Implementation of an HACCP model in foot and mouth disease control programmes.

The organisation and structure of the official Veterinary Services (OVS) are designed to meet a specific aim--the health certification of animal health, welfare and food safety in the production and processing stage. Disease prevention and control calls for programmes and projects that, depending on the characteristics of each disease, may involve any branch of the OVS, from the laboratory to field activities. For the purpose of this work, the model used is that of a country that is 'free from foot and mouth disease with vaccination' in accordance with the conditions stipulated in Chapter 8.8. of the World Organisation for Animal Health Terrestrial Animal Health Code. These conditions state that, to maintain this health status, a programme of monitoring and continuous control of the relevant variables must be implemented. This is achieved by applying good practice and identifying the critical control points in all processes, using a checklist that simplifies the task. The system that is developed can also serve as a guide for internal or external programme audits.

Advances in scrapie research.

The bovine spongiform encephalopathy (BSE) crisis clearly demonstrated the need to keep animal transmissible spongiform encephalopathies (TSE) under control in order to protect animal and human health. Scrapie is the most widespread TSE of livestock in the world. For this reason, health authorities in different countries have elaborated plans that aim towards scrapie eradication. The unusual nature of the scrapie agent and the fragmented status of scientific knowledge about it, along with the limitations of currently available diagnostic tools, make it unlikely that the objective of eradication will be achieved in the near future. Scientific research is focused on acquiring the knowledge that will improve the efficiency of these efforts.

Localization of foot and mouth disease virus RNA in tissue culture infected cells via in situ polymerase chain reaction.

Foot and mouth disease virus RNA was visualized in infected primary tissue culture cells by in situ PCR incorporating digoxigenin-labeled dUTP. The viral RNA polymerase gene was used as a target for amplification. Infected cells revealed cytoplasmic staining, predominantly perinuclear. The intensity of staining was in proportion to the degree of cytopathology observed and similar to the results obtained using immunoperoxidase staining. The in situ PCR technique for FMDV detection could be applied to formalin-fixed samples and be useful for the study of persistent infections.

Reconstitution of immunosuppression mice with mononuclear cells from donors sensitized to foot-and-mouth disease virus (FMDV).

Passive transfer experiments were performed to serve as a basis for analyzing the immune response of adult mice to FMDV infection. Animals were irradiated (750 rad: 1 lethal dose 50%) and reconstituted with allogeneic mononuclear cells from blood, spleen, thymus and peritoneal cavity from donors 2 and 8 days post-inoculation (p.i.). Donors were primed with 10 000 suckling mouse 50% lethal doses of FMDV strain O1 Campos. The following parameters were studied in recipient mice challenged with 10 000 suckling mouse 50% lethal doses of the same virus: (1) viremia; (b) FMDV neutralizing antibody titres; (c) sheep red blood cell (SRBC) hemagglutinating antibody titres. Viremia was substantially prolonged in irradiated control mice, which did not produce detectable antibodies to FMDV or SRBC. In contrast, the span of viremia was markedly shorter in animals reconstituted with cells obtained 8 days p.i. and its eclipse coincided with the onset of neutralizing antibody production. An equally efficient antibody response to the inoculation of SRBC was observed in these animals. No effect was detected after the transfer of cells obtained 2 days p.i. It is concluded that the humoral immune response plays a predominant role in the recovery from FMDV experimental infection in adult mice.

Foot-and-mouth disease virus (FMDV) experimental infection: susceptibility and immune response of adult mice.

Adult mice are susceptible to foot-and-mouth disease virus (FMDV) infection only under some experimental conditions. This paper report the results of pathogenesis studies on 4 different strains of mice (CF1, C3H, NIH-nude, BALB-c/J) infected with the cloned and uncloned 0(1)C strain of FMDV. High virus titers were detected in blood and pancreas 12-24 h after infection (p.i.); these persisted for up to 48 h p.i. in CF1 and BALB-c/J mice and 72 h p.i. in the two other mouse strains. Virus titers observed in other organs were lower than those found in blood. In pancreas, and occasionally in salivary glands, oropharynx, heart and testicles, viral antigen was detected by direct immunofluorescent assay. Circulating neutralizing antibodies appeared in CF1 and C3H mice at 72 and 96 h p.i. respectively, and their titers remained unchanged during the 30-day experimental period. Antibodies against viral infection-associated antigen (VIA) were detected for a shorter period. In animals irradiated with 1 LD 50 (total body irradiation), viremia persisted up to 14 days p.i. and a low antibody response was observed which began at the end of viremia. No differences in the response of mice to cloned or uncloned FMDV were observed.

