Evaluation of chickens infected with a recombinant virulent NDV clone expressing chicken IL4.

BACKGROUND: There is evidence that chicken IL4 (chIL4) functions similarly to its mammalian analogue by enhancing type 2 T helper (Th2) humoral immunity and promoting protection against parasitic infections; however, no studies have been performed to assess the effect of chIL4 on the pathogenesis of Newcastle disease (ND). To assess the role of chIL4 in velogenic NDV pathogenesis we created a vNDV infectious clone expressing chIL4. We hypothesized that co-expression of chIL4 during virus replication would result in decreased inflammation caused by the Th1 response and thereby increasing survival to challenge with vNDV. METHODS: To evaluate the effect of chIL4 during early infection with velogenic Newcastle disease virus (NDV) in chickens, recombinant NDV clones expressing either chIL4 (rZJ1-IL4) or a control expressing green fluorescent protein (rZJ1-GFP) were created by inserting an expression cassette in an intergenic region of the NDV genome. The pathogenesis of rZJ1-IL4 was assessed in 4-week-old specific pathogen free chickens. The extent of virus replication was evaluated by titration in mucosal secretions and immunohistochemistry in multiple tissues. Expression of chIL4 was confirmed in tissues using immunohistochemistry. RESULTS: Infection of birds with the rZJ1-IL4 resulted in successful viral replication in vivo and in vitro and generation of the chIL4 in tissues. All birds were clinically normal 2 DPI, with one bird in each group showing conjunctival swelling and enlarged spleens grossly. At 5 DPI, moderate or severe depression was observed in birds infected with rZJ1-GFP or rZJ1-IL4, respectively. Neurological signs and thymic atrophy were observed in one bird infected with rZJ1-IL4. Grossly, conjunctival swelling, mottled spleen and proventricular hemorrhages were observed at 5 DPI in one bird from each group. At 5 DPI, severe necrosis in the spleen, bursa and cecal tonsils were observed in birds infected with rZJ1-GFP, along with minimal evidence of chIL4 expression. In contrast, splenic atrophy, and moderate necrosis in the bursa and cecal tonsils were observed in birds infected with rZJ1-IL4. In addition, chIL4 signal was found in all tissues of rZJ1-IL4 birds at 5DPI. CONCLUSIONS: The production of chIL4 by a recombinant NDV strain resulted in the activation of the positive feedback loop associated with IL4 production. Insertion of chIL4 into NDV may decrease necrosis to lymphoid organs while increasing the severity of lymphoid atrophy and prolonged disease. However, with a low number of birds it is difficult to determine whether these results are significant to disease outcome.

Gene expression and linkage analysis implicate CBLB as a mediator of rituximab resistance.

Elucidating resistance mechanisms for therapeutic monoclonal antibodies (MAbs) is challenging, because they are difficult to study in non-human models. We therefore developed a strategy to genetically map in vitro drug sensitivity, identifying genes that alter responsiveness to rituximab, a therapeutic anti-CD20 MAb that provides significant benefit to patients with B-cell malignancies. We discovered novel loci with genome-wide mapping analyses and functionally validated one of these genes, CBLB, which causes rituximab resistance when knocked down in lymphoma cells. This study demonstrates the utility of genome-wide mapping to discover novel biological mechanisms of potential clinical advantage.

Pathogenesis of New Strains of Newcastle Disease Virus From Israel and Pakistan.

In the past few years, Newcastle disease virus (NDV) strains with epizootic characteristics belonging to subgenotypes VIIi and XIIIb emerged in the Middle East and Asia. In this study, 2 NDV strains-1 representative of subgenotype VIIi isolated in Israel (Kvuzat/13) and 1 representative of subgenotype XIIIb isolated in Pakistan (Karachi/07)-were characterized by intracerebral pathogenicity index and detailed clinicopathologic assessment. The intracerebral pathogenicity index values for Kvuzat/13 and Karachi/07 were 1.89 and 1.85, respectively, classifying these strains as virulent by international standards. In 4-week-old White Leghorn chickens, both strains caused 100% mortality within 4 (Kvuzat/13) and 5 (Karachi/07) days postinfection. Histopathology and immunohistochemistry for NDV nucleoprotein showed that both strains had wide systemic distribution, especially targeting lymphoid organs and mucosa-associated lymphoid tissues in the respiratory and intestinal tracts. Results of the animal experiment confirm that both Kvuzat/13 and Karachi/07 are highly virulent and behaved as velogenic viscerotropic NDV strains.

