Changes in Distribution and Numbers of CD4+ and CD8+ T‐lymphocytes in Lymphoid Tissues and Intestinal Mucosa in the Early Phase of Experimentally Induced Early Onset Mucosal Disease in Cattle

Mucosal disease (MD), one sequelae of bovine virus diarrhoea virus (BVDV) infection, causes severe lesions in lymphoid tissues and mucosal surfaces. Lesions are associated with the presence of cytopathogenic (cp) BVDV and initially characterized by apoptotic cell death. The objective of this investigation was to determine if this cell death is mediated only by the cp BVDV, which is known to induce apoptosis in cell culture or if immune‐mediated host reactions might also contribute. Early onset MD was experimentally induced in calves by inoculation of persistently viremic calves with a closely related cp BVDV. Calves were euthanized in the early phase of infection between days 5 and 13 post‐inoculation and tissues from tonsils, lymph nodes, Peyer's patches, jejunum and colon were collected. Presence of cp BVDV antigen was correlated with distribution of lymphocyte subpopulations in consecutive cryostat sections. In the lymphoid tissues, cp BVDV antigen was predominantly found in the lymphoid follicles. The increase of infected cells with time post‐inoculation was paralleled by a decrease of B‐lymphocytes and an increase of CD4+ T‐lymphocytes. An increased number of CD8+ T‐lymphocytes was seen in progressed lesions only. In the intestinal mucosa, initially multifocal, later diffuse infection with cp BVDV was accompanied by a multifocal or diffuse increase of CD4+ T‐lymphocytes, respectively. Numbers of IgA+ plasma cells and CD8+ T‐lymphocytes were decreased. The common change observed in lymphoid tissues and mucosa was the increase of CD4+ T‐lymphocytes in sites with lesions. This might indicate a cell‐mediated immune response to the cp BVDV. Besides their helper function to other cells of the immune system, activated CD4+ T‐lymphocytes might also exert cytotoxic activity, induce apoptosis in target cells via Fas/Fas ligand binding and thus contribute to the severity of tissue lesions in MD.     

Ultrastructural changes in follicles of small-intestinal aggregated lymphoid nodules in early and advanced phases of experimentally induced mucosal diseases in calves

OBJECTIVE: To investigate ultrastructural changes in follicles of small-intestinal aggregated lymphoid nodules (Peyer's patches) of calves with early and advanced phases of experimentally induced mucosal disease (MD). ANIMALS: Twenty 2.5- to 7-month-old Holstein-Friesian calves (11 females, 9 males). PROCEDURE: MD was induced in 13 of 18 calves that were persistently viremic with bovine viral diarrhea virus (BVDV). Eight of the 13 calves were euthanatized before the onset of clinical signs of MD, and 5 were euthanatized after becoming moribund with MD. Five persistently viremic calves and 2 calves without BVDV served as controls. Specimens of small-intestinal aggregated lymphoid nodules were prepared for transmission electron microscopy. RESULTS: The ultrastructure of follicles of small-intestinal aggregated lymphoid nodules from healthy calves was consistent with that in sheep. In the early phase of MD, changes were characterized by numerous apoptotic lymphocytes and macrophages with apoptotic bodies. In more advanced lesions, affected lymphoid follicles consisted of macrophages and variable numbers of follicular dendritic cells (FDC), whereas others did not contain FDC. In moribund calves, small follicles consisting predominantly of FDC and follicles with central cavities surrounded by macrophages, and few neutrophils were observed. CONCLUSIONS AND CLINICAL RELEVANCE: The ultrastructural changes in lymphoid follicles of small-intestinal aggregated lymphoid nodules indicate apoptosis of lymphocytes as an initial event. The development of small follicles consisting predominantly of FDC or the complete loss of follicular architecture in advanced phases of MD is determined by the intensity of apoptosis of lymphocytes, the capacity of the macrophages for uptake, and the reorganization of a stromal network.     

