Marek’s disease virus and skin interactions

Marek’s disease virus (MDV) is a highly contagious herpesvirus which induces T-cell lymphoma in the chicken. This virus is still spreading in flocks despite forty years of vaccination, with important economical losses worldwide. The feather follicles, which anchor feathers into the skin and allow their morphogenesis, are considered as the unique source of MDV excretion, causing environmental contamination and disease transmission. Epithelial cells from the feather follicles are the only known cells in which high levels of infectious mature virions have been observed by transmission electron microscopy and from which cell-free infectious virions have been purified. Finally, feathers harvested on animals and dust are today considered excellent materials to monitor vaccination, spread of pathogenic viruses, and environmental contamination. This article reviews the current knowledge on MDV-skin interactions and discusses new approaches that could solve important issues in the future.     

Investigation of Marek’s disease virus from chickens in central Ethiopia

Marek’s disease (MD) is a lymphoproliferative and neuropathic disease of domestic chickens and less commonly, turkeys and quails, caused by a highly contagious, cell-associated, oncogenic herpesvirus. In Ethiopia, MD is believed to be introduced with importation of exotic and crossbred to improve the poultry production and has been reported to be a potential threat to the poultry sector both in backyard and commercial farming systems. This study was aimed at isolation and molecular analysis of MD virus isolates circulating in chicken population in the central part of Ethiopia where commercial farms are populated. From September 2013 to January 2014, clinical and post-mortem examination were conducted on diseased chickens suspected of MD virus infection. Representative spleen and feather follicle samples were collected following sterile procedure, and infectious virus isolation was performed using primary chicken fibroblast cell culture. Cell culture inoculated with suspension of pathological samples developed characteristic MD virus cytopathic effect of rounding of the cells and small plaques. Further analysis of the virus was conducted by conventional PCR amplifying the ICP4 gene fragment from eleven tissue samples using MD virus specific primers. PCR products were further sequenced and analyzed. Nucleotide sequence similarity search of the local isolates resulted a high degree of sequence similarity with Gallid Herpes virus type 2 strain (Marek’s disease virus type 1, JN034558). To our knowledge, the present study is the first report conducted on virus isolation and molecular characterization of MD virus isolates circulated in Ethiopia. Eleven ICP4-like gene fragment (318 bp) sequences generated in the present study were uploaded in the public database (KU842366–76). Further research on virus isolation, genetic characterization, and infection dynamics is recommended targeting chickens of all age groups reared in different agro-ecological zones under different production system.

     

Effects of interferon-γ knockdown on vaccine-induced immunity against Marek’s disease in chickens

Interferon (IFN)-γ has been shown to be associated with immunity to Marek’s disease virus (MDV). The overall objective of this study was to investigate the causal relationship between IFN-γ and vaccine-conferred immunity against MDV in chickens. To this end, 3 small interfering RNAs (siRNAs) targeting chicken IFN-γ, which had previously been shown to reduce IFN-γ expression in vitro, and a control siRNA were selected to generate recombinant avian adeno-associated virus (rAAAV) expressing short-hairpin small interfering RNAs (shRNAs). An MDV challenge trial was then conducted: chickens were vaccinated with herpesvirus of turkey (HVT), administered the rAAAV expressing shRNA, and then challenged with MDV. Tumors were observed in 4 out of 10 birds that were vaccinated with HVT and challenged but did not receive any rAAAV, 5 out of 9 birds that were administered the rAAAV containing IFN-γ shRNA, and 2 out of 10 birds that were administered a control enhanced green fluorescent protein siRNA. There was no significant difference in MDV genome load in the feather follicle epithelium of the birds that were cotreated with the vaccine and the rAAAV compared with the vaccinated MDV-infected birds. These results suggest that AAAV-based vectors can be used for the delivery of shRNA into chicken cells. However, administration of the rAAAV expressing shRNA targeting chicken IFN-γ did not seem to fully abrogate vaccine-induced protection.     

Expression kinetics of chicken β2-microglobulin and Class I MHC in vitro and in vivo during Marek’s disease viral infections

Marek’s disease virus (MDV) is a highly cell-associated herpesvirus that causes a disease in chickens characterized by tumor formation and immunosuppression. The changes of major histocompatibility complex (MHC) expression in different MDV-infected cells are not completely understood. In this study, we investigated the expression of the Class I MHC and β2-microglobulin (β2m) genes in response to MDV infection at different time points by real-time PCR. In both in vitro and in vivo, the expression levels of Class I MHC and β2m genes were upregulated during early MDV infections in comparison to control cells; We also found that the expression of Class I MHC gene was downregulated in BudR (5-bromo-2′-deoxyuridine)-treated MSB1 cells at 48 h and MDV-infected chicken embryo fibroblast cells (CEF) at 120 and 168 h post infection (hpi); Furthermore, compared to control groups, Class I MHC and β2m expression levels were downregulated in peripheral blood lymphocytes (PBLC) from MDV-infected chickens at 14 and 28 days post infection (dpi); Interestingly, both Class I MHC and β2m gene expression levels increased again in PBLC from MDV RB1B-infected chickens at 35 dpi, in which MDV was in the latent or transformed infection stages. In addition, Class I MHC expression was clearly decreased in MDV-infected CEF at 120 hpi although β2m expression was significantly increased. These changes in Class I MHC and β2m gene expression might provide more insights into host-virus interaction.     

