Role of SPI-1 in the interactions of Salmonella Typhimurium with porcine macrophages

Salmonella Pathogenicity Island 1 (SPI-1) genes are indispensable for virulence of Salmonella Typhimurium in mice after oral challenge. These genes mediate invasion in intestinal epithelial cells and induce cell death in murine macrophages. The role of SPI-1 in the pathogenesis of Salmonella Typhimurium infections in food producing animals is not known. It was the aim of the present study to characterize the interactions of a porcine Salmonella Typhimurium field strain and its isogenic mutants in the SPI-1 genes hilA, sipA and sipB with porcine macrophages. SPI-1 was found to be important in the invasion of porcine pulmonary alveolar macrophages (PAM) and the induction of the formation of spacious phagosomes. Both early and delayed cytotoxicity were seen in PAM, but only the early cytotoxicity was SPI-1 dependent. Exposure of PAM to Salmonella Typhimurium induced the production of reactive oxygen species (ROS) and interleukin-8, but no differences were noticed between the induction mediated by the wild type strain and its SPI-1 mutant strains. In conclusion, invasion of porcine macrophages and the induction of early, but not delayed, cytotoxicity by Salmonella Typhimurium is SPI-1 dependent. SPI-1 dependent invasion, however, is not a prerequisite to induce a pro-inflammatory response.     

Salmonella Typhimurium induces SPI-1 and SPI-2 regulated and strain dependent downregulation of MHC II expression on porcine alveolar macrophages

ABSTRACT: Foodborne salmonellosis is one of the most important bacterial zoonotic diseases worldwide. Salmonella Typhimurium is the serovar most frequently isolated from persistently infected slaughter pigs in Europe. Circumvention of the host's immune system by Salmonella might contribute to persistent infection of pigs. In the present study, we found that Salmonella Typhimurium strain 112910a specifically downregulated MHC II, but not MHC I, expression on porcine alveolar macrophages in a Salmonella pathogenicity island (SPI)-1 and SPI-2 dependent way. Salmonella induced downregulation of MHC II expression and intracellular proliferation of Salmonella in macrophages were significantly impaired after opsonization with Salmonella specific antibodies prior to inoculation. Furthermore, the capacity to downregulate MHC II expression on macrophages differed significantly among Salmonella strains, independently of strain specific differences in invasion capacity, Salmonella induced cytotoxicity and altered macrophage activation status. The fact that strain specific differences in MHC II downregulation did not correlate with the extent of in vitro SPI-1 or SPI-2 gene expression indicates that other factors are involved in MHC II downregulation as well. Since Salmonella strain dependent interference with the pig's immune response through downregulation of MHC II expression might indicate that certain Salmonella strains are more likely to escape serological detection, our findings are of major interest for Salmonella monitoring programs primarily based on serology.     

A limited role for SsrA/B in persistent Salmonella Typhimurium infections in pigs

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.     

T-2 toxin induced Salmonella Typhimurium intoxication results in decreased Salmonella numbers in the cecum contents of pigs, despite marked effects on Salmonella-host cell interactions

