Listeria monocytogenes prevalence and genomic diversity along the pig and pork production chain.

The facultative intracellular bacterium Listeria monocytogenes (L. monocytogenes) is the causative agent of listeriosis, a severe invasive illness. This ubiquitous species is widely distributed in the environment, but infection occurs almost exclusively through ingestion of contaminated food. The pork production sector has been heavily affected by a series of L. monocytogenes-related foodborne outbreaks in the past around the world. Ready-to-eat (RTE) pork products represent one of the main food sources for strong-evidence listeriosis outbreaks. This pathogen is known to be present throughout the entire pig and pork production chain. Some studies hypothesized that the main source of contamination in final pork products was either living pigs or the food-processing environment. A detailed genomic picture of L. monocytogenes can provide a renewed understanding of the routes of contamination from pig farms to the final products. This review provides an overview of the prevalence, the genomic diversity and the genetic background linked to virulence of L. monocytogenes along the entire pig and pork production chain, from farm to fork.

Monophasic Variant of Salmonella Typhimurium Infection Affects the Serum Metabolome in Swine.

Salmonella is the most relevant foodborne zoonotic agent found in swine, and its presence in French herds is significant. Its carriage is asymptomatic, which makes it difficult to detect during rearing, thus increasing the risk of its presence on pork meat. Studies have shown that enteric infection in animals could be associated with changes in the serum metabolome composition, through the immune response or changes in the digestive microbiota composition. We hypothesized that these changes in the serum metabolome composition could be used as markers for the detection of asymptomatic animals infected by Salmonella. Using untargeted analysis by liquid chromatography coupled with mass spectrometry, we showed that significant differences in the composition of the serum metabolome could be detected between infected or noninfected animals both 1 and 21 days after experimental infection. This serum metabolome composition significantly changed during the 21 days postinfection in the infected animal groups, suggesting an evolution of the impact of infection with time. Despite this evolution, differences in the serum metabolome composition persisted between infected and noninfected animals 21 days after the initial infection. We also showed a possible difference between high-shedding and low-shedding animals 21 days postinfection. Finally, some of the variations in the metabolome were found to be significantly associated with variations of specific members of the fecal microbiota. Thus, excreting and asymptomatic animals, but also high-shedding animals, could be identified on the basis of their serum metabolome composition.

Comparative Analysis of Campylobacter jejuni and C. coli Isolated from Livestock Animals to C. jejuni and C. coli Isolated from Surface Water Using DNA Sequencing and MALDI-TOF.

This study evaluated the contribution of cattle, sheep, poultry and pigs to the contamination of surface water from rivers by Campylobacter jejuni and C. coli using MLST, cgMLST and considered MALDI-TOF MS as an alternative technique. The 263 strains isolated from cattle (n = 61), sheep (n = 42), poultry (n = 65), pigs (n = 60) and surface water (n = 35) were distributed across 115 sequence types (STs), 49 for C. jejuni and 66 for C. coli. Considering MLST data, 14.2%, 11.4% and 2.8% of the surface water strains could be attributed to cattle, poultry and sheep, respectively, none to pigs, and 85.7% were non-attributed. Analysis of cg-MLST data with STRUCTURE indicated that C. jejuni strains from water were predominantly attributed to poultry (93.5%), weakly to sheep (<1%) and 6.3% non-attributed, and that conversely, C. coli strains from water were predominantly non-attributed (94.3%) and 5.7% attributed to poultry. Considering the protein profiles with a threshold of 94% and 97% of similarity, respectively, strains from surface water could be attributed to poultry (31.4% and 17.1%), and to cattle (17.1% and 5.7%); 54.1% and 77.1% were non-attributed. This study confirmed these livestock animals might contribute to the contamination of surface water, with a level of contribution depending on the typing technique and the method of analysis. MALDI-TOF could potentially be an alternative approach for source attribution.

Quantification of indicator and pathogenic bacteria in manures and digestates from three agricultural biogas plants over a one-year period.

