IL-13 replaces IL-4 in development of monocyte derived dendritic cells (MoDC) of swine

Dendritic cells (DCs) are a critical aspect of innate immune responses in addition to initiating adaptive immunity. In vitro generation of monocyte derived dendritic cells (MoDC) by culturing cells in IL-4 and granulocyte/macrophage colony stimulating factor (GM-CSF) has been reported for multiple species including swine. However, IL-4 is not a prominent cytokine detected in the periphery of common breeds of swine such as Yorkshire pigs. In this study, we report the generation and characterization of porcine MoDC in vitro using porcine IL-13 and porcine GM-CSF. These cells have the predicted expression of Class II MHC and T cell costimulatory molecules, phagocytic capacity and the ability to process and present antigen. Critically, porcine IL-13/GM-CSF MoDC have the unique ability to stimulate a primary mixed lymphocyte response in vitro. The type I interferon response of these MoDC to poly I:C (TLR3 ligand), LPS (TLR4 ligand) and CpG (TLR9 ligand) was tested. Of these TLR agonists, LPS or CpG did not stimulate induction of type I interferons, but a strong response was observed to poly I:C. This analysis shows that the generation of MoDCs in IL-13 yields cells of equivalent phenotype and function as IL-4 generated DC. However, for swine, in vitro generation of MoDC in IL-13 is likely to induce a more physiological cell population to study given expression of IL-4 is lacking in the periphery of these animals.     

The role of dendritic cells in shaping the immune response

Dendritic cells are central to the initiation of primary immune responses. They are the only antigen-presenting cell capable of stimulating naive T cells, and hence they are pivotal in the generation of adaptive immunity. Dendritic cells also interact with and influence the response of cells of the innate immune system. The manner in which dendritic cells influence the responses in cells of both the innate and adaptive immune systems has consequences for the bias of the adaptive response that mediates immunity to infection after vaccination or infection. It also provides an opportunity to intervene and to influence the response, allowing ways of developing appropriate vaccination strategies. Mouse and human studies have identified myeloid, lymphoid and plasmacytoid dendritic cells. Studies in domesticated animals with agents of specific infectious diseases have confirmed the applicability of certain of the generic models developed from mice or from in vitro studies on human cells. In vivo and ex vivo studies in cattle have demonstrated the existence of a number of subpopulations of myeloid dendritic cells. These cells differ in their ability to stimulate T cells and in the cytokines that they produce, observations clearly having important implications for the bias of the T-cell response. Dendritic cells also interact with the innate immune system, inducing responses that potentially bias the subsequent adaptive response.     

Resilience of second year grazing cattle to parasitic gastroenteritis following negligible to moderate exposure to gastrointestinal nematode infections in their first year

The influence of gastrointestinal nematode infections on performance of four groups of female Holstein Friesian calves was monitored until the end of the second grazing season (SGS). In the first year three groups were grazed and one group (G4) was permanently housed. General and grazing management during the first grazing season (FGS) was arranged such that G1 acquired moderate infections, G2 low infections and G3 very low infections with gastrointestinal nematodes. These infections were monitored through faecal egg counts, differentiation of faecal larval cultures, pasture larval counts, serum pepsinogen values, ELISA with a recombinant Cooperia oncophora protein, weight gain, tracer worm counts and sentinel worm counts. In 1998 all four groups were grazed together as one herd from 23 April to 26 October and infections were monitored with the same techniques with the exception of sentinel calves.In the FGS weight gain was higher in G4 than in the other groups and higher in G3 (28. 6kg) than in G1. Weight gain of G2 was intermediate to G1 and G3 but did not significantly differ from either group. In the SGS weight gain in G4 was far less than in any other group and the mean weight at the end of the experiment was 41.9, 38.6 and 50.9kg lower than G3, G1 and G2, respectively. Though no significant differences were observed between G1, G2 and G3 at the end of the experiment it was obvious that the weight gain advantage of G3 over G1 at the end of the FGS had disappeared.Parasitological and serological findings in the SGS indicated that G3 and G4 had build up less immunity during the FGS compared to G1 and G2.The conclusion of the experiment is that resilience to parasitic gastroenteritis in the SGS depends on the level of exposure to nematodes in the FGS. However, problems with poor weight gain only will be expected when exposure is very low in the FGS and high in the SGS.     

