared Salmonella Enteritidis levels in eggs held at 65°F versus 45°F, Salmonella Enteritidis levels in the resulting egg products, and the risk of human salmonellosis from consumption of those egg products. For eggs stored 5 days at 65°F (following 36 h at 75°F [23.9°C] in the layer house), the mean level of Salmonella Enteritidis contamination is 30-fold higher than for eggs stored at 45°F. These increased levels of contamination lead to a 47-fold increase in the risk of salmonellosis from consumption of egg products made from these eggs, with some variation in the public health risk on the basis of the egg product type (e.g., whole egg versus whole egg with added sugar). Assuming that 7% of the liquid egg product supply originates from eggs stored at 65°F versus 45°F, this study estimates an additional burden of 3,562 cases of salmonellosis per year in the United States. A nominal range uncertainty analysis suggests that the relative increase in the risk linked to the storage of eggs at higher temperature estimated in this study is robust to the uncertainty surrounding the model parameters. The diversion of eggs from broiler production to human consumption under the current storage practices of 65°F (versus 45°F) would present a substantive overall increase in the risk of salmonellosis.

Assessment of a Novel Antimicrobial Peptide Against Clinically Isolated Animal Pathogens and Prediction of Its Thermal-Stability.

Multidrug resistance of pathogenic microorganisms is a common problem in the treatment of infections. Therefore, searching for new agents with antimicrobial activities appears to be essential. Thrombocidin-1 (TC-1) is an antimicrobial peptide (AMP) derived from platelets. The present study aimed to produce recombinant TC-1 (rTC-1) with His tag to evaluate its antimicrobial activity and also predict its thermal-stability through molecular dynamic (MD) simulations. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration of rTC-1 against bacterial isolates were determined. Considering the importance of thermal-stability of proteins in their therapeutic applications, thermal-stability of rTC-1 predicted through MD simulation during 25ns at two important temperatures including avian normal body temperature and water boiling temperature at sea level. MIC results revealed that rTC1 had the most and least potency against Salmonella enteritidis and Escherichia coli, respectively. The root-mean-square deviation of rTC-1 during 25ns MD simulations revealed that this protein is stable at avian normal body temperature (40°C) but might lose stability at water boiling temperature at sea level. rTC-1 as an AMP has a good potency against some bacterial pathogens especially Salmonella spp. and E. coli 0157:H7. rTC-1 can be used as an alternative for common antimicrobial agents used in clinical settings.

Antagonistic trait of Lactobacillus reuteri S5 against Salmonella enteritidis and assessment of its potential probiotic characteristics.

Salmonella enteritidis is an important foodborne pathogen that has caused multiple outbreaks of infection associated with poultry and egg consumption. Thus, the prevention and inhibition of Salmonella enteritidis infection are of great concern. Lactic acid bacteria have anti-pathogenic activity; however, their underlying mechanisms and modes of action have not yet been clarified. In this study, the antibacterial mechanism of Lactobacillus reuteri S5 (L. reuteri S5) against Salmonella enteritidis ATCC13076 (S. enteritidis ATCC13076) was studied by different methods. We found that L. reuteri S5 was able to form a stable biofilm formation, colonizing the entire intestinal tract of chickens. In addition, bacterial cultures and the cell-free supernatant (CFS) of L. reuteri S5 inhibited SE ATCC13076 growth, and this growth inhibition was also observed in the co-culture assay. This effect may be predominantly caused by antimicrobial metabolites produced by L. reuteri S5. Furthermore, treatment with the CFS of L. reuteri S5 resulted in a significant reduction in the expression of Salmonella virulence, motility and adhesion genes and a significant reduction in the motility ability and inhibitory effect on biofilm formation. In addition, the damage to the membrane structure and intracellular structure induced by the CFS of L. reuteri S5 could be observed on Transmission electron microscopy images and dodecyl sulfate, sodium salt (SDS)-Polyacrylamide gel electrophoresis confirmed the disruptive action of the CFS of L. reuteri S5 on the cytoplasmic membrane. Our findings demonstrate that L. reuteri S5, an intestinal Lactobacillus species associated with chicken health, is able to form biofilm and stably colonize chicken intestines. It also possesses anti-SE activity, preventing SE growth, inhibits the expression of SE genes involved in adhesion and invasion, virulence and cell membrane integrity, inhibits SE biofilm formation and motility, damages or destroys bacterial structures, and inhibits intracellular protein synthesis. L. reuteri S5 therefore has potential applications as a probiotic agent.