Interest in the conversion of manure in biogas via anaerobic digestion (AD) is growing, but questions remain about the biosafety of digestates. For a period of one year, we monitored the impact of three mesophilic agricultural biogas plants (BPs) mainly fed with pig manure (BP1, BP3) or bovine manure (BP2) on the physicochemical parameters, the composition of the microbial community and the concentration of bacteria (E. coli, enterococci, Salmonella, Campylobacter, Listeria monocytogenes, Clostridium perfringens, Clostridium botulinum and Clostridioides difficile). The BP2 digestate differed from those of the two other BPs with a higher nitrogen content, more total solids and greater abundance of Clostridia MBA03 and Disgonomonadacea. Persistence during digestion ranked from least to most, was: Campylobacter (1.6 to >2.9 log10 reduction, according to the BP) < E. coli (1.8 to 2.2 log10) < Salmonella (1.1 to 1.4 log10) < enterococci (0.2 to 1.2 log10) and C. perfringens (0.2 to 1 log10) < L. monocytogenes (-1.2 to 1.6 log10) < C. difficile and C. botulinum (≤0.5 log10). No statistical link was found between the reduction in the concentration of the targeted bacteria and the physicochemical and operational parameters likely to have an effect (NH3, volatile fatty acids and total solids contents, hydraulic retention time, presence of co-substrates), underlining the fact that the fate of the bacteria during mesophilic digestion depends on many interacting factors. The reduction in concentrations varied significantly over the sampling period, underlining the need for longitudinal studies to estimate the impact of AD on pathogenic microorganisms.

Effects of Major Families of Modulators on Performances and Gastrointestinal Microbiota of Poultry, Pigs and Ruminants: A Systematic Approach.

Considering the ban on the use of antibiotics as growth stimulators in the livestock industry, the use of microbiota modulators appears to be an alternative solution to improve animal performance. This review aims to describe the effect of different families of modulators on the gastrointestinal microbiota of poultry, pigs and ruminants and their consequences on host physiology. To this end, 65, 32 and 4 controlled trials or systematic reviews were selected from PubMed for poultry, pigs and ruminants, respectively. Microorganisms and their derivatives were the most studied modulator family in poultry, while in pigs, the micronutrient family was the most investigated. With only four controlled trials selected for ruminants, it was difficult to conclude on the modulators of interest for this species. For some modulators, most studies showed a beneficial effect on both the phenotype and the microbiota. This was the case for probiotics and plants in poultry and minerals and probiotics in pigs. These modulators seem to be a good way for improving animal performance.

Controlled Experimental Infection in Pigs with a Strain of Yersinia enterocolitica Harboring Genetic Markers for Human Pathogenicity: Colonization and Stability.

Yersinia enterocolitica (Ye) is one of the major causes of foodborne zoonosis. The BT4/O:3 bioserotype is most commonly isolated in human infections. Pigs are considered the main reservoir of Ye, and hence, understanding the dynamics of infection by this pathogen at the individual and group levels is crucial. In the present study, an experimental model was validated in Large White pigs infected with a BT4/O:3 strain. This study showed that Ye contamination in pigs may occur via the introduction of the bacteria not only by mouth but also by snout, with a colonization process consisting of three periods corresponding to three contamination statuses of pigs: P1, corresponding to the 24 h following ingestion or inhalation of Ye with the appearance of bacteria in tonsils or in feces; P2, from 2 days postinoculation (dpi), corresponding to expansion of Ye and colonization of the digestive system and extraintestinal organs associated with an IgG serological response; and P3, after 21 dpi, corresponding to regression of colonization with intermittent Ye detection in tonsils and feces. Although the inoculated strain persisted up to 56 dpi in all pigs, genetic variations with the loss of the gene yadA (a gene involved in human infection) and the emergence of two new multilocus variable-number tandem-repeat analysis (MLVA) profiles were observed in 33% of the 30 isolates studied. This experimental infection model of pigs by Ye provides new insights into the colonization steps in pigs in terms of bacterial distribution over time and bacterial genetic stability.

