Title Participants Abstract "A limited role for SsrA/B in persistent Salmonella Typhimurium infections in pigs" "Filip Boyen, Frank Pasmans, Filip Van Immerseel, Eirwen Morgan, Nadine Botteldoorn, Marc Heyndrickx, Jiri Volf, Herman Favoreel, Jean-Pierre Hernalsteens, Richard Ducatelle, Freddy Haesebrouck" "Virulence genes regulated by the SsrA/B system are indispensable for systemic disease in BALB/c mice. The role of this regulating system in the pathogenesis of Salmonella Typhimurium infections in pigs is not documented. In the present study, the interactions of Salmonella Typhimurium and an ssrA/B mutant were compared in vitro and in vivo. The ssrA/B mutant strain displayed decreased Salmonella Pathogenicity Island 2 (SPI-2) expression levels, showed a replication defect in mouse macrophages and was attenuated in a mouse model after oral inoculation. Using real time qRT-PCR and a porcine ileal loop model, it was shown that the ssrA/B mutant strain was not significantly attenuated in overall virulence and SPI-1 expression in specific. Flowcytometric analysis demonstrated that the ssrA/B mutant strain was defective in intracellular replication in porcine macrophages. After oral inoculation of piglets with the wild type strain or the ssrA/B mutant strain, the animals of both groups excreted Salmonella and were colonized by Salmonella to the same extent. In an intravenous mixed infection model, the ssrA/B mutant strain was defective in the colonization of several internal organs. These results suggest that the ssrA/B gene of Salmonella Typhimurium plays a limited role in the persistent intestinal colonization of pigs." "Investigating the meat pathway as a source of human nontyphoidal Salmonella bloodstream infections and diarrhea in East Africa" "John A Crump, Kate M Thomas, Jackie Benschop, Matthew A Knox, David A Wilkinson, Anne C Midwinter, Peninah Munyua, John B Ochieng, Godfrey M Bigogo, Jennifer R Verani, Marc-Alain Widdowson, Gerard Prinsen, Sarah Cleaveland, Esron D Karimuribo, Rudovick R Kazwala, Blandina T Mmbaga, Emanuel S Swai, Nigel P French, Ruth N Zadoks" "BACKGROUND: Salmonella Enteritidis and Salmonella Typhimurium are major causes of bloodstream infection and diarrheal disease in East Africa. Sources of human infection, including the role of the meat pathway, are poorly understood.METHODS: We collected cattle, goat, and poultry meat pathway samples from December 2015 through August 2017 in Tanzania and isolated Salmonella using standard methods. Meat pathway isolates were compared with nontyphoidal serovars of Salmonella enterica (NTS) isolated from persons with bloodstream infections and diarrheal disease from 2007 through 2017 from Kenya by core genome multi-locus sequence typing (cgMLST). Isolates were characterized for antimicrobial resistance, virulence genes, and diversity.RESULTS: We isolated NTS from 164 meat pathway samples. Of 172 human NTS isolates, 90 (52.3%) from stool and 82 (47.7%) from blood, 53 (30.8%) were Salmonella Enteritidis sequence type (ST) 11 and 62 (36.0%) were Salmonella Typhimurium ST313. We identified cgMLST clusters within Salmonella Enteritidis ST11, Salmonella Heidelberg ST15, Salmonella Typhimurium ST19, and Salmonella II 42:r:- ST1208 that included both human and meat pathway isolates. Salmonella Typhimurium ST313 was isolated exclusively from human samples. Human and poultry isolates bore more antimicrobial resistance and virulence genes and were less diverse than isolates from other sources.CONCLUSIONS: Our findings suggest that the meat pathway may be an important source of human infection with some clades of Salmonella Enteritidis ST11 in East Africa, but not of human infection by Salmonella Typhimurium ST313. Research is needed to systematically examine the contributions of other types of meat, animal products, produce, water, and the environment to nontyphoidal Salmonella disease in East Africa." "A limited role for SsrA/B in persistent Salmonella Typhimurium infections in pigs" "F. Boyen, Frank Pasmans, F. Van Immerseel, E. Morgan, N. Botteldoorn, Marc Heyndrickx, J. Volf, Herman Favoreel, Jean-Pierre Hernalsteens, Richard Ducatelle, Freddy Haesebrouck" "Virulence genes regulated by the SsrA/B system are indispensable for systemic disease in BALB/c mice. The role of this regulating system in the pathogenesis of Salmonella Typhimurium infections in pigs is not documented. In the present study, the interactions of Salmonella Typhimurium and an ssrA/B mutant were compared in vitro and in vivo. The ssrA/B mutant strain displayed decreased Salmonella Pathogenicity Island 2 (SPI-2) expression levels, showed a replication defect in mouse macrophages and was attenuated in a mouse model after oral inoculation. Using real time qRT-PCR and a porcine ileal loop model, it was shown that the ssrA/B mutant strain was not significantly attenuated in overall virulence and SPI-1 expression in specific. Flowcytometric analysis demonstrated that the ssrA/B mutant strain was defective in intracellular replication in porcine macrophages. After oral inoculation of piglets with the