Neuropathogenic Capacity of Lentogenic, Mesogenic, and Velogenic Newcastle Disease Virus Strains in Day-Old Chickens.

Strains of Newcastle disease virus (NDV) have different abilities to elicit neurologic signs. To determine the capacity of different NDV strains to replicate and cause lesions in the brain, independently of their peripheral replication, 1-day-old chickens were inoculated in the subdural space with 7 NDV strains of different virulence (4 velogenic, 2 mesogenic, 1 lentogenic). Velogenic strains induced severe necrotizing and heterophilic ventriculitis and meningitis, as well as edema of the neuroparenchyma, and replicated extensively in the nervous tissue by day 2 postinfection, as demonstrated by immunohistochemistry, when all infected birds died. Clinical signs, microscopic lesions, and viral replication were delayed (days 3 and 4 postinfection) with mesogenic strains. Velogenic and mesogenic NDV strains replicated mainly in neurons, and immunolabeling was first detected in surface-oriented areas (periventricular and submeningeal), possibly as a reflection of the inoculation route. The lentogenic NDV strain did not cause death of infected birds; replication was confined to the epithelium of the ependyma and choroid plexuses; and lesions consisted of lymphoid aggregates limited to the choroid plexuses. Results show that extensive NDV replication in the brain is typical of velogenic and mesogenic, but not lentogenic, NDV strains. In addition, this study suggests that differences in the rate of NDV replication in nervous tissue, not differences in neurotropism, differentiate velogenic from mesogenic NDV strains. This study indicates that intracerebral inoculation might be used as an effective method to study the mechanisms of NDV neuropathogenesis.

Pathologic characterization of genotypes XIV and XVII Newcastle disease viruses and efficacy of classical vaccination on specific pathogen-free birds.

To characterize the clinicopathologic features of recently described genotypes of Newcastle disease virus (NDV), 1 representative strain of genotype XIV and 2 of genotype XVII, all isolated from West Africa, were used to infect groups of ten 4-week-old specific pathogen-free chickens. The pathobiology of these 3 strains was compared to a South African NDV strain classified within genotype VII. All chickens infected with the 4 viruses died or were euthanized by day 4 postinfection due to the severity of clinical signs. Gross and histologic lesions in all infected chickens included extensive necrosis of lymphoid tissues (thymus, spleen, bursa of Fabricius, cecal tonsils, gut-associated lymphoid tissue), gastrointestinal necrosis and hemorrhages, and severe hemorrhagic conjunctivitis. Immunohistochemical staining revealed systemic viral distribution, and the most intense staining was in the lymphoid organs. Results demonstrate that the 3 West African strains from the previously uncharacterized genotypes XIV and XVII are typical velogenic viscerotropic NDV strains with lesions similar to the South African strain. Under experimental conditions, QV4 and LaSota NDV vaccine strains successfully protected chickens from morbidity and mortality against the genotype VII and one genotype XVII NDV strain, with no significant differences in the amount of virus shed when 2 vaccine schemes were compared.

Green algal peritonitis in 2 cows.

Peritonitis due to infections with green algae was diagnosed at slaughter (in Texas and South Dakota) in 2 cows. One cow also had a generalized lymphadenitis. The intralesional green algae were histologically similar to those previously associated with bovine lymphadenitis. Amplified and sequenced algal ITS2 genes had higher homology with the genus Scenedesmus than with Chlorella.

Spontaneous autoimmune gastritis and hypochlorhydria are manifest in the ileitis-prone SAMP1/YitFcs mice.

