Progression of histomonosis in commercial chickens following experimental infection with an in vitro propagated clonal culture of Histomonas meleagridis

Four commercial strains of chickens, namely, ISA brown leghorn (ISA), TETRA-SL brown (TETRA-SL), Lohmann brown (LB), and Lohmann LSL (LSL), were infected with a well-defined clonal culture of Histomonas meleagridis (H. meleagridis/Turkey/Austria/2922-C6/04) to investigate their susceptibility to histomonosis. Each group included 16 chickens, which were housed under the same conditions in separate pens. All chickens were infected with 10(4) histomonads orally and intracloacally at 14 days of age. No mortality or clinical signs were observed during the experiment in all birds. Three birds of each chicken strain were euthanatized on days 4, 7, 10, 14, and 21 postinfection. Incidence of histomonosis on the basis of cecal lesions was found to be 64.00% in TETRA-SL, 62.50% in LB, 53.12% in LSL, and 43.75% in ISA chickens. Fewer lesions were noticed in livers than in ceca, with an incidence of 15.62% in TETRA-SL, 9.37% in LB, and 3.12% in ISA chickens. No liver lesions were found in the LSL chickens. Statistical analysis revealed that there was no significant difference (P > 0.05) in susceptibility to experimental H. meleagridis infection based on cecal and liver involvement. Polymerase chain reaction (PCR) and immunohistochemistry were found to be reliable tools to confirm the presence of histomonads and changes in the ceca. However, some negative PCR results were recorded from the livers despite the presence of macroscopic lesions. Additionally, DNA of H. meleagridis was detected by PCR in a few of the lungs, but immunohistochemistry was negative. Nucleic acid of the protozoan parasite was not detected in samples from kidney, brain, spleen, or bursa of Fabricius. Altogether, the high susceptibility of commercial chicken lines to histomonosis could be demonstrated and characterized by severe lesions in the ceca and insignificant involvement of the liver, approaching a maximum on days 7-14 postinfection.     

Cryptosporidium baileyi: effects of intra-abdominal and intravenous inoculation of oocysts on infectivity and site of development in broiler chickens

Developmental stages of Cryptosporidium baileyi were observed on the epithelium of the larynx, trachea, primary and secondary bronchi, air sacs, bursa of Fabricius, and cloaca of 12 chickens inoculated intra-abdominally with oocysts. All 12 birds inoculated intra-abdominally developed clinical signs of respiratory disease and had gross lesions of airsacculitis at necropsy. Developmental stages of C. baileyi and clinical signs of disease were not observed in 12 chickens inoculated intravenously with oocysts. The response of chickens to intra-abdominal inoculation of oocysts was similar to responses recorded following intratracheal inoculation of oocysts in previous studies.     

Presence of Campylobacter jejuni in various organs one hour, one day, and one week following oral or intracloacal inoculations of broiler chicks

Day-old broiler chicks (n=30) were obtained from a commercial hatchery and inoculated, either orally or intracloacally, with a characterized strain of Campylobacter jejuni. At 1 hr, 1 day, and 1 wk after inoculation, broilers (n = 5) from the orally and intracloacally inoculated groups along with control birds (n=4) were humanely killed by cervical dislocation. The broilers from the control and treatment groups were aseptically opened, and the thymus, spleen, liver/gallbladder, bursa of Fabricius, and ceca were aseptically removed and individually analyzed for C. jejuni. Overall, C. jejuni was isolated after oral inoculation from 13% (10/ 75), 17% (13/75), and 28% (14/50) of the 1-hr, 1-day, and 1-wk samples, respectively. Campylobacter jejuni was isolated from 10% (4/ 40), 8% (3/40), 10% (4/40), 25% (10/40), and 40% (16/40) of the thymus, spleen, liver/gallbladder, bursa of Fabricius, and ceca samples, respectively. Following the intracloacal route of inoculation, C. jejuni was recovered from 32% (24/75), 8% (6/75), and 16% (8/50) of the 1-hr, 1-day, and 1-wk samples, respectively. Campylobacter jejuni was isolated from 5% (2/40), 5% (2/40), 5% (2/40), 45% (18/40), and 40% (16/40) of the thymus, spleen, liver/gallbladder, bursa of Fabricius, and ceca samples, respectively, for all sampling periods. Campylobacter spp. were not recovered from sample sites examined from the control broilers from trial one, trial two, or trial three samples examined after 1 hr and 1 day. However, one control sample was positive from the 1-wk sampling from repetition three; therefore, those data were omitted. The rapid movement of Campylobacter to internal organs following both oral and intracloacal inoculation may be significant, particularly if it persists in these organs as reservoirs throughout the 65-wk life cycle of breeding birds.     