An immuno-epidemiological model for Johne’s disease in cattle

To better understand the mechanisms involved in the dynamics of Johne’s disease in dairy cattle, this paper illustrates a novel way to link a within-host model for Mycobacterium avium ssp. paratuberculosis with an epidemiological model. The underlying variable in the within-host model is the time since infection. Two compartments, infected macrophages and T cells, of the within-host model feed into the epidemiological model through the direct transmission rate, disease-induced mortality rate, the vertical transmission rate, and the shedding of MAP into the environment. The epidemiological reproduction number depends on the within-host bacteria load in a complex way, exhibiting multiple peaks. A possible mechanism to account for the switch in shedding patterns of the bacteria in this disease is included in the within-host model, and its effect can be seen in the epidemiological reproduction model.     

A suspected case of Addison’s disease in cattle

A 4.75-year old Simmental cow was presented with symptoms of colic and ileus. The clinical signs and blood analysis resulted in the diagnosis of suspected primary hypoadrenocorticism (Addison’s disease). Although Addison’s disease has been frequently described in other domestic mammals, to our knowledge, this disease has not previously been reported in cattle.     

Antimicrobial susceptibility and molecular subtypes of Staphylococcus aureus isolated from pig tonsils and cow’s milk in China

This study investigated and compared the antimicrobial resistance patterns and ribotypes of Staphylococcus aureus isolated from pig tonsils and cow’s milk in China. A total of 90 isolates of S. aureus was included: 42 strains were isolated from tonsils of pigs and 48 from half-udder milk. The broth microdilution method and the double-disc diffusion test (D test) were used for antimicrobial susceptibility testing. The mecA gene for methicillin-resistant S. aureus (MRSA) and the ermA, ermB, ermC, and msrA genes for erythromycin-resistant strains were detected by polymerase chain reaction (PCR). The isolates were ribotyped with the Riboprinter system. The highest frequency of resistance was observed with clindamycin (91.1%), followed by penicillin (90.0%), and erythromycin (85.6%). All strains were susceptible to vancomycin and trimethoprim-sulfamethoxazole. The D test showed that 54.5% (42/77) of erythromycin-resistant isolates had the constitutive resistance phenotype and 45.5% (35/77) had the inducible resistance phenotype to clindamycin. A higher proportion of resistance to cephalosporins, macrolides, fluoroquinolones, and pleuromutilins was observed in pig isolates than in milk isolates (P < 0.05). The mecA gene was detected in all MRSA isolates; 89.6% of erythromycin-resistant strains harbored the ermC gene and 16.9% harbored the ermB gene. A total of 35 different ribogroups was found among the isolates investigated; 83.3% of pig strains belonged to 1 cluster with a similarity coefficient of 0.84. In contrast, 3 main clusters were observed among 68.8% of milk strains, which indicates a high degree of host specificity.     

A high-morbidity outbreak of Johne’s disease in game-ranched elk

Following an outbreak of Johne’s disease on an elk farm in northern Alberta, Canada, fecal culture, fecal polymerase chain reaction (PCR), and serum enzyme-linked immunosorbent assay (ELISA) tests were performed on individual animals. The magnitude of the outbreak is described and the challenges associated with poor test agreement, as well as herd management options, are discussed.     

Agent-based model for Johne’s disease dynamics in a dairy herd

Johne’s disease is an infectious gastrointestinal disease in ruminants caused by Mycobacterium avium subsp. paratuberculosis that causes diarrhea, emaciation, decreased milk production and eventually death. The disease is transmitted in utero and via milk and colostrums to calves, and fecal-orally to all age classes. Financial losses due to the disease are estimated to be over $200 million in the US dairy industry. The goal of this study was to evaluate the cost effectiveness of control measures based on diagnosis with a sensitive ELISA, EVELISA. An agent-based, discrete time model was developed to simulate Johne’s disease dynamics in a US dairy herd. Spatial aspects of disease transmission were taken into account by using six spatial compartments. The effects on disease prevalence were studied with and without transmission routes included in the model. Further, using the model, cost effectiveness of ELISA-based Johne’s disease control was evaluated. Using the parameters we collected and assumed, our model showed the initial prevalence of Johne’s disease (33.1 ± 0.2%) in the farm increased to 87.7 ± 1.7% in a 10 year-simulation. When ELISA-based control measures were included in the simulation, the increase in prevalence was significantly slowed down, especially when EVELISA was used. However, the level of the prevalence was still higher than the initial level after 10 year simulation even with the ELISA-based diagnostic intervention. The prevalence was further reduced when quarterly ELISA testing was included. The cost analysis showed that the quarterly ELISA and EVELISA testing could bring $44.8 and $51.5/animal/year more revenues, respectively, to a dairy farm.

