•BoHV1 unvaccinated buffaloes resulted gB-positive and gE-negative by ELISA.•Similar results are expected in animals vaccinated with a BoHV1 gE-deleted vaccine.•A BuHV1 was isolated from a buffalo ...resulted gB-positive and gE-negative by ELISA.•Experimental infection with this BuHV1 strain produced mild disease in buffalo.
During a serological survey, 157 out of 681 unvaccinated buffaloes resulted seropositive for bovine alphaherpesvirus 1 (BoHV1) glycoprotein B (gB) and seronegative for BoHV1 glycoprotein E (gE). These serological results were generally expected in animals vaccinated with a BoHV1 gE-deleted vaccine but not in unvaccinated animals. Seroneutralization tests on 36 selected sera detected neutralizing antibody titers more than three times higher for BuHV1 than for BoHV1. In order to investigate the virus, one of these buffaloes was injected with dexamethasone, and from nasal and vaginal swabs collected at different time points, a ruminant herpesvirus was isolated, characterized and also detected by PCR. Restriction enzyme analysis, sequencing and phylogenic analysis of gB and gD genes showed that the virus was genetically similar but not identical to BuHV1 strain b6. Intranasal inoculation of the virus in a healthy seronegative buffalo resulted in a mild and transient upper respiratory disease; the virus was isolated from clinical specimens and DNA was detected by PCR in nasal and vaginal swabs up to 9 days after infection. Further investigations should be aimed at sequencing the whole viral genome and at evaluating the host-range of this virus. Specific tests are needed to discriminate infections by different ruminant herpesviruses and to improve eradication programs of infectious bovine rhinotracheitis/infectious pustular vulvovaginitis in cattle.
In order to characterize the complete range of lesions, especially minimal, affecting mammary gland and viral antigen distribution and target cells using immunohistochemistry in naturally Visna/maedi ...(VM) 84 infected sheep were studied, forty-four from flocks with clinical cases (A) and 35 randomly sampled from two abattoirs (B) together with five negative controls (C). An immunocytochemistry technique was developed and further milk samples (n = 39) were used to study viral excretion, carrier cells and the role of milk and colostrum in the transmission of the disease.
All sheep from group C and three sheep from group B were negative to VM in tissue sections by histopathology, immunohistochemistry and PCR, and also in serum using ELISA. Several degrees of CD3 + lymphocytic interstitial mastitis were observed in groups A and B: minimal (+) n = 26 sheep; moderate (++), n = 32 and severe (+++), n = 12. No differences in lesion distribution were observed between groups A and B. Viral presence was confirmed by immunohistochemistry using two different antibodies and/or PCR in every tissue with lesions while serology was negative in six sheep with lesions. Two milk samples taken from milk tanks from two flocks from group A and fourteen milk samples from 29 infected sheep from group B were positive to VM (most of them from animals with moderate and severe lesions). Positivity was only found in macrophages, even in focal and minimal lesions, while no positivity was observed in epithelial or any other cells in either tissue and milk samples.
This new observation of the minimal lesions described in this work increased the prevalence of VM lesions in mammary gland up to 90.9% and VM should be considered as a differential diagnosis when minimal interstitial lesions are detected. A high prevalence of VM was observed in intensive milk-producing sheep, ELISA serology did not detect as positivity all infected animals, while histology, IHC or PCR showed higher sensitivity. The cytological technique developed was very useful in milk-cell studies using hematoxylin and eosin and immunocytochemistry. Viral detection in milk samples (16/39) confirms a potential but limited role of milk/colostrum in viral transmission.
The members of
Mycobacterium avium species, comprising
M. avium subsp.
paratuberculosis,
M. a. hominissuis,
M. a. avium,
M. a. silvaticum, are currently the most prevalent opportunistic pathogenic ...mycobacteria causing mycobacterial infection in animals and humans. The ability to distinguish between these subspecies is of relevance for proper diagnosis and control programmes of the diseases. The aim of this study was to design a fast and specific PCR strategy for the detection and differentiation of
M. avium subspecies from the solid plate cultures for use in routine veterinary diagnosis. We have developed a multiplex PCR based on IS
900, IS
901, IS
1245 and the
dnaJ gene. This method allows the detection of
M. a.
paratuberculosis,
M. a. hominissuis and
M. a. avium/
M. a. silvaticum in one PCR reaction and theoretically enables mixed infections of
M. a. paratuberculosis and
M. a. avium or
M. a. paratuberculosis and
M. a. hominissuis to be revealed. The sensitivity of this multiplex PCR is 10
3
CFU for each bacterial strain in one PCR reaction, which also enabled the use of this test directly for DNA isolated from the tissue of the heavily infected sheep.
