Exogenous Small Ruminant Betaretrovirus Envelope Protein Is Detected in Draining Lymph Nodes in Contagious Respiratory Tumors of Sheep and Goats.

Contagious respiratory tumors of sheep and goats are epithelial neoplasms of the lung and nasal cavities. They are associated with oncogenic betaretroviruses known as jaagsiekte sheep retrovirus and enzootic nasal tumor retrovirus of sheep and goats. We investigated the presence of the envelope protein (ENV) of these retroviruses in retropharyngeal and mediastinal lymph nodes using a specific monoclonal antibody by immunohistochemistry methods, single-labeled or combined with ovine B or T lymphocytes or macrophage cell markers. Samples of lymph nodes, fixed in formalin and zinc fixative, were obtained from paraffin-embedded material. Four groups of samples were used: 24 natural cases of ovine pulmonary adenocarcinoma (OPA), 13 of enzootic nasal adenocarcinoma of sheep (ENAS), 19 of enzootic nasal adenocarcinoma of goats (ENAG), and 14 control samples. ENV was detected by single labeling in cortical lymphoid follicles. Six of 24 OPA samples were positive and only in those from sheep with extensive neoplasia. Immunolabeling was detected in 5/13 ENAS and 10/19 ENAG samples. Positive labeling was found either in the intercellular spaces, membranes, or cytoplasm of cells in follicles. Control samples were not correspondingly labeled. Double immunohistochemistry demonstrated co-labeling of ENV and CD21 (B cells and follicular dendritic cells) in all samples, CD14 (macrophage) in OPA samples, and Pax-5 (B cells) in ENAG samples, but not with CD8 or CD4 (T lymphocytes). These results demonstrate the presence of betaretrovirus ENV proteins in nontumor cells in regional lymph nodes in sheep and goats with contagious respiratory tumors.

Presence of anti-Leishmania infantum antibodies in sheep (Ovis aries) in Spain.

Sandflies are the primary transmission vector for Leishmania spp parasite in endemic regions. The role of other animals, different from the dog, is under discussion in the leishmaniosis endemic countries. A limited number of reports have been published on the possible role of livestock in European countries for Leishmania maintenance and diffusion. The aim of the present study was to perform a serosurvey on sheep in areas of Spain that are endemic for zoonotic leishmaniosis and establish the possible role of sheep regarding Leishmania infantum infection in endemic areas. Three hundred and two serum samples were obtained from sheep and were evaluated for serological survey to detect L. infantum by using the in-house ELISA technique. Twenty-eight out of the 302 samples included in this study, were positive for L. infantum antibodies (9.27%). In the present study, a significant association was found between adult age and seropositivity (p = 0.006) and female gender and seropositivity (p = 0.02). This association has not been previously described in other European studies related to L. infantum infection in sheep. Our study reveals that domestic sheep in a European Mediterranean country are exposed to L. infantum. To our knowledge, this study demonstrates the presence of seropositive sheep in different regions of Spain for the first time. Further epidemiological studies focus on evaluating the rural cycle of this parasite to know if livestock could act as a potential reservoir are needed.

First Evidence of Leishmania infantum Antibodies in Sheep (Ovis aries) from Southern Germany.

In Europe, Leishmania infantum is the most prevalent Leishmania species, and this protozoan is transmitted by phlebotomine sandflies. A recent publication has shown that sheep harbor L. infantum antibodies. This raises questions about the epidemiological role of small ruminants. Therefore, sera from small ruminants located in two southern German federal states, Baden-Wuerttemberg (BW) and Bavaria (BAV), were analyzed with an ELISA to determine the presence of L. infantum antibodies. The species, sex and age (gimmer vs. ewe) were recorded, and a univariate analysis was conducted to determine possible associations. In total, seven sheep flocks (274 sheep/10 goats) from BW and seven sheep flocks (277 sheep/78 goats) from BAV were examined. In BW, four sheep from three flocks tested positive for L. infantum antibodies. In BAV, the same number of positive sheep were detected but in four flocks. The total seropositivity rate in sheep was 1.45%. All goats tested negative. No significant association (p > 0.05) was detected between Leishmania seropositivity and the variables evaluated. Our study reveals the exposure of sheep to L. infantum in a non-endemic area. Further investigation is needed to determine whether sheep can be used as sentinels to identify new phlebotomine habitats and Leishmania risk areas.

