The role of vaccine banks in resilience, response and recovery in respect of animal diseases.

Veterinary vaccine banks, also referred to as vaccine stockpiles or vaccine strategic reserves, play an important role in the response to infectious animal diseases, by assisting control of the disease and resilience in recovering from its effects. Vaccine banks have a part to play in both emergencies and control programmes. The concept of vaccine banks was initially established as a countermeasure to an animal disease emergency or outbreak or the introduction of a new disease. They have increasingly been used to prevent important diseases identified by Veterinary Authorities as requiring a well-planned control programme through vaccination. Vaccine banks can consist of physical or virtual reserves, or the maintenance of production capacity. Physical reserves comprise vaccine antigen or readyto-use vaccines. Virtual reserves include inventory management by vaccine manufacturers. Maintenance of production capacity encompasses management of the vaccine seed material, and the maintenance of knowledge by the manufacturers through continued research into the target vaccines. The establishment, maintenance and implementation of vaccine banks depend on a number of prerequisites, which include a strategy for the physical or virtual vaccine stockpile, a legislative framework, regulatory arrangements, effective supply-chain mechanisms and adequate surveillance systems. Through international solidarity, vaccine banks are available and accessible to countries that do not have their own reserves. The World Organisation for Animal Health and European Union vaccine banks perform this task. Moreover, vaccine banks have facilitated access to important public health vaccines, such as rabies vaccines. These issues are discussed below, as well as potential opportunities for public-private partnerships in different aspects of vaccine banks.

Les banques de vaccins vétérinaires, que l'on appelle également stocks de vaccins ou réserves stratégiques de vaccins, jouent un rôle important dans la réponse apportée aux maladies animales infectieuses, en contribuant à lutter contre la maladie elle-même et en édifiant la résilience lors du processus de récupération. Les banques de vaccins ont un rôle à jouer aussi bien dans les programmes d'urgence que dans les programmes de contrôle des maladies. Le concept de banque de vaccins est apparu initialement comme une contremesure face aux urgences zoosanitaires ou à l'apparition de nouveaux foyers ou de maladies émergentes. Par la suite, le recours aux banques de vaccins est allé croissant dans le cadre de la prévention d'importantes maladies identifiées par les autorités vétérinaires comme devant faire l'objet d'un programme de contrôle bien planifié reposant sur la vaccination. Les banques de vaccins peuvent consister en des réserves physiques ou virtuelles, ou renvoyer au maintien des capacités de production. Les réserves physiques sont constituées d'antigène vaccinal ou de vaccins prêts à l'emploi. Les réserves virtuelles résultent de la gestion des inventaires par les fabricants de vaccins. Le maintien des capacités de production recouvre la gestion des souches de semence primaire destinées à la production de vaccin, ainsi que l'entretien de la base de connaissances des fabricants à travers la recherche continue sur les vaccins cibles. La mise en place, le maintien et l'exploitation de banques de vaccins dépendent d'un certain nombre de conditions préalables, parmi lesquelles l'existence d'une stratégie misant sur les réserves physiques ou virtuelles de vaccins, mais aussi d'un cadre législatif, de dispositions réglementaires, de mécanismes efficaces régissant la chaîne d'approvisionnement et de systèmes de surveillance adéquats. Grâce à la solidarité internationale, les banques de vaccins sont accessibles et à la disposition des pays qui ne comptent pas de réserves propres. Les banques de vaccins de l'Organisation mondiale de la santé animale et de l'Union européenne remplissent cette fonction. En outre, les banques de vaccins ont facilité l'accès à des vaccins importants pour la santé publique, en particulier les vaccins antirabiques. L'auteur aborde ces enjeux ainsi que les possibilités éventuelles de création de partenariats public-privé couvrant différents aspects des banques de vaccins.

