The Amazing Immune System in the Neonatal Pig 新生猪惊人的免疫系统 Tom Molitor 汤姆莫利特博士 University of Minnesota 明尼苏达大学 Leman China, October 13-

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1 The Amazing Immune System in the Neonatal Pig 新生猪惊人的免疫系统 Tom Molitor 汤姆莫利特博士 University of Minnesota 明尼苏达大学 Leman China, October 13-15, 2013, Xi an

2 Amazing Immune system in the neonatal pig 新生猪惊人的免疫系统 : Outline 大纲 Differences in Immune system in the pig. 猪免疫系统的差异 Developmental Immunity in the pig 猪免疫功能的发育 Vaccination of the Neonatal Animal 新生动物的疫苗接种 Passive Immunity 被动免疫 : Pros and cons 优点和缺点 Cross-fostering affects immunity 交叉寄养影响免疫力 Maternal Interference? 母抗干扰? Summary 总结 -Applications 应用

3 Rationale 思考 / 对比 / 分享 PEDV, PRRSV, FMDV, PPV, Mycoplasma, Protection, recovery, dependent on the immune system. PEDV, PRRSV, FMDV, PPV, 支原体, 保护, 恢复, 对免疫系统的依赖 Maternal interference, ineffective vaccination, Immunosuppression, stress, disease, all resulting from an ineffective immune response. 母抗干扰, 无效疫苗接种, 免疫抑制, 应激, 疾病, 都源于无效的免疫反应 The Immune system is highly important for the health and welfare of swine. 免疫系统对猪的健康和福利来说是非常重要的 Swine have a number of profound differences in the Immune system including anatomical, physiological, cellular and from a functional response. 猪有一些免疫系统的深刻差异, 包括解剖, 生理, 细胞和功能反应 Your task is. 你的任务是...

4 Think/Pair/Share 思考 / 对比 / 分享 Provide at least three differences in the immune system of a pig compared to human (or the classic animal model-mice)? 相比于人类 ( 或经典的动物模型 - 老鼠 ), 提供至少三个猪免疫系统的不同点

5 Differences in Immune system 免疫系统的差异?

6 Development of Immunity 免疫力的发育 Fetus 胎儿 Neonate 新生儿 Weaned pig 断奶猪

7 Why does the piglet need maternal Immunity? 仔猪为什么需要母体的免疫力? Epitheliochorial placenta 上皮绒毛膜胎盘 The multi-layered placenta in pigs prevents the transit of maternal Ig into the fetus 多层猪胎盘防止母体 lg 运输到胎儿 (Butler, 1998; Salmon, 1999). The placenta is a sterile environment 胎盘是一个无菌的环境 Sterzl et al 1966, Porter 1988 Neonatal Piglet immunologically naïve 新生仔猪是无免疫的 Immune system develops ~70 d of 116 total of gestation for the pig, third trimester for all 猪免疫系在总共夫人 116 天妊娠统发育 70 天 New born is Immature, not incompetent 新生猪不成熟, 不是无能 Adkins et al 2005

8 Relationship of Placental Type to Maternal Immunity Transfer 胎盘类型与母体免疫传递的关系胎盘类型子宫内免疫传递初乳种类上皮绒毛膜内皮绒毛马, 牛, 猪, 羊狗, 猫绒毛膜受血灵长类动物, 啮齿类动物 How is immunity transferred?? 细胞介导的免疫转移?

Colostrum 初乳 The perfect Food-colostrum 完美的食物 - 初乳!

Clark in Colostrum life s first food: Bovine Colostrum rebuilds the immune

tissue, helps loose weight, burn fat, increase bone and lean muscle mass and

9 Colostrum 初乳 The perfect Food-colostrum 完美的食物 - 初乳!! Elixir of life 仙丹 ImmunePlus with colostrum 7505 初乳 7505 ImmunePlus Daniel Clark in Colostrum life s first food: Bovine Colostrum rebuilds the immune system, destroys viruses and bacteria and fungi, accelerates healing of the body tissue, helps loose weight, burn fat, increase bone and lean muscle mass and slows down and even reverses aging 丹尼尔克拉克在初乳生命的第一食物中提到 : 初乳重建免疫系统, 破坏病毒和细菌和真菌, 加速身体组织的愈合, 有助于减轻体重, 燃烧脂肪, 增加骨骼和肌肉质量, 减慢甚至逆转老化