Distribution of viral antigen and tissue lesions in persistent and acute infection with the homologous strain of noncytopathic bovine viral diarrhea virus.

Viral distribution and lesions were compared between calves born with persistent infection (PI) and calves acutely infected with the same bovine viral diarrhea virus (BVDV) isolate. Two PI calves from 1 dairy herd were necropsied. The PI viruses from these calves were isolated, characterized by sequencing, and found to be identical. This virus strain, designated BVDV2-RS886, was characterized as a noncytopathic (ncp) type 2 BVDV. To establish acute infections, BVDV2-RS886 was used to inoculate clinically healthy, seronegative calves which were 3 weeks to 3 months old. Nine calves received 10(6)-10(7) tissue culture infective dose of BVDV2-RS886 intranasally. Four additional age-matched animals served as noninfected controls. Infected calves were necropsied at 3, 6, 9, or 13 days postinoculation (dpi). Viral antigen was detected by immunohistochemistry in frozen sections, and lesions were evaluated in hematoxylin eosin-stained paraplast sections. In the PI calves, a wide distribution of viral antigen was found in all tissues and was not associated with lesions. In the acutely infected calves, viral antigen was widespread in lymphoid tissues at 6 dpi but had been mostly eliminated at 9 and 13 dpi. Depletion of lymphoid tissues was seen at 6, 9, and 13 dpi and repopulation at 9 and 13 dpi. In 1 of the calves at 13 dpi, severe arteritis was present in lymph nodes and myocardium. This comparison shows that an ncp BVDV strain that causes no lesions in PI animals is able to induce marked depletion of lymphoid tissues in calves with acute infection. Therefore, the failure to eliminate PI cattle from a herd causes problems not only in pregnant cattle but may also affect other age groups.     

Distribution of viral antigen and development of lesions after experimental infection of calves with a BVDV 2 strain of low virulence.

To examine the virus-host interaction in subclinical bovine viral diarrhea virus (BVDV) infections, the spread of a BVDV 2 strain of low virulence to different organs and the development of lesions were investigated. Eight colostrum-deprived, clinically healthy, 2-3-month-old calves were intranasally inoculated with 10(6) tissue culture infective dose of the naturally occurring BVDV 2 strain 28508-5 of low virulence, and 2 served as controls. Two calves each were euthanized at days 3, 6, 9, and 13 postinoculation (pi). Representative tissues were processed for histology and immunohistology. Signs of overt clinical disease were absent. However, a mild temperature elevation at days 7 or 8 pi and a moderate decrease of circulating lymphocytes occurred in all inoculated calves. The BVDV antigen was detected at day 3 pi in several lymphoid tissues. At day 6 pi, BVDV antigen was found widespread in lymphoid tissues and multifocally in intestinal epithelial cells but was associated with no or subtle lesions only. At day 9 pi, much less BVDV antigen was detectable, but there was severe depletion of lymphoid tissues. At day 13 pi, BVDV antigen had been cleared from most lymphoid tissues that were at variable phases of depletion and recovery. In conclusion, the BVDV strain of low virulence spread to lymphoid tissues and intestinal epithelial cells but was rapidly eliminated. Transient depletion of lymphoid tissues was followed by recovery.     

Distribution of viral antigen and development of lesions after experimental infection with highly virulent bovine viral diarrhea virus type 2 in calves

