Hog Cholera: V. Demonstration of the Antigen in Swine Tissues by the Fluorescent Antibody Technique

The fluorescent antibody technique was employed to detect hog cholera virus in tissue sections of various organs from experimentally infected swine. The method proved to be highly sensitive and infection could be detected in these animals as early as three days after inoculation with the virus. Best results were obtained when tissues were collected from young animals in advanced disease rather than from sows or from pigs in the early febrile phase. Tonsil, spleen and lymph node were the tissues of choice and were most satisfactory when removed from freshly killed animals rather than from those that had died.     

A sequential study of experimental infection of pigs with porcine circovirus and porcine parvovirus: immunostaining of cryostat sections and virus isolation

The sequential tissue distribution of virus was investigated using virus isolation and immunofluorescence tests in 1-day-old piglets inoculated with porcine circovirus 2 (PCV2) and/or porcine parvovirus (PPV). Enlarged mesenteric lymph nodes were seen in the pig inoculated with PCV2 alone and killed at 26 days post-inoculation (PI). One of the pigs inoculated with PCV2 and PPV and killed at 21 days PI had an enlarged liver. The pig killed at 26 days PI in this group had enlarged liver, kidneys and heart. Histopathological changes were seen in lymphoid tissues of the pigs inoculated with PCV2 alone and killed at 14 and 26 days PI. Similar, but more severe, lesions were observed in the pigs infected with PCV2 and PPV and killed from 10 days PI onwards. Histological lesions of nephritis, pneumonia and hepatitis were also apparent in these animals. Mild nephritis was also seen in the pigs infected with PPV alone and killed at 14 and 26 days PI. Moderate amounts of PPV antigen were detected in tissues from the pigs inoculated with PPV alone and killed at 14 days PI. Low levels of PCV antigen were detected, mainly in lymphoid tissues, in the pigs inoculated with PCV alone and killed at 14 days PI. Low to moderate amounts of PCV antigen were detected in a wider range of tissues in the pig in this group killed at 26 days PI. In the pigs inoculated with both viruses, PPV antigen was detected in tissues of pigs killed from 3 to 26 days PI with maximal amounts detected between 6 and 14 days PI. PCV2 antigen was detected in low to moderate amounts in the tissues of pigs killed at 14 days PI. Large amounts of PCV2 antigen were detected in most of the tissues from pigs in this group killed between 17 and 26 days PI. Virus isolation results for PCV2 generally correlated well with the results for immunofluorescent staining. PPV was isolated from almost all tissues from pigs inoculated with PCV2 and PPV, a much higher incidence of positive tissues than observed for immunofluorescent staining.     

牛传染性鼻气管炎病毒攻毒方式的对比研究

本试验旨在建立牛传染性鼻气管炎病毒的攻毒模型,明确病毒在牛体内的分布及确定最佳攻毒方式,建立牛传染性鼻气管炎的发病标准,用来评价IBRV LNM弱毒疫苗的保护效力。试验共使用健康断乳牛9头,设鼻内喷雾组、滴鼻组和对照共3组,每组3头牛。将实验室分离保存的IBRV LN01/08强毒株采用鼻内喷雾和滴鼻两种方式接种试验组动物后,连续14 d,每日观察临床症状,监测体温及采集鼻拭子,对收集的试验数据进行对比分析。选取临床发病不同时期剖杀动物,采取主要脏器进行病毒分离。结果显示,所有动物攻毒后有不同程度的临床表现,以鼻内喷雾组临床表现最为严重,剖检可见肺部病变明显。病毒主要分布在呼吸道和眼结膜组织中。研究结果显示,采用IBRV自然感染方式攻击动物,喷雾方法攻毒临床效果明显强于滴鼻方式,保证了临床发病模型的建立,可以用来IBRV疫苗免疫效果评价,为研制IBR疫苗提供前提基础。     

Study on the pathogenesis of bluetongue: replication of the virus in the organs of infected sheep.

The pathogenesis of bluetongue infection was studied by the titration of the virus in tissue samples taken from sheep inoculated subcutaneously in the auricula of the ear with 76 TC ID50 of the plaque-purified type 10 bluetongue virus. Tissue samples were taken from individual animals killed at daily intervals over a period of 11 days. The mean incubation time was 6.9 days and the first clinical sign was pyrexia. On the 4th day, bluetongue virus was demonstrated in the lymph nodes of the cephalic area, tonsils and spleen; viraemia became demonstrable on the 6th day post-inoculation and typical macroscopic lesions due to the virus were first observed on the 8th day. It was concluded that, post-infection, the virus entered the regional lymph nodes. From there it was disseminated via the lymph and/or the blood stream to the lymphoid tissues in other parts of the body where further replication occurred. From these primary sites the virus was carried via the blood stream and infected the majority of tissues. Humoral antibody, as detected by immunofluorescence, did not appear to have a direct influence on the concentration of virus in solid tissues. Persistence of the virus in infected sheep was not demonstrated when tissues were taken 6, 8 and 16 weeks after infection.     

