Do UK sheep farmers use orf vaccine correctly and could their vaccination strategy affect vaccine efficacy?

Orf, a parapoxvirus, is a zoonosis causing a contagious pustular dermatitis, and has a high morbidity in sheep worldwide. Despite a vaccine being available, orf prevalence in England is estimated to be 2 per cent in ewes and 20 per cent in lambs​​. There is concern that farmers are not complying with the vaccination guidelines and therefore the objective of this study was to investigate if orf vaccine is used correctly on sheep farms in the UK and to identify barriers and motivators of sheep farmers to use the vaccine. The survey was completed by 570 respondents. The results show several areas of concern; only 27 per cent of respondents used the correct site (axilla), 37 per cent of respondents would use orf vaccine up to a week after opening a vial (shelf life is eight hours), 33 per cent of respondents would vaccinate their ewes too close to lambing and 73 per cent of respondents did not separate vaccinated and non-vaccinated animals (both leading to infection risk for non-vaccinated animals). When vaccinating, only 53 per cent of respondents were wearing gloves and 31 per cent washed their hands just before and immediately after vaccination. Results demonstrate that orf vaccination is not carried out correctly on all UK sheep farms, which is likely to affect vaccine efficacy. A concern around vaccine efficacy, the 'hassle' of the scratch administration, the 'risk of making it worse' and the zoonotic risk when vaccinating were the most common barriers for using orf vaccine, highlighting the importance of veterinary advice when prescribing orf vaccine.

The re-emerging of orf virus infection: A call for surveillance, vaccination and effective control measures.

Orf disease is known to be enzootic among small ruminants in Asia, Africa, and some other parts of the world. The disease caused by orf virus is highly contagious among small ruminant species. Unfortunately, it has been neglected for decades because of the general belief that it only causes a self-limiting disease. On the other hand, in the past it has been reported to cause huge cumulative financial losses in livestock farming. Orf disease is characterized by localized proliferative and persistent skin nodule lesions that can be classified into three forms: generalized, labial and mammary or genitals. It can manifest as benign or malignant types. The later type of orf can remain persistent, often fatal and usually causes a serious outbreak among small ruminant population. Morbidity and mortality rates of orf are higher especially in newly infected kids and lambs. Application of antibiotics together with antipyretic and/or analgesic is highly recommended as a supportive disease management strategy for prevention of subsequent secondary microbial invasion. The presence of various exotic orf virus strains of different origin has been reported in many countries mostly due to poorly controlled cross-border virus transmission. There have been several efforts to develop orf virus vaccines and it was with variable success. The use of conventional vaccines to control orf is a debatable topic due to the concern of short term immunity development. Following re-infection in previously vaccinated animals, it is uncommon to observe the farms involved to experience rapid virus spread and disease outbreak. Meanwhile, cases of zoonosis from infected animals to animal handler are not uncommon. Despite failures to contain the spread of orf virus by the use of conventional vaccines, vaccination of animals with live orf virus is still considered as one of the best choice. The review herein described pertinent issues with regard to the development and use of potential effective vaccines as a control measure against orf virus infection.

Prevalence, risk factors and vaccination efficacy of contagious ovine ecthyma (orf) in England.

Orf is a viral disease found in English sheep flocks which can cause economic losses. It is a zoonosis with little epidemiological research available in the UK. In 2012, 3000 questionnaires were sent to English sheep farms in order to investigate the prevalence of orf, determine vaccination efficacy and to identify some of the potential risk factors. The usable response rate was 25.4 per cent. The usable farms (N=762 in the years 2011 and 2012) were used to model the percentage of animals affected on the farm, and the probability of a farm being found with the disease. The disease prevalence (DP) was standardised for the year and calculated as 1.88 per cent for ewes and 19.53 per cent for lambs. The disease risk ratio (RR) for the use of the vaccine was calculated as 2.04 for ewes and 0.75 for lambs, and therefore, the study found that lamb vaccination was beneficial (RR <1). Weed infestation and an increased number of orphan lambs were associated with increased cases of orf. We conclude that the DP in ewes and lambs affect each other, though the impact is higher for lambs in the presence of increasing prevalence in ewes. A short lambing season lowers the probability of a farm experiencing cases of orf. Vaccination was effective in lambs but not in ewes, though lambs benefitted when ewes were vaccinated (reduced orf prevalence in lambs born from vaccinated ewes), probably because any unvaccinated ewes may have been carriers that could spread the virus to the new-born lambs.