Electronic supplementary material

The online version of this article (doi:10.1186/s13567-015-0195-y) contains supplementary material, which is available to authorized users.     

Evaluation of clinical pathology parameters in fecal PCR-positive or PCR-negative goats for Johne’s disease

Tropical Animal Health and Production - Johne’s disease (JD) is an economically important infectious disease of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). This...     

Fluorescent tagging of VP22 in N-terminus reveals that VP22 favors Marek’s disease virus (MDV) virulence in chickens and allows morphogenesis study in MD tumor cells

Marek’s disease virus (MDV) is an alpha-herpesvirus causing Marek’s disease in chickens, mostly associated with T-cell lymphoma. VP22 is a tegument protein abundantly expressed in cells during the lytic cycle, which is essential for MDV spread in culture. Our aim was to generate a pathogenic MDV expressing a green fluorescent protein (EGFP) fused to the N-terminus of VP22 to better decipher the role of VP22 in vivo and monitor MDV morphogenesis in tumors cells. In culture, rRB-1B EGFP22 led to 1.6-fold smaller plaques than the parental virus. In chickens, the rRB-1B EGFP22 virus was impaired in its ability to induce lymphoma and to spread in contact birds. The MDV genome copy number in blood and feathers during the time course of infection indicated that rRB-1B EGFP22 reached its two major target cells, but had a growth defect in these two tissues. Therefore, the integrity of VP22 is critical for an efficient replication in vivo, for tumor formation and horizontal transmission. An examination of EGFP fluorescence in rRB-1B EGFP22-induced tumors showed that about 0.1% of the cells were in lytic phase. EGFP-positive tumor cells were selected by cytometry and analyzed for MDV morphogenesis by transmission electron microscopy. Only few particles were present per cell, and all types of virions (except mature enveloped virions) were detected unequivocally inside tumor lymphoid cells. These results indicate that MDV morphogenesis in tumor cells is more similar to the morphorgenesis in fibroblastic cells in culture, albeit poorly efficient, than in feather follicle epithelial cells.     

Tyzzer’s disease in foals: Retrospective studies from 1969 to 2010

Reports of 148 cases of Tyzzer’s disease in foals in central Kentucky were analyzed to identify features of the disease and factors associated with it. The records indicate that Tyzzer’s disease is a rapidly progressive, highly fatal hepatitis caused by Clostridium piliforme. Common clinical findings are lethargy, fever, anorexia, and icterus. Seizures, coma, and death may rapidly ensue. Laboratory findings are leukopenia, metabolic acidosis, hypoglycemia, and increased activity of hepatic enzymes. Diagnosis is primarily based on clinical signs and postmortem findings but a polymerase chain reaction (PCR) is now available to detect C. piliforme DNA in organs and feces. Disease occurred most frequently in foals between 9 and 30 days of age that were born in April to May and was associated with heavy rainfall in the spring and high protein and nitrogenous diets fed to nursing mares. The findings are consistent with the ingestion of C. piliforme in the feces of adult horses and overgrowth in the intestine of foals with a high level of nutrients in their intestine.     

Seroprevalence of porcine reproductive and respiratory syndrome,Aujeszky’s disease,and porcine parvovirus in replacement gilts in Thailand

The present study investigated the seroprevalence of porcine reproductive and respiratory syndrome virus, Aujeszky’s disease virus (ADV), and porcine parvovirus (PPV) in replacement gilts from selected five swine herds in Thailand. The study consisted of three parts. First, a retrospective data analysis on the seroprevalence of porcine reproductive and respiratory syndrome virus (PRRSV) and ADV glycoprotein I (gI) in gilts, sows, boars, nursery, and fattening pigs in five herds (n = 7,030). Second, a cross-sectional study on seroprevalence of PRRSV, ADV, and PPV (n = 200) in replacement gilts. Last, the seroprevalence of PRRSV, ADV, and PPV in gilts culled due to reproductive failure (n = 166). Across the herds, the seroprevalence of PRRSV and ADV was 79.3% and 5.3%, respectively. The cross-sectional study revealed that 87.5%, 4.0%, and 99.0% of the replacement gilts were infected with PRRSV, ADV, and PPV, respectively. In the gilts culled due to reproductive failure, the seroprevalence of PRRSV, ADV, and PPV was 73.5%, 28.3%, and 86.0%, respectively. Of these culled gilts, 75.5% had been infected with at least two viruses and 18.9% had been infected with all three viruses. It could be concluded that most of the replacement gilts were exposed to PRRSV (84%), PPV (97%), and ADV (4%) before entering the breeding house. PPV was an enzootic disease among the selected herds. The prevalence of ADV was higher in gilts culled due to reproductive disturbance than in the healthy gilts.     