The determination at housing of exposure to gastrointestinal nematode infections in first-grazing season calves

Various parameter estimates were assessed at housing in calves that had been exposed to gastrointestinal nematodes during a first grazing season. The analysis involved 41 groups of first grazing season (FGS) calves on 15 different farms in Belgium and comprised groups that had received chemoprophylactic treatment and untreated controls. Serum pepsinogen levels gave the clearest division between chemoprophylactic-treated calf groups (all were <2.6 U tyr), and untreated calf groups in which sub-clinical (range: 2.0–4.1 U tyr) and clinical infections (range 3.7–6.3 U tyr) occurred. There was also a tight relationship between individual pepsinogen values and adult Ostertagia burdens obtained at slaughter. In chemoprophylactic-treated groups there was a significant negative relationship between mean serum pepsinogen levels at housing and the proportion of the grazing season covered by different chemoprophylactic systems. Although only limited data on crude adult Ostertagia antigen ELISA were available, a good relationship between optical densities and estimated exposure was also found. The parasitological parameters, faecal egg counts and pasture Ostertagia larval counts at housing, and weight gain per day, gave less clear divisions among the three categories (chemoprophylaxis, sub-clinical and clinical). Distinguishing how much exposure a calf group has experienced during a first grazing season could help in designing more appropriate control measures for the FGS calves in the next year, assuring good protection and at the same time allowing sufficient exposure for the development of acquired immunity to Ostertagia, and for this serum pepsinogen is recommended.     

The effect of continuous drug exposure on the immune response of lambs challenged with drug-susceptible or drug-resistant nematode larvae

Groups of lambs either with or without controlled-release albendazole (ABZ) capsules (CRCs) were challenged twice weekly for 6 weeks with either drug-susceptible or drug-resistant Ostertagia circumcincta and Trichostrongylus colubriformis. Groups with and without CRCs remained unchallenged as controls. There was minimal establishment of drug-susceptible parasites of either species in those lambs with CRCs. However, drug-resistant parasites of both species established adult worm burdens in the presence of the capsules. The humoral immune response, as measured by the serum anti-larval (L₃) antibody (Ab) titre, was pooled for weeks 4–6 and compared for each group. With the exception of anti-T. colubriformis Ab in group 2, anti-L₃ Ab titres were significantly higher in all the parasite-challenged groups as compared to the control animals. Also, with the exception of anti-resistant O. circumcincta Ab levels in the CRC-treated animals (group 5), no significant difference was observed in Ab titres between the four groups challenged with either resistant or susceptible larvae. The results demonstrate the inability of CRCs to prevent establishment of drug-resistant parasites and that immune stimulation is not inhibited by the capsules.     

Phenotype and function of murine discrete Peyer's patch macrophage derived - dendritic cells

The phenotype and function of peritoneal cavity macrophage-derived dendritic cells (PEC-DC) was previously reported. In this study we have gone further in using our established culture system to generated discrete Peyer's patch dendritic cells (DPP-DC) from murine discrete Peyer's patch macrophages (DPP-M?), following stimulation with granulocyte macrophage colony stimulating factor (GM-CSF) plus interleukin 4 (IL-4) for 7 days. DPP-M? from murine small intestines were obtained by mechanical disruption of discrete Peyer's patches (DPP), followed by metrizamide density gradient centrifugation to remove Peyer's patch resident DC and debri, after which an overnight adherent step in tissue culture medium was carried out for macrophage enrichment. Characterization of the generated DPP-DC was carried out using well-established criteria of morphology, expression of membrane antigens and capacity for antigen presentation. Dendritic cells expressed DEC-205, F4/80 and CD34 at high levels, but exhibited very low CD11c levels. They were shown to present soluble protein antigen to CD3(+) spleen T cells. A comparison of the surface antigen expression in the progenitor DPP-M? population and the generated DPP-DC showed a significant decrease in MHC class II levels and a marked down regulation of the co-stimulatory molecule CD86 (B7-2). High expression of the haemopoietic progenitor marker CD34 indicates that the generated DC, possess a haemopoietic rather than myeloid origin. Taken together, these results may provide a better understanding of the complex network regulating mucosal immune responses.     

Duration of protective immunity against ovine haemonchosis following vaccination with the nematode gut membrane antigen H11

To establish for how long protective antibody levels may be maintained, lambs were vaccinated with the gut membrane antigen H11 and challenged with Haemonchus contortus 14, 84, 126 or 168 days later. Compared to controls, mean faecal egg counts of vaccinated lambs were reduced by 97 per cent, 99 per cent, 92 per cent and 86 per cent respectively. Total worm burdens at postmortem five weeks after infection were reduced by 87 per cent, 94 per cent, 92 per cent and 62 per cent respectively. In vaccinated lambs, antibody levels to H11 peaked at about 60 days after the first vaccination and were maintained for the duration of the experiment. There was evidence of secondary antibody responses to H11 following challenge.     

