Detection of Jaagsiekte sheep retrovirus in the peripheral blood during the pre-clinical period of ovine pulmonary adenomatosis.

The envelope protein (Env) of the Jaagsiekte sheep retrovirus (JSRV) is known to be a unique oncoprotein responsible for inducing ovine pulmonary adenocarcinoma (OPA). The objective of this study was to prepare a specific monoclonal antibody (mAb) against the JSRV Env protein using bioinformatic analysis. According to the structure and epitope prediction results of JSRV Env, the JSRV-Env572-615 antigen was prepared via peptide synthesis (amino acid sequence 572-615, denoted as JSRV-Env572-615). BALB/c mice were immunized to prepare the anti-JSRV-Env572-615 mAb. Spleen cells were fused with SP2/0 myeloma cells after being screened by indirect ELISA and cloned by limiting dilution. The specificity of mAb was evaluated by western blot analysis and immunohistochemistry assays. Western blot results showed that the JSRV Env protein was able to bind to mAb with high specificity. Immunohistochemistry assays demonstrated that the mAb was able to recognize JSRV Env in adenomatous hyperplasia of the lung. Furthermore, JSRV was detected in peripheral blood leukocytes during the pre-clinical period of OPA in 2 of the 25 sheep using this newly synthesized mAb. Therefore, this mAb may be a useful tool for the detection of JSRV in sheep.

Assessment of transthoracic ultrasound diagnosis of ovine pulmonary adenocarcinoma in adult sheep.

Ovine pulmonary adenocarcinoma (OPA), caused by Jaagsiekte sheep retrovirus (JSRV), is a disease of increasing concern in the sheep industry. There is no commercial antemortem test for OPA; therefore, an early evaluation phase study was undertaken to examine the accuracy of transthoracic ultrasound examination using a 5-6.5 MHz sector ultrasound machine widely available in veterinary practice in the UK to diagnose OPA. Restraint, preparation and examination time was restricted to five minutes per sheep to represent the cost limitations of commercial sheep farming. One hundred sheep were examined. All 41 cases identified with suspect OPA lesions during transthoracic ultrasound examination had the diagnosis confirmed at postmortem examination, while sheep without ultrasonographic changes characteristic of OPA had no gross lesions of OPA at postmortem examination. This demonstrates the specificity of transthoracic ultrasound for diagnosis of OPA. The authors propose that, in the absence of any other reliable preclinical diagnostic test, the use of transthoracic ultrasound examination should be considered for a second opinion on an initial diagnosis of OPA, for screening purchased adult flock replacements for OPA, or for screening sheep in a known OPA-affected flock. However, the authors emphasise that a negative scan cannot provide a guarantee that the animal is free of JSRV infection nor early OPA.

Surveillance of Jaagsiekte sheep retrovirus in sheep in Hokkaido, the northern island of Japan.

Surveillance of jaagsiekte sheep retrovirus (JSRV) infection was performed by polymerase chain reaction (PCR) of blood DNA samples collected from 40 sheep and goats in 10 different flocks in Hokkaido, the northern island of Japan. No exogenous (oncogenic) JSRV sequence was detected by PCR in these samples, while the ovine endogenous retrovirus sequence was successfully amplified in all samples. Our paper is the first demonstration of JSRV surveillance in Japan and shows no evidence of oncogenic JSRV infection in sheep and goats in Hokkaido.

Diagnostic accuracy of PCR for Jaagsiekte sheep retrovirus using field data from 125 Scottish sheep flocks.

Using a representative sample of Scottish sheep comprising 125 flocks, the sensitivity and specificity of PCR for Jaagsiekte sheep retrovirus (JSRV) was estimated. By combining and adapting existing methods, the characteristics of the diagnostic test were estimated (in the absence of a gold standard reference) using repeated laboratory replicates. As the results of replicates within the same animal cannot be considered to be independent, the performance of the PCR was calculated at individual replicate level. The median diagnostic specificity of the PCR when applied to individual animals drawn from the Scottish flock was estimated to be 0.997 (95% confidence interval [CI] 0.996-0.999), whereas the median sensitivity was 0.107 (95% CI 0.077-0.152). Considering the diagnostic test as three replicates where a positive result on any one or more replicates results in a positive test, the median sensitivity increased to 0.279. Reasons for the low observed sensitivity were explored by comparing the performance of the test as a function of the concentration of target DNA using spiked positive controls with known concentrations of target DNA. The median sensitivity of the test when used with positive samples with a mean concentration of 1.0 target DNA sequence per 25µL was estimated to be 0.160, which suggests that the PCR had a high true (analytical) sensitivity and that the low observed (diagnostic) sensitivity in individual samples was due to low concentrations of target DNA in the blood of clinically healthy animals.

[Enzootic nasal adenocarcinoma in a dairy sheep flock]. / Enzootische nasale Adenokarzinome in einem Milchschafbestand.

In a herd of dairy sheep several losses occurred due to a respiratory syndrome in combination with progressive wasting. Clinical and pathomorphological diagnostics of 3 sheep revealed the presence of cancerous masses in the nasal cavities. These neoplasms were identified as adenocarcinomas originating from the nasal mucosa. Etiologically, they were attributed to JRSV (Jaagsiekte Sheep Retrovirus) by detection of capsid protein 24 in western blot. The significance of the disease in Switzerland is discussed, also in the context of lung adenomatosis.

