应用吞噬性能选育矮小鸡的抗病群体

doi:10.3864/j.issn.0578-1752.2016.23.015

摘要/Abstract

摘要： 【目的】单核巨噬细胞在免疫系统中发挥着重要的作用，试验研究了对单核巨噬细胞吞噬性能的选择影响矮小鸡G1代抗传染性支气管炎病毒（IBV）的能力。【方法】在矮小鸡290日龄时，检测了G0代500只个体（母鸡400只，公鸡100只）的吞噬指数（PI），并根据PI的大小分为强吞噬力组（HPIG）和弱吞噬力组（LPIG）。建立2 ´ 2交配组合：HPIG ♂ ´ HPIG ♀（强公强母组），LPIG ♂ ´ HPIG ♀（弱公强母组），HPIG ♂ ´ LPIG ♀（强公弱母组），LPIG ♂ ´ LPIG ♀（弱公弱母组）。随机选择400只1日龄G1代雏鸡（每组100只，公母各半），其中360只采用滴鼻方式人工接种含 IBV M41病毒的鸡胚尿囊液，40只作为对照，连续观察14d，记录死亡数，制作石蜡切片进行H. E.染色，并通过红细胞凝集抑制试验（HI）测定15 d时存活鸡只的抗体效价。G1代鸡20周龄时，选择母鸡12只，其中强组母鸡后代6只，弱组母鸡后代6只，以病毒模拟物Poly I:C刺激体外培养的单核巨噬细胞，采用荧光定量PCR的方法测定细胞因子及主要组织相容性复合体（MHC）mRNA的表达水平。【结果】G0代不同个体对异源红细胞的吞噬能力差异显著，通过测定G0代个体的吞噬指数，建立交配组合，孵育得到G1代鸡。对G1代鸡的IBV攻毒试验的结果为强公强母组后代的死亡率（33.3±0.05）%显著低于弱公弱母组（55.6±0.05）%，其他两个组合后代的死亡率介于上述值之间，弱公强母组为(43.3±0.05)%，强公弱母组为(47.8±0.05)%；母鸡对后代的影响大于公鸡，强母组后代的死亡率（38.3±0.04）%显著低于弱母组（51.7±0.04）%。攻毒组在接种IBV M41病毒3 d后表现出咳嗽、呼吸困难、食欲减退、精神沉郁等临床症状，对病死鸡的气管及肾脏的H.E.染色结果可见典型的病灶，气管上皮细胞坏死脱落，肾小管上皮细胞发生空泡变性等，而对照组均无临床症状及组织病变。对198只攻毒后存活个体抗体滴度的测定结果显示强组母鸡后代抗体滴度（8.45±0.07）显著高于弱组母鸡后代的抗体滴度（8.10±0.08）。利用病毒模拟物Poly I:C刺激体外培养的单核巨噬细胞2 h后，强吞噬力个体（强组母鸡后代）细胞因子IFN γ和IL-1β的表达量分别是弱吞噬力个体（弱组母鸡后代）的5.14倍（P＜0.05）和2.41倍（P＜0.05）。强吞噬力个体MHCⅠ的表达量显著高于弱吞噬力个体，而MHCⅡ的表达量差异不显著。【结论】通过测定体外培养的单核巨噬细胞的吞噬性能，按照吞噬指数的高低建立交配组合，强吞噬力母鸡后代的攻毒死亡率显著低于弱吞噬力母鸡后代，且其抗体滴度、细胞因子（IFN γ、IL-1β）和MHCⅠ的表达量显著高于弱吞噬力母鸡后代，说明强吞噬力母鸡后代具有较强的抗IBV的能力。因此单核巨噬细胞的吞噬能力可以作为培育抗IBV品系的一种指标。