SAMP1/YitFcs mice serve as a model of Crohn's disease, and we have used them to assess gastritis. Gastritis was compared in SAMP1/YitFcs, AKR, and C57BL/6 mice by histology, immunohistochemistry, and flow cytometry. Gastric acid secretion was measured in ligated stomachs, while anti-parietal cell antibodies were assayed by immunofluorescence and enzyme-linked immunosorbent spot assay. SAMP1/YitFcs mice display a corpus-dominant, chronic gastritis with multifocal aggregates of mononuclear cells consisting of T and B lymphocytes. Relatively few aggregates were observed elsewhere in the stomach. The infiltrates in the oxyntic mucosa were associated with the loss of parietal cell mass. AKR mice, the founder strain of the SAMP1/YitFcs, also have gastritis, although they do not develop ileitis. Genetic studies using SAMP1/YitFcs-C57BL/6 congenic mice showed that the genetic regions regulating ileitis had comparable effects on gastritis. The majority of the cells in the aggregates expressed the T cell marker CD3 or the B cell marker B220. Adoptive transfer of SAMP1/YitFcs CD4(+) T helper cells, with or without B cells, into immunodeficient recipients induced a pangastritis and duodenitis. SAMP1/YitFcs and AKR mice manifest hypochlorhydria and anti-parietal cell antibodies. These data suggest that common genetic factors controlling gastroenteric disease in SAMP1/YitFcs mice regulate distinct pathogenic mechanisms causing inflammation in separate sites within the digestive tract.

Clinicopathological characterization in poultry of three strains of Newcastle disease virus isolated from recent outbreaks.

Newcastle disease is a severe threat to the poultry industry and is caused by Newcastle disease virus, a member of the genus Avulavirus, family Paramyxoviridae. The virus is rapidly evolving, and several new genotypes have been discovered in the past few years. Characterization of these strains is important to evaluate field changes, anticipate new outbreaks, and develop adequate control measures. Three Newcastle disease isolates (APMV-1/duck/Vietnam, Long Bien/78/2002, APMV-1/chicken/Australia/9809-19-1107/1998, and APMV-1/double-crested cormorant/USA, Nevada/19529-04/2005) from recent outbreaks were investigated via clinicopathological assessment, immunohistochemistry (IHC), in situ hybridization, virus isolation, and serology in experimentally infected 4-week-old chickens. Phylogenetic studies showed that Australia isolate belongs to class II genotype I, Long Bien to class II genotype VIId, and Nevada cormorant to class II genotype V. Even though all 3 viruses had a virulent fusion protein cleavage site and ICPI values greater than 1.5, they all differed in their ability to cause clinical signs, in their lesions, and in their viral distribution in body tissues. The Long Bien isolate showed the most severe clinicopathological picture and the most widespread viral distribution. The Australia and Nevada cormorant isolates had a milder pathological phenotype, with viral replication restricted to only a few organs. The variability in clinicopathological characteristics despite the similarity in ICPI suggests that full clinicopathological assessment is necessary to fully characterize new isolates and that there are differences in pathogenesis among viruses of different genotypes.

Lesion development and replication kinetics during early infection in cattle inoculated with Vesicular stomatitis New Jersey virus via scarification and black fly (Simulium vittatum) bite.

Vesicular stomatitis viruses are the causative agents of vesicular stomatitis, an economically important contagious disease of livestock that occurs in North, Central, and South America. Little is known regarding the early stages of infection in natural hosts. Twelve adult Holstein steers were inoculated with Vesicular stomatitis New Jersey virus (VSNJV) on the coronary bands (CB) of the feet via scarification (SC) or by VSNJV-infected black fly (Simulium vittatum) bite (FB). Three additional animals were inoculated on the neck skin using FB. Clinical disease and lesion development were assessed daily, and animals were euthanatized from 12 hours post inoculation (HPI) through 120 HPI. The animals inoculated in the neck failed to develop any clinical signs or gross lesions, and VSNJV was detected neither by in situ hybridization (ISH) nor by immunohistochemistry (IHC). Lesions on the CB were more severe in the animals infected by FB than by SC. In both groups, peak VSNJV replication occurred between 24 and 48 HPI in keratinocytes of the CB, as evidenced by ISH and IHC. There was evidence of viral replication limited to the first 24 HPI in the local draining lymph nodes, as seen through ISH. Successful infection via FB required logarithmically less virus than with the SC technique, suggesting that components in black fly saliva may facilitate VSNJV transmission and infection in cattle. The lack of lesion development in the neck with the same method of inoculation used in the CB suggests that specific characteristics of the CB epithelium may facilitate VSNJV infection.