Use of a local anaesthetic and antiseptic wound formulation for the treatment of lambs naturally infected with Orf virus.

Contagious ecthyma (CE) is a worldwide highly contagious zoonotic viral skin disease of sheep and goats. Treatment for Orf virus (ORFV) infection usually involves topical and oral antibiotics. An anaesthetic and antiseptic topical gel (Multisolfen® or Tri-Solfen®; MS®, Medical Ethics, Australia) has been documented as an efficacious therapy for lesions from mucosal and epithelial viral infections in ruminants. The present study tested a new treatment protocol of MS® for CE therapy on-farm in 150 lambs naturally infected with ORFV. Lambs were divided into three cohorts of 50 lambs each (C, D and E). Cohort C was treated with MS® 3 times with an interval of 3 days between treatments, cohort D was treated daily with hypochlorous acid, whilst cohort E served as untreated controls. The lambs were examined clinically every two days, weight measured weekly, with whole blood and sterile swabs from ORFV lesions collected for haematological analysis and specific ORFV PCR. Cohort C presented fewer lambs displaying ORFV-associated lesions than other cohorts at different times of the experiment. Further, lesions treated with MS® were milder compared with other cohorts. However, following cessation of therapy, most of the lambs again developed ORFV-associated lesions. No differences between cohorts were observed in weight, haematological and PCR results. These findings suggest that topical treatment with MS® is effective for CE in field conditions, especially in the first stages of the clinical course, although treatment with MS® may need to be extended a minimum of 4 weeks.

Use of a Local Anaesthetic/Antiseptic Formulation for the Treatment of Lambs Experimentally Infected with Orf Virus.

Contagious ecthyma is a highly transmissible eruptive viral disease of the skin and mucosa of sheep and goats distributed worldwide. The treatment of orf lesions is usually based on the use of antiseptics and antibiotics for the management of presumptive secondary infections, increasing risks of antimicrobial resistance. The wound dressing formulation Tri-Solfen® (TS) containing two local anaesthetics (lignocaine and bupivacaine), adrenaline and an antiseptic (cetrimide) in a gel formulation has been demonstrated to reduce suffering and enhance recovery in cattle and buffalo with oral and skin lesions due to foot-and-mouth disease virus infection and reduced the orf viral load in lambs. In the present study, experimental infection with the orf virus was conducted in 50 newborn lambs and 25 animals were treated after the presence of the first lesions with TS and repeated three days later. Daily clinical examination, haematological, serological, biomolecular and post-mortem analyses were conducted during 34 days after treatment. Results indicated that treatment had no effect on weight gain and clinical progression of the lesions. It was determined that seroconversion after experimental infection occurs 34 days after infection and suggested that the deep basal epithelial location of the orf lesions may have prevented the therapy from having altered the clinical course.

Jaagsiekte sheep retrovirus found in milk macrophages but not in milk lymphocytes or mammary gland epithelia of naturally infected sheep.

Jaagsiekte sheep retrovirus (JSRV) causes ovine pulmonary adenocarcinoma. JSRV can be transmitted via infected colostrum or milk, which contain somatic cells (SCs) harboring JSRV provirus. Nevertheless, the cell types involved in this form of transmission and the involvement of the mammary gland remain unknown. We separated adherent cells (macrophages and monocytes) by plastic adherence, and lymphocytes (CD4+ and CD8+ T cells, and B cells) by flow cytometry, from SCs in milk samples from 12 naturally infected, PCR blood test JSRV-positive, subclinical ewes. These cell populations were tested by PCR to detect JSRV provirus. The ewes were euthanized, and mammary gland samples were analyzed immunohistochemically to detect JSRV surface protein. We did not detect JSRV provirus in any milk lymphocyte population, but milk adherent cells were positive in 3 of 12 sheep, suggesting a potential major role of this population in the lactogenic transmission of JSRV. Immunohistochemistry did not reveal positive results in mammary epithelial cells, pointing to a lack of participation of the mammary gland in the biological cycle of JSRV and reducing the probability of excretion of free viral particles in colostrum or milk.

