A foot-and-mouth disease vaccine for swine.

An inactivated vaccine containing purified foot-and-mouth disease virus type O1, strain Brugge, emulsified with incomplete Freund's adjuvant was studied in swine. The antigen mass ranged from 0.02 to 416 mug in 0.25 ml of vaccine. At 90 days postinoculation (DPI) 33 to 100% of the swine which had been inoculated with 0.72% mug or larger amounts of antigen were protected against challenge. There was little protection at 182 DPI although the neutralizing titers obtained with 2.9, 34.6 and 416 mug doses of antigen were similar to those observed at 90 DPI. The 50% protective dose for swine was approximately 2.3 mug of antigen whether used in a freshly prepared state or after storage at 4 degrees C for 105 or 259 days. Significant protection was afforded when small volumes (0.1 and 0.5 ml) of vaccine were applied with a jet injector gun to the ear or neck of swine. Initial tissue reactions at the site of inoculation were minimal with these small doses of vaccine and generally disappeared ny 90 DPI.     

Selection of foot-and-mouth disease viruses by passage in bovine and swine kidney cell cultures

Two uncloned populations of foot-and-mouth disease virus, one pathogenic for adult mice and the other nonpathogenic, were passaged in cultures of primary bovine kidney (BK) cells and a line of pig kidney (MVPK) cells. Within 10 passages in MVPK cells, the nonpathogenic virus became pathogenic for adult mice, but similar passages of this virus in BK cells did not affect its pathogenicity. In contrast, passage of the pathogenic virus in MVPK cells resulted in a decrease in pathogenicity, but again passage in BK cells had no effect on this characteristics. Neither of the viruses changed in pathogenicity for infant mice during the four passages that were tested. The nonpathogenic virus passaged in BK cells was more infectious for BK than for MVPK cells, but after passage in MVPK cells, this virus was about equally infectious for the two types of cells. Infectivity of the pathogenic virus was relatively unchanged by passage in either type of cell. The parent nonpathogenic virus and the 10th BK cell passage of this virus were much more resistant to adsorption with homogenized mouse kidney than were the MVPK cell passages of nonpathogenic virus. The parent and passaged pathogenic viruses were readily adsorbed. The results demonstrated that passage of the two viruses in MVPK cells had a pronounced selective effect.     

Evaluation of infectivity and transmission of different Asian foot-and-mouth disease viruses in swine

Most isolates of foot-and-mouth disease virus (FMDV) display a broad host range. Since the late 1990s, the genetic lineage of PanAsia topotype FMDV serotype O has caused epidemics in the Far East, Africa, the United Kingdom, France, the Netherlands, and numerous other countries throughout Europe and Asia. In contrast, there are several FMDV isolates that exhibit a more restricted host range. A Cathay topotype isolate of FMDV serotype O from the 1997 epizootic in Taiwan (O/TAW/97) demonstrated restricted host specificity, only infecting swine. Methods used to evaluate infectivity and pathogenicity of FMDV isolates in cattle are well-documented, but there has been less progress studying transmission and pathogenicity of FMDV isolates in pigs. In previous studies designed to examine pathogenicity, various chimeric viruses derived from O/TAW/97 were intradermally inoculated in the heel bulb of pigs. Subsequent quantitative scoring of disease and evaluation of virus released into nasal secretions and blood was assessed. Here we prove the usefulness of this method in direct and contact inoculated pigs to evaluate infectivity, pathogenicity and transmission of different Asian FMDV isolates. Virus strains within the Cathay topotype were highly virulent in swine producing a synchronous disease in inoculated animals and were efficiently spread to in-contact naive pigs, while virus strains from the PanAsia topotype displayed more heterogeneous properties.     

Laboratory diagnosis of foot-and-mouth disease and swine vesicular disease in the years 1962-1988 in Greece]

During 26 years (1962-1988) 499 samples from FMD-suspicious cases were examined in Greece. These materials came from 348 (70%) cattle, 95 (19%) pigs and 56 (11%) sheep and goats. The cattle with 197 (72.4%) positive cases seems to play the most important roll in FMD. The different isolated virus types belonged in 60 cases to type A, in 187 to type O, in 14 to type C, in 6 to type SAT 1 and in 2 cases to type ASIA 1, respectively. SVD was isolated in 3 cases from the same area and at the same time. Most samples have been examined by means of CF, cell culture, unweaned mice or by a combined way of these assays. From 363 samples have examined: A) 148 by CF, B) 32 by CF and cell culture, C) 64 by CF and baby mice D) 80 by CF, cell culture baby mice E) 8 by cell culture, F) 18 in cell culture and baby mice and G) 13 by baby mice. Form these samples were found positive in the case A) 74 (50%), E) 3 (38%) and none in case G. On the other hand, the correlation of the positive samples in combined assays were in case B) 9:21, C) 9:1, D) 8:22:16 and the case F) 3:2 respectively. The D case shows that CF detected less positive cases than the cell culture did. For a reliable labor diagnosis of FMD every sample must be examined by more than one method.     