Effect of infectious bursal disease virus insult on iron, copper, and zinc concentration in liver, bursa of Fabricius, spleen, pancreas, and serum of chickens

The effect of a systemic disease on the dynamics of iron, zinc, and copper in chickens fed ad libitum was examined by infecting 10-day-old specific pathogen-free chickens with infectious bursal disease virus (IBDV). Liver, bursa of Fabricius, pancreas, spleen, and serum were sampled in 10 controls and 10 challenged chickens at 3-day intervals postinfection (PI) for 15 days. The samples were analyzed using atomic absorption spectroscopy. Serum levels were similar to that reported in the literature. Concentrations of iron and zinc did not change significantly in the pancreas, but there was an increase in copper in infected pancreatic tissue on days 9 and 15 PI. Iron concentration in the spleen showed a significant increase on days 6, 9, and 15 PI, whereas zinc was only significantly increased on day 15 PI. There was no significant change in copper concentrations in the spleens of infected chickens vs. controls. This finding is in line with previously reported data. The results showed that the liver was not a major tissue where iron and zinc were sequestered, as previous data have shown in mammals. Instead, the bursa of Fabricius had significantly increased levels of both iron and zinc in infected tissue vs. control tissue from 9 days PI on. Furthermore, the bursa had increased levels of copper in the latter portion of the study. These findings suggest that the bursa of Fabricius rather than the liver is the major organ for metallic ion sequestering during IBDV infection.     

禽网状内皮组织增生病病毒感染对SPF雏鸡法氏囊细胞Bcl-2和C-myc基因表达及凋亡的影响

【目的】 探究禽网状内皮组织增生病病毒(Reticuloendotheliosis virus,REV)感染禽类后导致免疫器官发生细胞凋亡的机理。【方法】 以1日龄SPF雏鸡为试验对象,将100只SPF雏鸡随机均分为REV感染组和未感染病毒的对照组,REV感染组雏鸡经腹腔感染500 μL REV稀释液,对照组雏鸡经相同途径注射等量灭菌生理盐水,于病毒感染后第1、7、14、21、28和42天,2组雏鸡随机各抽取5只,心脏采血处死雏鸡后快速摘取法氏囊。分别应用HE染色和病理切片成像系统测定分析法氏囊细胞核浆比,TUNEL细胞凋亡原位检测试剂盒测定凋亡细胞数,免疫组化法测定Bcl-2和C-myc阳性细胞数量,实时荧光定量PCR和ELISA法分别检测法氏囊Bcl-2和C-myc 基因mRNA表达和蛋白含量。【结果】 ①REV感染1日龄SPF雏鸡后21~42 d,其法氏囊淋巴细胞凋亡百分比显著或极显著高于对照组雏鸡(P<0.05;P<0.01);②SPF雏鸡感染REV后21和28 d,其法氏囊细胞核浆比显著低于对照组雏鸡(P<0.05);③REV感染SPF雏鸡法氏囊中Bcl-2和C-myc阳性细胞数在病毒感染后21和28 d显著高于对照组雏鸡(P<0.05);④REV感染SPF雏鸡后21 d,其法氏囊Bcl-2和C-myc 基因mRNA表达极显著高于对照组雏鸡(P<0.01)。⑤SPF雏鸡感染REV后,其法氏囊中Bcl-2蛋白含量较对照组雏鸡有不同程度的增加,其中21和28 d分别差异极显著(P<0.01)和显著(P<0.05),病毒感染组雏鸡的C-myc蛋白含量也始终高于对照组雏鸡,且21和28 d极显著增高(P<0.01)。【结论】 REV感染所致SPF雏鸡法氏囊细胞Bcl-2和C-myc的mRNA表达以及蛋白含量异常均与病毒感染导致的法氏囊细胞凋亡密切相关,而法氏囊细胞凋亡数量增加与REV感染引发的机体免疫机能抑制密切相关。     

Cryptosporidiosis of the bursa of Fabricius of chickens.

