An estimate of specificity for a Johne's disease absorbed ELISA in northern Australian cattle

OBJECTIVE: To estimate the specificity of an absorbed enzyme-linked immunosorbent assay kit for Johne's disease (JD) when used in mature cattle populations resident in northern Australia. DESIGN: Blood samples were collected from beef cattle in northern Queensland, the Northern Territory and northern Western Australia, and from dairy cattle in northern Queensland. The specificity of a serological test for JD was estimated by testing the blood samples with an absorbed ELISA kit. Further samples were collected from cattle with positive ELISA results to determine the presence or absence of infection with Mycobacterium avium subsp paratuberculosis. PROCEDURE: During 1995 and 1996, blood, tissue and gut contents were collected from beef cattle at abattoirs in Queensland and the Northern Territory; and blood and faecal samples were collected from dairy cattle in herds assessed to be most at risk for JD in northern Queensland. The blood samples were tested using an absorbed ELISA kit. Tissues and gut contents from beef cattle that had positive ELISA results were cultured for M. avium subsp paratuberculosis, and tissues were examined histologically. Faecal samples from dairy cattle with positive ELISA results were cultured for M. avium subsp paratuberculosis. RESULTS: Estimates of specificity for this absorbed ELISA in mature northern Australian cattle were 98.0% (97.0 to 98.8%, 95% CI) in beef cattle, and 98.3% (96.7 to 99.3%, 95% CI) in dairy cattle. CONCLUSION: Estimates of specificity in this study were lower for beef cattle from the Northern Territory and northern Western Australia and for dairy cattle from northern Queensland than those quoted from studies on cattle in southern Western Australia. This should be considered when serological testing using the JD ELISA is carried out on northern Australian cattle.     

An estimate of specificity for a Johne's disease absorbed ELISA in northern Australian cattle

Objective To estimate the specificity of an absorbed enzyme-linked immuno-sorbent assay kit^d for Johne's disease (JD) when used in mature cattle populations resident in northern Australia.
Design Blood samples were collected from beef cattle in northern Queensland, the Northern Territory and northern Western Australia, and from dairy cattle in northern Queensland. The specificity of a serological test for JD was estimated by testing the blood samples with an absorbed ELISA kit. Further samples were collected from cattle with positive ELISA results to determine the presence or absence of infection with Mycobacterium avium subsp paratuberculosis .
Procedure During 1995 and 1996, blood, tissue and gut contents were collected from beef cattle at abattoirs in Queensland and the Northern Territory; and blood and faecal samples were collected from dairy cattle in herds assessed to be most at risk for JD in northern Queensland. The blood samples were tested using an absorbed ELISA kit. Tissues and gut contents from beef cattle that had positive ELISA results were cultured for M avium subsp paratuberculosis , and tissues were examined histo-logically. Faecal samples from dairy cattle with positive ELISA results were cultured for M avium subsp paratuberculosis .
Results Estimates of specificity for this absorbed ELISA in mature northern Australian cattle were 98.0% (97.0 to 98.8%, 95% CI) in beef cattle, and 98.3% (96.7 to 99.3%, 95% CI) in dairy cattle.
Conclusion Estimates of specificity in this study were lower for beef cattle from the Northern Territory and northern Western Australia and for dairy cattle from northern Queensland than those quoted from studies on cattle in southern Western Australia. This should be considered when serological testing using the JD ELISA is carried out on northern Australian cattle.     