Copper Poisoning, a Deadly Hazard for Sheep.

Copper (Cu) is an essential microelement for animals. However, sheep are particularly susceptible to Cu intoxication, a deadly disease reported worldwide. The risk of developing this poisoning is higher in vulnerable breeds and in intensively managed lambs or milk sheep. Two types of Cu intoxication can occur depending on the chronic or acute exposure to Cu. In chronic Cu poisoning (CCP), the most common form, Cu is accumulated in the liver during a subclinical period. A low intake of Cu antagonists (molybdenum, sulphur, iron, or zinc) favours Cu accumulation. The sudden release of Cu into the blood causes acute haemolysis with anaemia, haemoglobinuria, jaundice and death within 1-2 days. Acute Cu poisoning is related to the accidental administration or ingestion of toxic amounts of Cu. Acute oral exposure to Cu causes severe gastroenteritis, shock and death. Collapse and death occur shortly after parenteral administration. The diagnosis is based on history, clinical, gross pathological, histological and toxicological findings. Treatment of sheep with severe clinical signs often has poor success but is very effective during the Cu accumulation phase. Different therapies, based on either chelating agents or Cu antagonists, have been used to treat and prevent CCP.

Comparative Study of the Use of Doxycycline and Oxytetracycline to Treat Anaplasmosis in Fattening Lambs.

Lamb icteric carcasses condemnation due to Anaplasma ovis is causing relevant economic losses. A comparative study was developed on the effects of different antibiotics to treat ovine anaplasmosis in fattening lambs. A total of 100 A. ovis naturally infected lambs were selected and randomly divided into four groups of 25 lambs: Group ID, treated with injectable doxycycline; Group OD, oral doxycycline; Group O, injectable oxytetracycline; and Group C, untreated animals for the control group. Clinical, haematological, and molecular analyses were performed before the treatment and 12 and 45 days after the beginning of the treatments, and carcass condemnation was followed after slaughter. The A. ovis bacterial load was high before the treatments in the four groups and decreased significantly 45 days after treatment in the ID and O Groups (p < 0.001). The parameters that were related to haemolysis showed similar results. At the abattoir, 15 out of the 47 examined carcasses were condemned; 7 of C Group, 6 of OD Group, 2 of O Group, and 0 of ID Group. It can be concluded that injectable doxycycline and oxytetracycline significantly reduce A. ovis bacterial load in blood and carcass condemnation at the abattoir. Further studies are needed in order to confirm these encouraging findings.

Neoplasia-Associated Wasting Diseases with Economic Relevance in the Sheep Industry.

We review three neoplastic wasting diseases affecting sheep generally recorded under common production cycles and with epidemiological and economic relevance in sheep-rearing countries: small intestinal adenocarcinoma (SIA), ovine pulmonary adenocarcinoma (OPA) and enzootic nasal adenocarcinoma (ENA). SIA is prevalent in Australia and New Zealand but present elsewhere in the world. This neoplasia is a tubular or signet-ring adenocarcinoma mainly located in the middle or distal term of the small intestine. Predisposing factors and aetiology are not known, but genetic factors or environmental carcinogens may be involved. OPA is a contagious lung cancer caused by jaagsiekte sheep retrovirus (JSRV) and has been reported in most sheep-rearing countries, resulting in significant economic losses. The disease is clinically characterized by a chronic respiratory process as a consequence of the development of lung adenocarcinoma. Diagnosis is based on the detection of JSRV in the tumour lesion by immunohistochemistry and PCR. In vivo diagnosis may be difficult, mainly in preclinical cases. ENA is a neoplasia of glands of the nasal mucosa and is associated with enzootic nasal tumour virus 1 (ENTV-1), which is similar to JSRV. ENA enzootically occurs in many countries of the world with the exception of Australia and New Zealand. The pathology associated with this neoplasia corresponds with a space occupying lesion histologically characterized as a low-grade adenocarcinoma. The combination of PCR and immunohistochemistry for diagnosis is advised.