Pathogenesis of a bovine enterovirus-1 isolate in experimentally infected calves.

The pathogenesis and virulence of Bovine enterovirus-1 (BEV-1) in cattle is largely unknown. Reports concerning its virulence suggest that there might be an association between BEV-1 infections and a range of diseases in cattle that vary from respiratory to enteric to reproductive disease and infertility. In the current study, the pathogenesis associated with acute infection of BEV-1 in calves experimentally inoculated with the Oklahoma isolate of BEV-1 was described. Although interpretation of the study was limited by lack of an effective control group, results suggest that an association between inoculation of BEV-1, virus localization, and the potential development of lesions in the brain and heart probably exists. In the experiment, BEV-1 virus localized to the terminal ileum, ileocecal and cecocolonic junctions, spiral colon, and ileocecal lymph nodes; BEV-1 virus was detected in the cytoplasm of enterocytes, lamina propria macrophages, endothelium, neurons of the submucosal and myenteric plexi, and lymphocytes of the submucosal lymphoid tissue. Although no clinical signs were noted following acute infection, BEV-1 was localized in the cerebellar white matter of a calf with encephalitis and in the heart of another calf with coronary arteritis. The current study suggests that the BEV-1 isolate is infectious to young calves and that BEV-1 potentially can have a similar pathogenesis to that observed in natural or experimental enterovirus infections in other species.

Immunohistochemical study of rabies virus within the central nervous system of domestic and wildlife species.

Immunohistochemistry using a commercial polyclonal antibody for lyssavirus was applied to 39 archival cases of rabies. Paraffin blocks from 13 different species were available, including 3 dogs, 4 cats, 1 pig, 6 cattle, 4 horses, 1 llama, 7 skunks (Mephitis mephitis), 7 raccoons (Procyon lotor), 1 bat (Myotis species), 1 white-tailed deer (Odocoileus virginianus), 1 bobcat (Lynx rufus), 2 gray foxes (Urocyon cinereoargenteus), and 1 red fox (Vulpes vulpes). All cases had previously been diagnosed as rabies using histopathology and/or fluorescent antibody testing. The immunohistochemistry technique successfully detected lyssavirus antigen in all cases. In species for which 3 or more samples were available, distributional trends were seen in 4 main brain regions: brainstem, cerebellum, hippocampus, and cerebrum. The best site for rabies virus detection in dogs and cats was the hippocampus. For cattle, viral antigen was most prominent in the brainstem, followed by the cerebellum. In horses, the cervical spinal cord and adjacent brainstem were the optimal sites for detecting rabies virus antigen. In raccoons and skunks, positive labeling was widely dispersed, so selection might be less important for these wildlife reservoir species. Immunohistochemistry should prove useful in enhancing the accuracy of rabies diagnosis through informed selection of brain sampling sites when composite sampling is not feasible. This immunohistochemical technique could provide reliable virus detection in formalin-fixed tissues in any potentially infected species.

Agricultural diseases on the move early in the third millennium.

With few exceptions, the diseases that present the greatest risk to food animal production have been largely similar throughout the modern era of veterinary medicine. The current trend regarding the ever-increasing globalization of the trade of animals and animal products ensures that agricultural diseases will continue to follow legal and illegal trade patterns with increasing rapidity. Global climate changes have already had profound effects on the distribution of animal diseases, and it is an inevitable reality that continually evolving climatic parameters will further transform the ecology of numerous pathogens. In recent years, many agricultural diseases have given cause for concern regarding changes in distribution or severity. Foot-and-mouth disease, avian influenza, and African swine fever continue to cause serious problems. The expected announcement of the global eradication of rinderpest is one of the greatest successes of veterinary preventative medicine, yet the closely related disease peste des petits ruminants still spreads throughout the Middle East and Asia. The spread of novel strains of bluetongue virus across Europe is an ominous indicator that climate change is sure to influence trends in movement of agricultural diseases. Overall, veterinary practitioners and investigators are advised to not only maintain vigilance against the staple disease threats but to always be sufficiently broad-minded to expect the unexpected.