Role of apoptosis in the pathogenesis of Asian and South American foot-and-mouth disease viruses in swine

In this study, we performed experiments to evaluate the extend of the process of apoptotic cell death by foot-and-mouth disease virus (FMDV). Apoptosis can also occur in some virus-infected cells, and ability of viruses to either inhibit or promote apoptosis may influence the pathologic outcome of infection. In this study, to determine if apoptosis plays a role in the outcome of FMDV infection in swine, we evaluated apoptosis in diseased tissues collected from pigs inoculated with two different stains of FMDV (O1 Campos and O Taiwan). And host cell DNA fragmentation in diseased tissue from animals which were infected with either virus was evaluated by occurrence of a laddering pattern characteristic of apoptosis. Infection of cultured keratinocytes from swine tongue failed to demonstrate apoptosis in the first few hours of infection, suggesting that cell-to-cell correlation between viral antigen and apoptotic changes, e.g. cytokine secretions by immune system cells, could be critical to initiating apoptosis. Consistent with this finding, we were able to detect the pro-inflammatory cytokine TNF-alpha in diseased tissues. A clear difference in the pathogenicity of the two different FMDV isolates to pigs was not demonstrated in our study.     

Pathogenesis of swine vesicular disease in pigs.

Pigs exposed to swine vesicular disease virus developed vesicular lesions by postinoculation day 2. Lesions first appeared on the coronary band and then on the dewclaw, tongue, snout, lips, and bulbs of the heels. The onset of viremia coincided with febrile response and the appearance of vesicles. Virus was isolated from the nasal discharge, esophageal-pharyngeal fluid, and feces as early as postinoculation day 1. Greater amounts of virus were isolated from samples collected during the first week of infection, and lesser amounts from samples collected during the second week. The appearance and the distribution of specific fluorescence in various tissues indicated that during the development of swine vesicular disease virus infection, the epithelial tissues were initially involved, followed by a generalized infection of lymph tissues, and subsequently, a primary viremia. Seroconversion was detectable as early as postinoculation day 4. A mild nonsuppurative meningoencephalomyelitis throughout the CNS was observed in both inoculated and contact-exposed pigs. The olfactory bulbs were most severely and were frequently affected, particularly in contact pigs. The most severe brain lesions were found in pigs 3 to 4 days after the onset of viremia; contact pigs showed more severe brain lesions than inoculated pigs. Microscopic changes were also found in the coronary band, snout, tongue, and heart.     

Riems foot-and-mouth disease oil emulsion vaccines for swine. 1. Development and testing of highly effective and well-tolerated foot-and-mouth disease vaccine for swine using Dessauer oil adjuvants

Swine plays a very particular role in FMD epizootiology. It is, therefore, absolutely necessary to have highly effective vaccines available for this species at all times. They have to ensure early buildup of long-lasting strong immunity even after one single application. Since the effectiveness of conventional adsorbate vaccines had proved to be insufficient, monovalent and trivalent oil emulsion vaccines were specifically developed of swine, using a GDR-made oil adjuvant. Stable immunity is very soon induced by them to endangered pig stock even against the immunologically problematic sub-types O1 and A5 after one single subcutaneous (s.c.) application of 2 ml (monovalent) or 5 ml (trivalent). Application establishes in s.c. connective tissue an oil emulsion depot that leads to formation of a vaccination granuloma. The immunocompetent cells identified in the latter are morphologically correlated to adjuvant action.     

Experimental swine vesicular disease, pathology and immunofluorescence studies.