Evolution of the Seroprevalence of Pestivirus and Respiratory Viral Infections in Spanish Feedlot Lambs.

The presence of respiratory viruses and pestiviruses in sheep has been widely demonstrated, and their ability to cause injury and predispose to respiratory processes have been proven experimentally. A longitudinal observational study was performed to determine the seroprevalence of bovine parainfluenza virus type 3 (BPIV-3), bovine respiratory syncytial virus (BRSV), bovine herpesvirus type 1 (BHV-1) and pestiviruses in 120 lambs at the beginning and the end of the fattening period. During this time, the animals were clinically monitored, their growth was recorded, and post-mortem examinations were performed in order to identify the presence of pneumonic lesions in the animals. Seroconversion to all viruses tested except BHV-1 was detected at the end of the period. Initially, BPIV-3 antibodies were the most frequently found, while the most common seroconversion through the analysed period occurred to BRSV. Only 10.8% of the lambs showed no detectable levels of antibodies against any of the tested viruses at the end of the survey. In addition, no statistical differences were found in the presentation of respiratory clinical signs, pneumonic lesions nor in the production performance between lambs that seroconverted and those which did not, except in the case of pestiviruses. The seroconversion to pestiviruses was associated with a reduction in the final weight of the lambs.

Absence of SARS-CoV-2 Antibodies in Natural Environment Exposure in Sheep in Close Contact with Humans.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the zoonotic causative agent of coronavirus disease 2019 (COVID-19) that has caused a pandemic situation with millions of infected humans worldwide. Among domestic animals, there have been limited studies regarding the transmissibility and exposure to the infection in natural conditions. Some animals are exposed and/or susceptible to SARS-CoV-2 infection, such as cats, ferrets and dogs. By contrast, there is no information about the susceptibility of ruminants to SARS-CoV-2. This study tested the antibody response in 90 ovine pre-pandemic serum samples and 336 sheep serum samples from the pandemic period (June 2020 to March 2021). In both cases, the animals were in close contact with a veterinary student community composed of more than 700 members. None of the serum samples analyzed was seroreactive based on an enzyme-linked immunosorbent assay (ELISA) using the receptor-binding domain (RBD) of the spike antigen. In this sense, no statistical difference was observed compared to the pre-pandemic sheep. Our results suggest that it seems unlikely that sheep could play a relevant role in the epidemiology of SARS-CoV-2 infection. This is the first study to report the absence of evidence of sheep exposure to SARS-CoV-2 in natural conditions.

Impact of Stress on Health and Final Weight in Fattening Lambs.

In order to determine whether the stress level had any influence on the health and final weight of Rasa Aragonesa fattening lambs, stress markers were measured throughout the fattening period in 80 feedlot lambs (group F) and in 40 lambs finished on the farm of birth (group C). The highest values of the stress indicators-neutrophil-to-lymphocyte-ratio (N/L), non-esterified fatty acids (NEFA), serum cortisol and fecal cortisol metabolites (FCM)-were recorded after the road transportation of group F to the feedlot. No differences between the groups were identified at the end of the adaptation period, but at the end of the fattening, statistically significant higher values for NEFA and serum cortisol were determined in group F. However, statistically significant differences between the groups were not found in the percentage of lambs with clinical signs of illness and ovine respiratory complex (ORC) lesions or in the final weight of the lambs. Independent of the location at which the lambs were fattened, those with FCM values at the beginning of the study (at the time of weaning) in the highest quartile developed a higher percentage of clinical signs of illness (45.5% vs. 32.1%, p > 0.05) and ORC lesions (45.5% vs. 10.7%, p < 0.01), and their average final weight was lower (24.36 kg vs. 27.40 kg, p = 0.001) than those with values in the lowest quartile. Stress experienced by lambs prior to finishing seems to be relevant for their further development, and FCM concentration at the time of weaning could be used as an indicator of health and productive performance of the lambs during the fattening period, regardless of finishing location.