ABSTRACT: The mycotoxin T-2 toxin and Salmonella Typhimurium infections pose a significant threat to human and animal health. Interactions between both agents may result in a different outcome of the infection. Therefore, the aim of the presented study was to investigate the effects of low and relevant concentrations of T-2 toxin on the course of a Salmonella Typhimurium infection in pigs. We showed that the presence of 15 and 83 μg T-2 toxin per kg feed significantly decreased the amount of Salmonella Typhimurium bacteria present in the cecum contents, and a tendency to a reduced colonization of the jejunum, ileum, cecum, colon and colon contents was noticed. In vitro, proteomic analysis of porcine enterocytes revealed that a very low concentration of T-2 toxin (5 ng/mL) affects the protein expression of mitochondrial, endoplasmatic reticulum and cytoskeleton associated proteins, proteins involved in protein synthesis and folding, RNA synthesis, mitogen-activated protein kinase signaling and regulatory processes. Similarly low concentrations (1-100 ng/mL) promoted the susceptibility of porcine macrophages and intestinal epithelial cells to Salmonella Typhimurium invasion, in a SPI-1 independent manner. Furthermore, T-2 toxin (1-5 ng/mL) promoted the translocation of Salmonella Typhimurium over an intestinal porcine epithelial cell monolayer. Although these findings may seem in favour of Salmonella Typhimurium, microarray analysis showed that T-2 toxin (5 ng/mL) causes an intoxication of Salmonella Typhimurium, represented by a reduced motility and a downregulation of metabolic and Salmonella Pathogenicity Island 1 genes. This study demonstrates marked interactions of T-2 toxin with Salmonella Typhimurium pathogenesis, resulting in bacterial intoxication.     

Oxidative and nitrosative responses of the chicken macrophage cell line MQ-NCSU to experimental Salmonella infection

Phagocytes limit replication or kill ingested organisms by producing toxic reactive oxygen and nitrogen species via NADPH oxidase and inducible nitric oxide synthase (iNOS). The present experiments were to investigate the production and the possible roles of superoxide, hydrogen peroxide (H2O2) and nitric oxide (NO) in the MQ-NCSU chicken macrophage cell line infected with Salmonella in vitro. After infection, intracellular Salmonella viable counts remained constant until 24 h post infection (PI) and started to decline from 48 h PI. Infection of cells with S. Typhimurium, S. Enteritidis and S. Gallinarum, as well as exposure to S. Enteritidis LPS induced low, but significant concentrations of superoxide 1 to 2 h PI, as determined by reduction of ferricytochrome c. There was no difference in superoxide production in infected cells and control cells after 4 h. Increased H2O2 was observed from cells infected with all the different Salmonella species between 2 and 3 h of infection. Nitrite was always greater in infected cells compared to uninfected cells at all times. However, Salmonella was not completely eliminated from the cells though these cells are capable of eliciting a noticeable oxidative burst response and great nitrosative responses, indicating that a strong oxidative burst (and other mechanism/s) is essential for the elimination of intracellular Salmonella.     

Salmonella enterica serotype typhimurium and S. Stanley differ in genomic evolutionary patterns and early immune responses in human THP-1 cell line and CD14+ monocytes

Salmonella Typhimurium and S. Stanley are the most prevalent serogroup B serovars to infect humans in Taiwan. The aim was to determine possible factors to influence the prevalence between S. Typhimurium and S. Stanley. Genotypes were determined by pulsed field gel electrophoresis (PFGE) analysis and the intracellular survival, phagocytosis, reactive oxygen species (ROS) production of human monocyte THP-1 cell and tumor necrosis factor-α(TNF-α), interleukin-6 (IL-6), and IL-1βexpression in peripheral blood CD14⁺ cells after infection were analyzed. 182 S. Stanley was clonal disseminated with main pulsotypes 2 from 2004 to 2007. Overall S. Typhimurium evolved more genotypes, while S. Stanley conserved in genotypes. Human blood CD14⁺ monocytes expressed TNF-α, IL-6 and IL-1β differently among serovars and bacterial conditions (live vs. killed). Live S. Stanley and S. Typhimurium suppressed the TNF-α and IL-6 expression compared to killed bacteria. However, live S. Typhimurium stimulated more IL-1β expression than the killed bacteria, but S. Stanley expressed similar IL-1β levels in both conditions. Furthermore, S. Stanley and S. Typhimurium differed in intracellular survival in the THP-1 cells, an early decrease for S. Stanley, not for S. Typhimurium. Additionally, higher reactive oxygen species (ROS) production in THP-1 cells was found agsinst S. Stanley infection, not found in S. Typhimurium. However, some isolates of S. Stanley could recover from early loss to become more in the monocytes than S. Typhimurium. Difference in phagocytized number, intracellular survival, ROS production and IL-1β expression may contribute to prevalence different between two serovars.     