关键词: 单核细胞, 巨噬细胞, 吞噬性能, 抗病性能, 矮小鸡

Abstract: 【Objective】 Monocytes-macrophages play an important role in the immune system. The effect of selection for monocytes-macrophages phagocytosis on infectious bronchitis virus (IBV) resistance in generation 1 (G1) of dwarf chickens was studied. 【Method】 The phagocytic index (PI) of 500 dwarf chickens (400 hens and 100 cocks) of generation 0 (G0) was tested at 290 d of age, and then the chickens were divided into high and low PI groups (HPIG and LPIG). 2 ´ 2 mating combinations were conducted: HPIG ♂ ´ HPIG ♀, LPIG ♂ ´ HPIG ♀, HPIG ♂ ´ LPIG ♀, LPIG ♂ ´ LPIG ♀. Four hundred G1 chickens (half in sex) equally from 4 mating groups were selected to IBV challenge at 1 d, 360 of which were artificially inoculated with allantoic fluid containing IBV M41 virus, while 40 as a control. Chickens were observed for 14 d and deaths were recorded. Paraffin sections were made and stained by hematoxylin-eosin (H. E.). Antibody titers of the survival chickens at 15 d were measured by the red cell agglutination inhibition test (HI). Twelve G1 chickens at 20 w equally from high and low PI groups were selected. Monocytes- macrophages were isolated and cultured, then challenged with Poly I:C. Expression of mRNA of cytokines and major histocompatibility complex (MHC) were tested by quantitative real-time PCR. 【Result】 Phagocytic ability of heterologous erythrocytes were different significantly in G0. G1 chickens were incubated according to mating groups according to PI of G0. Results of challenge in G1 showed that the mortality rate (33.3±0.05)% of progeny from HPIG ♂ ´ HPIG ♀ were significantly lower than that of progeny from LPIG ♂ ´ LPIG ♀(55.6±0.05)%. Mortality rates of progeny from LPIG ♂ ´ HPIG ♀ and HPIG ♂´ LPIG ♀were (43.3±0.05)% and (47.8±0.05)% respectively. Effect of hens on the offspring was greater than cocks. The mortality rate of progeny from HPIG ♀ was (38.3±0.04)%, which was significantly lower than that of progeny from LPIG ♀(51.7±0.04)%. Chickens showed clinical symptoms of cough, shortness of breath, loss of appetite and depression after challenged with IBV M41 for 3 d. Typical damages on the trachea and kidney of dead and sick chickens could be seen through H. E. staining. Epithelial cells appeared necrosis and empty bubble degeneration in tracheal, respectively. The control group showed no clinical symptoms and pathological changes. The antibody titers of 198 surviving challenged individuals showed that antibody titer of progeny from HPIG ♀ (8.45±0.07) was significantly higher than that of progeny from LPIG ♀(8.10±0.08). Expressions of IFN γ and IL-1β of high phagocytic chickens (progeny of HPIG ♀) was 5.14 times (P＜0.05) and 2.41 times (P＜0.05) higher than the low phagocytic chickens (progeny of LPIG ♀). Expression of MHC Ⅰ of high phagocytic chickens was significantly higher than that of low phagocytic chickens, while expression of MHCⅡ was not significant. 【Conclusion】 The experiment was performed with four mating combinations according to phagocytic index of monocytes-macrophages in vitro. Mortality rate of progeny of HPIG ♀ was significantly lower than that of progeny of LPIG ♀ after challenge, while the antibody titer and the expression of cytokines (IFN γ and IL-1β) and MHC Ⅰ were significantly higher. The results showed that the progeny of HPIG ♀ was more resistant to IBV than progeny of LPIG ♀. Therefore, the phagocytic ability of monocytes-macrophages could be an indicator for breeding IBV resistant lines.

Key words: monocyte, macrophage, phagocytosis, disease resistance, dwarf chicken

麻慧，韩红兵，宁中华，连正兴. 应用吞噬性能选育矮小鸡的抗病群体[J]. 中国农业科学, 2016, 49(23): 4628-4637.

MA Hui, HAN Hong-bing, NING Zhong-hua, LIAN Zheng-xing. Breeding of Disease Resistant Dwarf Chickens by Phagocytic Ability[J]. Scientia Agricultura Sinica, 2016, 49(23): 4628-4637.