Los bancos de vacunas veterinarias, también denominados «reservas de vacunas¼ o «reservas estratégicas de vacunas¼, cumplen una importante función en la respuesta a enfermedades animales infecciosas, ayudando a combatir la enfermedad y a conferir resiliencia para recobrarse de sus efectos. Los bancos de vacunas son un elemento que interviene tanto en situaciones de emergencia como en programas de lucha. Ideados en un principio como medida para contrarrestar una emergencia o un brote zoosanitarios o la penetración de una nueva enfermedad, también se han venido utilizando cada vez más para prevenir las enfermedades importantes que a juicio de las autoridades veterinarias exigen un programa bien planificado de lucha mediante vacunación. Los bancos de vacunas pueden contener reservas físicas o virtuales, o a veces tienen que ver más bien con el mantenimiento de los medios de producción. Las reservas físicas pueden albergar antígenos vacunales o vacunas listas para usar. Por «reserva virtual¼ se entiende la gestión de existencias por parte de los fabricantes de vacunas. La idea de «mantenimiento de los medios de producción¼, por su parte, corresponde a la gestión del material de siembra para fabricar vacunas y al mantenimiento del saber hacer del fabricante por medio de investigaciones continuas sobre determinadas vacunas. La creación, el mantenimiento y la explotación de bancos de vacunas dependen del cumplimiento de una serie de requisitos previos, en particular la existencia de un procedimiento aplicable a la reserva física o virtual de vacunas, un ordenamiento legislativo, disposiciones reglamentarias, mecanismos eficaces relativos a la cadena de suministro y sistemas adecuados de vigilancia. Gracias a la solidaridad internacional, los países que carecen de reservas propias pueden disponer y servirse de bancos de vacunas. Los bancos de vacunas de la Organización Mundial de Sanidad Animal y la Unión Europea cumplen esta función. Por otra parte, los bancos de vacunas han facilitado el acceso a vacunas importantes para la salud pública, como las antirrábicas. Los autores examinan todas estas cuestiones, así como posibles vías para establecer asociaciones publico-privadas en relación con diferentes aspectos de los bancos de vacunas.

Safety and immunogenicity of a live attenuated Rift Valley Fever recombinant arMP-12ΔNSm21/384 vaccine candidate for sheep, goats and calves.

Rift Valley fever (RVF) causes serious health and economic losses to the livestock industry as well as a significant cause of human disease. The prevention of RVF in Africa is a global priority, however, available vaccines have only been partially effective. Therefore, the objective of this study was to evaluate the safety and immunogenicity of a live, attenuated recombinant RVFV arMP-12ΔNSm21/384 nucleotide deletion vaccine candidate in domestic ruminants. Evaluation involved testing to determine the infectivity titer of the vaccine virus in Vero cells for industrial scale up vaccine production. Safety experiments were conducted to determine the potential of the vaccine virus to revert to virulence by serial passages in sheep, the possibility of virus spread from vaccinated sheep and calves to unvaccinated animals, and the potential health effects of administering overdoses of the vaccine to sheep, goats and calves. The immunogenicity of 3 doses of 104, 105 and 106 Tissue Culture Infectious Doses50% (TCID50) of the vaccine was assessed in 3 groups of 10 sheep and 3 groups of 10 goats, and doses of 105, 106 and 107 TCID50 was evaluated in 3 groups of 10 calves subcutaenous vaccintation. The results showed that the infectivity titer of the vaccine virus was 108.4 TCID50/ml, that the vaccine did not spread from vaccinated to un-vaccinated animals, there was no evidence of reversion to virulence in sheep and the vaccine overdoses did not cause any adverse effects. The immunogenicity among sheep, goats and calves indicated that doses of 104-106 TCID50 elicited detectable antibody by day 7 post-vaccination (PV) with antibody titers ranging from 0.6 log to 2.1 log on day 14 PV with sustained titers through day 28 PV. Overall, these findings indicated that the RVFV arMP-12ΔNSm21/384 vaccine is a promising candidate for the prevention of RVF among domestic ruminants.

Development of a low-dose fast-dissolving tablet formulation of Newcastle disease vaccine for low-cost backyard poultry immunisation.