Reduction in morbidity due to diarrhea in nursing beef calves by use of an inactivated oil-adjuvanted rotavirus-Escherichia coli vaccine in the dam.

An outbreak of neonatal diarrhea occurred among beef calves (2000 animals) from one large Argentinian farm in 1985. Rotavirus was detected in 78% (106/136) and enterotoxigenic Escherichia coli in 1.5% of the samples (2/136) obtained from sick calves. In comparison rotavirus was identified in only 1.6% (1/63) of the samples from clinically healthy calves. The rotavirus strain responsible for the outbreak was characterized as serotype 6 belonging to group A. In the following three years the protective capacity of a combined rotavirus-E. coli inactivated vaccine administered to the dams during the last third of the gestation period was evaluated on this farm by comparison of morbidity due to diarrhea in calves from vaccinated vs. placebo cows within the same year. The morbidity due to diarrhea among calves from dams in the vaccinated and placebo groups was 34% and 77%, respectively in 1986; 23% and 47% in 1987, and 15% and 34%, in 1988. In 1987 morbidity of diarrhea in calves born from vaccinated heifers was 54% and 74% in calves from placebo heifers. In 1988 morbidity from diarrhea was 41% and 54%, respectively among calves in these two groups. In all experiments, calves from heifers showed significantly greater morbidity than calves from cows. Differences in diarrhea morbidity between the vaccinated and placebo groups were statistically significant (P less than 0.05). Additional studies showed that the diarrhea had a significant influence (P less than 0.05) on the average live weight of the calves at weaning (5 to 7 months) with an average weight loss of 7.8 kg per calf among the calves affected with diarrhea.

Immune response to foot-and-mouth disease virus in an experimental murine model. II. Basis of persistent antibody reaction.

A murine model was used to study the mechanisms involved in the prolonged immune response to live and inactivated foot-and-mouth disease virus (FMDV). The antibody response elicited by the infection persisted throughout the entire life of the animal, while immunization with inactivated virus induced a transient response. The administration of inactivated virus in a water-in-oil emulsion increased antibody titres to values as high as those obtained by infection. There was a high correlation between neutralizing antibody titre and transfer of immunity with primed cells, and the protection afforded against challenge with infectious virus. It appears that the mechanism involved in the induction of prolonged immune memory in infected animals is not due to viral persistence. Nude mice infected with FMDV also evidenced a prolonged immune response, showing marked differences in antibody levels but equal effectiveness against challenge when nu/nu and nu/+ animals were compared. Furthermore, athymic and euthymic littermates were efficient in conferring protection when cells were transferred to irradiated animals. It is concluded that there is an effective, T-cell-independent, prolonged immune memory against FMDV in this murine model, and that the difference in the immune responses to live and inactivated virus is due mainly to differential antigenic processing rather than to a difference in the degree of sensitization of effector cells.

Detection of antibodies against foot-and-mouth disease virus using a liquid-phase blocking sandwich ELISA (LPBE) with a bioengineered 3D protein.

A liquid-phase blocking sandwich enzyme-linked immunosorbent assay (ELISA-3D) was developed to detect specific antibodies to the 3D protein in sera from foot-and-mouth disease (FMD) virus (FMDV)-infected animals. The assay uses a nonstructural 3D recombinant protein and two polyclonal antisera, one for capture (bovine) and the other for detector (guinea pig). The specificity of the assay was demonstrated by negative results with 101 sera of cattle from the FMD-free zone in Argentina and with bovine and porcine sera raised against various RNA and DNA viruses. The ELISA-3D was able to detect antibodies in cattle after natural or experimental infection with FMDV of A, O, or C types as early as 5 days postinfection and at later stages in persistently infected animals. Comparison of the results with those obtained with the routinely used agar gel immunodiffusion test and a previously described ELISA, both employing a partially purified virus-infection-associated antigen, shows that the ELISA-3D is highly sensitive and specific and gives reproducible results. Its use as a tool for monitoring viral activity and for certification of FMDV-free animals is recommended.

Assessment using ELISA of the herd immunity levels induced in cattle by foot-and-mouth disease oil vaccines.