0
/ / 推荐

导出引用管理器 EndNote|Reference Manager|ProCite|BibTeX|RefWorks

链接本文: https://www.chinaagrisci.com/CN/10.3864/j.issn.0578-1752.2016.23.015

https://www.chinaagrisci.com/CN/Y2016/V49/I23/4628

参考文献

[1] 陶家权, 刘乔然, 乔荣岑, 孙进忠, 张许科. 鸡传染性支气管炎的流行及防控. 中国家禽, 2009, 31(24):65-66. TAO J Q, LIU Q R, QIAO R C, SUN J Z, ZHANG X K. Epidemiology and control of chicken infectiou bronchitis. China Poultry, 2009, 31(24): 65-66. (in Chinese) [2] O'MAHONY D S, PHAM U, IYER R, HAWNT R, LILES W C. Differential constitutive and cytokine-modulated expression of human toll-like receptors in primary neutrophils, monocytes, and macrophages. Internationnal Journal of Medical Sciences, 2008, 5(1): 1-8. [3] METTERLEIN T, SCHUSTER F, TADDA L, HAGER M, ROEWER N, ANETSEDER M. Statins alter intracellular calcium homeostasis in malignant hyperthermia susceptible individuals. Cardiovascular Therapeutics, 2010, 28(6):356-360. [4] YONASH N, BACON L D, WITTER R L, CHENG H H. High resolution mapping and identification of new quantitative trait loci (qtl) affecting susceptibility to marek’s disease. Animal Genetics, 1999, 30(2):126-135. [5] ZHAO X L, HONAKER C F, SIEGEL P B. Phenotypic responses of chickens to long-term selection for high or low antibody titers to sheep red blood cells. Poultry Science, 2012, 91:1047-1056. [6] BEAUMONT C, CHAPUIS H, PROTAIS J, SELLIER N, MENANTEAU P, FRAVALO P, VELGE P. Resistance to Salmonella carrier state: Selection may be efficient but response depends on animal’s age. Genetics Research, 2009, 91:161-169. [7] CHENG H W, EICHER S D, CHEN Y, SINGLETON P, MUIR W M. Effect of genetic selection for group productivity and longevity on immunological and hematological parameters of chickens. Poultry Science, 2001, 80:1079-1086. [8] LYALL J, IRVINE R M, SHERMAN A, MCKINLEY T J, NUNEZ A, PURDIE A, OUTTRIM L, BROWN I H, ROLLESTON-SMITH G, SANG H, TILEY L. Suppression of avian influenza transmission in genetically modified chickens. Science, 2011, 331:223-226. [9] SWAGGERTY C L, PEVZNER I Y, FERRO P J, CRIPPEN T L, KOGUT M H. Association between in vitro heterophil function and the feathering gene in commercial broiler chickens. Avian Pathology, 2003, 32(5):483-488. [10] SWAGGERTY C L, FERRO P J, PEVZNER I Y, KOGUT M H. Heterophils are associated with resistance to systemic Salmonella enteritidis infection in genetically distinct lines of chickens. Fems Immunology and Medical Microbiology, 2005, 43:149-154. [11] SWAGGERTY C L, LOWRY V K, FERRO P J, PEVZNER I Y, KOGUT M H. Disparity in susceptibility to vancomycin-resistant Enterococcus organ invasion in commercial broiler chickens that differ in innate immune responsiveness. Food and Agricultural Immunology, 2005, 16:1-15. [12] SWAGGERTY C L, GENOVESE K J, HE H, DUKE S E, PEVZNER I Y, KOGUT M H. Broiler breeders with an efficient innate selection for resistangce against salmonella. Poultry Science, 2011, 90: 1014-1019. [13] SWAGGERTY C L, KAISER P, ROTHWELL L, PEVZNER I Y, KOGUT M H. Heterophil cytokine mRNA profiles from genetically distinct lines of chickens with differential heterophil-mediated innate immune responses. Avian Pathology, 2006, 35:102-108. [14] LI H, ZHANG Y, NING Z H, DENG X M, LIAN Z X, LI N. Effect of selection for phagocytosis in dwarf chickens on immune and reproductive characters. Poultry Science, 2008, 87(1):41-49. [15] SUN S F, PAN Q Z, HUI X, ZHANG B L, WU H M, LI H, XU W, ZHANG Q, LI J Y, DENG X M, CHEN J W, LIAN Z X, LI N. Stronger in vitro phagocytosis by monocytes-macrophages is indicative of greater pathogen clearance and antibody levels in vivo. Poultry Science, 2008, 87(9):1725-1733. [16] BASSOE C F, SMITH I, SORNES S, HALSTENSEN A, LEHMANN A K. Concurrent measurement of antigen- and antibody-dependent oxidative burst and phagocytosis in monocytes and neutrophils. Methods, 2000, 21(3):203-220. [17] 萨仁娜, 佟建明, 何春年, 高微微, 张琪. 海带岩藻聚糖及其级分在氧化应激条件下对肉鸡巨噬细胞免疫功能的影响. 