Two day old piglets were inoculated intravenously with 1 ml of swine vesicular disease virus UK-G 27-72 isolate. Using infectivity tests, immunofluorescent staining and gross and histopathological examination, pathogenesis of the infection was studied in tissue specimens collected daily from one through seven days postinoculation. Swine vesicular disease virus had a strong affinity for the epithelia of the tongue, snout, coronary band and lips, the myocardium and the lymphoid elements of the tonsil and the brain stem. The virus had the greatest affinity for the epithelium of the tongue. However, there was no evidence that the tongue was the initial replication site for swine vesicular disease virus. Prickle cells in the stratum spinosum appear to be the primary targets for the virus. The necrotic foci in the stratum spinosum appeared first, followed the next day by reticular degeneration and multilocular intraepidermal vesicular formation. In the digestive tract and most of the other visceral organs the short duration and sudden drop of the virus titres and the negative fluorescence and pathological findings suggest that these are not important sites for the replication of swine vesicular disease virus in this experiment. The virus was recovered from most of the central nervous tissue specimens. Although the piglets had significant central nervous system lesions, signs of impaired central nervous system function were not detected. However, subtle nervous signs could have been obscured by difficulties in locomotion resulting from severe lesions of the feet.     

口蹄疫病毒RT-PCR检测和分型方法的建立及初步应用

参考GenBank中各个血清型口蹄疫病毒3D、vp1、2A基因的标准序列，设计引物P1／P2和S1／S2。建立用于检测口蹄疫病毒及其利用引物S1／S2克隆片段同源性比较而确定血清型的RT-PCR方法。通过敏感性试验检测，2对引物均可以检测到10TCID50的病毒量；特异性试验的检测，2对引物对正常细胞、牛黏膜病病毒、猪瘟病毒、水疱性口炎病毒、牛传染性鼻气管炎病毒的检测结果均为阴性。利用该方法对病牛的流涎液体、水疱液体、舌皮组织、感染犊牛心脏等组织进行检测初步结果显示：该方法可以对O型和AsiaⅠ型口蹄疫病毒进行特异性检测，能够用于口蹄疫急性及亚临床感染的诊断及流行病学调查。     

Radial immuno-diffusion and serum-neutralisation techniques for the assay of antibodies to swine vesicular disease.

Pig sera were assayed for antibodies to swine vesicular disease virus by (a) the radial immuno-diffusion technique combined with autoradiography and (by serum neutralisation tests. The former was more sensitive and was used for initial screening of sera while the latter was used to obtain estimates of titres of positive sera. In a survey of 1759 sera collected at slaughterhouses there were 14 significant titres from a total of seven premises situated in localities where the disease had been known to occur, and it was concluded that this did not indicate wither widespread undetected disease or the occurence of inapparent infection in the pig population.     

口蹄疫病毒和水泡性口炎病毒二重RT-LAMP鉴别检测方法的建立

LAMP技术是一种快速新型的核酸检测技术,该技术利用4条引物在恒温下扩增目的 DNA,特异性好敏感性高。本试验旨在建立一种能同时鉴别诊断FMDV和VSV的二重RT-LAMP检测方法。根据口蹄疫病毒(FMDV)3D基因和水泡性口炎病毒(VSV)N基因的保守序列,设计了2套特异性引物,在每套引物的内引物中插入酶切位置EcoRⅠ,对反应条件进行了优化,建立了恒温快速的检测方法。结果显示:该方法特异性好,能检测到口蹄疫病毒的A,O,Asia1亚型和水泡性口炎的NJ和IND亚型,并与其他对照牛病原体不发生交叉反应;敏感性高,最低能够检测个100个FMDV病毒RNA和100个VSV病毒RNA;干扰性小,能同时检测两个模板的不同浓度组合。本试验建立的口蹄疫和水泡性口炎二重RT-LAMP方法具有简便、快速、特异、敏感等优点,可用于FMDV和VSV的临床检测和流行病学调查。     

Virus survival in slurry: analysis of the stability of foot-and-mouth disease, classical swine fever, bovine viral diarrhoea and swine influenza viruses

Farm slurry can be highly contaminated with viral pathogens. The survival of these pathogens within slurry is important since this material is often distributed onto farm land either directly or after heat treatment. There is clearly some risk of spreading pathogens in the early stages of an outbreak of disease before it has been recognized. The survival of foot-and-mouth disease virus, classical swine fever virus, bovine viral diarrhoea virus and swine influenza virus, which belong to three different RNA virus families plus porcine parvovirus (a DNA virus) was examined under controlled conditions. For each RNA virus, the virus survival in farm slurry under anaerobic conditions was short (generally ≤ 1 h) when heated (to 55°C) but each of these viruses could retain infectivity at cool temperatures (5°C) for many weeks. The porcine parvovirus survived considerably longer than each of the RNA viruses under all conditions tested. The implications for disease spread are discussed.