Impact of a Topical Anaesthesia Wound Management Formulation on Pain, Inflammation and Reduction of Secondary Infections after Tail Docking in Lambs.

We examined several procedures for surgical tail docking; with and without general anaesthesia (GA), including the use of a topical wound gel formulation to provide pain relief (PR) and improve healing after surgery, containing local anaesthetics lignocaine and bupivacaine, with cetrimide and adrenalin. Forty-four lambs were recruited into four equal cohorts: Groups A and C, the tail was excised with a scalpel without anaesthesia or stitches; Groups B and D, the tail was surgically excised and stitched under GA; Groups C and D wounds were immediately sprayed with PR. Behavioural observations identified that Groups A and C displayed significantly less pain-related behaviours than Groups B and D shortly after the procedure, especially if treated with PR. Similarly, the mean of days when animals showed no signs of wound infection was longer in the groups not undergoing stitching. Finally, treatment with PR appeared to reduce the cortisol response and avoided the elevation of serum amyloid A in lambs where the tail was excised without general anaesthesia. In conclusion, surgical tail-docking without GA but where wounds are immediately sprayed with PR, appears as an affordable and more welfare-appropriate method for conducting tail docking in lambs.

Effects of allergen-specific immunotherapy on peripheral blood regulatory T cells and serum concentrations of cytokines and immunoglobulins in horses with allergic dermatitis.

The aim of this study was to assess the effect of allergen-specific immunotherapy (ASIT) on the immunological responses of horses. Blood samples were taken from thirty-two horses with allergic dermatitis treated with ASIT and 10 healthy control horses at 0, 3, 6, 9 and 12 months to investigate the evolution of the percentage of regulatory T cells (Treg) in the peripheral blood and the serum levels of cytokines and immunoglobulins. Clinical improvement was appreciated by the majority of the horses' owners (56.6%). No effect of ASIT on CD4+CD25High Treg cells was found during the one year treatment period. No differences in the percentage of CD4+ T cells were observed between the groups, and no effects of ASIT over time were observed. The percentage of CD25+ T cells was always higher in the ASIT group (17.9 ±â€¯11.3%) than in the control group (7.3 ±â€¯4.4%, p < 0.001). We did not detect any effect of ASIT on the serum levels of TGF-ß, IL-10 and IFN-γ or on the serum concentrations of IgA and IgG4. A reduction in the serum levels of total IgE in the horses with allergic dermatitis was observed at the 6th month (p < 0.05), but increased again at the end of the study. The results indicate that immunotherapy was insufficient to induce significant changes that could indicate T cell tolerance, a shift in cytokine production to more protective Th1 cells. More studies are needed with new vaccine compositions and administration protocols to improve the immunological responses of the horses with allergic dermatitis.

Colostrum and milk can transmit jaagsiekte retrovirus to lambs.

Ovine pulmonary adenocarcinoma (OPA) is a contagious disease caused by jaagsiekte sheep retrovirus (JSRV). In the three studies performed, we have obtained data of the importance of colostrum/milk (C/M) in the transmission of JSRV. In the first study, a group of sheep from a flock with a long history of OPA, samples from colostrum and peripheral blood leucocytes (PBLs) were collected. Two specific PCRs (U3-LTR and env of the JSRV) were carried out. Using U3PCR 8/34 sheep were positive in colostrum whereas with envPCR 7/34 were positive. From these animals only one was positive with U3PCR in the PBLs. Evidence of the transmission of JSRV infection by C/M was obtained in two more separate studies. In the second study, PBLs from five lambs from JSRV+ ewes and two from JSRV-ewes were tested by the U3PCR. They were fed C/M by their mothers during 3 months and slaughtered 7 months after birth. Three out of five lambs from the JSRV+ sheep become PBL positive at 3-4 months old and the other two were also positive at 4-6 months of age. One lamb of the JSRV-sheep became also PBL positive at an age of 3 months. In the third study, a group of lambs from JSRV negative mothers were fed with C/M from JSRV+ sheep and housed in separate unit. For comparison, another group of the same origin and maintained in another different unit, were fed with C/M containing a JSRV virus preparation. All lambs were blood sampled monthly and JSRV infection was detected as early as 15 days and several times onwards in both groups. Control groups fed with C/M from JSRV free flock and JSRV blood test negative sheep were always negative. Together these results indicate that suckling is an important natural transmission route for JSRV.