10 Absorption of colostrum components is critical for piglet survival 吸收初乳成分是仔猪存活的关键 Epitheliochorial placenta 上皮绒毛膜胎盘 The piglet is immunologically naïve and immature 仔猪无免疫能力并且不成熟

11 Need for colostrum 需要初乳 : Colostrum deprivation and survival to 90 kg 无初乳和生存至 90kg 的关系出生断奶生存 24 小时断奶

12 Colostrum components 初乳成分免疫球蛋白溶菌酶乳过氧化物酶核苷酸水解抗体 K- 酪蛋白 α- 乳球蛋白结合咕啉粘蛋白奶中的乳铁蛋白抗分泌外源凝集素自由分泌成分低聚糖脂肪酸母体白细胞和细胞因子 scd14 补体及补体受体 β- 防御素 1 Toll 样受体双歧因子益生元 Colostrum 初乳 = first milk 首次乳汁 Hosea Blewett et al., 2008 Wagstrom et al., 2000 Salmon et al, 2008 Le Jan, 2004

13 Immunoglobin in sow colostrum 母猪初乳中的免疫球蛋白相对浓度 Straw et al., 1999 泌乳日哺乳期母猪乳浆中 IgG, IgM 及 IgA 浓度泌乳阶段 Klobasa et al., 1987

Approach 途径 Sow Periphery 母猪外周 Mammary Gland (MG) 乳腺 Piglet 仔猪 Samples 样品 üsow Blood 母猪血液 üsow Colostrum 母猪初乳 üpiglet blood pre colostrum ingestion 摄入初乳前仔猪血 üpiglet blood after colostrum

14 Approach 途径 Sow Periphery 母猪外周 Mammary Gland (MG) 乳腺 Piglet 仔猪 Samples 样品 üsow Blood 母猪血液 üsow Colostrum 母猪初乳 üpiglet blood pre colostrum ingestion 摄入初乳前仔猪血 üpiglet blood after colostrum ingestion 摄入初乳后仔猪血 Endpoints 终点 ütotal IgG and IgA via ELISA 通过 ELISA 总的 IgG 和 IgA ütotal mononuclear cells 单核细胞总数 ücd4, CD8, T lymphocyte phenotyping via flow cytometry 通过流式细胞仪 CD4, CD8, T 淋巴细胞表型

15 Maternal derived immunity 母源免疫 : In the sow vs the neonatal pig? 在母猪 vs 新生猪? Bandrick et al 2010 Immunoglobulin G and A in sow serum and colostrum 母猪血清和初乳中的免疫球蛋白 G 和 A 免疫球蛋白 Immunoglobulin (mg/ml) IgG IgA n=16 Variation: SEM 0 母猪血清 Sow serum Colostrum 初乳

Maternal immunoglobins are beneficial to the newborn pig 母体免疫球蛋白有利于仔猪 Maternally-derived antibody-mediated immunity (AMI) is protective 母体介导的免疫是有保护性的 PCV-2 猪圆环病毒 -2; Opriessnig et al.

16 Maternal immunoglobins are beneficial to the newborn pig 母体免疫球蛋白有利于仔猪 Maternally-derived antibody-mediated immunity (AMI) is protective 母体介导的免疫是有保护性的 PCV-2 猪圆环病毒 -2; Opriessnig et al., 2008 SIV 猪流感病毒 ; Loeffen et al., 2003 B. pertussis 百日咳博德特氏菌 ; Elahi et al., 2006 CSF 猪乙脑病毒 ; Parchariyanon et al., 1994 TGEV 猪传染性胃肠炎病毒 ; Sestek et al., 1996 Pseudorabies 伪狂犬病 ; Kritas et al., 1997 Janeway, 2005

17 Relative immunoglobin concentration in piglet serum pre and post colostrum ingestion 仔猪摄入初乳前后血清免疫球蛋白相对浓度 Ig 浓度 Ig concentration mg/ml IgG PS IgG AS IgA PS IgA PS PS= Pre Suckle 哺乳前 ; AS= After suckle 哺乳后 Neither IgG nor IgA were detectable by ELISA in PS serum 哺乳前血清用 ELISA 检测不到 IgG 或 IgA Bandrick et al 2010