OBJECTIVE: To correlate tissue distribution with development of lesions after experimental infection with a virulent strain of noncytopathic bovine viral diarrhea virus (BVDV) type 2 in calves. ANIMALS: Ten 14-day-old and two 2-month-old colostrum-deprived calves. PROCEDURE: Calves were intranasally inoculated with BVDV type-2 strain 1373 from an outbreak of clinically severe bovine viral diarrhea (BVD).Two 14-day-old calves served as noninfected controls. Two calves each were euthanatized on postinoculation days 3, 6, and 12, and 1 each on days 8, 9, 13, and 14. Tissues were collected for immunohistologic and histologic examination. RESULTS: Inoculated calves developed nonspecific clinical signs characterized by high fever and decreased numbers of leukocytes and thrombocytes. Viral antigen was detected focally in lymphoid tissues on day 3. On days 6, 8, 9, 12, and 14, viral antigen became increasingly widespread throughout organs and tissues. Viral antigen in lymphoid tissues was associated with severe depletion of all compartments. Lesions in other tissues were not well correlated with distribution of viral antigen. Depletion of lymphoid tissues was observed in a calf on day 13, but viral antigen had been cleared from most tissues and was detected in vascular walls only. CONCLUSIONS AND CLINICAL RELEVANCE: Infection with a virulent BVDV strain resulted in wide dissemination of viral antigen in host tissues. Severe lymphoid depletion developed in lymphoid tissues, whereas viral antigen was generally not associated with lesions in other tissues. Findings suggest that development of lesions in acute BVD is not solely a function of viral replication and is also attributable to host reaction to infection.     

Effect of infection with bovine viral diarrhea virus alone,bovine rotavirus alone,or concurrent infection with both on enteric disease in gnotobiotic neonatal calves

OBJECTIVE: To compare experimentally induced concurrent infection with bovine viral diarrhea virus (BVDV) and bovine rotavirus (BRV) with infection of either virus alone in calves. ANIMALS: Seventeen 1-day-old gnotobiotic calves. PROCEDURE: Calves were allotted to 8 treatments as follows: group 1, mock-infected control calves (n = 2); group 2, inoculated with BVDV on day 1 (2); groups 3, 5, and 7, inoculated with BRV on days 1 (2), 4 (1), or 7 (2), respectively; and groups 4, 6, and 8, inoculated with BVDV on day 1 and with BRV on days 1 (2), 4 (2), or 7 (4), respectively. Concentrations of BVDV in serum and ileal tissues were measured, and BRV shedding in feces was determined. Histologic examination and immunohistochemical analysis were conducted to detect lesions and viral antigens. RESULTS: Neonatal calves inoculated with BVDV alone or with BVDV on day 1 and BRV on day 7 developed villus atrophy and submucosal inflammation of the intestines. Concurrent BVDV and BRV infections acted synergistically in the intestinal tract, causing more severe enteric disease than infection with either virus alone. Severe lymphoid depletion was associated with BVDV infection in calves regardlesss of concurrent BRV infection. CONCLUSIONS AND CLINICAL RELEVANCE: Infection with BVDV played direct and indirect roles in enteritis in neonatal calves, causing villus atrophy in the duodenum and submucosal inflammation of the intestines. Also, BVDV potentiated effects of BRV. Concurrent infection with BVDV and BRV resulted in more severe enteric disease in neonatal calves than infection with BRV or BVDV alone.     

Experimental infection of calves with bovine viral diarrhea virus genotype II (NY-93).

To ascertain the virulence of bovine viral diarrhea virus (BVDV) genotype II, isolate NY-93 was inoculated intranasally into 3 calves, 2 of which were treated with a synthetic glucocorticoid prior to and after virus inoculation. Anorexia, fever (up to 42 C), dyspnea, and hemorrhagic diarrhea developed 6 days after intranasal inoculation with BVDV NY-93. The condition of all calves deteriorated further until the end of the study on day 14 postinoculation. The most significant postmortem macroscopic changes in all calves were limited to the gastrointestinal tract and consisted of moderate to severe congestion of the mucosa with multifocal hemorrhages. Microscopic lesions found in the gastrointestinal tract were similar to those observed in mucosal disease, including degeneration and necrosis of crypt epithelium and necrosis of lymphoid tissue throughout the ileum, colon, and rectum. The basal stratum of the epithelium of tongue, esophagus, and rumen had scattered individual necrotic cells. Spleen and lymph nodes had lymphocytolysis and severe lymphoid depletion. Severe acute fibrinous bronchopneumonia was present in dexamethasone-treated calves. Abundant viral antigen was detected by immunohistochemistry in the squamous epithelium of tongue, esophagus, and forestomachs. BVDV antigen was prominent in cells of the media of small arteries and endothelial cells. The presence of infectious virus in tissues correlated with an absence of circulating neutralizing antibodies. These findings highlight the potential of BVDV genotype II to cause severe disease in normal and stressed cattle.     