One Health surveillance-A cross-sectoral detection, characterization, and notification of foodborne pathogens.

Introduction: Several Proficiency Test (PT) or External Quality Assessment (EQA) schemes are currently available for assessing the ability of laboratories to detect and characterize enteropathogenic bacteria, but they are usually targeting one sector, covering either public health, food safety or animal health. In addition to sector-specific PTs/EQAs for detection, cross-sectoral panels would be useful for assessment of the capacity to detect and characterize foodborne pathogens in a One Health (OH) perspective and further improving food safety and interpretation of cross-sectoral surveillance data. The aims of the study were to assess the cross-sectoral capability of European public health, animal health and food safety laboratories to detect, characterize and notify findings of the foodborne pathogens Campylobacter spp., Salmonella spp. and Yersinia enterocolitica, and to develop recommendations for future cross-sectoral PTs and EQAs within OH. The PT/EQA scheme developed within this study consisted of a test panel of five samples, designed to represent a theoretical outbreak scenario. Methods: A total of 15 laboratories from animal health, public health and food safety sectors were enrolled in eight countries: Denmark, France, Italy, the Netherlands, Poland, Spain, Sweden, and the United Kingdom. The laboratories analyzed the samples according to the methods used in the laboratory and reported the target organisms at species level, and if applicable, serovar for Salmonella and bioserotype for Yersinia. Results: All 15 laboratories analyzed the samples for Salmonella, 13 for Campylobacter and 11 for Yersinia. Analytical errors were predominately false negative results. One sample (S. Stockholm and Y. enterocolitica O:3/BT4) with lower concentrations of target organisms was especially challenging, resulting in six out of seven false negative results. These findings were associated with laboratories using smaller sample sizes and not using enrichment methods. Detection of Salmonella was most commonly mandatory to notify within the three sectors in the eight countries participating in the pilot whereas findings of Campylobacter and Y. enterocolitica were notifiable from human samples, but less commonly from animal and food samples. Discussion: The results of the pilot PT/EQA conducted in this study confirmed the possibility to apply a cross-sectoral approach for assessment of the joint OH capacity to detect and characterize foodborne pathogens.

Inflammatory Responses Induced by the Monophasic Variant of Salmonella Typhimurium in Pigs Play a Role in the High Shedder Phenotype and Fecal Microbiota Composition.

Pigs infected with Salmonella may excrete large amounts of Salmonella, increasing the risk of spread of this pathogen in the food chain. Identifying Salmonella high shedder pigs is therefore required to mitigate this risk. We analyzed immune-associated markers and composition of the gut microbiota in specific-pathogen-free pigs presenting different shedding levels after an oral infection with Salmonella. Immune response was studied through total blood cell counts, production of anti-Salmonella antibodies and cytokines, and gene expression quantification. Total Salmonella shedding for each pig was estimated and hierarchical clustering was used to cluster pigs into high, intermediate, and low shedders. Gut microbiota compositions were assessed using 16S rRNA microbial community profiling. Comparisons were made between control and inoculated pigs, then between high and low shedders pigs. Prior to infection, high shedders had similar immunological profiles compared to low shedders. As soon as 1 day postinoculation (dpi), significant differences on the cytokine production level and on the expression level of several host genes related to a proinflammatory response were observed between high and low shedders. Infection with Salmonella induced an early and profound remodeling of the immune response in all pigs, but the intensity of the response was stronger in high shedders. In contrast, low shedders seroconverted earlier than high shedders. Just after induction of the proinflammatory response (at 2 dpi), some taxa of the fecal microbiota were specific to the shedding phenotypes. This was related to the enrichment of several functional pathways related to anaerobic respiration in high shedders. In conclusion, our data show that the immune response to Salmonella modifies the fecal microbiota and subsequently could be responsible for shedding phenotypes. Influencing the gut microbiota and reducing intestinal inflammation could be a strategy for preventing Salmonella high shedding in livestock. IMPORTANCE Salmonellosis remains the most frequent human foodborne zoonosis after campylobacteriosis and pork meat is considered one of the major sources of human foodborne infections. At the farm, host heterogeneity in pig infection is problematic. High Salmonella shedders contribute more significantly to the spread of this foodborne pathogen in the food chain. The identification of predictive biomarkers for high shedders could help to control Salmonella in pigs. The purpose of the present study was to investigate why some pigs become super shedders and others low shedders. We thus investigated the differences in the fecal microbial composition and the immune response in orally infected pigs presenting different Salmonella shedding patterns. Our data show that the proinflammatory response induced by S. Typhimurium at 1 dpi could be responsible for the modification of the fecal microbiota composition and functions observed mainly at 2 and 3 dpi and to the low and super shedder phenotypes.