Quantitative polymerase chain reaction: another tool to evaluate viable virus content in live attenuated orf vaccine.

A probe-based real-time polymerase chain reaction (PCR) assay based on the highly conserved DNA polymerase gene of orf virus (ORFV) for the quality control of attenuated orf vaccine is reported. Primary lamb testis (PLT) cells were infected with orf vaccine virus and harvested at a critical time point to obtain maximum viable virus content as determined by real-time PCR. DNA extracted from these harvests was subjected to real-time PCR. A critical time point for the harvesting of PLT cells infected with various log(10) dilutions of vaccine virus was found to be 42 h (highest slope of 3.335), which was obtained by comparing the slopes of standard curves of different time intervals. The assay was employed to evaluate viable virus content in different batches of orf vaccine. The titres estimated by real-time PCR and conventional TCID(50) were comparable with a correlation of 0.8169. Thus, the real-time PCR assay could provide an alternative method or supplementary tool to estimate live ORFV particles in attenuated orf vaccine.

Comparative study on three locally developed live orf virus vaccines for sheep in Saudi Arabia.

The epidemiology of orf virus infection in Saudi Arabia (SA) has been researched since 1990. The results obtained during this period indicate that the disease is widespread, has great economic impact and that no vaccine has been used against it. The present study compares the immunogenicity and protective efficacy of three locally developed live orf virus vaccines. Two of them differ in their passage history in Vero cell culture and the third was used as a virulent virus in glycerine buffer. To the best of the authors' knowledge, no similar comparative study has been conducted in the Middle East utilising three types of vaccines prepared from the same virus strain. Selection of the candidate seed orf virus and performance of the quality control tests were as laid out by the OIE for veterinary vaccine production. The vaccine seed virus was a field orf virus isolated from a previous orf outbreak in Saudi Arabia. A simple novel formula was developed to calculate the rate of reduction in the healing time (RHT %) in the challenged sheep. This allowed direct comparison of the efficacy of the three types of vaccines employed in the present study. The efficacy of each vaccine was tested on a cohort of local Noemi sheep.

Evaluation of homologous and heterologous protection induced by a virulent field strain of orf virus and an orf vaccine in goats.

OBJECTIVE: To evaluate cross protection provided by administration of contagious ecthyma vaccines against strains of orf virus in goats. ANIMALS: 126 Boer-Spanish crossbred goats (3 to 20 days old). PROCEDURES: 85 goats were vaccinated with a goat-derived contagious ecthyma vaccine. Of these, 41 were challenge exposed with the virus strain for the contagious ecthyma vaccine, 40 were challenge exposed with a more virulent field strain of orf virus, and 4 were lost to predation or died. Another 41 goats were vaccinated with a vaccine produced from a more virulent field strain of orf virus; of these, 18 were challenge exposed with the virus strain of the goat-derived contagious ecthyma vaccine, 18 were challenge exposed with the more virulent field strain of orf virus, and 5 were lost to predation or died. RESULTS: Vaccination with the goat-derived contagious ecthyma vaccine did not significantly reduce the number of goats with lesions or lesion severity caused by challenge exposure with the more virulent field strain of orf virus. Vaccination with the vaccine produced from the more virulent field strain of orf virus significantly reduced the number of goats with lesions attributable to challenge exposure with the virus strain of the goat-derived contagious ecthyma vaccine, but it failed to significantly reduce lesion severity. CONCLUSIONS AND CLINICAL RELEVANCE: Vaccination did not result in cross protection for the 2 strains of orf virus. This may have been attributable to antigenic differences and may be a factor in outbreaks of contagious ecthyma in vaccinated goats.

Development of orf virus as a bifunctional recombinant vaccine: surface display of Echinococcus granulosus antigen EG95 by fusion to membrane structural proteins.