18 泌乳日 Cells in Sow Colostrum 母猪初乳中的细胞 PMN Macrophage lymphocyte epithelial cell 中性粒细胞巨噬细胞淋巴细胞中性粒细胞巨噬细胞淋巴细胞上皮细胞上皮细胞 70-90% are T cells 70-90% 是 T 细胞 > 1x10 7 cells/ml Total number of cells/m 细胞总数 /ml Wagstrom et al., 2000

19 T lymphocyte distribution in sow blood and colostrum T 淋巴细胞在母猪血液和初乳中的分布 A Event counts CD4 CD8 Neg Blood Neg Blood Neg Colostrum Colostrum Colostrum Blood Black 黑色 : colostrum 初乳 Dark Gray 深灰色 : Blood 血 Light Gray 浅灰色 : negative control 阴性对照 Variation 变异 : SEM 均值标准误 B % mononuclear cells 单核细胞百分数 a,b % mononuclear cells a Sow blood b 母猪血 c a b CD8 CD4 Colostrum 初乳

20 Epithelial Surface 上皮表面 DAM 死亡的 killed Ig- dam 母猪 diff sow Ig- diff dam 牛 bovine Igbovine Lumen 管腔 Circulation 循环

21 Intestinal absorption of lymphoid cells in pigs 猪淋巴细胞的肠吸收 Maternal derived live cells 母源活细胞 ---yes 是 Allogeneic cells, other dam 异体细胞, 其他母猪 ---no 否 Heat killed maternal cells 热灭活母体细胞 ----no 否 Bovine derived cells in pigs 猪的牛源性细胞 ----no 否 Tuboly and Bernath 2002 Tuboly et al 1988 Williams P 1993 Reber et al 2008 Does the absorption of colostrum affect immunity? 初乳的吸收影响免疫力吗?

22 A. The lymphocyte migration across the neonatal intestine is source dependent 淋巴细胞在新生肠道转移是依赖来源的 Jejunum intestinal section 空肠段 A No cells 无细胞 B. Non mother 非母体 C. Mother cells 母体细胞 B. The transfer of lymphocytes across intestinal epithelium is colostral cell specific. 淋巴细胞肠上皮的转移具有初乳细胞特异性 A No cells 无细胞 B. PBMNC C. Mother cells 母体细胞 Elmubark et al 2013

Function of Colostral T lymphocytes 初乳 T 淋巴细胞功能 Innate Immunity 先天免疫 Proliferation 增殖 : Mitogens 有丝分裂原 Le Jan 1994* Williams 1993* Evans 1982* Mixed Leukocyte Reaction 混合淋巴细胞反应 Reber

, 2007* Le Jan 1996* Kohl et al 1982 + Th-2 response promoting TGF-, IL-4 Anti-inflammatory TGF-, IL-10 Pro-inflammatory TNF, IL-1, IL-6 References Species 参考物种 : * Swine 猪 # Bovine 牛

, 2007 # Macrophage 巨噬细胞 CD11a CD11c CD14 Toxicity 毒性 : Bactericidal/Fungicidal 杀菌 / 杀真菌 Ho & Lawson 1978 = Robinson et al 1978 = IL-2 Adaptive Immunity 适应性免疫 Delayed-Type

23 Function of Colostral T lymphocytes 初乳 T 淋巴细胞功能 Innate Immunity 先天免疫 Proliferation 增殖 : Mitogens 有丝分裂原 Le Jan 1994* Williams 1993* Evans 1982* Mixed Leukocyte Reaction 混合淋巴细胞反应 Reber et al 2005 # Th-1 response promoting IFN-, IL-12 Cytokine Production 细胞因子的产生 Ngyuen et al., 2007* Le Jan 1996* Kohl et al Th-2 response promoting TGF-, IL-4 Anti-inflammatory TGF-, IL-10 Pro-inflammatory TNF, IL-1, IL-6 References Species 参考物种 : * Swine 猪 # Bovine 牛 + Human 人 = Murine 鼠 Colostral Tlymphocyte 初乳 T 淋巴细胞 Activation of neonatal APC 新生儿 APC 的激活 Reber et al., 2008 # Donovan et al., 2007 # Macrophage 巨噬细胞 CD11a CD11c CD14 Toxicity 毒性 : Bactericidal/Fungicidal 杀菌 / 杀真菌 Ho & Lawson 1978 = Robinson et al 1978 = IL-2 Adaptive Immunity 适应性免疫 Delayed-Type Hypersensitivity 迟发型超敏反应 Schlesinger & Covelli, Proliferation 增殖 : Bacterial antigens 细菌抗原 Park et al, 1992* Viral antigens 病毒抗原 Ogra & Ogra 1978* Parmeley 1976* Activation of neonatal lymphocytes 新生儿淋巴细胞的激活 Reber et al., 2008 # Le Jan 1996* Bertotto et al., Sirota et al., 1995* MHC I CD25 CD26 B-cell:B 细胞 Isotype switching 同型交换