Occurrence and Diversity of Listeria monocytogenes Isolated from Two Pig Manure Treatment Plants in France.

The presence of Listeria monocytogenes in piggery effluents intended for irrigation crops may be a source of bacterial dissemination in agriculture. The occurrence and diversity of L. monocytogenes in the farm environment were examined in two pig manure treatment systems (S1 and S2). Samples collected over the course of one year consisted of manure, the liquid fraction of treated manure (lagoon effluent), and soil surrounding the lagoon. L. monocytogenes was enumerated using the Most Probable Number (MPN) method, serotyped by PCR, genotyped by pulsed-field gel electrophoresis (PFGE), and sequenced for multilocus sequence typing (MLST). L. monocytogenes was detected in 92% of manure samples and in approximately 50% of lagoon effluent and soil samples. Concentrations ranged between 5 and 103 MPN 100| |mL-1. Serogroups IIa, IIb, and IVb were identified. Diversity was high with 44 PFGE profiles (252 isolates) and 17 clonal complexes (CCs) (96 isolates) with higher diversity in manure at site S1 supplied by four farms. Some PFGE profiles and CCs identified in manure or in pig feces from a previous study were also detected in lagoons and/or soil, reflecting pig L. monocytogenes circulation throughout the manure treatment and in the vicinity of the sampling sites. However, some PFGE profiles and CCs were only found in the lagoon and/or in soil, suggesting an origin other than pigs. The present study highlights the limited ability of biological treatments to eliminate L. monocytogenes from pig manure. The persistence of some PFGE profiles and CCs throughout the year in the lagoon and soil shows the ability of L. monocytogenes to survive in this type of environment.

Characterization of Clostridioides difficile strains isolated from manure and digestate in five agricultural biogas plants.

Clostridioides difficile strains were isolated from manure and digestate samples from five biogas plants in France. The objective of this study was to characterize these isolates using PCR ribotyping, wgMLST, a multiplex PCR targeting genes encoding for the main virulence factors, i.e. tcdA, tcdB, cdtA and cdtB, and antimicrobial susceptibility assays. The 54 strains characterized were all positive for tcdA and tcdB and 83% (45/54) were positive for the binary toxin genes. PCR ribotypes 126 (59%) and 078 (37%) were predominant, and wgMLST analysis of 18 isolates showed close proximity of strains within a single biogas plant. Samples from the biogas plant supplied with cattle and poultry manure displayed the largest variety in PCR ribotypes. The in vitro activities of nine antimicrobial agents were determined. All the strains were susceptible to vancomycin and metronidazole, which are currently considered first-line treatments for C. difficile infection in humans. All the strains were resistant to clindamycin. The results of this study show that a high percentage of C. difficile strains present in the French biogas plants investigated are toxigenic strains from PCR ribotypes also commonly found in humans.

Longitudinal study describing time to Salmonella seroconversion in piglets on three farrow-to-finish farms.