Comparison of growth phase on Salmonella enterica serovar Typhimurium invasion in an epithelial cell line (IPEC J2) and mucosal explants from porcine small intestine

Salmonella Typhimurium DT104 is a zoonotic enteropathogen of increasing concern for human health. In this study, the influence of growth phase on invasiveness of a S. Typhimurium DT104 field isolate and two reference strains (SL1344 and ATCC 14028) was compared in IPEC J2 cells and mucosal explants from porcine ileum. Internalized bacteria were quantified by a gentamicin resistance assay. After 90 min of exposure to the apical aspect of epithelial monolayers or luminal surface of explants, internalization of all S. Typhimurium strains in mid-logarithmic phase of bacterial growth was comparable. Internalization of stationary phase bacteria was reduced relative to log phase bacteria, with DT104 exhibiting the greatest decrease. Growth phase-related differences in S. Typhimurium invasion are similar in porcine intestinal epithelial cells and mucosal explants, but may be greater in multidrug-resistant strains.     

Salmonella enterica serovar Typhimurium and Enteritidis infection of pigs and cytokine signalling in palatine tonsils

Pigs are considered as one of the major sources of zoonotic strains of Salmonella enterica for humans. Out of many S. enterica serovars, S. Typhimurium dominates in pigs, however, in several countries in Central Europe, S. Enteritidis is also quite frequent in pig herds. In this study we therefore compared the colonisation of pigs with S. Typhimurium and S. Enteritidis. We found that 3 weeks after infection S. Enteritidis 147 colonised the intestinal tract in higher quantities but was shed in faeces in lower quantities than S. Typhimurium 17C10. In a second experiment we found out that S. Enteritidis 147 and its SPI-1 and SPI-4 mutants increased proinflammatory cytokine (IL-1β and IL-8) signalling in the ileum 5 days post infection. On the other hand, independent of SPI-1 or SPI-4, S. Enteritidis 147 suppressed expression of IL-18, MCP1, TLR2, CD86, IL-7, IL-10 and IL-15 in the palatine tonsils. The suppression of cytokine signalling may facilitate the initial colonisation of the palatine tonsils by Salmonella. Moreover, immune suppression may also influence pig resistance to opportunistic pathogens and Salmonella infection in pigs thus may become an issue not only in terms of pork contamination but also in terms of affecting the immunological status of pig herds.     

Interaction between Salmonella typhimurium and phagocytic cells in pigs. Phagocytosis, oxidative burst and killing in polymorphonuclear leukocytes and monocytes

Interactions between Salmonella typhimurium and peripheral blood leucocytes from healthy, Salmonella-free pigs were investigated in vitro. Both granulocytes and monocytes phagocytized FITC-labelled heat-killed Salmonella bacteria as shown by flow cytometry. Phagocytosis in whole blood and isolated leucocytes was measured as acquired fluorescence in the leukocytes and was both time and dose related. Living, serum-opsonized Salmonella bacteria induced a dose-dependent oxidative burst in PMNs and monocytes as measured by luminol-enhanced chemiluminescence (LC). When opsonized in normal serum the Salmonella bacteria, in the range of 2-5 x 10(7) cfu, induced a LC response in monocytes comparable to the level of responses induced by phorbol myristate acetate (PMA) and opsonized zymosan, and the Salmonella-induced response was only marginally reduced by superoxide dismutase (SOD). Intracellular killing of Salmonella by monocytes was assessed from plate colony counts of lysed monocytes and showed that Salmonella typhimurium was able to survive and proliferate in adherent monocytes in vitro despite a reduction in intracellular cfu during the first hour's incubation in cells from some pigs. Experiments with the exhaustion of oxidative burst in non-adherent monocytes were performed by prestimulation with PMA, heat-killed Salmonella or buffer. Prestimulation with PMA led to a strong reduction in oxidative burst induced by living opsonized Salmonella bacteria, whereas prestimulation with heat-killed bacteria gave rise to an enhanced response. In these experiments intracellular killing of the added living Salmonella gave variable results, in that monocytes from two out of three pigs showed no essential change in intracellular bactericidal activity, but with cells from one pig a less pronounced bactericidal activity was found after prestimulation with PMA.     