In vitro erythrophagocytosis by cultured macrophages stimulated with extraneous substances and those isolated from the blood, spleen and bone marrow of Boran and N'Dama cattle infected with Trypanosoma congolense and Trypanosoma vivax

A standard radioactive chromium (51Cr) release assay was used to assess the in vitro phagocytosis and lysis of bovine erythrocytes by cultured splenic, bone marrow and peripheral blood monocyte-derived (PBM) macrophages isolated from healthy and Trypanosoma congolense and T. vivax-infected cattle of the Boran and N'Dama breeds. Recombinant cytokines (rHuTNF-alpha and rBolFN-gamma) and non-acid-dialysed peripheral blood mononuclear cell (PBMNC) culture supernatants stimulated these PBM for enhanced activities. The stimulants caused increases in the rate of erythrocyte phagocytosis and lysis by cultured PBM in a concentration-dependent manner. But very high stimulant concentrations caused deceased in vitro erythrophagocytosis. However, bacterial lipopolysaccharide (LPS) and acid-dialysed PBMNC culture supernatants did not cause any increase in cultured PBM erythrophagocytosis. In vitro erythrocyte phagocytosis and lysis by splenic, bone marrow and peripheral blood monocyte (PBM)-derived macrophages of Boran breed of cattle infected with Trypanosoma congolense increased from 14 days post-infection (DPI) onwards and thereafter maintained at various levels above pre-infection. Cultured splenic macrophages showed the greatest erythrocyte destruction capability while PBM-derived macrophages was the least. The rates of in vitro erythrocyte phagocytosis and lysis were higher with the cultured PBM of the Boran than those of the N'Dama cattle during T. congolense infection. The rate of in vitro erythrocyte destruction was however, similar in both groups of cattle during T. vivax infection. These results correlated positively with the dynamics and degree of anaemia developed by these groups of animals during both T. congolense and T. vivax infections. Cattle infected with T. congolense and T. vivax developed varying degrees of normocytic normochromic anaemia during infection. Boran cattle developed a more severe anaemia, and had to be treated with diminazine aceturate, than N'Dama cattle during T. congolense infection. Both breeds of cattle developed a milder but similar degree of anaemia during T. vivax infection. None of the animals were treated. The results of this study indicated a role of in vivo macrophage stimulatory factors, notably cytokines such as TNF-alpha and IFN-gamma in host's serum, as well as parasite antigens, which may act singly or in concert, in the process of enhanced erythrocyte destruction, hence anaemia by the mononuclear phagocytic system (MPS) during bovine trypanosomosis.     

Experimental production of fatal mucosal disease in cattle

Three outbreaks of mucosal disease were investigated. Careful examination of 47 cattle that were persistently viraemic with bovine virus diarrhoea virus (BVDV) revealed no clinical disease, no or low levels of BVDV antibody and only non-cytopathic virus in their blood. The four animals with mucosal disease all showed clinical disease and both cytopathic and non-cytopathic virus in their blood. Following post mortem examination, there were particularly high levels of cytopathic virus in gut tissue. A hypothesis for the induction of mucosal disease is suggested. It states that animals become persistently infected with non-cytopathic virus following in utero infection and when, in post natal life, they become superinfected with a cytopathic virus, then mucosal disease ensues. The experimental reproduction of mucosal disease in support of this hypothesis is described.     

Cattle remain immunocompetent during the acute phase of foot-and-mouth disease virus infection

ABSTRACT: Infection of cattle with foot-and-mouth disease virus (FMDV) results in the development of long-term protective antibody responses. In contrast, inactivated antigen vaccines fail to induce long-term protective immunity. Differences between susceptible species have also been observed during infection with FMDV, with cattle often developing persistent infections whilst pigs develop more severe symptoms and excrete higher levels of virus. This study examined the early immune response to FMDV in na?ve cattle after in-contact challenge. Cattle exposed to FMDV were found to be viraemic and produced neutralising antibody, consistent with previous reports. In contrast to previous studies in pigs these cattle did not develop leucopenia, and the proliferative responses of peripheral blood mononuclear cells to either mitogen or third party antigen were not suppressed. Low levels of type 1 interferon and IL-10 were detected in the circulation. Taken together, these results suggest that there was no generalised immunosuppression during the acute phase of FMDV infection in cattle.     