Immunophenotype and functional properties of feline dendritic cells derived from blood and bone marrow

Dendritic cells (DCs) are a heterogeneous population of cells of fundamental importance in initiating innate as well as specific immune responses. The identity and function of DCs in the cat are unknown, although they are likely pivotal in the response to infection. In this study, feline DCs were derived by 3-10-day culture of adherent blood mononuclear cells (PBMCs) and bone marrow mononuclear cells (BMMCs) in the presence of IL 4 and GM-CSF. BMMC consistently yielded a greater number of DCs than PBMC, and there were fewer macrophages than DC from both compartments. DCs expressed a distinct constellation of surface molecules, which included CD1a, CD1b, and CD1c, CD11b, CD14, and 2-3-fold higher levels of MHC class I and II molecules than co-cultured macrophages or fresh blood monocytes. DCs displayed typical cytoplasmic processes, limited non-specific esterase activity, and acquired antigen by phagocytosis, pinocytosis, and binding to specific receptors. Cytokine-exposed cells induced proliferation of allogeneic lymphocytes. Thus, the cells derived by these culture conditions had markers and functions analogous to immature myeloid DCs. Availability of feline DCs will enable investigation of their role in infectious disease and their potential therapeutic application.     

Expression and function of Toll-like receptors on dendritic cells and other antigen presenting cells from non-human primates

Antigen presenting cells (APCs), especially dendritic cells (DCs), play a crucial role in immune responses against infections by sensing microbial invasion through Toll-like receptors (TLRs). In this regard, TLR ligands are attractive candidates for use in humans and animal models as vaccine adjuvants. So far, no studies have been performed on TLR expression in non-human primates such as rhesus macaques. Therefore, we studied the TLR expression patterns in different subsets of APC in rhesus macaques and compared them to similar APC subsets in human. Also, expression was compared with corresponding DC subsets from different organs from mice. Here we show by semi-quantitative RT-PCR, that blood DC subsets of rhesus macaque expressed the same sets of TLRs as those of human but substantially differed from mouse DC subsets. Macaque myeloid DCs (MDCs) expressed TLR3, 4, 7 and 8 whereas macaque plasmacytoid DCs (PDCs) expressed only TLR7 and 9. Additionally, TLR expression patterns in macaque monocyte-derived dendritic cells (mo-DCs) (i.e., TLR3, 4, 8 and 9), monocytes (i.e., TLR4, 7, and 8) and B cells (i.e., TLR4, 7, 8, and 9) were also similar to their human counterparts. However, the responsiveness of macaque APCs to certain TLR ligands partially differed from that of human in terms of phenotype differentiation and cytokine production. Strikingly, in contrast to human mo-DCs, no IL-12p70 production was observed when macaque mo-DCs were stimulated with TLR ligands. In addition, CD40 and CD86 phenotypic responses to TLR8 ligand (poly U) in mo-DCs of macaque were higher than that of human. Despite these functional differences, our results provide important information for a rational design of animal models in evaluating TLR ligands as adjuvant in vivo.     

Exsheathment of Ostertagia ostertagi infective larvae following exposure to bovine rumen contents derived from low and high roughage diets

The objective of this study was to characterize the exsheathment kinetics of Ostertagia ostertagi infective larvae (L3) following in vivo exposure to bovine rumen contents derived from low and high roughage diets. O. ostertagi L3 were placed in disposable dialysis bags and incubated for various time points between 0 and 360 min in the rumen of a fistulated steer maintained on a 71% grain diet or a 100% grass diet. The maximum percentage of exsheathed L3 was observed 120 min post-exposure to grass-derived rumen contents, while maximum exsheathment for L3 exposed to grain-derived rumen contents did not occur until 360 min. This work provides the first report of the in vivo exsheathment kinetics for O. ostertagi in its bovine host. Results of this study also support earlier reports that rumen pH may affect the exsheathment efficiency of abomasal trichostrongylids.     