Mitsuokella jalaludinii inhibits growth of Salmonella enterica serovar Typhimurium

Salmonella continues to be a significant human health threat, and the objective of this study was to identify microorganisms with the potential to improve porcine food-safety through their antagonism of Salmonella. Anaerobic culture supernatants of 973 bacterial isolates from the gastrointestinal tract and feces of swine were screened for their capacity to inhibit the growth of Salmonella enterica serovar Typhimurium. Growth inhibition of 1000-fold or greater was observed from 16 isolates, and 16S rRNA sequencing identified the isolates as members of the genera Mitsuokella, Escherichia/Shigella, Anaerovibrio, Selenomonas, and Streptococcus. Four isolates were identified as Mitsuokella jalaludinii, and the mechanism of Salmonella Typhimurium growth inhibition by M. jalaludinii was further investigated. M. jalaludinii stationary phase culture supernatants were observed to significantly inhibit growth, and featured the production of lactic, succinic, and acetic acids. Aerobic and anaerobic S. Typhimurium growth was restored when the pH of the culture supernatants (pH 4.6) was increased to pH 6.8. However, S. Typhimurium growth in fermentation acid-free media was the same at pH 4.6 and pH 6.8 - indicating a synergistic effect between fermentation acid production and low pH as the cause of S. Typhimurium growth inhibition. Furthermore, exposure of S. Typhimurium to M. jalaludinii culture supernatants inhibited Salmonella invasion of HEp-2 cells by 10-fold. The results identify M. jalaludinii as a possible inhibitor of Salmonella growth and invasion in swine, and thus a potential probiotic capable of improving food safety.     

Porcine neutrophil function in the presence of virulent and avirulent Salmonella choleraesuis

Porcine polymorphonuclear leukocytes (PMNLs) may be activated by bacteria to begin phagocytosis followed by oxidative and non-oxidative mechanisms of killing. The purpose of this study was to identify differences between virulent and avirulent Salmonella choleraesuis (S. choleraesuis) strains, 38 and 9 respectively, in their interactions with porcine PMNLs using five different assays. (1) Staphylococcus aureus (S. aureus) ingestion was determined by exposure of porcine PMNLs to a mixture of S. choleraesuis and 125I labeled S. aureus. There was a 2.98% and 22.20% decrease in S. aureus ingestion by mouse-avirulent S. choleraesuis 9 and mouse-virulent S. choleraesuis 38 respectively. (2) Iodination of proteins was done by exposing zymosan stimulated porcine PMNLs to S. choleraesuis in the presence of 125I and measuring its incorporation into porcine PMNL proteins. This assay indicated a 73.7% and 74.7% decrease in iodination by S. choleraesuis 9 and S. choleraesuis 38, respectively. (3) Cytochrome c reduction was performed by using porcine PMNLs, zymosan, and S. choleraesuis to determine the bacterial effect on superoxide anion production. S. choleraesuis 9 and S. choleraesuis 38 inhibited superoxide anion production by 78.0% and 92.6%, respectively. (4) Lactoferrin release from porcine PMNLs was measured by an ELISA using the supernatant from the cytochrome c assay. Results indicate a 52.0% and 61.0% increase in lactoferrin release by S. choleraesuis 9 and 38 respectively. (5) The bactericidal assay was performed by counting cfus of S. choleraesuis after preliminary incubation with porcine PMNLs, followed by killing of extracellular S. choleraesuis and lysis of porcine PMNLs. Survival of S. choleraesuis 9 and E. coli (control) were 7.50% and 1.37%, respectively, in contrast to 52.62% survival of the virulent S. choleraesuis 38. These results indicate that both strains inhibited protein iodination and caused a slight increase in lactoferrin release, but the virulent S. choleraesuis 38 inhibited S. aureus ingestion, cytochrome c reduction, and survived porcine PMNL killing more effectively than the avirulent S. choleraesuis 9.     