Quantitative assessment of tolerance response to stress after exposure to oregano and rosemary essential oils, carvacrol and 1,8-cineole in Salmonella Enteritidis 86 and its isogenic deletion mutants ∆dps, ∆rpoS and ∆ompR.

This study assessed the influence of rpoS, dps and ompR genes on the tolerance response of Salmonella Enteritidis 86 (SE86) to homologous and heterologous stressing agents after exposure to essential oils (EOs) from Origanum vulgare L. (oregano; OVEO) and Rosmarinus officinalis L. (rosemary; ROEO) and their major constituents (ICs), carvacrol (CAR) and 1,8-cineole (CIN), respectively, by modelling the log reduction over time. Minimum inhibitory concentration values of OVEO (1.25 µL/mL), CAR (0.62 µL/mL), ROEO (20 µL/mL) and CIN (10 µL/mL) against SE86 were always one-fold higher than those against ∆dps, ∆rpoS and ∆ompR mutants. Exposure to the same concentration of OVEO, CAR, ROEO or CIN caused higher reductions (up to 2.5 log CFU/mL) in ∆dps, ∆rpoS and ∆ompR mutants than in SE86 in chicken broth. In assays with homologous stressing agents, ompR, dps and rpoS influenced the tolerance to OEs or ICs. After adaptation to OVEO, CAR, ROEO and CIN, osmotolerance and acid tolerance of SE86 were influenced by rpoS gene, while thermotolerance of SE86 was influenced by ompR. Tolerance of SE86 to sodium hypochlorite after adaptation to OEs or ICs was influenced by rpoS and dps. These findings quantitatively describe for the first time the influence of rpoS, dps and ompR genes on the tolerance of Salmonella Enteritidis to OVEO, CAR, ROEO and CIN.

Economic evaluation of the development of a phage therapy product for the control of Salmonella in poultry.

Poultry products are one of the major transmission media of Salmonella enteritidis to humans. A promising alternative to reduce the load of Salmonella in poultry are bacteriophages. Elsewhere, a mixture of six bacteriophages has been used successfully, but large-scale production would be necessary to supply potential poultry market and costs analyses have not been calculated yet. For this, a powerful tool to predict production costs is bioprocess modeling coupled with economic analyses. This work aims to model the scaled-up production of a six bacteriophages mixture based on a laboratory/pilot-scale production using Biosolve Process. For the model construction, a combination of experimental and reported data was applied, in which different production alternatives and the range of 1-100% of the Colombian poultry market (at broiler's farm and slaughterhouse) were analyzed. Results indicate that the best cost-effective process configuration/scale is to use one bioreactor (156 L) for the six bacteriophages, then a 0.45 µm filtration for removal of biomass, and a 0.22 µm filtration for sterility; this to supply the 35% of the market size for broiler farms (equivalent to 210 million chickens). This configuration gives a production cost per chicken of US$ 0.02. Additionally, a sensitivity analysis and a theoretical contrast for understanding the impact that titer and recovery have on production scale determined that titer affects the most the cost and requires optimization. The present works serves as a first, and required, approach for the development of phage therapy products that are alternatives to present-day pathogens control strategies.

[Episodes of nontyphoidal salmonellosis in adult patients of the Valladolid West Area in 2017: Evaluation of the suitability of the microbiological culture request and the prescribed treatment]. / Episodios de salmonelosis no tifoidea en pacientes adultos del Área Valladolid Oeste en 2017: Evaluación de la idoneidad de la petición del cultivo microbiológico y del tratamiento pautado.