The parapoxvirus, orf virus (ORFV) causes superficial skin lesions in infected sheep. Unattenuated ORFV is used globally to vaccinate against orf. Recombinant poxviruses are proven delivery systems and we investigated strategies to express the immunogenic Echinococcus granulosus peptide EG95 from ORFV with the aim of developing a recombinant bivalent vaccine. EG95 is an oncosphere protein of the cestode E. granulosus, a parasite responsible for causing cystic hydatid disease in a wide range of hosts including humans and grazing animals such as sheep. Recombinant viruses were produced in which EG95 was expressed by itself or fused to ORFV envelope-associated structural proteins 10 kDa and F1L. Infection studies in sheep showed that specific antibodies were produced against ORFV and EG95 and that the antibody levels against EG95 were comparable to that of animals immunized with purified EG95 in Quil A adjuvant, an immunization regime that is known to afford protection. A single exposure to the dual vaccine has potential for protecting lambs against orf and for priming against EG95 so as to respond strongly to a later injection of EG95 protein.

Orf virus DNA vaccines expressing ORFV 011 and ORFV 059 chimeric protein enhances immunogenicity.

BACKGROUND: ORFV attenuated live vaccines have been the main prophylactic measure against contagious ecthyma in sheep and goats in the last decades, which play an important role in preventing the outbreak of the disease. However, the available vaccines do not induce lasting immunity in sheep and goats. On the other hand, variation in the terminal genome of Orf virus vaccine strains during cell culture adaptation may affect the efficacy of a vaccine. Currently, there are no more effective antiviral treatments available for contagious ecthyma. RESULTS: We constructed three eukaryotic expression vectors pcDNA3.1-ORFV011, pcDNA3.1-ORFV059 and pcDNA3.1-ORFV011/ORFV059 and tested their immunogenicity in mouse model. High level expression of the recombinant proteins ORFV011, ORFV059 and ORFV011/ORFV059 was confirmed by western blotting analysis and indirect fluorescence antibody (IFA) tests. The ORFV-specific antibody titers and serum IgG1/IgG2a titers, the proliferation of lymphocytes and ORFV-specific cytokines (IL-2, IL-4, IL-6, IFN-γ, and TNF-α) were examined to evaluate the immune responses of the vaccinated mice. We found that mice inoculated with pcDNA3.1-ORFV 011/ORFV059 had significantly stronger immunological responses than those inoculated with pcDNA3.1-ORFV011, pcDNA3.1-ORFV059, or pcDNA3.1-ORFV011 plus pcDNA3.1-ORFV059. Compared to other vaccine plasmids immunized groups, pcDNA3.1-ORFV011/ORFV059 immunized group enhances immunogenicity. CONCLUSIONS: We concluded that DNA vaccine pcDNA3.1-ORFV011/ORFV059 expressing ORFV011 and ORFV059 chemeric-proteins can significantly improve the potency of DNA vaccination and could be served as more effective and safe approach for new vaccines against ORFV.

Investigation and analysis of a human orf outbreak among people living on the same farm.

Human orf is a viral zoonotic infection caused by Parapoxvirus. The skin lesions of human orf can be misdiagnosed as cutaneous anthrax leading to overtreatment and also fear. This study was conducted to analyze an outbreak which led to deaths among kids and lambs in the same flock, and skin lesions in some persons who were living on the same farm that were initially diagnosed as cutaneous anthrax by a practitioner. Eight patients with skin lesions and eleven persons who had no skin lesion were considered as patients and control groups, respectively. The cultures obtained from the lesions of all patients were negative for Bacillus anthracis. The diagnosis of skin lesions was done by clinical findings, histopathological examination and PCR as human orf. To be under 20 years of age, direct contact with the animals, and contact with flayed skin of sick animals were the risk factors for human orf (Odds Ratio 7.5; 95% Confidence Interval 1.02-54.54, OR 12.25; 95% CI:1.3-100.9, OR 16.67; 95% CI:1.65-148.20, respectively). Orf should be kept in mind in the differential diagnosis of skin lesions resembling anthrax. For control and prevention of orf, transmission routes should be known; good hand hygiene and other personal protective measures have to be implemented.

Ovine diseases. Orf.