Characterization of porcine dendritic cell response to Streptococcus suis

ABSTRACT: Streptococcus suis is a major swine pathogen and important zoonotic agent causing mainly septicemia and meningitis. However, the mechanisms involved in host innate and adaptive immune responses toward S. suis as well as the mechanisms used by S. suis to subvert these responses are unknown. Here, and for the first time, the ability of S. suis to interact with bone marrow-derived swine dendritic cells (DCs) was evaluated. In addition, the role of S. suis capsular polysaccharide in modulation of DC functions was also assessed. Well encapsulated S. suis was relatively resistant to phagocytosis, but it increased the relative expression of Toll-like receptors 2 and 6 and triggered the release of several cytokines by DCs, including IL-1β, IL-6, IL-8, IL-12p40 and TNF-α. The capsular polysaccharide was shown to interfere with DC phagocytosis; however, once internalized, S. suis was readily destroyed by DCs independently of the presence of the capsular polysaccharide. Cell wall components were mainly responsible for DC activation, since the capsular polysaccharide-negative mutant induced higher cytokine levels than the wild-type strain. The capsular polysaccharide also interfered with the expression of the co-stimulatory molecules CD80/86 and MHC-II on DCs. To conclude, our results show for the first time that S. suis interacts with swine origin DCs and suggest that these cells might play a role in the development of host innate and adaptive immunity during an infection with S. suis serotype 2.     

Cellular immune response of lymph nodes from dogs following the intradermal injection of a recombinant antigen corresponding to a 66 kDa protein of Echinococcus granulosus

A recombinant polypeptide (referred to as EgA31), which represents a 66kDa protein, was prepared from an Echinococcus granulosus cDNA library. In order to assess its potential to induce cellular immune responses, dog popliteal and prescapular lymph nodes were sensitized with this recombinant polypeptide. Subpopulations of lymphocytes were then analyzed by flow cytometry and immunohistochemistry on lymph node sections. Five days after the sensitization, the paracortical areas of the lymph nodes appeared hypertrophic, the number of CD3+, CD4+, CD8+ and CD5+ cells increased, the number of B-cells began to augment and some secondary follicles occurred, and a number of CD4+ cells appeared in germinal centers. Many large secondary follicles and a significantly augmented number of CD5+ cells in cords of medullae were observed 10 days after the sensitization. These active cellular responses strengthen the interest for further studies on the development of a vaccine with this recombinant polypeptide.     

Rapid infection in market-weight swine following exposure to a Salmonella typhimurium-contaminated environment

OBJECTIVE: To evaluate the possibility of swine becoming infected with Salmonella Typhimurium when housed for 2 to 6 hours in an environment contaminated with Salmonella, similar to a lairage situation prior to slaughter. ANIMALS: 40 crossbred market pigs with an approximate body weight of 92 kg. PROCEDURE: Five trials were conducted (8 pigs/trial) in simulated lairage conditions. Superficial inguinal, ileocecal, and mandibular lymph nodes, cecal contents, distal portion of the ileum, and fecal samples were obtained from each pig after 2 (n = 10), 3 (10), and 6 (5) hours of exposure to an environment contaminated with feces defecated by 10 pigs intranasally inoculated with nalidixic acid-resistant Salmonella Typhimurium (chi4232). In addition, 5 control pigs that were not exposed were also evaluated in the same manner. RESULTS: Feces deposited on the floor by intranasally inoculated swine were mixed with water to form slurry with a resulting load of approximately 10(3) colony-forming units of Salmonella Typhimurium/g of material. Eight of 10, 6 of 10, and 6 of 6 pigs exposed to the slurry for 2, 3, or 6 hours, respectively, had positive results for at least 1 sample when tested for the specific strain of Salmonella Typhimurium. CONCLUSIONS AND CLINICAL RELEVANCE: Pigs can become infected during routine resting or holding periods during marketing when exposed to relatively low amounts of Salmonella organisms in the preslaughter environment. Intervention at this step of the production process may have a major impact on the safety of pork products.     

The requirement for early exposure of Haemonchus contortus larvae to Bacillus thuringiensis for effective inhibition of larval development

The potential for a nematocidal Bacillus thuringiensis (Bt) to target the free-living larval stages of Haemonchus contortus was examined using in vitro larval development and migration assays. Bt toxicity in larval development assays decreased as the time period between egg hatch and initial exposure to the Bt was increased; a time lag of 48 h resulted in a 350-fold increase in the IC(50) (from 2.6 ng/ml to 910 ng/ml). The effects on larval migration largely paralleled the effects on larval development, indicating that the larvae which reached the infective stage after exposure to Bt were generally as 'fit' as control worms in terms of migration ability. However, a comparison of the two assays also showed the presence of a level of Bt exposure which showed significantly more toxicity in migration assays than development assays, indicating that, in some cases, fully developed Bt-exposed larvae were less able to migrate than controls, and hence may be compromised in their ability to infect sheep. The rapid decrease in toxicity when exposure to the Bt is delayed highlights a significant issue concerning the use of Bt for control of the free-living larval stages of animal-parasitic nematodes. Targeting the larvae by delivering bacterial spores to the faeces through the host animal's digestive tract would require the spores to germinate upon defecation, grow through a vegetative phase, to then produce crystal toxin protein upon subsequent sporulation. This period of bacterial development will introduce a time lag between worm egg hatching and initial exposure of the larvae to the Bt, which, as demonstrated in the present study, may allow the worm larvae to develop to late larval stages which are relatively insensitive to the toxin.     