BACKGROUND: Pigs are frequently colonised with Salmonella enterica, and this constitutes a major risk for human salmonellosis. The infection can be assessed by the serological response of pigs to S enterica. A longitudinal study was undertaken on-farm to correctly describe this serological response and investigate factors associated with age at Salmonella seroconversion. METHODS: Three pig farms and in each farm three successive batches were considered. Per batch, 40 piglets were selected at random from 10 sows (four piglets per sow). Blood was sampled from sows one week after farrowing and from piglets at weeks 1, 6, 10, 14, 18 and 22 and at the slaughterhouse. Salmonella antibodies were detected in serum using a commercial ELISA test. Factors related to farm characteristics, batch management system, porcine reproductive and respiratory syndrome infection, and sows' Salmonella serological status were recorded to assess their effect on age at seroconversion. RESULTS: At week 1 after farrowing, 96.5 per cent of the sows had antibodies against Salmonella. The serological results of piglets at weeks 1 and 6 only were positively correlated with those of the sows. The average age at Salmonella seroconversion was 137±2.2 days (confidence interval at 95 per cent). The first seroconversions occurred from weeks 10 to 14, but most of the pigs (54.6 per cent) were seropositive at the end of the fattening period, with variations between farms and batches (28.9-75.7 per cent). Herd/farm was significantly associated with age at seroconversion. CONCLUSION: This longitudinal study allowed the authors to follow precisely the evolution of Salmonella seroconversion from maternity to slaughterhouse and confirm the relationship between the seroconversion of sows and serology of their piglets. Moreover, factors related to farm practices and management as a whole are more influential than individual factors (at the pig level) on age at Salmonella seroconversion.

Evaluation of the occurrence of sporulating and nonsporulating pathogenic bacteria in manure and in digestate of five agricultural biogas plants.

The number of agricultural biogas plants has been increasing in the past decades in some European countries. Digestates obtained after anaerobic digestion (AD) of manure are usually spread on agricultural land; however, their hygiene status regarding pathogens posing public health and/or animal health challenges has been poorly characterized up to now in France. In this study, three replicates of manure and digestate were collected from five farm biogas plants receiving animal manure in order to assess the occurrence and concentrations of sporulating (Clostridium botulinum, Clostridioides difficile, Clostridium perfringens) and nonsporulating (Listeria monocytogenes, thermotolerant Campylobacter spp., Salmonella, Escherichia coli, enterococci) bacteria. Concentrations of E. coli, enterococci, and C. perfringens in digestates ranged from 102 to 104 , 104 to 105 , and <103 to 7 × 105  CFU/g, respectively. Salmonella and C. difficile were detected in manure and digestate from the five biogas plants at concentrations ranging from <1.3 to >7 × 102  MPN/g and from 1.3 to 3 × 102  MPN/g, respectively. Thermotolerant Campylobacter, detected in all the manures, was only found in two digestates at a concentration of cells ranging from <10 to 2.6 × 102  CFU/g. Listeria monocytogenes and C. botulinum were detected in three manures and four digestates. The bacterial counts of L. monocytogenes and C. botulinum did not exceed 3 × 102 and 14 MPN/g, respectively. C. botulinum type B was detected at very low level in both the manure and digestate of farm biogas plants with no botulism history. The levels of pathogenic bacteria in both manure and digestate suggested that some bacteria can persist throughout AD.

Experimental infection of pigs by Salmonella Derby, S. Typhimurium and monophasic variant of S. Typhimurium: Comparison of colonization and serology.

Salmonella serovars Derby, Typhimurium and the monophasic variant of Salmonella Typhimurium are the most frequently isolated serovars in pigs in France. To compare the excretion patterns, seroconversion to Salmonella and contamination of the organs of pigs inoculated with strains of all three serovars, we conducted an experimental trial with 28 SPF piglets. Four were used as a negative control, while the other 24 were divided equally into three groups. Each group was inoculated at 7 weeks of age with a different strain: S. Derby (SDb), S. Typhimurium (ST), and the monophasic variant of S. Typhimurium (mST). Fecal and blood samples were collected twice a week up until necropsy, on 21 days post-inoculation (DPI) for half of each group and 49 DPI for the remaining piglets. During necropsy, the tonsils, mesenteric lymph nodes and various intestinal contents were collected from each pig. Salmonella bacteria were quantified in CFU/g by a bacteriological method, and levels of Salmonella antibodies were measured using an ELISA Kit. Piglets inoculated with mST continuously excreted Salmonella in their feces throughout the trial. For each of the other serovars, one piglet was Salmonella-negative on one DPI. The quantity of Salmonella excreted was statistically different between the group inoculated with ST and mST (p < 0.05), but no differences were found between the other serovars. The tonsils, cecum and jejunum were the most contaminated organs in all groups. Seroconversion for all the piglets was completed by different DPI: 28 for ST, 31 for mST and 38 for SDb. No major differences were found in terms of excretion and colonization among the studied serovars.