24 Post natal protection 出生后保护 The first post natal protection of piglet is provided by colostrum and milk. 产后仔猪的首次保护由初乳和奶提供 This protection is the function of timing and quality of colostrum 保护受时间和初乳质量影响

25 IgG and IgA in piglet serum piglet before and 24 h after colostrum ingestion 摄入初乳前和 24 h 后仔猪血清 IgG 和 IgA 免疫球蛋白 Immunoglobulin (mg /ml) IgG IgA n=35 0 Pre-suckling 哺乳前 After-suckling Variation: SEM 哺乳后

26 T lymphocyte subsets in piglet blood before and 24 h after colostrum ingestion 摄入初乳前和 24 h 后仔猪血液中的 T 淋巴细胞亚群百分数 Percent (%) PBMC CD8 CD4 p<0.01 Variation: SEM 0 Pre-suckling 哺乳前 After-suckling 哺乳后 4.4 +/- 0.73x10 6 cells/ml 7.0 +/- 0.71x10 6 cells/ml*

27 Conclusions 结论 Maternally-derived Ig and lymphocytes are transferred to piglets 母源 Ig 和淋巴细胞传递至仔猪 There is no selectivity in Ig transfer into piglets Ig 转运至仔猪无选择性 There is selectivity in lymphocyte transfer into piglets 淋巴细胞转运至仔猪有选择性 Colostral lymphocytes are functional 初乳淋巴细胞是功能性的

28 Integration of immunity 综合免疫力 : transfer from sow to piglet 从母猪转移到仔猪 Sow Periphery 母猪外周 Mammary Gland (MG) 乳腺 Piglet 仔猪 Immune component 免疫成分 Location in the sow/ transfer to the MG 母猪体内部位 / 转移至乳腺 Transfer to the MG 转移至乳腺 Transfer to the piglet/ location in the piglet 转移至仔猪 / 仔猪体内部位 Antibodies 抗体 (IgG, IgM, IgA): blood & MG 血 & 乳腺 IgG, IgM, IgA from so serum; concentrated via Fc- R in MG IgG, IgM, IgA 来自血清, 通过 FcR 集中在乳腺中 IgA, IgG, IgM locally produced in the MG under hormonal control 在激素控制下 IgA, IgG, IgM 在乳腺局部生产 Transfer via intestinal FcRn; Peripheral or local protection 通过肠道 FcRn 转移 ; 周围或局部保护 Bourne & Curtis 1973; Porter 1988; Stokes & Bourne 1989; Schnulle and Hurley, 2003; Watson, 1980; Stirling et al., 2005 Lymphocytes 淋巴细胞 (T & B): mucosae 粘膜 MadCam-1, CCL25 VCAM-1, CCL28 Unknown transfer mechanism; Mucosal & Lymph node 未知的传输机制 ; 粘膜及淋巴结 Tuboly et al, 1988; Harp and Moon, 1988; Kleinman and Walker, 1979; Salmon, 2000; Bourges et al, 2008; Wilson and Butcher, 2004

Maternal cells are functional in the recipient: specific immunity 母体细胞对受体是功能性的 : 特定免疫 Humans 人 : tuberculin sensitivity (DTH) transferred from mom to offspring 结核菌素敏感性 (DTH) 从母亲转移到后代

29 Maternal cells are functional in the recipient: specific immunity 母体细胞对受体是功能性的 : 特定免疫 Humans 人 : tuberculin sensitivity (DTH) transferred from mom to offspring 结核菌素敏感性 (DTH) 从母亲转移到后代 Schlesinger and Covelli 1977 Mice 鼠 : transfer of CMI sensitivity (DTH and lymphoproliferation) to nematode and fungal antigens CMI 敏感度 (DTH 和淋巴组织增生 ) 转移到线虫和真菌抗原 Rifkind Kumar Pigs 猪 :???