The immunisation of backyard poultry is critical for maintaining healthy flocks to provide nutrition and income for low-resource farmers worldwide. A vaccine presentation for flocks of less than 50 birds could make it more affordable and accessible, increasing uptake and impact. Fast-dissolving tablets (FDT) of Newcastle disease virus (NDV) vaccine were produced by freeze drying the LaSota NDV strain combined with excipients into tablets containing a small number of doses and packaged in polymer blister sheets. The NDV-FDT vaccine maintained virus stability for more than six months at 4°C, based on plaque assay and egg infectivity dose data. Stability was further confirmed in a challenge study, where the tablet vaccine elicited a strong immune response and provided 100 per cent protection to vaccinated chickens infected with a virulent strain of NDV. The vaccine tablet can be diluted in water (no needle or syringe required) and administered either in drinking water or with a dropper via an intraocular and/or intransal route. Results indicate that FDTs containing a small number of doses are a feasible presentation for backyard poultry farmers. The compact packaging of the FDTs will also provide cost savings in storing and distributing the vaccine in the cold chain.

Animal trypanosomosis: making quality control of trypanocidal drugs possible.

African animal trypanosomosis is arguably the most important animal disease impairing livestock agricultural development in sub-Saharan Africa. In addition to vector control, the use oftrypanocidal drugs is important in controlling the impact of the disease on animal health and production in most sub-Saharan countries. However, there are no internationally agreed standards (pharmacopoeia-type monographs or documented product specifications) for the quality control of these compounds. This means that it is impossible to establish independent quality control and quality assurance standards for these agents. An international alliance between the Food and Agriculture Organization of the United Nations, the International Federation for Animal Health, the Global Alliance for Livestock Veterinary Medicines, the University of Strathclyde and the International Atomic Energy Agency (with critical support from the World Organisation for Animal Health) was established to develop quality control and quality assurance standards for trypanocidal drugs, with the aim of transferring these methodologies to two control laboratories in sub-Saharan Africa that will serve as reference institutions for their respective regions. The work of the international alliance will allow development of control measures against sub-standard or counterfeit trypanocidal drugs for treatment of trypanosome infection. Monographs on diminazene aceturate (synonym: diminazene diaceturate), isometamidium chloride hydrochloride, homidium chloride and bromide salts and their relevant veterinary formulations for these agents are given in the annex to this paper. However, the authors do not recommend use of homidium bromide and chloride, because of their proven mutagenic properties in some animal test models and their suspected carcinogenic properties.

Vaccination for the control of Rift Valley fever in enzootic and epizootic situations.

Vaccination continues to be the most effective way to control Rift Valley fever (RVF), a zoonotic insect-borne viral disease of livestock. The irregular, cyclical and persistent nature of RVF in its occurrence in enzootic situations suggests that the vaccination strategy to be considered for these regions should be different from what is envisaged for free from risk regions. Currently available RVF vaccines have been extensively used for the control of the disease. However, these vaccines have shortcomings that have encouraged many research groups to develop new vaccine candidates that would address a large number of the current challenges, and be suitable for use both in disease-free regions and in different contingency and emergency preparedness strategies. The characteristics of different RVF vaccines and vaccination strategies are discussed in this report.

A high oxfendazole dose to control porcine cysticercosis: pharmacokinetics and tissue residue profiles.

Oxfendazole (OFZ) is efficacious for porcine cysticercosis at 30 mg/kg. OFZ is not registered to be used at this dose. The assessment of the OFZ and metabolites [(fenbendazole sulphone (FBZSO2), fenbendazole (FBZ)] plasma pharmacokinetic and tissue residue profiles after its oral administration to pigs and the withdrawal period for human consumption were reported. Forty-eight pigs allocated into two groups received OFZ (30 mg/kg) orally as a commercial (CF) or as experimental formulation (SMF). Samples (blood, muscle, liver, kidney and fat) were collected over 30 days post-treatment and analyzed by HPLC. OFZ was the main compound recovered in plasma, followed by FBZSO2 and low FBZ concentrations. OFZ AUC0-LOQ (209.9±33.9 µg·h/ml) and Cmax (5.40±0.65 µg/ml) parameters for the CF tended to be higher than those for the SMF (AUC0-LOQ: 159.4±18.3 µg h/ml, Cmax: 3.80±0.35 µg/ml). The highest total residue (OFZ+FBZSO2+FBZ) concentrations were quantified in liver, followed by kidney, muscle and fat tissue. FBZSO2 residue levels were the highest found in muscle (0.68±0.39 µg/g) and fat (0.69±0.39 µg/g). In liver and kidney the highest residues corresponded to FBZ (5.29±4.36 µg/g) and OFZ (2.86±0.75 µg/g), respectively. A withdrawal time of 17 days post-treatment was established before tissues are delivered for human consumption.