Bronchoalveolar immune defense in cattle exposed to primary and secondary challenge with bovine viral diarrhea virus

To evaluate immune defense mechanisms against bovine viral diarrhea virus (BVDV), four calves received primary and secondary intrabronchial infections with the cytopathic, type I Singer strain of BVDV. The cellular and humoral responses to these site-specific infections with BVDV were monitored by sequential bronchoalveolar lavages (BAL) conducted prior to infection (day 0, non-infected controls) and on days 4, 7, 10, 17 (day 31, secondary infection), 35, 38, 41, 48 and 62 post-infection. Peak quantities of BVDV were recovered from BAL on day 4. BVDV clearance from the lung was complete between days 17 and 31. Immune clearance of BVDV from the lower airways upon secondary infection was swift, within 4 days, and sustained throughout a 1-month period. Total numbers of BAL CD4(+) and CD8(+) T-lymphocytes increased >200-fold by day 10, and increased to levels >70-fold higher than background by 4 days after a secondary BVDV infection. gammadelta(+) T-lymphocytes increased 100-fold by day 7 and remained at levels at least 10-fold higher than pre-infection throughout the study. B-lymphocytes increased to levels 30-fold greater than pre-infection levels by day 10, and further increased to levels 100-fold higher following secondary BVDV infection. Activation (defined by the phenotype CD25(+)/CD62L(-)) and memory (defined by the phenotype CD45RO(+)/CD45R(-)) profiles of lymphocytes in the lower airways were characterized. Activated subpopulations of CD4(+) and CD8(+) cells increased nearly 300- and 150-fold, respectively, by day 10, and to levels 100- and 50-fold 4 days after the secondary infection. Memory subpopulations of CD4(+) and CD8(+) cells increased to levels 170- and 120-fold, respectively, by day 10, and to levels approximately 400- and 300-fold, respectively, 7 days after the secondary infection. The primary antibody response consisted of increased titers of anti-BVDV-specific IgA in bronchoalveolar lavage fluid (BALF). A strong secondary antibody response with high levels of anti-BVDV-specific IgA and IgG in BALF before day 4 post-secondary BVDV infection, likely contributed, along with cellular defenses, to the rapid clearance of virus from the lung upon secondary exposure. These results demonstrate that primary infection of the bovine lung with BVDV initiates cell-mediated immune responses capable of clearing the virus within 2-3 weeks. Furthermore, populations of immune-activated and memory T-lymphocytes, combined with BVDV-specific antibody production, contribute to rapid BVDV clearance upon secondary exposure to the virus.     

BVD virus antigens in tissues of persistently viraemic, clinically normal cattle: implications for the pathogenesis of clinically fatal disease

The cellular events involved in precipitation of the clinically fatal outcome of an infection with bovine viral diarrhoea virus (BVDV) remain unresolved, though it is now known that this course of the infection, Mucosal Disease (MD), only occurs in calves persistently infected with non-cytopathic BVDV. In studies aimed at elucidating the pathogenesis of MD, the distribution of BVDV antigens and infectious virus in tissues of persistently infected, clinically normal calves was investigated. Virus antigen was detected in most tissues, in epithelial and immune cells. No signs of an inflammatory response were detected and cytopathological changes were subtle or absent. The infection may nevertheless create a cell-environment which will enhance replication of cytopathic virus. Variations in the clinical, pathomorphologies and virological appearance of MD-cases may depend on both the host-reactions, including virus-induced immunopathology, and the virus-strain combinations in a putative mixed infection.     