Immune responses of infected and vaccinated Hereford cattle to antigens of the cattle tick, Boophilus microplus

Responses of infested and vaccinated Hereford cattle to Boophilus microplus antigens were measured by enzyme-linked immunosorbent assay (ELISA), lymphocyte blastogenesis assay (LBA) and intradermal skin tests. Responses against soluble salivary gland extracts (SGS), salivary gland membrane (SGM), soluble gut extracts (GS), gut membrane (GM), soluble larval extracts (LS) and larval membrane (LM) antigens were tested. In one experiment, cattle infested with up to 160,000 ticks had positive cellular responses to SGS and significant antibodies against LM, GM, SGM, and SGS. Cellular responses to Concanavalin A were not depressed following infestation. Cattle vaccinated with GM, using Quil A as adjuvant, had positive cellular responses to gut and salivary gland antigens and significant antibody responses to all antigens tested. The antibody levels of vaccinated cattle were significantly higher than the antibody levels of infested cattle (P less than 0.05). In a second experiment, immune responses of cattle infested with 40,000 ticks were studied during 38 days. Cellular responses in LBA to several tick antigens were transiently elevated and significant levels of antibody were measured against LM, GM, SGM and SGS, from day 25 (P less than 0.05). Infested cattle had positive skin reactions following intradermal injection of larval and adult tick antigens (P less than 0.05).     

Accelerated binding of autoantibody to red blood cells with increasing anaemia in cattle experimentally infected with Theileria sergenti

Anaemia is the most important clinical manifestation in cattle infected with Theileria sergenti. In order to determine the mechanism of red blood cells (RBC) destruction in anaemic cattle, we studied the binding of autoantibody (IgG) to RBC during the development of anaemia in T. sergenti infection. The low levels of IgG-bound RBC before the development of anaemia were triggered in proportion with the progression of anaemia and parasitaemia. Our results suggest an accelerated destruction of RBC in anaemic cattle by IgG-dependent phagocytosis.     

Blockade of bovine PD-1 increases T cell function and inhibits bovine leukemia virus expression in B cells in vitro

Programmed death-1 (PD-1) is a known immunoinhibitory receptor that contributes to immune evasion of various tumor cells and pathogens causing chronic infection, such as bovine leukemia virus (BLV) infection. First, in this study, to establish a method for the expression and functional analysis of bovine PD-1, hybridomas producing monoclonal antibodies (mAb) specific for bovine PD-1 were established. Treatment with these anti-PD-1 mAb enhanced interferon-gamma (IFN-γ) production of bovine peripheral blood mononuclear cells (PBMC). Next, to examine whether PD-1 blockade by anti-PD-1 mAb could upregulate the immune reaction during chronic infection, the expression and functional analysis of PD-1 in PBMC isolated from BLV-infected cattle with or without lymphoma were performed using anti-PD-1 mAb. The frequencies of both PD-1⁺ CD4⁺ T cells in blood and lymph node and PD-1⁺ CD8⁺ T cells in lymph node were higher in BLV-infected cattle with lymphoma than those without lymphoma or control uninfected cattle. PD-1 blockade enhanced IFN-γ production and proliferation and reduced BLV-gp51 expression and B-cell activation in PBMC from BLV-infected cattle in response to BLV-gp51 peptide mixture. These data show that anti-bovine PD-1 mAb could provide a new therapy to control BLV infection via upregulation of immune response.     

Boophilus microplus: passage of bovine immunoglobulins and albumin across the gut of cattle ticks feeding on normal or vaccinated cattle

Cattle were vaccinated with antigens from adult female Boophilus microplus and haemolymph was collected from female ticks which had engorged on these animals and on matched control cattle. Radio-immunoassay for bovine plasma proteins in haemolymph from ticks fed on control cattle showed low concentrations of IgG1 and albumin. There was a significant increase in bovine plasma proteins passing across the gut in ticks fed on vaccinated cattle, with an average of 150 times more albumin and four to five times more IgG1 in the haemolymph. Ticks with obviously damaged gut had the highest concentrations of bovine plasma proteins but apparently undamaged ticks from vaccinated cattle also had elevated protein concentrations.     