Study on the efficacy and safety of different antigens and oil formulations of infectious coryza vaccines containing an NAD-independent strain of Avibacterium paragallinarum.

The present study was designed to assess and compare three different formulations of the new Onderstepoort infectious coryza (IC) quadrivalent vaccine, which contain an NAD-independent strain of Avibacterium paragallinarum (previously known as Haemophilus paragallinarum), and a commercial IC vaccine, not containing an NAD-independent strain, for their safety and ability to protect chickens of varying ages against virulent challenges with four different serovars of A. paragallinarum, including the NAD-independent strain of the C-3 serovar. Four groups of 140 chickens each were vaccinated at the age of 17 weeks and revaccinated at the age of 19 weeks with each of the four vaccine formulations. A similar sized group of non-vaccinated chickens was used as control. Two rounds of challenge were conducted: a group of chicken in each vaccination group was challenged between 31 and 35 weeks of age, while another group was challenged between 51 and 55 weeks of age. The "in-contact" challenge model was used in this experiment. For each vaccination group, the four challenge strains representing four local serovars were used in each challenge round. The efficacy of the vaccines was compared based on overall protection levels obtained and the duration of protection. The safety of the different vaccines was determined by the severity of post-vaccination reactions. The need for the incorporation of the NAD-independent strain in the vaccine was evidenced by the low protection level against NAD-independent challenge recorded in the group of birds vaccinated with the commercial vaccine. The results obtained confirmed not only the variation in virulence of different South African serovars, with serovar C-3 being the most virulent and serovar B having almost no virulence but also the age related increase in susceptibility. The importance of a suitable formulation of the vaccine is discussed.

Evaluation of different adjuvants for foot-and-mouth disease vaccine containing all the SAT serotypes.

Foot-and-mouth disease (FMD) is an economically important disease of cloven-hoofed animals that is primarily controlled by vaccination of susceptible animals and movement restrictions for animals and animal-derived products in South Africa. Vaccination using aluminium hydroxide gel-saponin (AS) adjuvanted vaccines containing the South African Territories (SAT) serotypes has been shown to be effective both in ensuring that disease does not spread from the endemic to the free zone and in controlling outbreaks in the free zone. Various vaccine formulations containing antigens derived from the SAT serotypes were tested in cattle that were challenged 1 year later. Both the AS and ISA 206B vaccines adjuvanted with saponin protected cattle against virulent virus challenge. The oil-based ISA 206B-adjuvanted vaccine with and without stimulators was evaluated in a field trial and both elicited antibody responses that lasted for 1 year. Furthermore, the ISA 206 adjuvanted FMD vaccine protected groups of cattle against homologous virus challenge at very low payloads, while pigs vaccinated with an emergency ISA 206B-based FMD vaccine containing the SAT 1 vaccine strains were protected against the heterologous SAT 1 outbreak strain.

Alternative vaccination against equine botulism (BoNT/C).