Long-term management of the liver transplant patient: recommendations for the primary care doctor.

No official document has been published for primary care physicians regarding the management of liver transplant patients. With no official source of reference, primary care physicians often question their care of these patients. The following guidelines have been approved by the American Society of Transplantation and represent the position of the association. The data presented are based on formal review and analysis of published literature in the field and the clinical experience of the authors. These guidelines address drug interactions and side effects of immunosuppressive agents, allograft dysfunction, renal dysfunction, metabolic disorders, preventive medicine, malignancies, disability and productivity in the workforce, issues specific to pregnancy and sexual function, and pediatric patient concerns. These guidelines are intended to provide a bridge between transplant centers and primary care physicians in the long-term management of the liver transplant patient.

Effect of heparin on synthesis of short chain collagen by chondrocytes and smooth muscle cells.

The treatment of embryonic chick chondrocyte cultures with heparin results in a decrease in collagen synthesis. One of the collagens synthesized by hypertrophic chondrocytes, specifically type X collagen, may play an important role in cartilage mineralization and endochondral ossification. Recently a new short chain collagenous component was found in cultures of rat vascular smooth muscle cells (Majack, R. A., and P. Bornstein, 1985, J. Cell Biol., 100: 613-619). The present study was initiated to investigate heparin's effect on type X collagen in embryonic chick chondrocytes and to further evaluate the nature of the short chain component synthesized by rat vascular smooth muscle cells. Different tissues may respond differently to the administration of heparin. In chondrocyte cultures heparin decreased both total collagen synthesis as well as the synthesis of type X collagen. There was an accumulation of collagen precursors, found principally in the cell layer compartment, which appeared to be the result of heparin's inhibition of the NH2-terminal protease. In cultures of rat vascular smooth muscle cells heparin was found to increase the synthesis of a short chain collagenous component as previously reported. However, comparison with a type X collagen standard showed this to be different from type X. In all cases, the effect of heparin on collagen chain precursors, chondrocyte type X synthesis, and synthesis of a vascular smooth muscle short chain collagen was shown to be reversible. Similar effects were obtained by adding chondroitin sulfate to chondrocytes, suggesting a role for extracellular matrix components in the modulation of collagen synthesis. These findings are consistent with the concept of a group of short chain collagens with type X collagen being unique to hypertrophic chondrocytes.

13-cis-retinoic acid: inhibition of bladder carcinogenesis in the rat.

Transitional cell and squamous cell cancer of the bladder was induced in Wistar/Lewis female rats by direct instillation of N-methyl-N-nitrosourea into the bladder. Feeding of the synthetic retinoid, 13-cis-retinoid acid, inhibited the incidence and extent of bladder cancer in these rats, even when 13-cis-retinoic acid administration was begun after completion of the carcinogen treatment.

13-cis-Retinoic acid: inhibition of bladder carcinogenesis induced in rats by N-butyl-N-(4-hydroxybutyl)nitrosamine.

Transitional cell carcinoma was induced in the bladders of male Fischer rats by 12 oral doses of the carcinogen N-butyl-N-(4-hydroxybutyl)nitrosamine. Feeding of 13-cis-retinoic acid after completion of carcinogen treatment diminished the number and severity of cancers and other proliferative lesions of the bladder.

Essential veterinary education in the cultural, political and biological complexities of international trade in animals and animal products.