Light-microscope and electron-microscope studies of a coccidial organism found in the bursa of Fabricius from 3 chickens clearly established the parasite as belonging to the family Cryptosporiidae. Hyperplasia and heterophil infiltration were associated with the presence of organisms attached to the microvillus border of epithelial cells lining the plicae of the bursa of Fabricius. Although there were no clinical signs or gross lesions common to the 3 cases described, all had similar histologic lesions in the epithelium lining the bursa of Fabricius.     

Pathological lesions following an experimental intoxication with aflatoxin B1 in broiler chickens

A follow-up of chickens dosed orally over 21 days with 0.2 and 3 micrograms of aflatoxin B1 (AFB1) g-1 of bodyweight daily and their subsequent recovery 10 days after withdrawal of contaminated food was conducted. Vacuolation of liver cells during the initial days of the intoxication and cellular depletion in the follicle medulla of the bursa of Fabricius were the lesions which appeared first and persisted during the recovery phase in both groups of intoxicated animals. The intensity of these lesions and their persistence was related to the dose of aflatoxin ingested. A significant reduction in the bodyweight and absolute weights of liver, bursa of Fabricius spleen and thyroid was observed in the higher dose group.     

鸡传染性法氏囊病的病理学研究

人工接种２８日龄非免疫鸡传染性法氏囊病病毒（ＩＢＤＶ）后,对感染鸡的法氏囊、胸腺、脾、盲肠扁桃体、哈德氏腺、肝、肾进行病理组织学检查。感染后４８ｈ,法氏囊淋巴组织最早出现坏死且长久存在。其他淋巴器官的病变出现较迟,程度轻微且恢复较快。ＩＢＤＶ单抗免疫荧光检测,法氏囊及其他淋巴器官中均检测到病毒,接种后１２ｈ法氏囊中即检出病毒,持续时间也最长（攻毒后１２ｄ）,其次是盲肠扁桃体（攻毒后８ｄ）。攻毒１３ｄ以后,上述器官均未检测到病毒。法氏囊粘膜上皮的扫描电镜观察,攻毒后２ｄ,上皮细胞肿胀,微绒毛减少或消失。攻毒后３ｄ,局部上皮细胞坏死、脱落,并向整个粘膜层扩展,攻毒后１０ｄ,上皮层基本修复。     

Ghrelin免疫反应阳性细胞在鸡外周器官中的定位分布与发育性变化

本试验应用免疫组织化学方法和显微图像分析技术,研究了2、16、30、44、58日龄岭南黄鸡外周器官中Ghrelin免疫反应阳性细胞的定位分布与发育性变化。结果表明,在鸡腺胃、肠、心、肝、脾、肺、肾、胰、脑垂体、肾上腺、胸腺、法氏囊中均可观察到Ghrelin免疫阳性反应。其阳性细胞类型包括:腺胃腺小管的内分泌细胞,肠道黏膜上皮和肠腺内的内分泌细胞,肠道黏膜下层和肌层间的神经丛,心内膜下层的蒲肯野纤维,肝血窦中的枯否氏细胞,脾的巨噬细胞和网状细胞,肺的巨噬细胞,肾脏肾小囊壁层的扁平上皮细胞和脏层的足细胞、球内系膜细胞和球旁复合体,胰腺的胰岛细胞,腺垂体的部分嗜酸性细胞和嗜碱性细胞,肾上腺的嗜铬细胞,胸腺上皮细胞、胸腺小体和巨噬细胞,法氏囊黏膜上皮和小结相关上皮内的内分泌细胞、囊小结的网状细胞、巨噬细胞、皮质髓质交界处的上皮细胞等。2~30日龄鸡,随着日龄的增长,各器官中的Ghrelin表达量增加,44、58日龄则有所下降。     