REASON FOR PERFORMING STUDY: In Europe the incidence of botulism in horses has increased in the last decade due to the growing popularity of haylage feeding. Recombinant vaccines are safer and less expensive to produce and are generally better tolerated than toxoids. OBJECTIVES: To investigate whether the recombinant C-terminal half of the heavy chain of the botulinum neurotoxin C (Hc BoNT/C) in combination with an immunstimulatory adjuvant is an appropriate vaccine candidate for horses by testing its efficacy to induce neutralising antibodies and by comparing its immunogenic properties and adverse reactions to a commercial toxoid vaccine. Formation of oedema and local pain reactions were assessed. ELISA and Western blot assay against Hc BoNT/C and testing of neutralising antibody induction in a mouse protection assay were used to evaluate the immune response. RESULTS: With the recombinant vaccine, only minor local swelling with full recovery after 5 days was noted after brisket injections. The toxoid vaccine produced local, painful reactions with longer recovery periods of up to 2 weeks. Horses vaccinated with either vaccine induced neutralising antibodies after the second booster vaccination, while seroconversion on ELISA and Western blot to Hc BoNT/C was apparent after the first recombinant vaccination, and at various time points in the vaccination schedule in horses that received commercial toxoid vaccine. CONCLUSION: The recombinant vaccine showed fewer adverse reactions compared to the only commercially available vaccine but induced similar concentrations of neutralising antibodies. There was no correlation between the serological response to Hc BoNT/C and the neutralising capacity of serum. POTENTIAL RELEVANCE: Recombinant Hc BoNT/C is an appropriate vaccine candidate to stimulate production of neutralising antibodies against botulinum neurotoxin C in horses and creates only minor local reactions at the injection site.

Veterinary vaccines and their use in developing countries.

The burden of infectious diseases in livestock and other animals continues to be a major constraint to sustained agricultural development, food security, and participation of developing and in-transition countries in the economic benefits of international trade in livestock commodities. Targeted measures must be instituted in those countries to reduce the occurrence of infectious diseases. Quality veterinary vaccines used strategically can and should be part of government sanctioned-programmes. Vaccination campaigns must be part of comprehensive disease control programmes, which, in the case of transboundary animal diseases, require a regional approach if they are to be successful. This paper focuses on the salient transboundary animal diseases and examines current vaccine use, promising vaccine research, innovative technologies that can be applied in countries in some important developing regions of the world, and the role of public/private partnerships.

Discrimination between sheep-associated and wildebeest-associated malignant catarrhal fever virus by means of a single-tube duplex nested PCR.

A single-tube duplex nested polymerase chain reaction (sdn-PCR) was developed for the detection of and discrimination between ovine herpesvirus-2 (OvHV-2) and alcelaphine herpesvirus-1 (AIHV-1). These viruses respectively cause sheep- and wildebeest-associated malignant catarrhal fever (SA-MCF and WA-MCF). In the first step of the sdn-PCR, two primers with high annealing temperatures based on conserved regions of the tegument genes were used for DNA amplification. In the second step, two primer sets based on variable regions of the respective OvHV-2 and AIHV-1 genes and with annealing temperatures > 11 degrees C below the primers used in the first step, were used. Internal regions of different sizes from amplicons produced in the first step were amplified. This single-tube test obviates the need for two separate assays to detect both viral types, thereby reducing time, labour and cost.

Foot and mouth disease in Mali: the current situation and proposed control strategies.

Two main reasons prompted the authors to write this paper. First, outbreaks of foot and mouth disease (FMD) have occurred repeatedly in Mali and neighbouring countries during the last decade. Secondly, there is a pressing need for control strategies, since the first molecular epidemiological studies of FMD virus in West Africa have demonstrated that FMD transmission across national boundaries is common in this region. The authors discuss the FMD outbreaks that occurred during the period of 1980 to 1996, which were reported to the Central Livestock Office in Mali by field veterinarians. The outbreaks in 1980 and 1982 were confined to the regions of Kayes and Gao, respectively. Between 1991 and 1992, outbreaks occurred in Segou, Sikasso and Bamako. In 1996, FMD outbreaks were reported in cattle populations throughout Mali, except in Kidal in the Sahara desert, where temperatures reach 45 degrees C. High mortality was reported in young animals, while morbidity approached 100% in adult cattle.

Foot-and-mouth disease control using vaccination: South African experience.