PCR examination of bronchoalveolar lavage samples is a useful tool in pre-clinical diagnosis of ovine pulmonary adenocarcinoma (Jaagsiekte).

Ovine pulmonary adenocarcinoma (OPA) is a contagious lung tumour of sheep caused by Jaagsiekte sheep retrovirus (JSRV). The disease is a particular problem in flocks in many parts of the world. The aim of the study was to assess screening methods for individual animals as a prelude to future eradication trials. Results of histological examination were used as the standard to evaluate the relative sensitivity and specificity of an established heminested polymerase chain reaction (PCR) test for JSRV proviral DNA from blood and bronchoalveolar lavage (BAL) samples. PCR results from tissue samples are included as control data. PCR testing of blood samples was found to have an estimated sensitivity of only 10% (95% confidence interval (CI) 3-20) while the sensitivity of the PCR test on BAL samples was 89% (CI 79-96) in comparison to the results of histological examination. We conclude that PCR testing of BAL samples is an effective confirmatory test for sheep with suspected clinical OPA. It is also a useful tool for the pre-clinical identification of individual infected sheep within an infected flock and therefore may prove beneficial in future control or eradication programmes.

A PCR technique for the detection of Jaagsiekte sheep retrovirus in the blood suitable for the screening of ovine pulmonary adenocarcinoma in field conditions.

Ovine pulmonary adenocarcinoma (OPA) is a naturally occurring contagious lung neoplasia caused by jaagsiekte sheep retrovirus (JSRV). Although no specific circulating antibodies against the virus can be detected in infected sheep, JSRV proviral DNA sequences can be found in peripheral blood leukocytes (PBLs) in clinically affected and in a proportion of in contact animals. In this study, existing hemi-nested PCR procedure is compared with a new one-step PCR technique that was developed to minimise potential DNA contamination and reduce sample and reagent handling. Different blood preparations were assessed and the best results were achieved on DNA prepared from buffy coat. The sensitivity of this PCR was lower in JSRV infected sheep without lesions of OPA than in clinically affected sheep, which indicate that this PCR may not be not fully appropriate for screening of individual sheep, but rather to provide results at flock level. This PCR is the only currently available blood test for detection of JSRV infected sheep and may be useful in epidemiological studies and in control programmes of OPA.

Lack of a specific immune response against a recombinant capsid protein of Jaagsiekte sheep retrovirus in sheep and goats naturally affected by enzootic nasal tumour or sheep pulmonary adenomatosis.

Enzootic nasal tumour (ENT) and sheep pulmonary adenomatosis (SPA) are two contagious adenocarcinomas of the respiratory tract of sheep and goats. Both diseases are associated with related, but distinct, type-D-retroviruses (ENTV and JSRV respectively). No evidence of circulating antibodies has been described in animals affected by either ENT or SPA using antigens from natural sources. We evaluated the usefulness of a recombinant JSRV capsid protein (JSRV-CA) as antigen to study the antibody responses of animals naturally affected by ENT or SPA, using immunoblotting. Positive reactions were detected in the sera of both affected and unaffected sheep and goats. The reactivity was abolished completely by absorption with the GST fusion partner but not by JSRV-CA, suggesting that it was not specific. The results support prior observations indicating that sheep and goats infected by JSRV and ENTV do not develop specific humoral responses to these retroviruses.

Coexistence of pulmonary adenomatosis and progressive pneumonia in sheep in the central sierra of Peru.

A survey of sheep with chronic respiratory tract disease was conducted in the central sierra of Peru. Histopathologic examinations coupled with an agar-gel immunodiffusion test for ovine progressive pneumonia (OPP) revealed that sheep pulmonary adenomatosis and OPP were present in these flocks. Of 80 sheep examined, 22 had lesions of sheep pulmonary adenomatosis, and 4 had metastases to regional lymph nodes. Four sheep had lesions consistent with OPP and 9 had lesions indicating the coexistence of both diseases. The agar-gel immunodiffusion test revealed that at least 26% of the sheep had been exposed to the OPP (or an antigenically similar) virus. A variety of other respiratory tract diseases complicated the evaluations of these sheep, including verminous pneumonia, hydatid disease, lung abscesses, and other nonspecific acute and chronic pneumonias.

[Pulmonary adenomatosis (jaagsiekte) in imported sheep (author's transl)]. / Longadenomatose (jaagziekte bij geïmporteerde schapen.

Pulmonary adenomatosis was diagnosed in a flock of Scottish Half-Bred sheep imported eighteen months previously. The differences between pulmonary adenomatosis (jaagsiekte) and chronic progressive pneumonia (zwoegerziekte) are stressed. These differences are clinical as well as morbid-anatomical and histological in character. This is the first time that pulmonary adenomatosis was reported as occurring in sheep in the Netherlands. The disease has not been reported in indigenous breeds of sheep.

[Ovine pulmonary adenomatosis]. / Plicní adenomatóza ovcí

Lung adenomatosis was histologically demonstrated in seven sheep coming from one flock in western Bohemia. On the basis of morphological characteristics and biological properties the disease can be considered as a neoplasma. It is capable of forming metastases and calls forth a systemic reaction of the organism. Lund adenomatosis can be experimentally transferred and the present knowledge speaks in favour of virus etiology of the disease.