Comparative investigation of tissue alterations and distribution of BVD-viral antigen in cattle with early onset versus late onset mucosal disease

Tissue alterations and distribution of BVDV antigen were examined in nine cattle with early onset and five cattle with late onset mucosal disease (MD) to evaluate the possibility to differentiate between the two disease entities. MD was induced by inoculation of persistently viremic cattle with different strains of cytopathogenic BVDV. Animals which developed early onset MD became moribund approximately 2 weeks post-inoculation (pi); animals with late onset MD 42-115 days pi. All animals were euthanized and necropsied when moribund. Macroscopic lesions were found in the upper and lower digestive tract of cattle with early and late onset MD. In cattle with late onset MD, lesions in the oral cavity were generally milder and in the intestinal tract they were not only associated with GALT, but frequently affected the mucosa outside. Histologically, the abrupt changes between hyperplastic and atrophic areas of mucosa were striking in the cattle with late onset MD. This corresponded with the multifocal distribution of areas of mucosa in which intense staining for BVD-virus antigen could be demonstrated. In both courses of MD, a severe depletion of Peyer's patches was noted, but only in late onset MD, there was a complete loss of architecture. The most distinctive difference was the presence of vascular lesions which were observed in all five cattle with late onset MD, but in none of the animals with early onset MD. The vasculopathy was characterized by segmental necrosis of vascular walls and lymphohistiocytic perivasculitis in arterioles and small arteries in the submucosa of the intestine.     

牛病毒性腹泻病毒感染牛外周血单核细胞对CD80和CD86 mRNA转录的影响

用非致细胞病变（noncytopathic,NCP）和致细胞病变（cytopathic,CP）型牛病毒性腹泻病毒（BVDV）感染临床健康BVDV检测阴性的荷斯坦奶牛外周血单核细胞（PBMC）,利用实时荧光定量PCR技术对感染后共刺激分子CD80和CD86mRNA转录水平的变化进行定量分析。结果表明,在NCP型BVDV感染牛PBMC后CD80在4h（P〈0.05）和12,24h（P〈0.01）出现2次转录高峰,CD86在6h（P〈0.05）出现转录高峰;CP型BVDV感染后,CD80在24h（P〈0.05）出现转录高峰,CD86在6h（P〈0.05）出现转录高峰。尽管CD80在NCP型BVDV感染后呈现较复杂的动态变化,但结果提示NCP型和CP型BVDV感染均可导致牛PBMC的共刺激分子CD80和CD86基因转录在感染早期明显受到抑制,PBMC的抗原呈递能力受到影响。     

Malignant catarrhal fever induced by Alcelaphine herpesvirus 1 is characterized by an expansion of activated CD3+CD8+CD4- T cells expressing a cytotoxic phenotype in both lymphoid and non-lymphoid tissues

ABSTRACT: Alcelaphine herpesvirus 1 (AlHV-1) is carried by wildebeest asymptomatically. It causes a fatal lymphoproliferative disease named wildebeest-derived malignant catarrhal fever (WD-MCF) when cross-species transmitted to a variety of susceptible species of the Artiodactyla order. WD-MCF can be reproduced experimentally in rabbits. In a previous report, we demonstrated that WD-MCF induced by AlHV-1 is associated with a severe proliferation of CD8+ T cells in the lymphoid tissues. Here, we further studied the mononuclear leukocytic populations in both the lymphoid (throughout the infection and at time of euthanasia) and non-lymphoid (at time of euthanasia) organs during WD-MCF induced experimentally in rabbits. To reach that goal, we performed multi-colour flow cytometry stainings. The results obtained demonstrate that the development of WD-MCF correlates in peripheral blood with a severe increase of CD8+ cell percentages; and that CD3+CD8+CD4- T cells were the predominant cell type in both lymphoid and non-lymphoid organs at time of euthanasia. Further characterization of the mononuclear leukocytes isolated from both lymphoid and non-lymphoid tissues revealed that the CD8+ T cells express high levels of the activation markers CD25 and CD44, produce high amount of gamma-interferon (IFN-γ) and perforin, and showed a reduction of interleukin-2 (IL-2) gene expression. These data demonstrate that the development of WD-MCF is associated with the expansion and infiltration of activated and cytotoxic CD3+CD8+CD4- T cells secreting high amount of IFN-γ but low IL-2.     