Jaagsiekte sheep retrovirus is not detected in human lung adenocarcinomas expressing antigens related to the Gag polyprotein of betaretroviruses.

A proportion of human lung adenocarcinomas (hLACs) express an antigen related to the major capsid protein (CA) of Jaagsiekte sheep retrovirus (JSRV), a Betaretrovirus that causes a transmissible lung cancer in sheep. In this study, we have investigated whether JSRV or related betaretroviruses are expressed in hLACs. Results obtained indicate that JSRV is not associated with human lung adenocarcinomas. However, a proportion of hLACs reacted positively in immunohistochemistry with antibodies specific towards different domains of the JSRV Gag suggesting that a bona fide retrovirus antigen could be expressed in these tumours. Further studies will be necessary to ascertain whether the detection of antigens cross-reacting with betaretrovirus Gag antisera in some hLACs is due to expression of a human endogenous retrovirus or, more unlikely, of an uncharacterized exogenous retrovirus.

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.

Cells infected with Jaagsiekte sheep retrovirus are detected in the bone marrow of asymptomatic sheep.

Ovine pulmonary adenocarcinoma (OPA) is a transmissible lung cancer caused by Jaggsiekte sheep retrovirus (JSRV). It is difficult to identify animals infected with JSRV but are clinically healthy. The virus does not induce a specific antibody response and, although proviral DNA sequences of JSRV can be found in mononuclear blood cells, the detection is inconsistent. The aim of this study was to investigate the presence of JSRV in the bone marrow of infected sheep and develop a more consistent screening method. Immunohistochemical examination of bone marrow samples from 8 asymptomatic JSRV-infected sheep revealed the presence of positively labelled cells. However, JSRV could not be detected by a highly sensitive polymerase chain reaction (PCR) in bone marrow aspirates periodically collected from these animals. Results suggest that JSRV-infected cells may be present in the bone marrow of symptomless animals, but the number is below the detectable level for PCR. Therefore, this technique does not seem to be helpful for preclinical diagnosis of OPA.

L'adénocarcinome pulmonaire ovin (OPA) est un cancer pulmonaire transmissible causé par le rétrovirus ovin de Jaggsiekte (JSRV). Il est difficile d'identifier les animaux infectés par le JSRV mais qui sont cliniquement en santé. Le virus n'entraine pas la production d'anticorps spécifiques et, bien que des séquences d'ADN provirales de JSRV peuvent être retrouvées dans les mononucléaires du sang, la détection est inconstante. L'objectif de la présente étude était d'examiner la présence de JSRV dans la moelle osseuse de moutons infectés et de développer une méthode de tamisage plus constante. L'examen par immunohistochime d'échantillons de la moelle osseuse de huit moutons asymptomatiques mais infectés par JSRV a révélé la présence de cellules positivement marquées. Toutefois, le JSRV ne put être révélé par une épreuve d'amplification en chaine par la polymérase (PCR) très sensible à partir d'aspirations de la moelle osseuse récolées périodiquement à partir de ces animaux. Les résultats suggèrent que les cellules infectées par JSRV peuvent être présentes dans la moelle osseuse d'animaux asymptomatiques, mais le nombre se situe sous le seuil détectable pas PCR. Ainsi, cette technique ne semble pas utile pour le diagnostic préclinique d'OPA.(Traduit par Docteur Serge Messier).