Globalisation has changed the veterinary profession in many ways and academic institutes may need to re-tool to help future professionals deal with the changes in a successful and productive way. The remarkably expanded and expanding volume of trade and traffic in animals and animal products means that to be effective veterinarians must grasp some of the complexities inherent in this trade. Being able to engage productively in cross-cultural dialogue will be important in negotiations over livestock shipments and also within the context of the delivery of medical services to companion animals in societies that are becoming increasingly diverse. Understanding the political landscapes that influence trade decisions will help to expedite agreements and facilitate the transfer of goods and materials that involve animal health. Disease emergence will continue to occur, and an awareness of the factors responsible and the response measures to undertake will help to contain any damage.

Localization of fibropapilloma-associated turtle herpesvirus in green turtles (Chelonia mydas) by in-situ hybridization.

Fibropapilloma-associated turtle herpesvirus (FPTHV) is the presumed aetiological agent of sea turtle fibropapillomatosis (FP). Intralesional DNA and RNA of the virus have been detected by polymerase chain reaction (PCR) and reverse transcriptase-PCR (RT-PCR), respectively, but the exact location and distribution of the virus within the tumours have not been addressed. In this study, in-situ hybridization (ISH) was used to investigate viral transcriptional activity and localization of FPTHV. Twenty-five tumours were obtained from the skin or conjunctiva of 105 green turtles (Chelonia mydas) examined on two islands in Puerto Rico (Culebra and Culebrita). These lesions comprised 19 fibropapillomas and six fibromas. FPTHV mRNA transcripts were detected by ISH in three fibropapillomas, with positive reactions confined to the nuclei of clusters of epithelial cells. Viral DNA was detected by riboprobe ISH combined with denaturation in 14 tumours, including both fibropapillomas and fibromas. Signals were confined to the nuclei of acanthotic epithelial cells and were not seen in the subepithelial fibrous areas of the tumours. These results suggest that FPTHV is present in epithelial cells and transcriptionally active in fibropapillomas.

Pathology and Distribution of Velogenic Viscerotropic Newcastle Disease Virus in the Reproductive System of Vaccinated and Unvaccinated Laying Hens (Gallus gallus domesticus) by Immunohistochemical Labelling.

This study investigated the pathological changes in the reproductive system of laying hens that lead to the poor egg production and quality in Newcastle disease (ND) and the distribution of the virus in the system. Two hundred and forty Isa-Brown pullets were divided randomly into vaccinated and unvaccinated groups (n = 120 each). The vaccinated group was given Hitchner B1 vaccine at 1 day of age, La Sota vaccine at 4 weeks of age and Komarov vaccine at 9 and 16 weeks of age. At the peak of egg production, the laying hens (32 weeks old) were assigned randomly into four groups (n = 60): VC, vaccinated with ND vaccines and inoculated intramuscularly with velogenic viscerotropic ND virus (vvNDV); VU, vaccinated unchallenged; UC, unvaccinated challenged; and UU, unvaccinated unchallenged. UC hens showed depression, diarrhoea and later torticollis. Mortality in UC hens was 90%. VC hens showed mild anorexia. The body weights of the UC hens were significantly (P <0.05) lower than those of UU hens. VC and UC hens showed a significant (P <0.05) drop in egg production. Only UC hens produced abnormal eggs and initially had swollen, oedematous, hyperaemic oviducts followed by atrophy and shortening of the reproductive tract with atresia of the ovarian follicles. The histopathological changes were of necrosis of the epithelium and secretory glands. VC hens showed mild inflammatory changes in the oviduct. Immunohistochemical labelling showed extensive presence of the virus in the ovary, infundibulum, magnum, isthmus, uterus and vagina of UC hens and in the ovary of VC hens. These changes will be the cause of serious egg production problems, especially in vaccinated layers in countries where vvNDV is enzootic.

Statistical aspects of extrapolation of dichotomous dose-response data.

A mathematical model based on the hypothesis that carcinogenesis is a multistage process was proposed for the statistical problem of extrapolating the results of bioassay experiments performed at high dose levels of the carcinogenic risk at lower dose levels. This general multistage model had extrapolation characteristic similar to those of the more specific models that are commonly used; a graphic technique based on statistical likelihood procedures provided a measure of the uncertainty in this extrapolation problem. The examples used were 1) exposure to DES and mammary carcinoma and 2) exposure to DMN and liver carcinoma.