SPI-1-encoded type III secretion system of Salmonella enterica is required for the suppression of porcine alveolar macrophage cytokine expression

ABSTRACT: Genes localized at Salmonella pathogenicity island-1 (SPI-1) are involved in Salmonella enterica invasion of host non-professional phagocytes. Interestingly, in macrophages, SPI-1-encoded proteins, in addition to invasion, induce cell death via activation of caspase-1 which also cleaves proIL-1β and proIL-18, precursors of 2 proinflammatory cytokines. In this study we were therefore interested in whether SPI-1-encoded type III secretion system (T3SS) may influence proinflammatory response of macrophages. To test this hypothesis, we infected primary porcine alveolar macrophages with wild-type S. Typhimurium and S. Enteritidis and their isogenic SPI-1 deletion mutants. ΔSPI1 mutants of both serovars invaded approx. 5 times less efficiently than the wild-type strains and despite this, macrophages responded to the infection with ΔSPI1 mutants by increased expression of proinflammatory cytokines IL-1β, IL-8, TNFα, IL-23α and GM-CSF. Identical macrophage responses to that induced by the ΔSPI1 mutants were also observed to the infection with sipB but not the sipA mutant. The hilA mutant exhibited an intermediate phenotype between the ΔSPI1 mutant and the wild-type S. Enteritidis. Our results showed that the SPI-1-encoded T3SS is required not only for cell invasion but in macrophages also for the suppression of early proinflammatory cytokine expression.     

SYBR Green Ⅰ实时PCR对猪细小病毒体外复制动态分析

根据已经发表的猪细小病毒（PPV）的VP2基因序列,设计2对特异性引物,建立了检测PPV的SYBR GreenⅠ实时PCR方法。该方法最小检出量为12个PPV拷贝。模板稀释度在108范围内呈良好的线性关系,与猪圆环病毒2型、猪繁殖与呼吸综合征病毒、乙型脑炎病毒、猪瘟病毒、伪狂犬病病毒无交叉反应。应用本方法对PPV在体外感染细胞后的复制动态进行了观察,并绘制了病毒的体外增殖曲线。数据换算为每瓶中细胞内、外病毒拷贝数,结果显示细胞外病毒含量在接毒初始的36h逐渐下降,随后开始逐步增加;接毒后84h培养液中的病毒含量（1.739×1010拷贝）逐渐超过细胞内的病毒含量（1.321×1010拷贝）;在接毒后108h培养液中病毒含量达到峰值（7.626×1010拷贝）,随即病毒含量开始快速下降。细胞内病毒粒子在接毒后24h内为对数增长期,然后为缓慢增长期,至接种后72h达到复制峰值（1.425×1010拷贝）,并维持至108h。与病毒复制动态变化相对应的细胞病变是从细胞聚集、开始形成空斑到约80%的细胞病变产生,108h之后随着细胞的大量死亡,细胞内、外病毒数量都开始急剧减少。     

Cytokine response of porcine cell lines to Salmonella enterica serovar typhimurium and its hilA and ssrA mutants