Comparative studies on N''Dama and zebu cattle following repeated infections with Trypanosoma congolense

Twenty N'Dama and eight zebu cattle were inoculated intradermally with bloodstream forms of a cloned strain of Trypanosoma congolense originating from East Africa. All inoculated cattle became parasitaemic. Zebus showed consistently higher levels of parasitaemia and lower packed red cell volume (PCV) percentages than did N'Damas. Three of the eight zebus required treatment when high numbers of trypanosomes were present in the blood and PCV values dropped below 15 per cent. None of the N'Dama cattle needed treatment. Statistical analysis was performed on the data to assess the variability of parasitaemia and PCV levels before and during infection of the N'Dama cattle. The variation in PCV values was large between individuals during the early stages of the disease and diminished as infection continued. After trypanocidal drug treatment and a recovery period of 14 months, the same animals were inoculated intradermally with T congolense bloodstream forms isolated and cloned in The Gambia. Differences in susceptibility to the ensuing disease were apparent when comparing N'Dama and zebu cattle. Five zebu cattle needed trypanocidal drug treatment, while none of the N'Damas needed drug intervention. Ranking the 20 infected N'Damas according to average PCV levels revealed that the animals responded similarly to both infections.     

Studies on the pathogenesis of bovine ephemeral fever in experimental cattle III. Virological and biochemical data

In an attempt to define the nature of the response of cattle to ephemeral fever infection, a number of indicators of inflammation were monitored during clinical disease. The total Ca, Zn, Fe, Cu, glucose and phosphate in plasma, together with blood ammonia, were assayed relative to changes in the rectal temperature. Ca_T levels fluctuated markedly and hypocalcaemia occurred in 4 of 8 cattle. Plasma Zn and Fe values fell while plasma Cu levels rose markedly in all cattle. Mean levels of serum NH₃ of 20–30 μmol 1⁻¹ rose to a peak value of 56 μmoll⁻¹. Plasma glucose levels rose to a peak of 4.6 ± 0.5 mMl⁻¹ and the plasma phosphate levels fell from 2.4 ± 0.1 mMl⁻¹ to 1.17 ± 0.2 mMl⁻¹ during fever. Values of pCO₂ fell from a mean of 46.9 ± 3.6 mmHg to 36.4 ± 3.1 mmHg and coincided with a rise in pH. Virus was isolated 73 h (± 23) after inoculation and persisted until 130 h (± 21). The common role of these parameters in generalised inflammation and ephemeral fever is discussed.     

Neospora caninum in persistently infected, pregnant cows: spontaneous transplacental infection is associated with an acute increase in maternal antibody.

Nine cows which were naturally and persistently infected with Neospora caninum were housed and observed intensively throughout pregnancy. No recrudescence of a latent infection was detected by PCR tests on maternal blood but fetal infection, implying a recrudescence of maternal parasitosis, was associated with a marked increase in maternal antibody. The increase occurred in the second half of pregnancy in five cows which infected their calves, and before mid-pregnancy in one cow which aborted. There was no change in the avidity of the antibody, which remained high and characteristic of long-term infection. In three infected cows that gave birth to uninfected calves there was no marked increase in maternal antibody. Antigen-specific interferon gamma responses of peripheral blood mononuclear cells were observed in all the infected cattle but they did not vary significantly either during pregnancy, or whether the cows did or did not infect their calves, although the responses were consistently higher in the latter. There was no change in the plasma concentrations of cortisol or acute phase proteins associated with the recrudescence of the parasite. Three uninfected cows housed with the infected cows remained uninfected throughout the experiment. No immunosuppressive event was detected which might have provoked parasite recrudescence but the acute antibody rise associated with transplacental infection provides a valuable, non-invasive marker for further studies to investigate the cause and consequences of parasite recrudescence in N caninum infection in cattle.     

Bovine trypanosomiasis: effect on the immune response of the infected host.