传染性法氏囊病病毒超强毒感染SPF鸡免疫器官中CD4+和CD8+T淋巴细胞动态分布

利用免疫组织化学染色对传染性法氏囊病病毒（IBDV）超强毒LX株感染SPF鸡免疫器官中CD4^ 和CD8^ T淋巴细胞的动态分布进行了研究。超强毒LX株接种2周龄SPF雏鸡，在其法氏囊、脾脏、胸腺、盲肠扁桃体、骨髓和哈氏腺中均可检出IBDV抗原的存在和CD4^ 与CD8^ T淋巴细胞的数量改变。在法氏囊中，CD4^ 淋巴细胞主要存在于淋巴滤泡间隙和滤泡皮质，而CD8^ T淋巴细胞则丰在于整个淋巴滤泡和滤泡间隙，并且CD8^ T淋巴细胞数量明显多于CD4^ T淋巴细胞，在接种后14d仍未见CD4^ 和CD8^ T淋巴细胞数量减少。脾脏中CD4^ T淋巴细胞主要存在于外周小动脉淋巴鞘或散在，而CD8^ T淋巴细胞则多存在于外周小动脉淋巴鞘和红髓。接种后胸腺中CD4^ 和CD8^ T淋巴细胞在皮质中减少，但在髓质增多，尤其是CD8^ T淋巴细胞数明显多于CD4^＋T淋巴细胞。盲肠扁桃体中CD4^ 和CD8^ T淋巴细胞主要存在于发生中心，尤其是CD8^ T淋巴细胞数比CD4^ T淋巴细胞明显多。骨髓和哈氏腺中也可见CD4^ 和CD8^ T淋巴细胞，而且CD8^ T淋巴细胞更多。在这些淋巴器官中，病毒损伤部位出现CD4^ 和CD8^ T淋巴细胞的迁入聚集，表明T淋巴细胞可能参与IBDV超强毒的免疫致病过程。     

Characterization of protection from systemic infection and disease by use of a modified-live noncytopathic bovine viral diarrhea virus type 1 vaccine in experimentally infected calves

OBJECTIVE: To evaluate protection against systemic infection and clinical disease provided by use of a modified-live noncytopathic bovine viral diarrhea virus (BVDV) type 1 vaccine in calves challenged with NY-1 BVDV. ANIMALS: 10 calves, 5 to 7 months of age. PROCEDURES: Calves were allocated (n = 5/group) to be nonvaccinated or vaccinated SC on day 0 with BVDV type 1 (WRL strain). Calves in both groups were challenged intranasally with NY-1 BVDV on day 21. Calves' rectal temperatures and clinical signs of disease were recorded daily, total and differential WBC and platelet counts were performed, and serum neutralizing antibody titers against NY-1 BVDV were determined. Histologic examinations and immunohistochemical analyses to detect gross lesions and distribution of viral antigens, respectively, were performed. RESULTS: After challenge exposure to NY-1 BVDV, nonvaccinated calves developed high rectal temperatures, increased respiratory rates, viremia, leukopenia, lymphopenia, and infection of the thymus. Vaccinated calves did not develop high rectal temperatures or clinical signs of respiratory tract disease. Vaccinated calves appeared to be protected against systemic replication of virus in that they did not develop leukopenia, lymphopenia, viremia, or infection of target organs, and infectious virus was not detected in peripheral blood mononuclear cells or the thymus. CONCLUSIONS AND CLINICAL RELEVANCE: The modified-live BVDV vaccine protected calves against systemic infection and disease after experimental challenge exposure with NY-1 BVDV. The vaccine protected calves against infection and viremia and prevented infection of target lymphoid cells.     