Salmonella enterica serovar Typhimurium (S. Typhimurium) is a facultative intracellular bacterium which can infect and colonize pigs. After contact with enterocytes and macrophages, S. Typhimurium induces production of cytokines thus triggering the innate immune response. In this study we evaluated the cytokine response of two porcine cell lines, IPI-2I and 3D4/31, of epithelial or macrophage origins, respectively, to the wild-type S. Typhimurium and its hilA and ssrA mutants. We observed that the 3D4/31 cell line essentially did not respond to S. Typhimurium infection when a medium with foetal calf serum was used. However when the 3D4 cell line was incubated overnight in the presence of porcine serum, it efficiently responded to the wild-type strain and the ssrA mutant but not to the noninvasive hilA mutant as measured by mRNA quantification of TNF-alpha, IL-8 and GM-CSF by the real-time RT-PCR. In IPI-2I, all the cytokines were also induced by the wild-type S. Typhimurium and the ssrA mutant although the induction of TNF-alpha was lower than that induced by the wild-type strain. The hilA mutant was unable to induce any of the cytokines tested. The ssrA mutant can therefore be considered as more suitable for further vaccine development as the stimulation of innate immune response is important for animal protection against a challenge with virulent strains.     

Characterization of a Salmonella choleraesuis isolate after repeated neutrophil exposure.

Salmonella choleraesuis strain 38 (glycerol-positive fermentation) was repeatedly exposed to porcine neutrophils in an attempt to mimic in vivo conditions of the host immune system. After phagocytosis, viable intracellular S choleraesuis were isolated and the process was repeated at least 5 times. A fifth-passage strain-38 neutrophil-adapted clone, 38PMNa-5X, was isolated, and was compared with the parent wild-type strain 38 for changes. Strain 38PMNa-5X had increased resistance to killing by hydrogen peroxide and phagocyte killing by porcine neutrophils, as measured by 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide reduction. Strain 38PMNa-5X was less invasive than the parent strain on Vero cell monolayers, and had been cured of a 50-kb plasmid. The 50-kb plasmid was marked with bacteriophage mini-Mu (kanamycin resistant) and was reinserted into strain 38PMNa-5X. Strain 38PMNa-5X was avirulent in mice, but the isolates with reinserted plasmids had intermediate resistance to neutrophil and hydrogen peroxide killing and had restored invasiveness and mouse virulence. Differences in complement sensitivity and enzymatic activity were not observed between the strains.     

Fragment Bb of bovine complement factor B: stimulatory effect on the microbicidal activity of bovine monocytes.

We have previously shown that the Bb fragment of bovine complement factor B activates bovine monocytes and neutrophils. The activation was demonstrated by the enhanced uptake of 3H-deoxyglucose. To investigate the potential effect of fragment Bb on the microbicidal activity of bovine monocytes, a direct method was used. This method involves an initial ingestion period at 37 degrees C followed by repeated washing. The decrease in the total number of viable intracellular Staphylococcus aureus during the reincubation of the bacteria with bovine monocytes determines the intracellular killing. Maximal intracellular killing was seen when the monocytes containing the ingested S. aureus was incubated with fresh bovine serum (mean +/- SEM = 73.4 +/- 1.4%). On incubation of the monocytes, containing the ingested bacteria with heat-inactivated bovine serum, 32.5 +/- 0.7% of the intracellular bacteria were killed. When affinity-purified bovine factor Bb was added to the heat-inactivated serum, the intracellular killing capacity was almost restored (65.8 +/- 1.5%). When monocytes were incubated with medium alone, they killed 22.4% of the intracellular microorganisms. When fragment Bb (25 micrograms/mL) was added to the medium, the intracellular killing of S. aureus doubled (46 +/- 1.29%). We conclude that the Bb fragment of bovine complement factor B stimulates bovine monocytes in their microbicidal activity.     