South Africa has zoned status from the Office International des Epizooties (OIE) with the largest part of the country being foot-and-mouth disease (FMD)-free without vaccination. Outbreaks in this zone are handled differently from outbreaks in the control zones, which do not affect the export status of the country. However, the different socio-economic groupings need to be considered when reaching control decisions and in this regard, the country has been challenged with unique foot-and-mouth disease (FMD) control options. Vaccination has been shown to be effective both in ensuring that disease does not spread from the endemic to the free zone, as well as controlling outbreaks in the free zone. New adjuvants that claim to illicit longer lasting immunity have been tested with antigens derived from the SAT serotypes and animals were challenged one year post vaccination to determine the level of protection. However, even with vaccines that provide immunity for more than a year, an annual vaccination campaign will most probably not be acceptable in the buffer zone where calving occurs throughout the year.

Vaccination in the control of bluetongue in endemic regions: the South African experience.

The eradication of bluetongue virus (BTV) from endemic regions of Africa is virtually impossible due to the role played by the widely distributed Culicoides spp. of midge vectors and the ubiquitous distribution of reservoir species. In endemic areas attempts can only be made to limit the occurrence of bluetongue (BT) disease and its economic impact through vaccination. Despite several potential problems (teratogenicity, risk of reassortment and reversion to virulence of the attenuated viral strains), epidemiological and recent molecular data support the fact that the live attenuated vaccine that has been used for decades in enzootic regions, provides a safe and efficacious means to control the disease in regions of southern Africa, as well as other areas of the world.

The use of vaccination in the control of bluetongue in southern Africa.

The eradication of bluetongue virus (BTV) from endemic regions of Africa is virtually impossible due to the role played by widely distributed Culicoides spp. midge vectors and the ubiquitous distribution of vertebrate reservoir species. In endemic areas, attempts can only be made to limit the occurrence of bluetongue (BT) disease and its economic impact through vaccination. Despite several potential problems (teratogenicity, risk of reassortment, and reversion to virulence of the attenuated viral strains), the current live-attenuated vaccine, produced by Onderstepoort Biological Products (OBP), South Africa, has been used for decades in enzootic regions, and has been shown to provide a safe and efficacious means to control the disease in regions of southern Africa, as well as other areas of the world.

Validation of an indirect enzyme-linked immunosorbent assay for the detection of antibody against Brucella abortus in cattle sera using an automated ELISA workstation.

An automated indirect enzyme-linked immunosorbent assay (I-ELISA) for the serological diagnosis of bovine brucellosis was developed and validated in-house. A total of 4,803 cattle sera from South Africa (n = 3,643), Canada (n = 652), Germany (n = 240), France (n = 73) and the USA (n = 195) was used. The South African panel of sera represented 834 sera known to be positive by the Rose Bengal test (RBT), serum agglutination test (SAT) and complement fixation test (CFT), 2709 sera that were negative by CFT, and 100 sera from animals vaccinated with a standard dose of Brucella abortus strain 19. Overseas sera were obtained from reference non-vaccinated brucella-free cattle (n = 834), naturally infected (n = 72), experimentally infected (n = 71), and vaccinated animals (n = 83). Also 100 sera collected from cattle in Canada and known to be positive by competitive ELISA (C-ELISA) were used. The intermediate ranges ("borderline" range for the interpretation of test results) were derived from two-graph receiver operating characteristics analysis. The lowest values of the misclassification cost-term analysis obtained from testing overseas panels, covered lower I-ELISA cut-off PP values (0.02-3.0) than those from local panels (1.5-5.0). The relatively low cut-off PP values selected for I-ELISA were due to the fact that the positive control used represents a very strong standard compared to other reference positive sera. The greater overlap found between negative and positive cattle sera from South Africa than that between reference overseas panels was probably due to the different criteria used in classifying these panels as negative (sera from true non-diseased/non-infected animals) or positive (sera from true diseased/infected animals). The diagnostic sensitivity of the I-ELISA (at the optimum cut-off value) was 100% and of the CFT 83.3%. The diagnostic specificity of I-ELISA was 99.8% and of the CFT 100%. Estimate of Youden's index was higher for the I-ELISA (0.998) than that for the CFT (0.833). Analysis of distribution of PP values in sera from vaccinated and naturally infected cattle shows that in vaccinated animals all readings were below 31 PP where in infected ones these values represented 43%. Therefore, it appears that I-ELISA could be of use in identifying some naturally infected animals (with values > 31 PP), but more sera from reference vaccinated and infected animals need to be tested to further substantiate this statistically. Of 834 sera positive by RBT, SAT and CFT, 825 (98.9%) were positive in the I-ELISA. Compared to C-ELISA the relative diagnostic sensitivity of the I-ELISA was 94% and of the CFT 88% when testing 100 Canadian cattle sera. Of 258 South African cattle sera, of which 183 (70.9 %) were positive by the I-ELISA and 148 (57.4 %) by the CFT, 197 (76.4%) were positive by C-ELISA when re-tested in Canada. One has to stress, however, that Canadian C-ELISA has not been optimised locally. Thus, the C-ELISA was probably not used at the best diagnostic threshold for testing South African cattle sera. This study shows that the I-ELISA performed on an automated ELISA workstation provides a rapid, simple, highly sensitive and specific diagnostic system for large-scale detection of antibodies against B. abortus. Based on the diagnostic accuracy of this assay reported here, the authors suggest that it could replace not only the currently used confirmatory CFT test, but also the two in-use screening tests, namely the RBT and SAT.