30 Maternally-derived antigen specific CMI is transferred to the neonate? 母体衍生的抗原特异性 CMI 被转移到新生儿? 母体猪肺炎支原体特异性细胞免疫被动转移至仔猪

DTH validation DTH 验证 A DxT (mm 2 ) PHA-DTH responses in piglets

Control 阴性对照 : Saline 盐水 1 0 24 36 48 D Positive Control 阳性对照 :

hyopneumoniae -DTH responses in piglets 仔猪猪肺炎支原体 DTH 反应 7 6

31 DTH validation DTH 验证 A DxT (mm 2 ) PHA-DTH responses in piglets 仔猪 PHA-DTH 反应 Nonvaccinated Vaccinated C Negative Control 阴性对照 : Saline 盐水 D Positive Control 阳性对照 : PHA B M. hyopneumoniae -DTH responses in piglets 仔猪猪肺炎支原体 DTH 反应 7 6 Nonvaccinated Vaccinated DxT (mm 2 ) E Experimental 实验 : M. hyopneumoniae 肺炎支原体 Time post injection (hours)

32 M. hyopneumoniae specific immunity in piglets 仔猪猪肺炎支原体特异性免疫 S/P ratio M. hyopneumoniae-specific antibodies 猪肺炎支原体特异性抗体 S/P 比值 PS AS * V sow N sow M. hyopneumoniae-specific proliferation 猪肺炎支原体特异性扩增 M. hyopneumonaie control (%) 猪肺炎支原体对照 Pre-suckling 哺乳前 After-suckling 哺乳后 Pre-suckling 哺乳前 After-suckling 哺乳后 Time at piglet blood sampling 仔猪采血时间 Variance: SE M. hyopneumoniae-immunity is transferred to piglets of vaccinated sows 猪肺炎支原体免疫力转移给了免疫母猪的仔猪 *

33 M. hyopneumoniae DTH responses in 3 d-old pigs 3 日龄猪的猪肺炎支原体 DTH 反应 DTH lesion size (mm) Maternal M. hyopneumoniae CMI is functional in the recipient 母体肺炎支原体 CMI 对受体是功能性的

34 Conclusions 结论 Maternal M. hyopneumoniae immunity is transferred to piglets 母体肺炎支原体免疫力转移到仔猪 Maternal M. hyopneumoniae CMI participates in the neonatal immune response 母体肺炎支原体 CMI 参与了新生猪的免疫反应

35 Vaccination of Neonates: pros and cons 新生猪接种的利弊 Traditional dogma 传统的教条 : neonatal and functional immature and maternal immunity is immunosuppressive 新生的, 功能不成熟的和母体免疫是免疫抑制 Need to be revisited 需要修订 : Vaccination of Neonates: Problems and Issues 新生猪接种疫苗 : 问题与建议 Douglas Hodgins and Patricia Shewan Vaccine 30:

36 Obstacles to Neonatal vaccination 新生猪接种疫苗的障碍 Corticosteriod associated with birth 与出生相关的皮质类固醇 Maternal cytokines eg TGF-b and IL-12 母体细胞激素, 如 TGF-b 和 IL-12 Nyugen et al 2007 Immunocompetent but Immunological Naïve 免疫功能正常, 但免疫缺乏 No exposure to antigens 没有接触抗原 Complement low 补体低 Microflora 菌群 Colonization with normal flora-increase in IgG, IgA 定植正常菌群 - IgG,IgA 增加 -Butler et al 2000 Gnotobiotic pig with E.coli---T and T independent Ag 无菌猪给大肠杆菌 - T 和 T 独立抗原 Germfree No response 无菌 - 无反应 Butler et al 2002 Adjuvant CpG enhanced response in gnotobiotic pigs CpG 佐剂增强无菌猪的反应 Butler et al 2005

37 Mucosal immune system 黏膜免疫系统 Gut flora 肠道菌群 Development of mucosal immune system in neonate driven by commensial microbial flora 微生物菌群驱动的新生猪黏膜免疫系统的发育 (Rothkotte etal 1991, Butler et al 2000) Pathogen associated molecular patterns 病原相关分子模式 (eg MDP, LPS) Substitute for gut flora in driving immunosresponsiveness 替代肠道菌群驱动免疫反应 (Butler et al 2005) Microflora will prove to be both essential for immune development and a approach in which to modulate responses 微生物群落将被证明是免疫发育所必须, 也是一种调节反应的方法