Characterization of protection against systemic infection and disease from experimental bovine viral diarrhea virus type 2 infection by use of a modified-live noncytopathic type 1 vaccine in calves

OBJECTIVE: To evaluate protection resulting from use of a modified-live noncytopathic bovine viral diarrhea virus (BVDV) type 1 vaccine against systemic infection and clinical disease in calves challenged with type 2 BVDV. ANIMALS: 10 calves, 5 to 7 months of age. PROCEDURES: Calves were allocated (n = 5/group) to be nonvaccinated or vaccinated SC on day 0 with BVDV 1 (WRL strain). Calves in both groups were challenged intranasally with BVDV type 2 isolate 890 on day 21. Rectal temperatures and clinical signs of disease were recorded daily, and total and differential WBC and platelet counts were performed. Histologic examinations and immunohistochemical analyses to detect lesions and distribution of viral antigens, respectively, were performed. RESULTS: After challenge exposure to BVDV type 2, nonvaccinated calves developed high rectal temperatures, increased respiratory rates, viremia, leukopenia, lymphopenia, and infection of the thymus. Vaccinated calves did not develop high rectal temperatures or clinical signs of respiratory tract disease. Vaccinated calves appeared to be protected against systemic replication of virus in that they did not develop leukopenia, lymphopenia, viremia, or infection of target organs, and infectious virus was not detected in peripheral blood mononuclear cells or the thymus. CONCLUSIONS AND CLINICAL RELEVANCE: The modified-live BVDV type 1 vaccine protected against systemic infection and disease after experimental challenge exposure with BVDV type 2. The vaccine protected calves against infection and viremia and prevented infection of target lymphoid cells.     

Comparative virulence of isolates of bovine viral diarrhea virus type II in experimentally inoculated six- to nine-month-old calves

OBJECTIVE: To determine the comparative virulence of 5 isolates of bovine viral diarrhea virus (BVDV) type II by inoculating 6- to 9-month-old beef calves with isolates originating from the tissues of cattle affected with naturally occurring, transient, acute, nonfatal infections or naturally occurring, peracute, fatal infections. ANIMALS: 22 calves that were 6 to 9 months old. PROCEDURE: The study used BVDV isolates 17011, 713, and 5521 that originated from fetuses aborted from cows with transient, nonfatal, acute BVDV infections and isolates 23025 and 17583 that originated from the tissues of cattle with peracute, fatal BVDV infections. Calves were allotted to 6 groups (1, mock-infected control calves [n = 2]; 2, inoculated with BVDV 17011 [4]; 3, inoculated with BVDV 713 [4]; 4, inoculated with BVDV 5521 [4]; 5, inoculated with BVDV 23025 [4]; and 6, inoculated with BVDV 17583 [41]. Rectal temperatures and clinical signs of disease were recorded daily. Total and differential WBC and platelet counts were performed. Histologic examination and immunohistochemical analysis were conducted to detect lesions and distribution of viral antigens, respectively. RESULTS: Calves inoculated with BVDV 23025 or 17583 developed more severe clinical signs of disease (fever and diarrhea), more severe lymphopenia, and more severe lesions (alimentary epithelial necrosis, lymphoid depletion, and BVDV antigen deposition in lymphatic tissues), compared with calves inoculated with BVDV 713, 5521, or 17011. CONCLUSIONS AND CLINICAL RELEVANCE: Relative severity of experimentally induced infections corresponded to severity of clinical signs of naturally occurring infections with respective BVDV isolates.     