Yearling cattle were inoculated with a recently isolated field strain of Trypanosoma congolense. Dinitrophenylated ovalbumin, a bacteriophage, or bovine parainfluenza-3 virus injected into the cattle during the first 5 weeks of infection resulted in peak serum antibody titers lower but not much lower than those produced by noninfected cattle. Primary and secondary antibody responses of inoculated cattle required more time to reach peak titers. Peripheral blood lymphocyte concentrations decreased to 42% of preinfection base-line value 1 1/2 weeks after the onset of T congolense infection and thereafter plateaued at less than 75% of base-line value. Lymphocyte cultures prepared from infected and noninfected cattle gave no marked differences in [(3)H]thymidine uptake after stimulation with phytohemagglutinin or pokeweed mitogen. Differences in lymphpocyte responsiveness were not noticed in one-way mixed lymphocyte culture reactions, using mitomycin C-treated lymphocytes from noninfected cattle as stimulator cells.     

Epidemiology of Dictyocaulus viviparus in Louisiana (U.S.A.)

An epidemiological investigation was conducted during a 1-year period on a permanent pasture naturally contaminated with Dictyocaulus viviparus and grazed by a varying number of yearling cattle. Seasonal variation in pasture infectivity to cattle was monitored by monthly slaughter of tracer calves, slaughter of pairs of resident yearlings at 30-60-day intervals, herbage larval recovery and by counts of first stage larvae in feces (modified Baermann technique) of resident cattle. A clinical outbreak of dictyocauliasis occurred during January-March 1986 and was associated with peak levels of pasture infectivity. Carrier animals were considered responsible for the survival of infection over summer. Although soil samples were taken regularly on a monthly basis to study the epidemiological importance of the soil as a source of infection, infective larvae were not recovered at any time. The epidemiological pattern observed in the present study provides basic information on the factors involved in infection and diseases outbreaks under sub-tropical conditions.     

牛环形泰勒虫病二温式PCR诊断方法的建立及初步应用

为建立特异、敏感、快速的二温式PCR诊断方法,根据牛环形泰勒虫裂殖子表面抗原(tams1)基因,设计了一对特异性引物,扩增出大小为154bp基因片段,经克隆、测序分析,与已知基因序列的相似性为96%。用建立的牛环形泰勒虫病二温式PCR诊断方法,对从新疆牛环形泰勒虫病流行地区采集的50份全血样品进行诊断,阳性率为88%,而血涂片检出的阳性率只有58%。经试验验证,该方法具有特异性高、敏感性强、重复性和稳定性好等优点。表明本试验所建立的二温式PCR诊断方法可用于牛环形泰勒虫病的临床诊断、隐性感染检测和流行病学调查。     

Infectivity of bovine leukaemia virus infected cattle: an ELISA for detecting antigens expressed in in vitro cultured lymphocytes.

A simple ELISA is described for quantifying expression of bovine leukaemia virus (BLV) antigens in short-term cultures of peripheral blood lymphocytes (PBL) isolated from infected cattle. The PBL-ELISA demonstrated that antigen expression levels in infected cattle could vary by more than 50-fold. Inoculation of sheep with dilutions of lymphocytes from two BLV-infected cattle, differentiated in the PBL-ELISA by 50 to 100-fold, suggested that antigen expression levels were correlated with infectivity. Haematological data indicated that increased antigen expression in PBL cultures was associated with an increased number of circulating B-lymphocytes, irrespective of whether or not an animal had lymphocytosis. This supported the hypothesis that BLV-infected cattle that are PBL-ELISA positive are more infectious and may present a greater risk of transmitting the disease. The applicability of the PBL-ELISA to a field situation was assessed with 98 BLV-infected cattle from three commercial dairy herds with infection prevalences of 11%, 23% and 47%. Similar percentages (49%, 50% and 52%) of PBL-ELISA positive cattle were identified among those infected cattle available for testing in the three herds. An additional 22 infected cattle from an experimental herd were tested to assess the stability of antigen expression levels over an 8 month period. Fewer (27%) of these cattle were identified as PBL-ELISA positive and antigen expression levels were generally lower than those observed in the commercial herds. Antigen expression levels in the experimental herd remained stable over the period of the study. The potential of the PBL-ELISA to assist in BLV eradication programs by identifying those seropositive cattle with the greatest potential to transmit infection is discussed.     