Orf virus is an epitheliotropic DNA parapoxvirus with a worldwide distribution that induces acute pustular lesions in the skin of sheep, goats and man. Genetic mapping and sequencing of the orf virus genome have revealed that orf virus has a typical poxvirus distribution of genes, with those essential for viral DNA synthesis, replication and packaging located in the central region, and those involved in virulence concentrated in the terminal regions. The immune and inflammatory response to orf virus infection in the skin and local lymph is vigorous and typical of an anti-viral response, involving CD4+ helper and CD8+ cytotoxic T cells, interferons and antibodies. In spite of this, the virus can repeatedly infect sheep. Host acquired immunity involving CD4+ T cells and interferons is effective in controlling the extent of viral replication, but does not prevent reinfection. Several virus putative virulence genes have been identified. These are: viral homologues of ovine vascular endothelial growth factor (VEGF); ovine IL-10; vaccinia virus E3L interferon resistance gene; and in addition a viral activity that inhibits the cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF). These may be responsible for rescuing orf virus, at least temporarily, from host immunity and aiding viral replication in epidermal cells.

Contagious ecthyma in the live sheep export industry.

OBJECTIVE: To investigate control options for contagious ecthyma (scabby mouth) in Australian sheep exported live to the Middle East. DESIGN: Prevalence, vaccination and modelling studies. PROCEDURE: One hundred and forty weaner sheep (less than 1 year old) on each of 106 farms in Western Australia (WA) and 18 farm groups of adult wethers received at a WA commercial feedlot were examined for lesions of scabby mouth. Sheep on a total of 26 farms in 3 States were divided into treatment and control groups for the vaccination study. A simple deterministic compartmental model was developed to establish which parameters had the greater effect on disease prevalence. RESULTS: The proportion of farms with evidence of scabby mouth in weaner sheep was 23.6% and, on those farms with the disease, the overall prevalence was 6.1%. At the feedlot, 4 out of 18 farm groups had 5 or more sheep with lesions on arrival. The overall prevalence in the 4 diseased groups was 5.2%. Sheep vaccinated on farm before trucking to the feedlot had a lower prevalence of scabby mouth at the end of simulated shipping than controls. The main determinant of scabby mouth prevalence was the proportion of sheep immune to the disease. CONCLUSION: A program of vaccination for scabby mouth will reduce the prevalence of disease during live export. However, using current technology it is not possible to deliver shipments of sheep to the Middle East that are guaranteed completely free of scabby mouth.

Microbiological research at the Centre for Tropical Veterinary Medicine (CTVM).

The nature of the research carried out by the staff of the Microbiology Section of the Centre for Tropical Veterinary Medicine (CTVM) at home and abroad is illustrated by precis of projects on rinderpest, orf in goats and sheep, bovine dermatophilosis, contagious caprine pleuropneumonia, tick-borne fever in goats and sheep, bovine petechial fever, and caprine cowdriosis.

Tissue culture-propagated orf virus vaccine protects lambs from orf virus challenge.

Twenty, eight-day-old specific pathogen-free (SPF) lambs were vaccinated by a single scarification approximately 4 cm in length on the inner right thigh with a double-pronged applicator. The titre of live virus in the vaccine was 10(7.2) TCID50/ml and the estimated dose per lamb was 0.04 ml. Three months and six months later 10 of the vaccinated lambs and five age-matched unvaccinated control specific pathogen free lambs were challenged by a single scarification with virulent virus on the inner left thigh in the same way. After the vaccination all 20 lambs developed lesions characteristic of orf virus infection that had largely resolved four weeks later, when they all had reciprocal ELISA antibody titres > or = 3200 that persisted in all but one of them until they were challenged. After the challenge, the development of lesions in the vaccinated and unvaccinated sheep was compared daily for four weeks by means of a clinical scoring system. Both groups of vaccinated lambs had significantly lower (P < 0.01) total clinical scores after challenge at three months and six months than the unvaccinated lambs.

[Behavior of Orf virus in permissive and nonpermissive systems]. / Verhalten von Orf-Virus in permissiven und nichtpermissiven Systemen.