38 MDI influences neonatal immune development MDI 影响新生儿免疫发育 Sow Periphery 母猪外周血 Mammary Gland (MG) 乳腺 Piglet 仔猪 Pros of maternal immunity in the neonate 新生猪母体免疫的优势 Contributes to the neonatal immune repertoire 有助于新生猪免疫保留 Immediate immune protection 直接来源免疫保护 Antigen neutralization & opsonization in vivo 抗原中和 & 体内调理作用 Activate neonatal leukocytes 新生猪白细胞激活 May allow for memory CMI responses in the neonate 可以允许新生猪 CMI 反应记忆 May allow for memory AMI responses in the neonate 可以允许新生猪 AMI 反应记忆 Cons of maternal immunity in the neonate 新生猪母体免疫的劣势 Masks B-cell epitopes 掩盖 B- 细胞抗原决定基 Potential interference with AMI responses AMI 反应的潜在干扰 Potential interference with CMI responsescmi 反应的潜在干扰 MDI (AMI) may not always beneficially affect development in the neonate MDI(AMI) 可能并不总是对新生猪的发育有有利影响

39 Pseudorabies 伪狂犬病 : Wittan and Ohlinger, 1987 Mycoplasma hyopneumoniae 猪肺炎支原体 : Haesebrouk et al., 2006 Swine influenza virus (SIV) 猪流感病毒 Loeffen et al., 2003 Bordetella bronchiseptica 支气管败血波氏杆菌 : Kono et al., 1994 Classical swine fever (CSF) 猪瘟 Klinkenberg et al., 2002

40 Swine Influenza virus 猪流感病毒 Classical Swine Fever 猪瘟 Kitikoon et al., 2006; Surdahat and Damrongwatanapokin, 2003 Pseudorabies 伪狂犬病 Bouma, 1998; Wittman and Ohlinger, 1987 Measles 麻疹 Gans, 2001 BVDV Endsley, 2004 Lymphocytic choriomeningitis virus 淋巴细胞性脉络丛脑膜炎病毒 Seiler, 1998

5 w 3 w 0 d 7 d 14 d 21 d Sow vaccination 疫苗接种 Blood (0 &

vaccination and blood collection 仔猪接种疫苗和血液收集 DTH and blood

Vaccination status 母猪接种情况 Vaccinated 接种 (10) Nonvaccinated

vaccination status 仔猪疫苗接种状态 Vaccinated 接种 (V)

41 5 w 3 w 0 d 7 d 14 d 21 d Sow vaccination 疫苗接种 Blood (0 & 24 hr) and colostrum collection 血 (24 小时 ) 和初乳采集 Piglet vaccination and blood collection 仔猪接种疫苗和血液收集 DTH and blood collection DTH 和采血 DTH and blood collection DTH 和采血 Sow Vaccination status 母猪接种情况 Vaccinated 接种 (10) Nonvaccinated 未接种 (10) 20 (V V) 20 (N V) 20 (V N) 20 (N N) Piglet vaccination status 仔猪疫苗接种状态 Vaccinated 接种 (V) Nonvaccinated 未接种 (N) Commercial Farm: Prairie Land Pork in MN; not a naïve herd

42 M. hyopneumoniae antibodies in piglets 仔猪猪肺炎支原体抗体 day old piglets 14 日龄仔猪, 7 dpv 21 day old piglets 21 日龄仔猪, 14 dpv 3.5 S/P 比值 S/P ratio N N N V V V V N N N N V V V V N Piglet Treatment 仔猪处理 Sow & Piglet treatment 母猪及仔猪处理 ; N: Nonvaccinated 未接种 ; V: Vaccinated 接种 ; dpv: days post vaccination 接种后天数 ; Variance: SD 方差 No evidence of AMI stimulation following piglet vaccination 没有证据表明仔猪疫苗接种后 AMI 激活

43 M. hyopneumoniae control 猪肺炎支原体 - 对照 (%) M. hyopneumoniae-specific proliferation 猪肺炎支原体特异性扩增 14 day old piglets 14 日龄仔猪, 7 dpv a day old piglets 21 日龄仔猪, 14 dpv a b b a 0 d14 N N N V V V V N -2 0 d 21 N N N V V V V N Piglet Treatment 仔猪处理 Sow & Piglet treatment 母猪及仔猪处理 ; N: Nonvaccinated 未接种 ; V: Vaccinated 接种 ; dpv: days post vaccination 接种后天数 ; Variance: SE 标准误 Evidence of CMI stimulation following piglet vaccination 仔猪疫苗接种后 CMI 激活的证据