The development of a liquid-phase blocking sandwich ELISA (LPBE) to measure antibodies (Ab) produced in cattle with the O, A and C foot-and-mouth disease virus (FMDV) types of commercial vaccines used in Argentina is described. The test was specific: 99% of naïve cattle sera (n = 130) gave titres below log10 = 1.2, and none had a titre above log10 = 1.5. Comparative studies with serum neutralization test (SNT) using sera from cattle which received one or more vaccine doses is reported. The overall rank correlation coefficient (Spearman's rho, rs) between SNT and LPBE were highly significant (rs > 0.67, P < 0.0001) for all vaccine strains. LBPE Ab titres on sera collected 90 days post vaccination were compared with results of cattle protection tests by applying a logistic regression. The minimum Ab titres at which 85% and 75% of the cattle were protected for each FMDV type were determined in order to interpret field Ab data in terms of protection. Application of this method allows large scale serological examinations to monitor antibody levels in vaccinated animals as an indirect indicator of the FMD control program status in the field. Its use in the evaluation of commercial batches of FMD vaccine is discussed.

Avian adenoviruses--distribution of infection in commercial flocks of fowls in Argentina.

Group-specific avian adenovirus antibodies were surveyed in broiler, layer, and breeder chickens in three poultry-producing areas of Argentina: north, middle, and south. The method used was the agar double-immunodiffusion test. Of the flocks examined, 89% had antibodies to avian adenovirus, with the percentages higher in layers and breeders than in broiler chickens. A close relation was found between poorer hygienic condition and high adenovirus antibodies values.

Viral antibodies in bovine fetuses in Argentina.

In order to establish the prevalence of viral infections of the bovine fetus in Argentina, a serological survey for antibodies against viral agents currently affecting cattle in this country was conducted. Antibodies against foot-and-mouth disease virus (FMDV), bovine herpesvirus-1 (BHV-1), bovine leukaemia virus (BLV), bovine rotavirus (BRV), bovine coronavirus (BCV), bovine viral diarrhoea virus (BVDV) and parainfluenza-3 (PI-3) were investigated in a total of 315 fetal serum samples. Conventional techniques were used: indirect immunofluorescence (FMDV, BHV-1, BVDV and BCV), radial immunodiffusion (BLV), ELISA (BRV) and haemagglutination inhibition (PI-3). Antibodies against BHV-1, BVDV and PI-3 were detected in samples from fetuses in the second and third trimester of gestation, with a prevalence of 1.21 per cent (two of 165), 2.03 per cent (four of 197) and 5.08 per cent (nine of 177), respectively. Either antibodies or non-antibody factors able to bind to BRV and BCV antigens were detected with a prevalence of 2.44 per cent (five of 205) and 4.54 per cent (five of 110), respectively. In addition, 14.68 per cent of non-specific inhibitors of PI-3 mediated haemagglutination were found. No seropositives against FMDV and BLV were detected.

Nucleic acid hybridisation and polymerase chain reaction in the diagnosis of infectious animal diseases.

The authors describe and summarise the use of nucleic acid hybridisation and polymerase chain reaction (PCR) technologies in the diagnosis of animal diseases. PCR is a powerful biological tool which enables exponential enzymatic amplification in vitro of a given deoxyribonucleic acid sequence. This technique is currently used to study the molecular pathogenesis of many infectious diseases and also for diagnosis. PCR is usually more sensitive than conventional methods and does not require complex facilities, and will therefore soon become the preferred technology of laboratory diagnosticians, field veterinarians and health officers.

Bovine spongiform encephalopathy in South America: a regional preventive approach.

Bovine spongiform encephalopathy (BSE) is a neurodegenerative disease of cattle caused by prions that was first described in the United Kingdom (UK) in 1986. The BSE epizootic that commenced in the UK in the 1980s has since spread into other countries in Europe and Asia through exports of contaminated meat-and-bone meal or infected cattle. Over the past few years, other emerging or reemerging diseases have spread into previously free countries or regions through international trade. This negative effect of globalisation means that to implement successful preventive and strategic programmes to safeguard animal health, such programmes must, as a priority, take a regional approach. Global thinking, regional planning and local performance constitute the key factors for the successful control of animal diseases. In South America, initial preventive actions against BSE were adopted in 1989. Further measures adopted since then and based on new scientific and technical findings, have led to the demonstration that the region is free of BSE. These early preventive actions have reliably protected the region from importing BSE-infected material. An integral part of the project to determine the BSE status of South America was the training of personnel, the incorporation of technology and the provision of updated information through close relationships with international organisations and prominent international researcher workers. Regional activities aimed at harmonising BSE prevention programmes, producing objective and transparent data on the equivalence of regional BSE status and facilitating regional and international trade have recently been launched. Maintaining the BSE-free status of the region must be given high priority by the beef agro-industrial sectors.