Prevalence and spectrum of Johne’s disease lesions in cattle slaughtered at two abattoirs in Kampala, Uganda

This study was conducted to determine the prevalence and characteristics of Johne’s disease (JD) lesions in Ugandan cattle slaughtered at two of the main abattoirs in Kampala. Ileocaecal junction and the associated lymph nodes of 1,022 cattle were examined for gross and microscopic lesions, followed by Ziehl Neelsen staining of the tissues bearing lesions. Confirmation of Mycobacterium avium subsp. paratuberculosis infection was done in some of the tissues using culture and IS900 PCR. The lesions were then described, characterised and tabulated. Characteristic Johne’s disease granulomas were found in 4.7% of the samples examined, derived from Zebu, Ankole longhorn, Friesian breeds of cattle and their crosses. Lesions were found both in the lymph nodes and ileocaecal junction mucosa. The lesions tended to be more severe in the lymph node than in the mucosa. There were also some unique and atypical lesions found in association with Johne’s disease granulomas. The diagnostic values of various gross lesions and criteria of lesion classifications and diagnosis are revisited and discussed based on the findings of this study. The prevalence of Johne’s disease lesions among slaughtered cattle in Kampala’s two abattoirs indicates that the disease is well established in the cattle population in the country. The diverse manifestations in lesions of JD need to be considered when making histological diagnosis in tissues where the disease is suspected.     

Breeding for resistance to footrot – the use of hoof lesion scoring to quantify footrot in sheep

So that genetic studies can be undertaken on footrot in sheep, it is necessary that a reliable and repeatable method to categorise the phenotype is available. This paper summarises the methods used and results obtained from 1600 hoof lesion scores of 100 mixed-age ewes independently scored twice by two trained operators. Using a 5-pont scale describing the severity of foot lesions, residual correlations were used to assess agreement between scorers and scoring occasions. Data were analysed using both zero-1 and continuous data methods. The average prevalence of any score >0 was 15%, and of scores >1 was 12%. The residual correlation between scorers for SUM_FR was 0.87 and between scoring occasions it was also 0.87, indicating high repeatability or agreement both within and between scorers. No significant differences were detected between scorers or between scoring occasions for any of the traits analysed, or different analytical methods used.     

Complete nucleotide sequence analysis of the oncogene “Meq” from serotype 1 Marek’s disease virus isolates from India

1. A study was undertaken to characterise the oncogene Meq at the molecular level for three serotype 1 Marek’s disease virus (MDV) field isolates from vaccinated poultry flocks which had encountered a Marek’s disease outbreak in the southern part of India. The isolates were named Ind/TN/11/01, Ind/KA/12/02 and Ind/TN/12/03. The oncogene Meq was amplified by PCR and sequenced.

2. The isolates were shown to have a homology for the Meq gene of 99.1–99.8% with various isolates from China and 98.5–99.2% with isolates from Europe and the USA. Alignment analysis of the nucleotide sequences showed that nucleotide mutations at 5 different positions in the Meq gene displayed perfect regularity in MDVs circulating in the southern part of India, which could be considered as features of field MDVs recently prevalent in this area.

3. In addition, the mutation in the Meq gene at positions 251, 260 and 437 was unique and coincides with very virulent strains from China GX0101, GXY2 and a Hungarian strain ATE. The mutation at positions 283 and 300 was unique and coincides with the very virulent strain ATE of Hungary. There were also single nucleotide mutations at positions 155 (A–T), 369 (A–C), 462 (C–A) and 548 (C–T) observed in the isolate Ind/TN/12/03.

4. Phylogenetic analysis of Meq sequences revealed that field MDVs in this area evolved independently but have similarities with very virulent strains from China, and that Meq has more similarities with the very virulent Hungarian strain.     

Veterinarians’ perspective on a voluntary Johne’s disease prevention program in Ontario and western Canada

A survey was conducted to assess the beliefs of veterinarians on Johne’s disease (JD) and their attitudes towards the Canadian, risk assessment based, JD prevention program. The veterinarians surveyed believed Mycobacterium avium subsp. paratuberculosis may have zoonotic potential, liked the risk assessment based program, and thought it could lead to the prevention of other on-farm diseases.