Phage displayed peptides and anti-idiotype antibodies recognised by a monoclonal antibody directed against a diagnostic antigen of Mycoplasma capricolum subsp. capripneumoniae.

A monoclonal antibody (Mab 4.52) raised against Mycoplasma capricolum subsp. capripneumoniae (Mccp) cell lysate was used as a template to obtain substitute antigens recognised by its paratope. Two approaches were investigated: a 17-mer random peptide library displayed on the surface of a filamentous phage was screened by panning on the immobilised Mab 4.52 and anti-idiotype antibodies were generated by immunising a chicken with the F(ab')(2) fragments of the antibody. Analysis of the peptide sequences displayed by the isolated phages identified two peptides. Both contained two cysteine residues and had identical or similar amino acids in positions 5 (P), 8 (I/L) and 13 (L). The fusion phages were also recognised by Mab 4.52 in enzyme-linked immunosorbent assay (ELISA) and binding was shown by surface plasmon resonance. One of the peptides was a markedly better inhibitor (67%) of the binding of Mab 4.52 to its original antigen than the other (20%) at 1mg/ml. After absorption, to remove isotypic and allotypic reactivities, the anti-idiotype IgY was specifically recognised by Mab 4.52 in ELISA and was able to inhibit its binding to the original antigen, whereas anti-idiotype antibodies raised against a bluetongue virus-specific antibody had no effect. In spite of unequivocal binding of the anti-idiotype antibodies and the fusion phages to the paratope of Mab 4.52, goat antisera appeared not to react with either of the surrogate antigens. In contrast, the test sera bound to the original antigen suggesting that Mab 4.52 does not recognise exactly the same antigenic site as antibodies in the goat antisera.

Evaluation of two PCR-based procedures for typing Clostridium perfringens.

Two polymerase chain reaction (PCR)-based procedures for typing Clostridium, perfringens, which affects most domestic animals, were compared and evaluated for efficiency as substitute to the guinea-pig intradermal test routinely used in our laboratory, namely a multiplex PCR and a protocol based on the individual amplification of gene sequences specific for each toxin. Reference isolates of C. perfringens types A, B, C and D as well as cultures from clinical specimens were tested. The sensitivity and specificity of the PCR was confirmed on reference isolates. There was similarity in results on 43 of the 46 samples typed by all 3 methods. Clear results were obtained by PCR on 5 clinical samples that showed either equivocal or weak skin reactions in guinea-pigs. The multiplex PCR protocol, in combination with the evaluation of bacterial growth, is a better alternative to in vivo toxin typing, since C. perfringens can only be incriminated as cause of a disease when it is present in large numbers in the intestine.