[Seroepidemiologic indicators for the evaluation of campaigns for the control of foot-and-mouth disease]. / Indicadores seroepidemiológicos para la evaluación de las campañas de control de fiebre aftosa.

A sero-epidemiological survey was conducted in two districts in Argentina between 1993 and 1995, to provide additional information on the epidemiology of foot and mouth disease (FMD) in Argentina and to assess the level of immunity in cattle populations, and the circulation of FMD virus. As part of the final stage of this survey, a comparison was made of the results obtained by the enzyme-linked immunosorbent assay (ELISA) and agar gel immunodiffusion techniques. Levels of population immunity against the four types of virus included in the vaccine increased progressively during the period of the survey until, in 1995, at the end of the vaccination period, the percentage of animals possessing adequate levels of protection was approximately 77% in yearlings, and more than 94% in cattle over one year old. During the three-year study, there was a clear tendency for viral activity to diminish, until in 1995 when between 3% and 0.6% were positive to the agar gel immunodiffusion test for the antigen associated with viral infection. By contrast, the ELISA detected antibody in about five times as many animals. The authors show how the increase in the level of population immunity was accompanied by a fall in viral activity.

[Prevention of and attention to emergencies in South America]. / Prevención y atención de emergencias en sudamérica.

The authors review the policies designed to prevent and deal with animal health emergencies which have been implemented in countries of South America. They describe the evolution of the epidemiological situation of the continent, the new arrangements for international trade in animals and products of animal origin arising from the creation of the World Trade Organization (WTO), and the consequences of such developments for livestock production in South America. Veterinary systems used to prevent and deal with emergencies in the eleven OIE Member Countries on the continent are described, together with emerging problems which confront the Veterinary Services of the continent, namely: exotic diseases, abnormal occurrence of endemic diseases subject to control programmes, faults in food-safety mechanisms, diseases which have an environmental impact, and problems connected with animal welfare. The emergencies which present the greatest risk to South America are foot and mouth diseases, transmissible spongiform encephalopathies, the porcine reproductive and respiratory syndrome, food poisoning, Newcastle disease and fowl plague. Other problems are the appearance of new strains of existing agents, and the presence of resistant individuals among species of bacteria or harmful arthropods. The authors emphasise the need to co-ordinate the prevention of emergencies with development work at the international level, particularly regional and international agreements, harmonization of procedures, progress in animal health and public health, risk analysis, etc. These systems and methods of prevention have a contribution to make in enhancing the potential of animal production in South America, and the adoption of stricter health and quality standards, according to criteria established by the WTO Agreement on the Application of Sanitary and Phytosanitary Measures.

Detection of virus infection-associated antigen and 3D antibodies in cattle vaccinated against foot and mouth disease.

The analysis of sera obtained from animals vaccinated or revaccinated with inactivated vaccines against foot and mouth disease (FMD) virus showed that these vaccines induced antibodies against the virus infection-associated (VIA) antigen, detectable by agar gel immunodiffusion (AGID). The present study evaluates the antibody response to protein 3D and the VIA antigen (VIAA) of FMD virus induced by different vaccines in a group of 51 calves. This response was detected using AGID and a liquid-phase blocking sandwich enzyme-linked immunosorbent assay (ELISA) for anti-3D antibodies (ELISA-3D). No anti-VIAA or anti-3D antibodies were detected after the initial vaccination. Following revaccination, animals giving positive results were detected by both methods. This immune response disappeared 60-120 days post-revaccination (dprv) according to the AGID method, and 90-180 dprv when ELISA-3D was used. Samples of oesophageal-pharyngeal fluid obtained from animals that remained positive for anti-VIAA antibodies at 90-120 dprv gave negative results for viral isolation, indicating that the transitional antibody response induced by the vaccine was due to the presence of non-structural antigens in the vaccine and not to viral infection. These results indicate that the ELISA-3D method could be used as a complementary method for sero-epidemiological studies as an indirect indicator of viral activity, as long as the age and vaccination status of the animals being sampled are taken into consideration.

[Analysis of risk factors associated with bovine spongiform encephalopathy in Argentina]. / Análisis de los factores de riesgo asociados a la encefalopatía espongiforme bovina en Argentina.