An overview of the immunogenicity and effectiveness of current human rabies vaccines administered by intradermal route.

Pre- as well as post-exposure prophylaxis plays an important role in controlling the number of deaths associated with human rabies. Rabies vaccines, classically injected intramuscularly, are now also administered by intradermal (ID) route. Vaccines to be administered by the ID route should meet the same quality, safety and efficacy specifications as vaccines for intramuscular (IM) use. The aim of this paper is to provide information based on publicly available data regarding the ID use of rabies vaccines and to identify potential needs for further analysis of the potency, immunogenicity and effectiveness of rabies vaccines administered by this route. A first literature search, focused on the immunogenicity of rabies vaccines given by ID route, identified 338 publications in the period 1997-2018, 40 of which were included in our analyses. A second search investigating the effectiveness of ID vaccination resulted in 371 hits for the period 2007-2018, of which 13 suitable publications were retained. The immunogenicity of current rabies vaccines was analyzed in 3 ways: proportion of subjects reaching the antibody threshold of 0.5 IU/ml after ID vaccination, relationship between potency and immunogenicity of the vaccine given intradermally, and comparison of antibody responses after IM or ID vaccination. Overall, vaccines administered intradermally were found immunogenic. Post-exposure prophylaxis by ID route appeared at least as immunogenic as by IM regimens. By contrast, ID pre-exposure prophylaxis trended towards lower antibody titers than IM vaccination, but the observation was not associated with any clinical relevance. Vaccine effectiveness was assessed by investigating survival after exposure. Data from more than 30,000 patients who sought rabies post-exposure prophylaxis did not indicate that current vaccines administered by ID route lack efficacy. These results support current recommendations for ID vaccination against rabies. However, published data on ID performance were associated with significant weaknesses that future research should better address.

Diversity of Yersinia enterocolitica isolated from pigs in a French slaughterhouse over 2 years.

The pig is one of the main reservoirs of Yersinia enterocolitica strains pathogenic to humans. A description of the Y. enterocolitica population in this reservoir, and accurate discriminatory techniques for typing isolates are needed for prevention, outbreak investigation, and surveillance. This study investigates the genetic diversity of pathogenic Y. enterocolitica isolates obtained from pig tonsils in a French pig slaughterhouse in 2009 (S1) and 2010 (S2). The use of Pulsed-Field Gel Electrophoresis (PFGE) and MLVA as typing techniques was also compared and evaluated. First, a total of 167 isolates (12 of biotype 3 recovered during S1, and 155 of biotype 4 recovered during S1 and S2) were typed by PFGE using the XbaI enzyme. MLVA was then tested on all the biotype 3 isolates in addition to 70 selected biotype 4 isolates recovered over the 2 years. PFGE generated two specific XbaI-PFGE profiles for biotype 3 isolates. Nine XbaI profiles were obtained for biotype 4, with a higher diversity (ID = 0.599) than biotype 3 (ID = 0.167). Two out of the nine XbaI profiles were reported during both surveys and at different months. MLVA improved the differentiation between isolates; the index of diversity reached 0.621 and 0.958, respectively, for biotype 3 (three MLVA types) and biotype 4 (32 MLVA types). The MLVA types for biotype 4 differed over the two surveys, but some isolates with different MLVA types were genetically closely related. This study provides an initial evaluation of the genetic diversity of Y. enterocolitica strains isolated from pigs in France. We show that some PFGE profiles are maintained in the pig production sector, and, through MLVA, that part of the Y. enterocolitica population remained genetically close over the two years. MLVA proved its effectiveness as a tool for investigating pathogenic Y. enterocolitica strains isolated from pigs.