OBJECTIVE: Gastroenteritic salmonellosis is still the second cause diagnosed of infectious diarrhea, most of these clinical pictures are mild and self-limited and therefore the use of antibiotics is limited to few cases. The aim of the study was to describe the episodes of diarrhea caused by Salmonella enterica subsp. enterica, assessing the suitability of the request and the use of antibiotics according to the criteria included in the methodology. METHODS: A retrospective, descriptive, observational study was conducted, collecting data from the clinical history. RESULTS: A total of 122 episodes were included. The reason for consultation was diarrhea, which generated a greater demand in the Hospital Emergency Services (42.6%). The most frequent serotypes isolated were Enteritidis (53.3%), and Typhimurium (40.2%). The adequate request of the stool was 90.2%. Antibiotic was prescribed in 64.6% (79) of the episodes, most patients under 65 years (58 episodes), the average age was 48.43 years. They were treated mainly with ciprofloxacin and azithromycin, in 57 and 14 episodes, respectively. The average duration of antibiotic treatment was 6 days. There was an adequate use of antibiotics in 49.1% of episodes. When the origin of the request was the Hospital Emergency Service, it was inadequate in 63.5% (33) of them. It was inadequate in 60.0% (39) of episodes when ser. Enteritidis was isolated. Almost half, 48.85% (42) of the 58 episodes in which antibiotics were prescribed among those under 65 (86), were treated without being indicated. CONCLUSIONS: Training actions should be implemented focused on optimizing the management of antibiotics in this entity.

Invasive Nontyphoidal Salmonella Disease in Africa.

Nontyphoidal salmonellae (NTS) are a major cause of invasive (iNTS) disease in sub-Saharan Africa, manifesting as bacteremia and meningitis. Available epidemiological data indicate that iNTS disease is endemic in much of the region. Antimicrobial resistance is common and case fatality rates are high. There are well-characterized clinical associations with iNTS disease, including young age, HIV infection, malaria, malnutrition, anemia, and sickle cell disease. However, the clinical presentation of iNTS disease is often with fever alone, so clinical diagnosis is impossible without blood culture confirmation. No vaccine is currently available, making this a priority area for global health research. Over the past ten years, it has emerged that iNTS disease in Africa is caused by distinct pathovars of Salmonella Typhimurium, belonging to sequence type ST313, and Salmonella Enteritidis. These are characterized by genome degradation and appear to be adapting to an invasive lifestyle. Investigation of rare patients with primary immunodeficiencies has suggested a key role for interferon gamma-mediated immunity in host defense against NTS. This concept has been supported by recent population-based host genetic studies in African children. In contrast, immunoepidemiological studies from Africa indicate an important role for antibody for protective immunity, supporting the development of antibody-inducing vaccines against iNTS disease. With candidate O-antigen-based vaccines due to enter clinical trials in the near future, research efforts should focus on understanding the relative contributions of antibody and cell-mediated immunity to protection against iNTS disease in humans.

Operational burden of implementing Salmonella Enteritidis and Typhimurium cluster detection using whole genome sequencing surveillance data in England: a retrospective assessment.

Since April 2014 all presumptive Salmonella isolates received by Public Health England (PHE) have been characterised using whole genome sequencing (WGS) and the genomic data generated used to identify clusters of infection. To inform the implementation and development of a national gastrointestinal infection surveillance system based on WGS we have retrospectively identified genetically related clusters of Salmonella Enteritidis and Salmonella Typhimurium infection over a one year period and determined the distribution of these clusters by PHE operational levels. Using a constrained WGS cluster definition based on single nucleotide polymorphism distance, case frequency and temporal spread we demonstrate that the majority of clusters spread to multiple PHE operational levels. The greatest investigative burden is on national level staff investigating small, geographically dispersed clusters. We also demonstrate that WGS identifies long-running, slowly developing clusters that may previously have remained undetected. This analysis also indicates likely increased workload for local health protection teams and will require an operational strategy to balance limited human resources with the public health importance of investigating small, geographically contained clusters of highly related cases. While there are operational challenges to its implementation, integrated cluster detection based on WGS from local to international level will provide further improvements in the identification of, response to and control of clusters of Salmonella spp. with public health significance.