wild type strain or the ssrA/B mutant strain, the animals of both groups excreted Salmonella and were colonized by Salmonella to the same extent. In an intravenous mixed infection model, the ssrA/B mutant strain was defective in the colonization of several internal organs. These results suggest that the ssrA/B gene of Salmonella Typhimurium plays a limited role in the persistent intestinal colonization of pigs." "Mycotoxins in pig feed: influence on the gut and interaction with Salmonella Typhimurium infections" "Elin Verbrugghe, Virginie Vandenbroucke, Frank Pasmans, Joline Goossens, Patrick De Backer" "Animal sources of antimicrobial-resistant bacterial infections in humans : a systematic review" "Christina Fastl, Helena Cardoso de Carvalho Ferreira, Sara Babo Martins, João Sucena Afonso, Carlotta di Bari, Narmada Venkateswaran, Sara Monteiro Pires, Lapo Mughini-Gras, Ben Huntington, Jonathan Rushton, David Pigott, Brecht Devleesschauwer" "Investigation of the concurrent colonization with Campylobacter and Salmonella in poultry flocks and assessment of the sampling site for status determination at slaughter" "Geertrui Rasschaert, K. Houf, J Van Hende, L. De Zutter" "Fifty six broiler flocks and 20 laying hen and breeder flocks were sampled in six slaughterhouses for the presence of Campylobacter and Salmonella. Samples were taken from three different sites of the gastrointestinal tract, namely from the crop, the duodenum and the ceca. The prevalence of flocks colonized with Campylobacter and Salmonella was determined and an association between the concurrent colonization with these two pathogens was investigated. Furthermore, the best sampling site for status determination at the slaughterhouse level was evaluated. Of the broiler flocks, 73% were colonized with Campylobacter, whereas 13% were Salmonella-positive at slaughter. Concerning the laying hen and breeder flocks, all flocks were colonized with Campylobacter and 65% of the flocks were Salmonella-positive. No association was found between Campylobacter and Salmonella occurrence in broiler flocks. Since all laying hen and breeder flocks were colonized with Campylobacter, no association between the concurrent colonization with the two pathogens could be determined. At the slaughterhouse level, sampling only the duodena was sufficient to determine the Campylobacter status of poultry flocks, whereas the three sampling sites had to be analyzed to detect all flocks colonized with Salmonella." "Effect of farm type on within-herd Salmonella prevalence, serovar distribution, and antimicrobial resistance" "Geertrui Rasschaert, J Michiels, D Arijs, Christa Wildemauwe, Stefaan De Smet, Marc Heyndrickx" "Salmonella represents a major challenge to the pig industry, as pork presents a risk for human salmonellosis. In this study, we have examined the effect of farm type on the prevalence of fattening pigs shedding Salmonella on 12 farms at risk for harboring Salmonella. On six open (grow-to-finish) and six closed (farrow-to-finish) farms, the prevalence of pigs shedding Salmonella was determined on two occasions approximately 2 months apart. The serovar, phage type, and antimicrobial resistance of the obtained Salmonella isolates were determined. On all farms, pigs shedding Salmonella were detected on at least one of the two sampling days. The mean within-herd prevalence was 7.8%. Closed farms were two times less likely to have pigs shedding Salmonella than open farms. On open farms, the odds of finding Salmonella shedding in pigs were 1.9 times higher when sampling was performed at slaughter age than when samples were taken halfway through the fattening period. Salmonella enterica serovar Typhimurium was the most predominant serotype, with a prevalence of 62 to 63% on both farm types. Of all the Salmonella Typhimurium isolates, 65% had the tetraresistant profile ASSuT (ampicillin, streptomycin, sulfonamide, and tetracycline) with or without additional resistance to trimethoprim-sulfonamide. Phage type DT120 seemed to be especially associated with this antimicrobial-resistant profile. The prevalence of Salmonella Typhimurium isolates showing resistance to ampicillin, streptomycin, tetracycline, sulfonamide, trimethoprim-sulfonamide, and lincomycin hydrochloride and spectinomycin sulfate tetrahydrate was significantly higher on open farms than on closed farms." "Persistent Salmonella Enteritidis environmental contamination on layer farms in the context of an implemented national control program with obligatory vaccination" "Isabelle Dewaele, H Van Meirhaeghe, Geertrui Rasschaert, M Vanrobaeys, E De Graef, Lieve Herman, R Ducatelle, Marc Heyndrickx, Koen De Reu" "The aim of this study was to closely examine the Salmonella enterica serovar Enteritidis environmental contamination on persistently positive layer farms in Belgium during successive laying cycles. All of the farms were required to vaccinate their layers under the national control program for Salmonella. Seven farms with previous or current Salmonella Enteritidis contamination were monitored during different stages of