Effect of Titanium Dioxide Film Thickness on Photocatalytic and Bactericidal Activities Against Listeria monocytogenes.

Structural, microstructural and bactericidal surface properties of TiO2 -coated glass substrates elaborated by reactive Radiofrequency sputtering are investigated. As pathogenic bacteria in biofilms are a major concern in food industries due to their growing resistance to cleaning and sanitizing procedures, the development of photoactive surfaces exhibiting bactericidal properties is acknowledged as an effective approach to tackle bacterial contaminations. Our principal aim concerns the study of the photoactive top-layer thickness impact (from 80 nm to ~500 nm) on Listeria monocytogenes. Structural characterization of the TiO2 layers demonstrates that anatase and rutile phases are both present, depending on the film thickness. Photocatalytic activity of the samples has been evaluated through the degradation of aqueous methylene blue (MB) solutions under UVA light illumination for various time periods. The results show an efficiency rating increase according to TiO2 film thickness up to a threshold value close to 400 nm. Moreover, a significant decrease of the adherent bacteria number is observed after 20 min of UVA illumination. The quantitative study of the bactericidal activity associated with scanning electron microscopy observations of the postprocess bacteria damaged cells demonstrates the efficiency of the 240-nm-thick TiO2 coating sample. The results are correlated with the production of hydroxyl radicals during the process of photocatalysis.

Colonization of Pigs Experimentally Infected with a Monophasic Variant of Salmonella Typhimurium.

The monophasic variant of Salmonella Typhimurium is highly prevalent in human and in pork. However, little is known about colonization dynamics and serology in pigs. We orally inoculated 24 seven-week-old piglets with 109 CFU/pig of a porcine strain of monophasic Salmonella Typhimurium in an experimental trial. Three groups of eight piglets were orally inoculated and monitored for 21, 49, or 84 days post-inoculation until necropsied. From 3 days post-inoculation to necropsy, individual feces were sampled twice weekly and blood once weekly. At necropsy, the tonsils, mesenteric lymph nodes, and the contents of the duodenum, jejunum, ileum, and cecum were collected from each pig. We determined the number of CFU/g in all the samples and measured also Salmonella antibodies in OD% in all blood samples. At different times during the trial, we tested by MLVA (Multilocus Variable Number Tandem Repeat Analysis) the genomic stability of the strain after passing through the intestinal tract. Salmonella was continuously excreted by pigs, ranging from 1.4 to 5.8 log10 CFU/g. At necropsy, Salmonella was present in all samples, but the tonsils were particularly infected. Salmonella antibodies were detected in five pigs 7 days post-inoculation. At 49 days post-inoculation, all the pigs were seropositive. We observed new MLVA types for 3.3% of the isolates tested over the trial. Our study allowed us to show the serovar's ability to persist in pigs after infection up to 84 days post-inoculation. We demonstrated that Salmonella seroconversion appeared earlier than in naturally infected pigs and that the strain's genome can evolve after passing through the digestive tract of pigs.

Population Genetic Structure of Listeria monocytogenes Strains Isolated From the Pig and Pork Production Chain in France.