Atrophy of the primary lymphoid organs of meat pigeons in Italy associated with circoviruslike particles in the bursa of Fabricius

During a survey effected in a meat pigeon slaughterhouse of central Italy, atrophy of primary lymphoid organs (bursa of Fabricius and thymus) and hypoplasia of bone marrow were observed. Histologic, ultrastructural, and hematologic examinations were performed on a total of 80 randomly selected 30-day-old meat pigeons. By histologic studies, lymphocytic depletion and necrosis with cyst formations in the bursa of Fabricius were detected in all subjects that showed thymus and bursa atrophy at necropsy. Basophilic intranuclear inclusions were also observed in bursal cells. After ultrastructural studies, these inclusions were proved to be viral particles resembling circoviruslike particles in morphology and size. Severe lymphocytic depletion of the bursa was plausibly associated with the presence of the viral particles.     

谷氨酰胺对肉鸡免疫器官胚后发育的影响

160只1日龄艾维茵肉仔鸡随机分成4组。分别饲喂添加0％、0．2％、0．4％和0．8％谷氨酰胺（Gln）饲粮28d。每周末取鸡24只，每组6只，颈静脉放血致死，取胸腺、法氏囊和脾脏，称重．制作切片．光镜观察，研究基础日粮中添加Gln对肉鸡免疫器官胚后发育的影响。结果显示：（1）添加0．8％Gln组1、4周龄和添加0．4％Gln组2、4周龄胸腺重量显著高于对照组（P〈0．05），添加0．8％Gln组2周龄极显著高于对照组（P〈0．01）；添加0．4％Gln组1、2、4周龄和添加0．8％Gln组4周龄法氏囊重量显著高于对照组（P〈0．05），添加0．8％Gln组1、2周龄极显著高于对照组（P〈0．01）；试验各组脾脏重量均高于对照组。添加0．8％Gln组4周龄显著高于对照组（P〈0．05）。（2）试验组胸腺小叶皮质增厚，淋巴细胞密集，髓质比例减小。胸腺小体减少、减小。试验组法氏囊皱襞内淋巴滤泡增多，皮质增厚．淋巴细胞增多。试验组脾脏脾小结增多、增大；动脉周围淋巴鞘增厚；椭球增多、增大．细胞排列疏松．鞘毛细血管管腔变大．内皮细胞增高。间隙增大。添加0．8％Gln对肉鸡免疫器官发育影响最为明显。结果表明，日粮中添加Gln对肉鸡中枢淋巴器官胚后发育的促进作用优于外周淋巴器官。并有延缓法氏囊退化作用。本试验从组织学角度证明Gln能够促进机体淋巴细胞的增殖分化及免疫器官的胚后发育，进而提高机体的免疫功能。     

B cell and macrophage response in chicks after oral administration of Salmonella typhimurium strains

Despite the fact that, in a number of countries, vaccination programmes are extensively used to control Salmonella infection in poultry, information on the immune mechanisms, especially the cellular response, is still needed. The aim of the study was to characterise the B cell and macrophage response in caecum (IgA+, IgM+, IgG+ cells, macrophages), bursa of Fabricius (IgM+ cells, macrophages), and spleen (IgM+ cells) of chicks after oral administration of a non-attenuated Salmonella (S.) typhimurium wild-type strain (infection) or an attenuated commercial live S. typhimurium vaccine strain (immunisation) to day-old chicks as compared to non-treated control birds using immunohistochemistry and image analysis. In caecum, higher counts of IgM-secreting cells were detected in infected animals compared with the controls from day 5 until day 12 of age. In contrast, in treated groups, IgA-secreting cells were found in higher numbers only between day 8 and 12 of age. Infected birds showed a higher number of IgA+ cells in spleen and bursa of Fabricius compared to the controls. In the bursa of Fabricius of immunised and infected birds, a depletion of strongly stained IgM+ cells and macrophages was established between day 5 and 9 indicating a possibly special and independent role of this organ during the immunological reaction against Salmonella organisms. The results suggest that IgM- and IgA-secreting cells are of importance in the caecal immune response of chickens against Salmonella strains. Immunised chickens always showed a weaker immune reaction compared to infected animals. Present findings regarding the B cell reaction within avian caeca prove a participation of both humoral and cellular immunity in defence against Salmonella strains. Immunohistochemical examination of the cellular response (B cells and macrophages) in relevant organs of chickens may be an important tool to evaluate the immunogenic characteristics of potential Salmonella live vaccine candidates.     