Rapid detection of Salmonella in raw chicken breast using real-time PCR combined with immunomagnetic separation and whole genome amplification.

We presented the first attempt to combine immunomagnetic separation (IMS), whole genome amplification by multiple displacement amplification (MDA) and real-time PCR for detecting a bacterial pathogen in a food sample. This method was effective in enabling real-time PCR detection of low levels of Salmonella enterica Serotype Enteritidis (SE) (∼10 CFU/g) in raw chicken breast without culture enrichment. In addition, it was able to detect refrigeration-stressed SE cells at lower concentrations (∼0.1 CFU/g) in raw chicken breast after a 4-h culture enrichment, shortening the detection process from days to hours and displaying no statistical difference in detection rate in comparison with a culture-based detection method. By substantially improving performance in SE detection over conventional real-time PCR, we demonstrated the potential of IMS-MDA real-time PCR as a rapid, sensitive and affordable method for detecting Salmonella in food.

Treatment of Salmonella-Contaminated Eggs and Pork with a Broad-Spectrum, Single Bacteriophage: Assessment of Efficacy and Resistance Development.

Numerous studies have assessed the efficacy of phage-based methods to inhibit Salmonella contamination in food products. As with most antibacterials, bacteria can develop resistance to phage in vitro. Here, we applied a single broad-spectrum Salmonella phage, vB_SalS_SJ_2 (SJ2; 108 PFU; MOI = 10), to Salmonella-contaminated meat and eggs to quantify the development of resistance in actual food matrices. Treatment with a single phage significantly reduced Salmonella Typhimurium contamination in both ground pork and liquid egg at various time points. Similarly, the same phage significantly reduced Salmonella Enteritidis in both food matrices. Efficacy was temperature dependent as larger reductions were seen at higher temperatures (21°C) versus lower temperatures (4°C) at 24 h. Following phage treatment, over 10,000 Salmonella isolates were examined for resistance to the treatment phage. The percentages of phage-resistant Salmonella (either serovar) recovered from phage-treated versus untreated pork did not differ. Conversely, significantly (p < 0.05) higher percentages of phage-resistant Salmonella Typhimurium (92.50% vs. 0.56% of control) and Salmonella Enteritidis (50.83% vs. 0.56% of control) isolates were observed in phage-treated versus untreated egg samples after incubation at room temperature for 48 h. Taken together, these data indicate that the food matrix may influence the emergence of phage resistance with resistance developing more rapidly in foods with less complex microbial communities. Future studies will focus on the impact the development of resistance in production and processing settings may have on the efficacy of phage treatments for longer term biocontrol of pathogens.

Distinct Salmonella Enteritidis lineages associated with enterocolitis in high-income settings and invasive disease in low-income settings.

An epidemiological paradox surrounds Salmonella enterica serovar Enteritidis. In high-income settings, it has been responsible for an epidemic of poultry-associated, self-limiting enterocolitis, whereas in sub-Saharan Africa it is a major cause of invasive nontyphoidal Salmonella disease, associated with high case fatality. By whole-genome sequence analysis of 675 isolates of S. Enteritidis from 45 countries, we show the existence of a global epidemic clade and two new clades of S. Enteritidis that are geographically restricted to distinct regions of Africa. The African isolates display genomic degradation, a novel prophage repertoire, and an expanded multidrug resistance plasmid. S. Enteritidis is a further example of a Salmonella serotype that displays niche plasticity, with distinct clades that enable it to become a prominent cause of gastroenteritis in association with the industrial production of eggs and of multidrug-resistant, bloodstream-invasive infection in Africa.