Differential regulation of porcine beta-defensins 1 and 2 upon Salmonella infection in the intestinal epithelial cell line IPI-2I

Intestinal epithelial cells represent the first line of defence against pathogenic bacteria in the lumen of the gut. Besides acting as a physical barrier, epithelial cells orchestrate the immune response through the production of several innate immune mediator molecules including beta-defensins. Here, we establish the porcine intestinal cell line IPI-2I as a new model system to test the regulation of porcine beta-defensins 1 and 2. Gene expression of both defensins was highly upregulated by foetal calf serum components in normal growth medium. In serum-free medium, baseline expression remained low, but pBD-2 gene expression was increased 10-fold upon infection with Salmonella Typhimurium. Arcobacter cryaerophilus and Salmonella Enteritidis, pathogenic bacteria with comparable adhesion and invasion characteristics, failed to increase pBD-2 mRNA levels. Heat killed or colistin-treated Salmonella Typhimurium had no effect, showing that the upregulation of pBD-2 was dependent on the viability of the Salmonella Typhimurium. Gene expression of pBD-1 was regulated differently since an increase in pBD-1 mRNA was observed by Salmonella Enteritidis infection. We conclude that the IPI-2I cells can serve as a new model to study porcine beta-defensin regulation and that pBD-1 and pBD-2 are differentially regulated in this cell line.     

Comparison of intestinal invasion and macrophage response of Salmonella Gallinarum and other host-adapted Salmonella enterica serovars in the avian host

The purpose of this investigation was to study the host specific infection of Salmonella Gallinarum in chickens and to determine the contribution of intestinal invasion and macrophage survival in relation to systemic infection in the host. This was carried out by comparing the kinetics of infection of S. Gallinarum to that of other Salmonella host-adapted (S. Cholerae-suis, S. Dublin and S. Typhimurium) and host-specific (S. Pullorum and S. Abortus-ovis) serovars. Establishment of the rate of colonisation in intestinal tissue, bursa and systemic sites was carried out by oral infection in day-old and week-old birds. Salmonella Gallinarum was the only serovar capable of causing systemic infection in chickens, however, general colonising ability in the intestine and bursa demonstrated no apparent selective advantage for S. Gallinarum. Further quantification of gastrointestinal invasion was carried out using ligated loops in the small intestine. Invasion in the jejunum of the chicken intestine over 3h demonstrated that Salmonella Typhimurium invasion was statistically higher (P<0.01) when compared with S. Gallinarum. Specific sites of high lymphoid tissue concentration in the chicken, including the bursa of Fabricius and caecal tonsils, were also targeted in invasion assays to investigate possible areas of tissue tropism. S. Typhimurium demonstrated significantly higher (P<0.01) invasion at these sites when compared with S. Gallinarum. Infection of chicken macrophages with S. Gallinarum did not demonstrate increased multiplication and survival intracellularly when compared with other Salmonella serotypes. The only difference seen was with S. Abortus-ovis, which demonstrated a significantly lower (P<0.05 to 0.001) intracellular survival. Together these data suggest that although S. Gallinarum host specificity in the chicken correlates with systemic infection, intestinal and lymphoid tissue invasion in the bursa and caeca, and macrophage survival does not influence this outcome.     

Salmonella (sero)types and their resistance patterns in pig faecal and post-mortem samples.

The purpose of this survey was to take stock of porcine Salmonella isolates derived from faecal and post-mortem samples over a 4-year period. Salmonella was isolated by direct inoculation on BGA(NO)-plates (faeces, intestinal content) or sheep blood agar (organs). Antimicrobial susceptibility was tested by the agar diffusion method. Salmonella was isolated in 4.2% of all porcine submissions received at the Animal Health Service. A total of 1305 salmonellae were isolated from a total of 1279 submissions from 1008 different herds. Salmonella Typhimurium was the most frequently isolated serotype (88%), and Salmonella Typhimurium DT104 was the most frequently isolated phagetype (17.2% of Salmonella Typhimurium). Resistance to antimicrobials occurred in 47.3% of all isolates, mainly those of the multiresistant phagetype DT104. Other pathogens were isolated in more than 50% of the submissions. In cases of clinical diarrhoea, multiple pathogens may be involved and therapy and preventive measures should be adjusted accordingly.