Visna/Maedi virus (VMV), a small ruminant lentivirus responsible for lymphoproliferative pneumonia, encephalitis, arthritis and/or mastitis in sheep, has been detected in different non-lymphoid ...organs. However, only a few investigations have been carried out in lymphoid tissues. In this study, some lymphoid tissues and lymph node draining or non-draining VMV target organs from five sheep infected experimentally by the respiratory route three years previously were investigated. Archival samples of spleen, red bone marrow, caudal mediastinal lymph nodes, mammary lymph nodes, popliteal lymph nodes and mesenteric lymph nodes were tested by PCR for the presence of proviral DNA. Popliteal and mesenteric lymph node samples were tested also by immunohistochemical staining of the viral capsid antigen p28. The proviral DNA was detected by PCR in all the lymphoid tissue samples from the infected sheep. The viral antigen was stained in mononuclear cells in popliteal and mesenteric lymph nodes of the infected sheep. Although the lymph nodes draining the classical target organs seem to be more infected than the others, both the viral capsid antigen and the proviral DNA were present also in lymph nodes draining non-target organs, such as the mesenteric lymph nodes. These findings show the presence of VMV in different lymphoid tissues in the late stages of infection and suggest a potential role of these tissues as a site for viral reservoir and replication, even three years after infection.
Abstract An outbreak of strangle-like disease involving 26 horses farmed in central Italy was investigated by clinic examination, endoscopy, cytology, bacteriology and polymerase chain reaction ...(PCR). At weekly interval, a total of three nasal swabs and one guttural pouches lavage fluid (GPLF) were collected, and no Streptococcus equi subsp. equi carrier was found. Some horses showed upper airways disease and endoscopic signs of pharyngeal lymphoid hyperplasia of different grade and/or abnormal endoscopic appearance of guttural pouches. Streptococcus dysgalactiae subsp. equisimilis was isolated from 14 horses while S. equi subsp. zooepidemicus was isolated from six horses. PCR confirmed the biochemical and serological identification of all isolates and was positive in 10 bacteriological negative samples. The absence of S. equi and the frequent detection of S. equisimilis and S. zooepidemicus suggest that beta-haemolytic streptococci other than S. equi could be the causative agent of strangle-like disease.
Maedi visna virus (MVV) vertical transmission in sheep via infected colostrums is a very important route of infection in lambs. To verify colostral transmission and to study early viral entry in ...lambs, colostrum samples, and small intestine and mesenteric lymph nodes of lambs born from experimentally infected ewes were examined by histopathology, immunohistochemistry (IHC) and in situ hybridisation (ISH) studies. In particular, newborn lambs were naturally fed maternal colostrum and humanely killed at 10, 24, 48, 72, 96
h and 7 and 10 days after birth; two caesarian-derived lambs served as uninfected controls. No lesions suggestive of MVV infection were found, but marked immunoreactions for MVV capsid antigen (CA, p28) were detected in lambs fed maternal colostrum and in macrophages cultured from colostrum. IHC results in lambs suggest an initial viral absorption by intestinal epithelial cells at the tip of the villi, passage to mononuclear cells in the lamina propria and involvement of ileum Peyers’ patches and mesenteric lymph nodes, with different staining patterns depending on infection times. ISH on intestinal sections of the 72
h lamb revealed the presence of proviral DNA in epithelial cells at the tip of the villi, suggesting a role for these cells in early MVV replication. The results contribute to knowledge about the pathogenesis of ovine lentivirus infection suggesting that the small intestine and mesenteric nodes are the sites of entry and propagation of MVV in lambs fed colostrums from infected ewes.
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