the laying period and after cleaning and disinfection (CD). Environmental samples, including from the equipment and vermin, were taken in the henhouse and egg-collecting area. Dilutions were performed to define the degree of Salmonella Enteritidis contamination. Eggshells, egg contents, and ceca were also tested for Salmonella. At the end of the first sampled laying period, 41.6% of the environmental samples were contaminated with Salmonella Enteritidis. After CD, the prevalence dropped to 11.4%. On average, the prevalence in the second laying period increased again: 17.8, 18.4, and 22.3% at the onset, middle, and end of the lay period, respectively. After CD before the third laying period, the prevalence decreased to 6.6% and stabilized at the onset of lay (6.3%). During lay, as well as after CD, a wide variety of contaminated environmental samples were found; for example, in the henhouse, in the egg-collecting area, on mobile equipment and in or on vermin. In the henhouse during laying, the most recurrent and highly contaminated sites were the overshoes, floor, manure belt, and hen feces. The egg-collecting area had a significantly higher number of contaminated samples compared with that of the henhouse. For both sites, the floor appeared to be the most suitable sampling site to estimate the Salmonella Enteritidis status of the farms. Eggshell and egg content contamination varied between 0.18 and 1.8% and between 0.04 and 0.4%, respectively. In total, 2.2% of the analyzed ceca contained Salmonella Enteritidis. This study revealed that Salmonella Enteritidis is present in the environment of persistently Salmonella Enteritidis-contaminated layer farms, demonstrated that in many cases Salmonella Enteritidis contamination was not eliminated after CD, and identified the egg-collecting area as a critical point on most farms." "The impact of prebiotics and salmonellosis on apparent nutrient digestibility and Salmonella typhimurium var. Copenhagen excretion in adult pigeons (Columba livia domestica)" "G.P.J. Janssens, Sam Millet, F. Van Immerseel, J. De Buck, M Hesta" "The effects of lactose or fructo-oligosaccharide (FOS) supplementation on the excretion of salmonellae, apparent digestibilities and excreta consistency were studied. Thirty-two male pigeons (Columba livia domestica) were randomly divided into 4 equal groups: 3 of 4 groups were orally infected with 10(9) Salmonella Typhimurium var. Copenhagen, after being offered a drinking water supplement of 2% FOS, 2% lactose, or no supplement, respectively, for 2 wk. Pigeons in the fourth group were not challenged with S. Typhimurium and remained unsupplemented. Initially, FOS increased water intake, resulting in more watery excreta. After infection, supplementation showed no major effects on S. Typhimurium excretion, nitrogen retention, or apparent nutrient digestibilities, although lactose--and to a lesser extent FOS--improved apparent fiber digestibility during recovery from the S. Typhimurium infection. The excreta consistency of all pigeons returned to normal when recovering from the Salmonella infection. In this trial, neither FOS nor lactose was successful in tempering the negative aspects of Salmonella infection in pigeons. Nevertheless, it should be stated that future investigations should clarify the importance of duration and level of prebiotic supplementation and infection level." "Limited genetic diversity and gene expression differences between egg- and non-egg-related Salmonella Enteritidis strains" "Nadine Botteldoorn, Els Van Coillie, J. Goris, Hadewig Werbrouck, Veerle Piessens, C Godard, Patsy Scheldeman, Lieve Herman, Marc Heyndrickx" "Salmonella Enteritidis strains of egg- and non-egg-related origin were characterized molecularly to find markers correlated with the egg-contaminating property of Salmonella Enteritidis. Isolates were examined by random amplified polymorphic DNA (RAPD), plasmid profiling and phage typing. Furthermore, the presence of 30 virulence genes was tested by PCR. In genetic fingerprinting and gene content, only small differences between the strains were found and no correlation was observed with the origin (egg-related versus non-egg-related). A major RADP group was present in both egg- and non-egg-related strains, but other smaller RAPD groups were present as well in both categories of strains. Phage types PT4 and PT21 were predominant. Differential mRNA expression levels of fimA and agfA under conditions of growth simulating the conditions during egg formation were determined by real-time RT-PCR. Although differences in fimA and agfA expression levels were observed between the strains, these could not be correlated with the origin of the strains (egg-related versus non-egg-related). The highest expression levels of agfA and fimA were only found in two non-egg-related strains, which seemed to be correlated with the presence of a 93 kb plasmid instead of the 60 kb virulence plasmid. Our results seem to indicate only a limited role for at least type I fimbriae (encoded by fim operon) in egg contamination by Salmonella Enteritidis."