禽淋巴细胞性白血病的诊断与病毒亚群鉴定

采用病理组织学方法对某商品蛋鸡场送检的发病鸡进行实验室诊断,在肿胀的肝、脾、肾和法氏囊组织切片中观察到成淋巴细胞增生病变。使用禽白血病A、B亚群抗体检测试剂盒对血清样本进行间接ELISA试验,抗体阳性率达到44.4%(12/27)。采集12只病死鸡的肝脏材料提取DNA样本,11份样本中扩增出ALV-A亚群特异性的病原核酸(691bp),检出率达到91.6%(11/12)。将扩增的目的基因克隆与测序,截取gp85基因片段可变区序列与ALV-A、B、C、D和E亚群参考毒株的相应序列进行比较,同源性分别达到89.0-89.6%、67.1-67.7%、69.1-70.1%、69.1-69.5%和70.9-72.0%。结果证实本次疫病是由ALV-A亚群病毒引起的淋巴细胞性白血病。     

ChMDA5通路参与传染性法氏囊病病毒致雏鸡法氏囊损伤的研究

传染性法氏囊病病毒（IBDV）为dsRNA病毒,可造成雏鸡法氏囊损伤进而发生免疫抑制;MDA5是能够特异性识别dsRNA病毒的模式识别受体。为研究鸡MDA5（chMDA5）信号通路在IBDV致雏鸡法氏囊病理损伤中的作用,试验选取50只14日龄SPF雏鸡随机分为IBDV感染组和空白对照组,每组25只,IBDV感染组雏鸡通过点眼、滴鼻感染IBDV JIC7株病毒液,0.6 mL·只^-1,空白对照组雏鸡经相同途径给予相同剂量无菌PBS,感染IBDV后第1、4、7、21及35天采集雏鸡法氏囊。采用qRT-PCR方法检测法氏囊中IBDV载量,chMDA5及chMDA5信号通路衔接蛋白（chIPS-1）、转录因子（chIRF3和chNF-κB）、下游产物细胞因子（chIFN-β、chTNF-α、chIL-1β、chIL-6） mRNA水平变化;间接免疫荧光法检测chMDA5蛋白表达变化,传统病理学方法检查法氏囊病理组织学变化。结果发现,雏鸡感染IBDV后,其法氏囊中chMDA5、chIPS-1、chIRF3、chNF-κB、chIFN-β、chTNF-α、chIL-1β、chIL-6的表达量均显著高于对照组,且法氏囊组织发生形态损伤,上述变化趋势与IBDV载量变化基本一致。结果表明,雏鸡法氏囊chMDA5及其信号转导通路可被IBDV激活,参与到IBDV感染雏鸡法氏囊损伤与抗损伤过程中。     

鸡胚法氏囊的组织发育

本试验通过对孵化至１３－２１日龄的鸡胚法氏囊进行组织学的连续性动态观察,较为详细地描述了早日龄鸡胚法氏囊的组织学分化发育过程。实验结果显示,直至出壳胶鸡胚法氏囊尚未形成较完整的能组织起 体液免疫促进作用的组织基础结构,其完善的形态结构需在后天环境中分化和发育。     

Pathogenicity of recent isolates of infectious bursal disease virus in specific-pathogen-free chickens: protection conferred by an intermediate vaccine strain

The pathogenicity of recent isolates of infectious bursal disease virus and the protection conferred against them by a commercial vaccine strain of intermediate virulence were examined in specific-pathogen-free chickens. Based on clinical signs, mortality, and macroscopic lesions in susceptible chickens, the isolates designated as A-Delmarva and U-28 were distinct from a previously known serotype I virulent isolate (Edgar). Histopathological analysis of the bursa of Fabricius did not establish differences between the field isolates. Although the vaccine strain produced some degree of bursal damage in antibody-free chickens, it was significantly less severe than the damage caused by the field isolates. The active immune response induced by vaccination was cross-protective against the pathological effects produced by the different isolates used in this study.     