Listeria monocytogenes is an ubiquitous pathogenic bacterium, transmissible to humans through the consumption of contaminated food. The pork production sector has been hit hard by a series of L. monocytogenes-related food poisoning outbreaks in France. An overview of the diversity of strains circulating at all levels of the pork production chain, from pig farming (PF) to finished food products (FFP), is needed to identify the contamination routes and improve food safety. Until now, no typing data has been available on strains isolated across the entire pig and pork production chain. Here, we analyzed the population genetic structure of 687 L. monocytogenes strains isolated over the last 20 years in virtually all the French départements from three compartments of this production sector: PF, the food processing environment (FPE), and FFP. The genetic structure was described based on Multilocus sequence typing (MLST) clonal complexes (CCs). The CCs were obtained by mapping the PFGE profiles of the strains. The distribution of CCs was compared firstly between the three compartments and then with CCs obtained from 1106 strains isolated from other food production sectors in France. The predominant CCs of pig and pork strains were not equally distributed among the three compartments: the CC37, CC59, and CC77 strains, rarely found in FPE and FFP, were prevalent in PF. The two most prevalent CCs in the FPE and FFP compartments, CC9 and CC121, were rarely or never detected in PF. No CC was exclusively associated with the pork sector. Three CCs (CC5, CC6, and CC2) were considered ubiquitous, because they were observed in comparable proportions in all food production sectors. The two most prevalent CCs in all sectors were CC9 and CC121, but their distribution was disparate. CC9 was associated with meat products and food products combining several food categories, whereas CC121 was not associated with any given sector. Based on these results, CC121 is likely able to colonize a larger diversity of food products than CC9. Both CCs being associated with the food production suggests, that certain processing steps, such as slaughtering or stabilization treatments, favor their settlement and the recontamination of the food produced.

No Clear Differences between Organic or Conventional Pig Farms in the Genetic Diversity or Virulence of Campylobacter coli Isolates.

To evaluate the impact of pig farm management on the genetic diversity and on the virulence of Campylobacter coli, we characterized isolates from 19 organic pig farms (62 isolates) and from 24 conventional pig farms (58 isolates). The 120 C. coli isolates were typed using pulsed field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) and the presence of nine virulence genes was screened using real-time PCR. The capacity of adhesion and invasion of 61 isolates (32 from organic and 29 from conventional farms) were then tested on human intestinal Caco-2 cells. A total of 59 PFGE types and of 50 sequence types (STs) were identified. Twelve PFGE types and nine STs, accounting for 34 and 41.6% of the isolates, respectively, were common between the two production systems with ST854 dominating (18.3% of the isolates). Twenty-nine PFGE types and 25 STs were only found in isolates from organic farms, and 18 PFGE types and 16 STs from conventional farms. No significant differences were found in diversity despite the differences in rearing systems, except at the locus level for the glnA, gltA, and uncA genes. All isolates, regardless of their origin, carried the ceuE, iam, ciaB, and flaA genes and more than 95% of the isolates carried the cadF and cdtABC genes. No significant differences were found in pathogenicity between the two farming systems. The pathogenicity of the C. coli isolates was low compared to C. jejuni control strains tested. The plasmid gene virb11 was detected in only 13 isolates from organic farms; these isolates showed greater invasion capacity than those without this gene. Our study indicates that pig farm management does not significantly affect the diversity and the virulence of Campylobacter coli isolated from pigs. The common genotypes between conventional and organic farms may indicate that some genotypes are adapted to pigs.

Campylobacter coli in Organic and Conventional Pig Production in France and Sweden: Prevalence and Antimicrobial Resistance.

The purpose of the study was to evaluate and compare the prevalence and antimicrobial resistance of Campylobacter coli in conventional and organic pigs from France and Sweden. Fecal or colon samples were collected at farms or at slaughterhouses and cultured for Campylobacter. The minimum inhibitory concentrations of ciprofloxacin, nalidixic acid, streptomycin, tetracycline, erythromycin, and gentamicin were determined by microdilution for a total of 263 French strains from 114 pigs from 50 different farms and 82 Swedish strains from 144 pigs from 54 different farms. Erythromycin resistant isolates were examined for presence of the emerging rRNA methylase erm(B) gene. The study showed that within the colon samples obtained in each country there was no significant difference in prevalence of Campylobacter between pigs in organic and conventional productions [France: conventional: 43/58 (74%); organic: 43/56 (77%) and Sweden: conventional: 24/36 (67%); organic: 20/36 (56%)]. In France, but not in Sweden, significant differences of percentages of resistant isolates were associated with production type (tetracycline, erythromycin) and the number of resistances was significantly higher for isolates from conventional pigs. In Sweden, the number of resistances of fecal isolates was significantly higher compared to colon isolates. The erm(B) gene was not detected in the 87 erythromycin resistant strains tested.