44 14 day-old piglets 14 日龄仔猪, 7dpv 21 day-old piglets 21 日龄仔猪, 14dpv Lesion size 病灶大小 (mm) N N N V V N V V N N N V V N V V Piglet treatment 仔猪处理 Sow & Piglet treatment 母猪及仔猪处理 ; N: Nonvaccinated 未接种 ; V: Vaccinated 接种 ; dpv: days post vaccination 接种后天数 Evidence of CMI memory response following piglet vaccination 仔猪疫苗接种后 CMI 记忆反应的证据

45 Conclusions 结论 7 day-old piglets respond to vaccination 7 日龄仔猪接种疫苗的反应 Maternally-derived cells are functional and participate in the antigen specific immune response in piglets 母体源性细胞是功能性的, 并参与仔猪抗原特异性免疫反应 No evidence that passively transferred immunity interferes with CMI responses in piglets 没有证据表明被动转移免疫力干扰仔猪 CMI 反应

46 Transfer of maternally-derived immunity (MDI) from colostrum to the piglet 自初乳将母源免疫力转移至仔猪 Effects of management Cross fostering? 管理交叉寄养的效应? Sow Periphery 母猪外周 Mammary Gland (MG) 乳腺 Piglet 仔猪

47 The effect of cross-fostering on transfer of maternal Mycoplamsa hyopneumonia-specific immune transfer 交叉寄养对母源猪肺炎支原体特异性免疫转移的影响 Bandrick et al Vet Record 2011

48 Sow vaccination 母猪疫苗接种 PS blood collection 哺乳前采血 AS blood collection 哺乳后采血 DTH sample injectiondt H 样品注入 DTH measurement DTH 测量 5 weeks 3 weeks 0h 6h 12h 18h 24h 36h 48h 72h Sow blood collection 母猪采血 piglet movement 仔猪移动 piglet movement 仔猪移动 piglet movement 仔猪移动 piglet movement 仔猪移动 Endpoints 终点 : Sows 母猪 : Serum-Antibody quantitation via ELISA 通过 ELISA 血清抗体定量 Piglets 仔猪 : Delayed-type hypersensitivity (DTH) 迟发型超敏反应 Serum-Antibody quantitation via ELISA 通过 ELISA 血清抗体定量 Piglet age (hrs) 仔猪年龄 ( 小时 )

49 Figure 2. Mycoplasma hyopneumoniae-specific antibodies in sows 图 2. 母猪猪肺炎支原体特异性抗体 Figure 2A. Mycoplasma hyopneumoniae-specific antibodies in sow serum 图 2A. 母猪血清中猪肺炎支原体特异性抗体 S:P ratio *** *** 0.0 Pre-vaccination 接种前 Nonvaccinated 未接种 Vaccinated 接种 Sow M. hyopneumoniae vaccination status 母猪猪肺炎支原体疫苗接种状态 Figure 2B. Mycoplasma hyopneumoniae-specific antibodies in sow colostrum 图 2B. 母猪初乳中的猪肺炎支原体特异性抗体 S:P ratios:p 比 *** Nonvaccinated 未接种 Vaccinated 接种 Sow M. hyopneumoniae vaccination status 母猪猪肺炎支原体疫苗接种状态

50 A DTH orthogonal diameterdth 正交外径 (mm) Figure 3. Mycoplasma hyopneumoniae-specific DTH responses in piglets at 3 d of age 图 3. 猪肺炎支原体特异性 DTH 反应, 仔猪 3 日龄 B Non-cross-fostered 非交叉培育 Vaccinated 接种 x Nonvaccinated 未接种 a 3 b 2 b 1 b 0 a a Vaccinated Nonvaccinated Dam vaccination status Dam 疫苗接种状况 Time at cross-fostering 交叉寄养时间 DTH orthogonal diameterdth 正交外径 (mm) C DTH orthogonal diameterdth 正交外径 (mm) Vaccinated 接种 x Vaccinated 接种 b b a a Time at cross-fostering 交叉寄养时间 D DTH orthogonal diameterdth 正交外径 (mm) Nonvaccinated 未接种 x Vaccinated 接种 Time at cross-fostering 交叉寄养时间