中国农业科学, 2008, 41(10): 3256-3263. SA R N, TONG J M, HE C N, GAO W W, ZHANG Q. Effect of fucoidan and fraction isolated from laminaria japonica on immunity of broiler macrophages under oxidation stress condition. Scientia Agricultura Sinica, 2008, 41(10): 3256-3263. (in Chinese) [18] NILSSON A, OLDENBORG P A. Cd47 promotes both phosphatidylserine- independent and phosphatidylserine-dependent phagocytosis of apoptotic murine thymocytes by non-activated macrophages. Biochemical and Biophysical Research Communications, 2009, 387(1): 58-63. [19] SOLANO G I, BAUTISTA E, MOLITOR T W, SEGALES J, PIJOAN C. Effect of porcine reproductive and respiratory syndrome virus infection on the clearance of haemophilus parasuis by porcine alveolar macrophages. Canadian Journal of Veterinary Research-revue Canadienne de Recherche Veterinaire, 1998, 62(4):251-256. [20] HAMAL K R, BURGESS S C, PEVZNER I Y, ERF G F. Maternal antibody transfer from dams to their egg yolks, egg whites, and chicks in meat lines of chickens. Poultry Science, 2006, 85(8):1364-1372. [21] GRINDSTAFF J L. Maternal antibodies reduce costs of an immune response during development. Journal of Experimental Biology , 2008, 211(Pt 5):654-660. [22] Reid J M, Arcese P, Keller L F, Hasselquist D. Long-term maternal effect on offspring immune response in song sparrows melospiza melodia. Biology Letters, 2006, 2(4):573-576. [23] WIGLEY P, HULME S, ROTHWELL L, BUMSTEAD N, KAISER P, BARROW P. Macrophages isolated from chickens genetically resistant or susceptible to systemic salmonellosis show magnitudinal and temporal differential expression of cytokines and chemokines following salmonella enterica challenge. Infection and Immunity, 2006, 74(2):1425-1430. [24] SWAGGERTY C L, PEVZNER I Y, HE H, GENOVESE K J, NISBET D J, KAISER P, KOGUT M H. Selection of broilers with improved innate immune responsiveness to reduce on-farm infection by foodborne pathogens. Foodborne Pathogens and Disease, 2009, 6(7): 777-783. [25] 黄琴, 黄怡, 崔志文, 李雅丽, 李卫芬, 余东游. 鼠李糖乳杆菌对巨噬细胞先天性免疫应答的调节作用. 中国农业科学, 2012, 45(8): 1621-1626. HUANG Q, HUANG Y, CUI Z W, LI Y L, LI W F, YU D Y. Modulation of lactobacillus rhamnosus on innate immune responses in macrophages. Scientia Agricultura Sinica, 2012, 45(8): 1621-1626. (in Chinese) [26] 王继英, 王彦平, 郭建凤, 王怀中, 林松, 张印, 武英. 仔猪外周血中内参基因的筛选及细胞因子和受体的表达水平. 中国农业科学, 2015, 48(7): 1437-1444. WANG J Y, WANG Y P, GUO J F, WANG H Z, LIN S, ZHANG Y, WU Y. Selection of reference genes and determination of cytokines and receptor mRNA expression in peripheral blood of piglets. Scientia Agricultura Sinica, 2015, 48(7): 1437-1444. (in Chinese) [27] CAUTHEN A N, SWAYNE D E, SEKELLICK M J, MARCUS P I, SUAREZ D L. Amelioration of influenza virus pathogenesis in chickens attributed to the enhanced interferon-inducing capacity of a virus with a truncated ns1 gene. Journal of Virology, 2007, 81(4): 1838-1847. [28] NIU M, HAN Y, LI W. Baculovirus up-regulates antiviral systems and induces protection against infectious bronchitis virus challenge in neonatal chicken. International Immunopharmacology, 2008, 8(12): 1609-1615. [29] PEI J, SEKELLICK M J, MARCUS P I, CHOI I S, COLLISSON E W. Chicken interferon type i inhibits infectious bronchitis virus replication and associated respiratory illness. Journal of Interferon and Cytokine Research, 2001, 21(12):1071-1077. [30] HE H, GENOVESE K J, KOGUT M H. Modulation of chicken macrophage effector function by th1/th2 cytokines. Cytokine, 2011, 53:363-369. [31] FLESCH I E, HESS J H, HUANG S, AGUET M, ROTHE J, BLUETHMANN H, KAUFMANN S H. Early interleukin 12 production by macrophages in response to mycobacterial infection depends on interferon gamma and tumor necrosis factor alpha. Journal of Experimental Medicine, 1995, 181(5):1615-1621. [32] FINKELMAN F, KATONA I, MOSMANN T R, COFFMAN R L. Ifn-gamma regulates the isotypes of ig secreted during in vivo humoral immune responses. The Journal of Immunology, 1988, 140(4): 1022-1027. [33] CHEESEMAN J H, KAISER M G, CIRACI C, KAISER P, LAMONT S J. Breed effect on early cytokine mrna expression in spleen and cecum of chickens with and without salmonella enteritidis infection. Developmental and Comparative Immunology, 2007, 31(1): 52-60. [34] BACON L D, HUNTER D B, ZHANG H M, BRAND K, ETCHES R. Retrospective evidence that the mhc (b haplotype) of chickens influences genetic resistance to attenuated infectious bronchitis vaccine strains in chickens. Avian Pathology, 2004, 33(6):605-609. [35] ZINKERNAGEL R. On cross-priming of mhc class i-specific ctl: Rule or exception? European Journal of Immunology, 2002, 32(9): 2385-2392.