Effect of oral administration of Propionibacterium acnes on growth performance, DTH response and anti-OVA titers in goat kids

Immunostimulants are susbstances that stimuli the response of effector cells to activate the immune response such as antigen uptake, cytokine release or antibody response. These substances can increase resistence to infection by different types of microorganisms, reducing dependence of antibiotics used in livestock animals. Recent reports have demonstrated the positive effect of Propionibacterium acnes (P. acnes) to control animal diseases. In this study, we evaluated the effect of the non-specific immunostimulant P. acnes on immunological functions and growth performance in goat kids. Twenty five goat kids served as control group (A) and another 25 animals received P. acnes being the experimental group (B). Kids were challenged with ovalbumin (OVA) to assess humoral immunity. To assess in vivo cell immunity, delayed type hypersensitivity (DTH) test with phytohemagglutinin (PHA) was used, clinical signs and body weight were recorded each week until 9 weeks of age when the experiment ended. Blood samples were obtained to analyze serum proteins fractions and anti-OVA specific antibodies. No clinical signs of disease and no differences (p>0.05) on body weight between groups were recorded (7.32±0.81 kg in group A, 7.13±0.65 kg in group B). Goat kids from group B had more total protein (59.8±5g/l) and albumin levels (32.8±3.3g/l) than goat kids from group A (56.6±5.7 g/l, 29.6±3.9 g/l respectively) (p<0.05). DTH response in goat kids from group B on day 42 was higher (p<0.05) than group A. At day 63, goat kids from group receiving P. acnes had higher percentage (85.4) of anti-OVA IgM titers (p<0.05) than control group (57.7). In conclusion, the results showed that oral administration of P. acnes to goat kids improved some aspects of the immune system of the animals and it could be used to control goat diseases.

Roles of the Ras-MEK-mitogen-activated protein kinase and phosphatidylinositol 3-kinase-Akt-mTOR pathways in Jaagsiekte sheep retrovirus-induced transformation of rodent fibroblast and epithelial cell lines.

Jaagsiekte sheep retrovirus (JSRV) is the causative agent of ovine pulmonary adenocarcinoma (OPA), a transmissible lung cancer of sheep. The virus can induce tumors rapidly, and we previously found that the JSRV envelope protein (Env) functions as an oncogene, because it can transform mammalian and avian fibroblast cell lines. (N. Maeda, Proc. Natl. Acad. Sci. USA 98:4449-4454, 2001). The molecular mechanisms of JSRV Env transformation are of considerable interest. Several reports suggested that the phosphatidylinositol 3-kinase/Akt pathway is important for transformation of mammalian fibroblasts but not for chicken fibroblasts. In this study, we found that Akt/mTOR is involved in JSRV transformation of mouse NIH 3T3 fibroblasts, because treatment with the mTOR inhibitor rapamycin reduced transformation. We also found that H/N-Ras inhibitor FTI-277 and MEK1/2 inhibitors PD98059 and U0126 strongly inhibited JSRV transformation of NIH 3T3 fibroblasts, suggesting that the H/N-Ras-MEK-mitogen-activated protein kinase (MAPK) p44/42 pathway is necessary for the transformation. In RK3E epithelial cells, the MEK1/2 inhibitors also eliminated transformation, but FTI-277 only partially inhibited transformation. It was noteworthy that p38 MAPK inhibitors enhanced JSRV transformation in both fibroblasts and epithelial cells. Treatment of transformed cells with p38 inhibitors both increased levels of phospho-MEK1/2 and phospho-p44/42 and induced rapid enhancement of the transformed phenotype. Immunohistochemical staining of tumor tissues from naturally and experimentally induced OPA and naturally occurring enzootic nasal adenocarcinoma revealed strong activation of MAPK p44/42 in all cases examined. However, p38 activation was not generally observed. These results indicate that signaling through two pathways (in particular, H/N-Ras-MEK-MAPK and, to a lesser extent, Akt-mTOR) is important for JSRV-induced transformation and that p38 MAPK has a negative regulatory effect on transformation, perhaps via MEK1/2 and p44/42.