Dogs were immunized i.m. with attenuated poxvirus vaccines (vaccinia virus, Orf-virus) and a bovine herpesvirus-1 (BHV-1) vaccine. After intradermal (i.d.) application of the vaccine viruses a specific delayed type hypersensitivity (DTH) reaction of the skin occurred only with vaccinia virus. The i.d. application of Orf-virus caused a short-term, non-specific inflammatory reaction of the skin, even in dogs not immunized with Orf-virus. Out of 30 sera from Orf-virus immunized beagles (n = 4) only eight were found reactive to Orf-virus in a competition ELISA. Three sera from dogs not Orf-virus immunized but skin-tested with the virus contained low antibody titers. Using indirect immunofluorescence (IIF) in flow cytometry, the existence of Orf-virus antigens was examined on the surface and in the cytoplasm of permissive (BFK and Vero)- and questionable permissive MDCK cells. The canine kidney MDCK cell line was found to be non-permissive for Orf-virus replication; the occurrence of an Orf-(ecthyma contagiosum) like disease in dogs is unlikely.

[Recent information about the orf virus of small ruminants]. / Neue Erkenntnisse über das Orf-Virus der kleinen Wiederkäuer.

New scientific findings in the field of immunobiology and diagnosis of parapoxvirus ovis (Orf-virus) as the causal agent of a zoonosis are presented. The adaptation of Orf-virus to cell lines and its in vitro multiplication without difficulties offer the possibility for extensive studies into the biology of parapoxviruses. The development of monoclonal antibodies (MAB) against an attenuated Orf-virus strain (D-1701) led to the elaboration of a simplified, cheap and highly sensitive "antigen detection ELISA" as a diagnostic tool. Moreover the MAB's can be used to identify parapoxvirus in cells of infected tissues (indirect immuno-peroxidase technique) which is an important precondition for more detailed surveillance of the pathogenesis of parapoxvirus infections. The immunobiological behavior of the Orf-virus in its natural hosts (small ruminants) as well as in non-susceptible species (e.g. laboratory animals) is explained by the course of cellular and humoral immune reactions following infection or vaccination. The problems of combat measures against contagious ecthyma (scabby mouth) of sheep and the human Orf-virus infections are discussed.

Vaccination of sheep with cell culture grown orf virus.

Orf virus, derived from contagious pustular dermatitis (scabby mouth) lesions in sheep, was adapted to cell culture and subsequently evaluated as a potential vaccine for sheep. The traditional vaccine virus, prepared from the infected scabs of orf virus lesions in sheep, was used to vaccinate sheep by scratching with an applicator (mounted pins) dipped in virus. Less than 10 TCID50 (50% tissue culture infectious doses) of virus was required to produce large lesions (greater than 5 mm diameter) which developed during a period of 10 to 14 d prior to onset of healing which was complete by 28 to 30 d. A serum neutralising antibody response was also detected and protection against challenge by application of virulent virus to abraded skin was demonstrated in that challenge lesions developed and healed more quickly (14 d against 30 d). However, cell culture-adapted virus required more than 10(5) TCID50 to induce even small lesions (less than 2 mm diameter). An antibody response could not be detected and no evidence of protection against challenge with virulent virus was demonstrated. In contrast, a recent field isolate has yielded a cell culture-adapted virus preparation that readily infects sheep, produces large lesions, detectable antibody and protects against challenge. This isolate is distinct from the traditional vaccine strain on the basis of restriction enzyme analysis but provides cross-protection in sheep inmmunisation and challenge studies. These results demonstrate that a cell culture produced scabby mouth vaccine is feasible.

[Ecthyma of sheep and goats]. / Ecthyma der Schafe und Ziegen.

Contagious pustular dermatitis (CPD, contagious ecthyma, Orf) is a highly contagious viral disease afflicting sheep and goats. The infectious agent is a parapoxvirus which is widespread and also contagious for humans (zoonosis). Recently the disease in sheep and goats is marked by increased incidence and severe cases which cause many losses especially among lambs in fattening farms. The immunity of once infected animals is based on cellular defense mechanisms which are very unstable. The most suitable method for the prevention of ecthyma is parenteral vaccination with attenuated live vaccines. Effective vaccination programmes should enclose the whole animal population at intervals of 6-8 months.