Epidemiological studies conducted in the United Kingdom have revealed the risk factors involved in the epidemic of bovine spongiform encephalopathy (BSE). This has facilitated a detailed study of risk factors associated with the possible introduction of BSE into Argentina. An analysis has been made of the numbers and geographical distribution of cattle in the country, the structure of the Veterinary Services responsible for controlling animal diseases, the way in which cattle and sheep are slaughtered, and the use of slaughter waste in the feed industry. The results of this analysis form the basis of a discussion of whether scrapie or BSE could become endemic in Argentina through contaminated feed or another route. The authors conclude that Argentina may be regarded as free from BSE, and that the importation of infected bovines is the sole potential risk for introduction of BSE in the future.

Bovine spongiform encephalopathy surveillance in Argentina.

Bovine spongiform encephalopathy (BSE) is a new disease of cattle first described in the United Kingdom in November 1986. BSE belongs to the scrapie-related group of diseases. The epidemiological studies performed in the United Kingdom demonstrate that the BSE epidemic was caused by feeding cattle with ruminant-derived protein contaminated by a scrapie-like agent. Until June 1994, the disease had been detected in indigenous cattle in Ireland, Switzerland and France. Three cases reported in Germany, two in the Sultanate of Oman, and single cases in the Falkland Islands (Islas Malvinas), Denmark, Portugal and Canada occurred in animals imported from the United Kingdom. Several countries have implemented surveillance programmes analysing the risk factors involved in the epidemic. An analysis of risk factors conducted in Argentina shows that it is highly unlikely that BSE or scrapie exist in the country, or will arise via feed in the future. As a continuation of the analysis of risk factors, a surveillance programme was implemented in the field and in abattoirs. Specialised personnel were trained in the clinical, histopathological and biochemical detection of the disease through a network of laboratories which covered 85% of the total cattle population and 100% of the high-risk group (dairy cows over five years of age). By using a statistical procedure with reference to the bovine population in nine provinces, 1,019 brains from animals belonging to the high-risk group were selected and studied by histopathological and biochemical analyses for BSE detection. The results were negative in all cases. It can be concluded from this analysis (with a sensitivity of detection of 2.95 per 1,000, and 95% statistical confidence) that Argentina may be regarded as BSE-free, and that the importation of infected animals or by-products may represent the sole potential source of introduction of BSE infection into the country in the future.

Incidence, epidemiology and control of bovine virus diarrhoea virus in South America.

Outbreaks of diarrhoea associated with mucosal erosions of the mouth, tongue and digestive tract, clinically diagnosed as bovine virus diarrhoea-mucosal disease (BVD-MD), have been reported in Argentina and Brazil since the 1960's. However, primary isolation of the virus of BVD-MD is fairly recent, occurring in 1974 for Brazil, 1984 for Argentina, 1985 for Chile and 1981 for Colombia. In Argentina both cytopathogenic and non-cytopathogenic BVD virus strains have been identified. Elsewhere in South America this differentiation does not seem to have been carried out. Serological surveys have confirmed the existence of BVD virus infection in six countries (Argentina, Brazil, Chile, Colombia, Peru and Uruguay), with an incidence rate ranging between 37 and 77% of cattle in the areas surveyed. Diarrhoea in calves between 3 and 18 months of age, often associated with mucosal erosions, has been the most commonly observed syndrome. In some cases an upper respiratory tract involvement was described. In one epizootic, in the Sabana de Bogota plateau of Colombia, reproductive failure associated with abortions or birth of weak calves was the main clinical syndrome.

Detection of bovine herpesvirus type 1 via an immunoperoxidase method, using monoclonal antibodies.

The sensitivity and specificity of the immunoperoxidase technique, using monoclonal antibodies, for the detection of bovine herpesvirus type 1 (BHV-1) was assessed and compared with viral isolation methods. In this study, BHV-1 antigens were detected in impression smears of brain obtained from calves in which BHV-1 was isolated. False-positive results were not observed after double-blind examination. Preliminary identification of isolates as antigenic variants was possible by use of 3 monoclonal antibodies reactive with neurotropic and/or pneumotropic strains of BHV-1. Results were consistent with previous work in which characterization was performed by use of immunofluorescense and ELISA. The immunoperoxidase technique, using monoclonal antibodies, was determined to be specific and sensitive, compared with viral isolation, for the diagnosis of BHV-1 encephalitis. In addition, it has operative advantages in that the assay does not require tissue culture facilities, and results can be obtained within hours after specimens are obtained.