Morphology of the oviduct fluke,Prosthogonimus ovatus,isolated from Indonesian native chickens and histopathological observation of the infected chickens

Prosthogonimus ovatus infection was detected in 5 of 130 chickens in the oviduct and 4 chickens in the bursa of Fabricius. Scanning electron microscopy (SEM) revealed that the spines of the P. ovatus were densely distributed on the cuticula of the entire dorsal surface of body, but on the ventral surface, they were densely present to the level of ventral sucker but gradually decreased in density posteriorly, and they could not be seen in the posterior 1/3 area. The spines were finger-shaped and denticulate at the tip. Histopathological examination showed that polypous elevations, degeneration and exfoliation of the mucosal epithelium were detected in the bursa of Fabricius possibly by the suction of flukes, in addition to the stratification of the mucosal epithelium, and interstitial cell infiltration.     

Immunosuppressive potential and pathogenicity of a recent isolate of infectious bursal disease virus in commercial broiler chickens

At 15 days of age and in the presence of measurable levels of maternal antibody against infectious bursal disease virus serotype I (1:170 virus-neutralization geometric mean titer), a recent isolate (U-28) and a prototype virulent isolate (Edgar) of the same virus caused subclinical infections in commercial broiler chickens. Isolate U-28 caused a significant reduction in the size of the bursa of Fabricius, whereas the Edgar isolate produced splenomegaly. Both isolates reduced the serological response to Newcastle disease virus. The experimental immunosuppressive potential and pathogenicity of isolate U-28 in broiler chickens confirms the role of this virus in recent infectious bursal disease outbreaks.     

Gamma/delta T cell response of chickens after oral administration of attenuated and non-attenuated Salmonella typhimurium strains

Poultry represents an important source of Salmonella infection in man. Despite intensive research on immunity, little is known about the involvement of T cell sub-populations in the immunological response of chickens against infection with non-host-adapted Salmonella (S.) serovars. In this study, the T cell composition of blood lymphocytes (CD4(+)CD8(+); CD4(+)CD8(-); CD4(-)CD8(+); CD8(+)TcR1(+); CD8(-)TcR1(+), CD8(+)TcR1(-)) after oral administration of the non-attenuated S. typhimurium wild-type strain 421 (infection) or the attenuated vaccine strain Salmonella vac((R)) T (immunization) to day-old chicks was investigated and compared with non-treated chickens by flow cytofluorometry. Additionally, the occurrence of T cell sub-populations (CD4(+); CD8(+); TcR1(+)(gammadelta); TcR2(+)(alphabeta(1))) in ceca, spleen and bursa of Fabricius of the birds was studied immunohistologically. Blood samples and tissues were examined between days 1 and 12 of age.Chicks inoculated with S. typhimurium 421 or Salmonella vac((R)) T showed significantly elevated percentages of CD8(+)TcR1(+) in blood on days 7, 8 and 9, or on day 8 in comparison to control animals. The CD4 to CD8 cell ratio was about 3:1 in infected animals on day 5 of age. In the organs of treated chicks the numbers of CD8(+)(gammadelta) and TcR1(+)(gammadelta) cells had markedly increased on days 4 and 5 in ceca, 8 and 9 in the bursa and 9 and 12 in the spleen. Moreover, infected or vaccinated birds revealed larger quantities of CD4(+) and TcR2(+) T cells in ceca on days 4 and 5. As shown by double staining, the TcR1(+) cells in the organs of infected animals additionally carried the CD8 antigen.In conclusion, immunization of day-old chicks with the attenuated Salmonella live vaccine strain resulted in the same changes in T cell composition as seen after infection with the non-attenuated Salmonella wild-type strain, but at a lower level. The remarkable increase of CD8(+)TcR1(+)(gammadelta) double positive cells in treated birds indicates an important role of this cell sub-population in the immunological defense of chickens against Salmonella exposure.