The control of Parafilaria bovicola transmission in South Africa

Ivermectin treatment of all cattle on a badly infected farm failed to interrupt the transmission of P. bovicola, even though ovipositional blood spots were drastically reduced in numbers for an entire summer season following treatment. Regular weekly to fortnightly dipping of all cattle in 50 ppm deltamethrin immediately reduced vector fly numbers to less than 1 fly per cow face. Sustained dipping for 9 months effectively reduced P. bovicola transmission from approximately 50% to less than 2%. However, cessation of fly control led to a return to predipping P. bovicola infection levels. Ovipositional blood spot counts and the ELISA technique for evaluating P. bovicola infection in a herd were compared and were both effective methods. Best results for the blood spot method, however, are obtained in spring at the peak of the bleeding season whereas the ELISA method does not have this limitation.     

Trypanosoma theileri: a literature review and report of incidence in New York cattle

The incidence of infection in adult dairy cattle in New York State with Trypanosoma (Megatrypanum) theileri (Laveran 1902) was determined by culturing buffy coats of peripheral blood samples in tissue culture growth media. Three sample groups of cattle were studied and revealed an overall rate of trypanosome infection of 44.4% (67 of 151) as determined by a single culture. Fifty-seven cows, representing four herds from three different geographic locations in the Southern-tier region of the state, comprised the first group. The infection rates of these herds ranged from 56.5 to 100%. The second group consisted of 81 cows randomly selected from animals admitted to the Large Animal Clinic of the New York State College of Veterinary Medicine. Trypanosoma theileri was cultured from 25.9% (21 of 81) of these animals. Repeated blood cultures from a third group of thirteen yearling heifers during the spring and summer revealed an increase in the infection rate from 0% in May, to 66.7% (8 of 12) in September, with 76.9% (10 of 13) positive at least once during this period. These findings are compared with the reported incidence of bovine T. theileri infections in other areas of the United States and in other countries.     

The measurement of serum haemoglobin reactive protein in selected groups of cattle and its use in clinical practice

Abstract

The presence of serum haemoglobin reactive protein (HRP) as an indicator of infection has been suggested. The serum HRP levels in varying groups of cattle were measured. A reference range, established on 233 cattle, of 0–10?mol/1 (as methaemoglobin binding capacity) was used. Forty eight of 104 cows that had recently aborted had raised serum HRP levels. Of 37 cows that had recently aborted, 26 had a leucocyte picture typical of bacterial infection and 18 of these had raised serum HRP levels. Eleven cows with a normal leucocyte picture had normal serum HRP levels. Seventeen cattle, with clinical infection and leucocyte pictures typical of bacterial infection, all had raised HRP levels. Of 133 cattle in which infection was considered a possibility by the attending veterinarian, 53 had an abnormal leucocyte picture with a raised serum HRP level. The mechanisms of a raised serum HRP level are unknown but, at the clinical level, its association with pyogenic infection make it a useful test in the confirmation of pyogenic infection.     

牛源粪肠球菌人工感染BALB/c雌鼠肾脏组织过程中PRNP和PRND的表达变化研究

为了研究牛源粪肠球菌感染对小鼠PRNP和PRND基因表达水平的影响,用牛源粪肠球菌感染小鼠,在1～7 d内解剖小鼠,制作病理组织革兰氏染色及石蜡切片HE染色,最后通过荧光定量PCR检测感染后1～7 d内PRNP和PRND的表达情况.结果表明:牛源粪肠球菌感染BALB/c雌鼠后,肾脏组织病变较明显,实时荧光定量检测确实对...