51 Figure 4. Mycoplasma hyopneumoniae-specific antibodies in piglets at 24 hrs of age 图 4. 猪肺炎支原体特异性抗体, 仔猪 24 小时龄 A Non-cross-fostered 非交叉培育 B Vaccinated 接种 x Nonvaccinated 未接种 S:P ratios:p 比 a S:P ratios:p 比 b b b 0.5 b Vaccinated 接种 Nonvaccinated 未接种 Dam vaccination statusdam 疫苗接种状况 0.5 a Time at cross-fostering 交叉寄养时间 C 3.5 Vaccinated 接种 x Vaccinated 接种 D 3.5 Nonvaccinated 未接种 x Vaccinated 接种 S:P ratios:p 比 S:P ratios:p 比 a b Time at cross-fostering 交叉寄养时间 0.5 c 0.0 d Time at cross-fostering 交叉寄养时间

52 Conclusions 结论 Cross-fostering affects transfer of maternallyderived CMI and AMI differently 交叉寄养不同地影响母源性 CMI 和 AMI 传递 Sow immune status should be uniform 母猪免疫状态应该是一致的 Cross-foster pigs after 12 at least 12 hours 至少 12 小时后交叉寄养猪

53 Amazing Immune system in Neonatal pig Summary 惊人的新生猪免疫系统总结 Swine are different in their immune system and response compared to humans and mice. 猪与人和鼠相比免疫系统和反应是不同的 Both AMI and CMI is transferred to piglets in colostrum; there is selectivity in lymphocyte transfer AMI 和 CMI 都通过初乳被转移至仔猪, 淋巴转移有选择性 Maternally-derived CMI is functional in the recipient 母源性 CMI 对受体是功能性的 Piglets respond to vaccination with CMI responses in the face of MDI 面对 MDI, 仔猪以 CMI 反应来回应疫苗接种 No evidence of passive interference with CMI responses from MDI at least in M. hyopneunominae 至少在猪支原体肺炎没有来自 MDI 的 CMI 反应被动干预的证据

54 Amazing Immune system summary cont 惊人的免疫系统总结续 Important consideration 重要的考虑因素 Vaccination programs 疫苗接种计划 Pay attention to maternally-derived CMI 注意母体源性 CMI CMI is an important response and may be stimulated even when there is a lack in AMI responses CMI 是一个重要的反应, 且可能在 AMI 反应缺乏时被激活 Cross-fostering timing and piglet management 交叉寄养时间和仔猪管理

55 Question 问题 : Using this information--what are the further challenges and opportunities? 使用此信息进一步的挑战和机遇是什么?

56 Acknowlegement 致谢 University of Minnesota Collaborators 明尼苏达大学的合作者 Meggan Bandrick DVM PhD Gamal Elmubark MD PhD Mike Olin PhD Sam Baidoo, PhD Mike Murtaugh PhD Funding support 资金支持 Minnesota Pork Board National Pork Board NIH T32 (Bandrick) Zoetis

57 Future directions 未来的发展方向 -- Wellness Testing 健康测试 Acute Phase proteins 急性时相蛋白 Immunoglobulins 免疫球蛋白 General, Ag specific 总体的, 抗原特异的 Immune cell population- 免疫细胞群 numbers, function 数字, 功能 Cell mediated responses 细胞介导的反应 ELISPOT, general, Ag-specific 酶联免疫斑点, 总体的, 抗原特异的 Cytokine measures 细胞因子测量

Mycoplasma hyopneumoniae and its role in the Porcine Respiratory Disease Complex 猪肺炎支原体及其在猪呼吸系统综合疾病中的作用 Dr. Maria Pieters 玛丽亚皮特斯博士

Mycoplasma hyopneumoniae and its role in the Porcine Respiratory Disease Complex 猪肺炎支原体及其在猪呼吸系统综合疾病中的作用 Dr. Maria Pieters 玛丽亚皮特斯博士 Outline 纲要 Porcine Respiratory Disease Complex (PRDC) 猪呼吸道疾病综合征 (PRDC)

The Amazing Immune System in the Neonatal Pig 新生猪惊人的免疫系统 Tom Molitor 汤姆 莫利特博士 University of Minnesota 明尼苏达大学 Leman China, October 13-

The Amazing Immune System in the Neonatal Pig 新生猪惊人的免疫系统 Tom Molitor 汤姆莫利特博士 University of Minnesota 明尼苏达大学 Leman China, October 13-