Differential cytokine gene expression in CD4+ and CD8+ T cell subsets of calves

The distinct patterns of cytokine expression in CD4+ and CD8+ T cells are well understood in mice and humans. However, little information is available about cytokine expression in bovine CD4+ and CD8+ T cells. In this study, mRNA expression of 19 different cytokines was analyzed in CD4+ and CD8+ T cells of calves with or without Concanavalin A (Con A) stimulation. CD4+ and CD8+ T cell populations were enriched to 98% purity by positive selection using magnetic cell sorting (MACS). CD4+ T cells spontaneously expressed the mRNAs of interleukin-1alpha (IL-1alpha), IL-1beta, IL-2, IL-6, IL-7, IL-8, IL-10, IL-18, IFN-gamma, TNF-alpha, TNF-beta and TGF-beta, and augmented the mRNA expression of IL-10, IFN-gamma and TNF-beta after Con A stimulation. The mRNAs of IL-3, IL-4, IL-5, IL-13 and GM-CSF were newly expressed in Con A-stimulated CD4+ T cells. CD8+ T cells displayed spontaneous mRNA expression of IL-6, IL-18, TNF-alpha, TNF-beta and TGF-beta, and newly expressed the mRNA of IL-2, IL-7, interferon-gamma (IFN-gamma) and GM-CSF after Con A stimulation. It was found that CD4+ T cells expressed the mRNA of 17 cytokines except for IL-12 and IL-15, while CD8+ T cells expressed only the mRNA of 9 cytokines after Con A stimulation. The profile of cytokine mRNA expression was substantially different in the CD4+ and CD8+ T cells of calves, indicating that CD4+ T cells can be distinguished from CD8+ T cells by the cytokine gene expression of IL-1alpha, IL-1beta, IL-3, IL-4, IL-5, IL-8, IL-10 and IL-13. Differential cytokine expression between CD4+ and CD8+ T cells serve to interpret an individual function of T cell subsets in the immune system of calves.     

Distribution of bovine virus diarrhoea viral antigens in the central nervous system of cattle with various congenital manifestations.

Distribution of bovine viral diarrhoea virus (BVDV) antigens in the central nervous system (CNS) of 26 cattle persistently BVDV infected, 11 cattle with mucosal disease (MD), and 32 calves with congenital brain malformations was studied using monoclonal antibodies against BVDV epitopes. In persistently infected cattle and in cattle with MD, a widespread infection of neurons was present. Predilection sites for BVDV antigens were the cerebral cortex and the hippocampus. In calves with congenital encephalopathies, viral antigen-containing neurons could only be detected in the CNS of four animals. From the topographical distribution of BVDV antigens in these four postnatal cases with end-stage lesions, no conclusions could be drawn concerning the pathogenesis of BVDV-induced encephalopathies.     

Hepatic immune response in calves during acute subclinical infection with bovine viral diarrhoea virus type 1

Eight colostrum-deprived calves aged 8-12 weeks were inoculated intranasally with a non-cytopathic strain of bovine viral diarrhoea virus (BVDV) genotype-1 and the effects on the hepatic immune response were studied. Two calves were sacrificed at each of 3, 6, 9 and 14 days post-inoculation (dpi) and two uninoculated animals were used as negative controls. BVDV was detected in hepatic macrophages and monocytes from 3 to 14dpi and in Küpffer cells (KCs) from 6 to 14dpi. Increases in the numbers of MAC387(+) KCs and monocytes, but not interstitial macrophages, differentiated by morphological features, were evident in the liver following inoculation with BVDV. There was a substantial increase in the number of monocytes positive for tumour necrosis factor (TNF)-α, but only small increases in the numbers of TNF-α(+) KCs and interstitial macrophages and interleukin (IL)-6(+) monocytes, KCs and interstitial macrophages. There was an increase in the number of interstitial CD3(+) T lymphocytes in the liver, but no substantial changes in the numbers of circulating CD3(+) T lymphocytes, interstitial or circulating CD4(+) or CD8(+) T lymphocytes, or CD79αcy(+) B lymphocytes. Serum haptoglobin and serum amyloid A increased transiently at 12dpi. Upregulation of some pro-inflammatory cytokines by hepatic macrophages is evident in subclinical acute BVDV type 1 infection in calves.