藏北高寒草甸土壤线虫群落结构对增温的响应

为揭示增温对高寒草甸土壤线虫群落的影响,利用OTC模拟短期和长期增温对藏北高寒草甸土壤线虫群落进行比较研究。结果表明,短期和长期增温改变了土壤线虫的群落组成,增加了双垫刃属(Ditylenchus)和丽突属(Acrobeles)丰度。长期增温导致食真菌类线虫丰度显著增加,但各处理间食细菌类线虫、植物寄生类线虫、杂食/捕食类线虫丰度以及cp1-5类群的丰度和属数量无显著差异(P0.05)。短期和长期增温均降低了土壤线虫的多样性和均匀度,其中2015年短期增温处理显著降低了其多样性。2015年和2016年短期和长期增温土壤显著降低了线虫数量,较对照分别降低了34.45%、32.09%和25.34%和22.66%。各处理样地间MI(Maturity index)、NCR(Nematode channel ratio)、PPI(Plant parasite index)和WI(Wasilewska index)指数无显著差异(P0.05),且均表现出WI1,NCR0.5,表明增温对高寒草甸的健康状态影响不大,土壤有机质矿化途径主要由食细菌和真菌线虫参与。环境因素与土壤线虫数量冗余分析表明,植物总盖度、莎草科盖度、土壤温湿度、细菌和真菌数量对土壤线虫数量影响达到显著水平(P0.05),增温通过改变植物、土壤理化性质和微生物数量等环境因子而影响高寒草甸土壤线虫群落组成。     

The response of broilers' adiposity to testosterone after embryonic exposure to androgen and tamoxifen

1. The effects of early exposure of heavy breed (HB) chicks to an anti-oestrogen (tamoxifen--TAM) and to an androgen which cannot be aromatised (5 alpha-dihydrotestosterone--DHT) on subsequent adiposity and its response to testosterone were studied. 2. Embryonic TAM administration reduced adiposity in females but not in males at 8 weeks of age. Embryonic DHT produced similar responses but to a lesser extent. 3. Testosterone propionate (TP) administration during growth had no effect on adiposity in any of the treated groups but TP reduced adiposity in males which had been exposed to DHT at the embryonic stage.     

圆叶决明对镉胁迫的生理响应

圆叶决明是我国20世纪80年代从澳大利亚引进的豆科牧草,适合亚热带地区广泛种植。为探明圆叶决明的耐镉性,本文采用添加外源有效态镉的土壤盆栽试验方法,研究不同浓度(0,1,2,4,8 mg/kg)镉胁迫对圆叶决明CIP86134生长发育及其生理响应机制。结果表明,镉胁迫抑制圆叶决明的生长发育,降低其生物产量,且随镉浓度的升高,抑制作用加强。当镉处理浓度为4 mg/kg时,地下部、地上部生物产量与对照比显著下降,分别为对照的59.69%和63.77%。镉胁迫抑制圆叶决明叶绿素a、叶绿素b和类胡萝卜素的合成,镉浓度升高,叶绿素含量逐渐降低,处理之间达显著差异水平。镉处理浓度为12 mg/kg时,SOD、POD、CAT 3类保护酶协调一致,圆叶决明生长正常,地下部、地上部生物产量与对照比差异不显著;镉浓度升高至4 mg/kg,SOD酶活性降低,MDA含量升高,POD和CAT酶被诱导激活,缓解了镉胁迫对圆叶决明的伤害,生物产量虽显著下降,但未中毒死亡;镉浓度升高至8 mg/kg时,MDA含量显著增加,细胞保护酶系统失调,圆叶决明至收获时全部干枯死亡。圆叶决明吸收的镉主要积累在地下部,迁移系数仅为0.1520.234。在供试土壤添加外源有效态镉的条件下,圆叶决明能忍耐2 mg/kg以下的镉胁迫,可作为南方红壤地区矿山镉污染土壤的生态修复植物类型。