Infection of the Bovine Udder with Bovine Herpesvirus

Infectious bovine rhinotracheitis — infectious pustular vulvovaginitis (bovine herpesvirus) grown in tissue culture was used as inoculum in trials to infect the lactating bovine udder. Six experiments were undertaken in which one or more quarters were infused with 1 ml. of tissue culture fluids containing 10⁶ to 10⁷ tissue culture infectious doses (TCID) of virus. In four of the experiments the inoculated quarters showed marked evidence of infection in the form of acute inflammation, swelling, reduced milk secretion and profound changes in the physical appearance of the milk. In each case virus was recovered in high titres in the milk from about the second until the tenth to fifteenth days following exposure. Uninfected quarters remained normal in appearance and virus could not be recovered from the milk.

In three of the experiments it was shown that serum and milk antibodies appeared shortly after the disappearance of virus from the milk. One experiment involving two animals showed that about 1000 TCID of virus were required to produce infection. In one experiment a cow having a pre-inoculation serum titre for bovine herpesvirus proved resistant to infection.

The experiments indicate that the bovine udder is readily susceptible to bovine herpesvirus in non-immune animals, and that the virus produces an acute, limited infection leading to a temporary disfunction of the gland. It appears that natural invasion of the udder through the teat canal is not readily accomplished by the virus.

     

Study of the Practicability of Various Diagnostic Methods in the Demonstration of Swine Fever Virus of High and Low Virulence in Organs of Experimentally Infected Pigs

Inoculation experiments were performed with two German strains of swine fever virus. In Experiment I, ten pigs of Danish Landrace were inoculated with a strongly virulent strain, while in Experiment II ten pigs were inoculated with weakly virulent virus.The animals were killed at varying times after inoculation and organs taken out for examination by means of fluorescent antibodies (FA), by the complement fixation test (CFT), and by the agar gel diffusion test (AGT). Cryostat sections of tonsils, spleen and lymph nodes were examined by FA staining. Tissue suspensions from the same organs were inoculated into primary pig kidney tissue cultures, which were also stained with FA. Antigen produced from spleen tissue was used in the CFT and pieces of pancreas tissue in the AGT.The strongly virulent virus could be demonstrated easily by FA in all the inoculated pigs, both by direct staining of cryostat sections and by staining of inoculated tissue cultures. The CFT and AGT were positive when the tested organs originated from animals killed at a more advanced stage of the disease.While the weakly virulent virus could be demonstrated by FA staining of tissues from eight of the ten pigs in Experiment II, virus was found in none of these animals by the FA tissue culture method. The CFT was positive in one case and the AGT in three cases.In both experiments it was found that FA staining of cryostat sections of tonsils was a particularly suitable method for the demonstration of virus.The results are discussed and compared with recent German and American studies.     

Malignant catarrhal fever. I. Response of American cattle to malignant catarrhal virus isolated in Kenya.

Fifty-three American cattle were inoculated with malignant catarrhal fever virus isolated from a wildebeest in Kenya. Three animals showed the mild form of the disease and recovered, and 47 showed the severe form of the disease. The other three did not react. Of the 47 cattle, 28 died, 16 were killed for the collection of specimens and three recovered. The incubation period for the 47 cattle ranged from 16 to 29 days and the course of the fatal disease for 28 cattle averaged three to 23 days. Virus titration of specimens from nine infected steers yielded a mean titer of 10(4)/TCID50 per gm for lymph nodes, 10(3) TCID50 per mL for buffy coats and 10(2.3) TCID50 per gm for spleens. Smaller amounts of virus were found in the liver, kidneys, adrenals and thyroids. Malignant catarrhal fever virus was also found in nasal secretions and saliva of viremic cattle. Viral infectivity was shown in bovine buffy coat cells stored at 4 degrees C for two days but was immediately destroyed upon freezing even when glycerine or dimethylsulfoxide was added. Viral particles were not found in infected animal tissues by electron microscopy. The disease was successfully transmitted in steers by intratracheal intubation and by aerosol inhalation but not by contact.     

Pathology of Sarcocystis campestris infection in Richardson''s ground squirrels (Spermophilus richardsoni).

Richardson's ground squirrels were infected with 1500 or 9000 sporocysts of Sarcocystis campestris from badgers. No lesions were found in animals killed one to three days postinfection (pi). Hepatitis and phlebitis of hepatic veins were present in animals killed between four and eight days pi. No meronts were detected in these animals, but the lesions suggested that a generation of merogony occurred in the hepatic veins. Meronts were found in endothelial cells in many tissues beginning on day 9 pi. They were most numerous on day 10 pi, and less so on day 11, and subsequently. Meronts were most numerous in the lung; none was found in liver or spleen. Four of ten squirrels infected with 1500 sporocysts in one trial died between days 11 and 13 pi. There were petechial hemorrhages in skeletal muscle, lung, serosal membranes, and brain in these animals, with microscopic evidence of pulmonary, myocardial, and brain injury. One animal infected with 9000 sporocysts had petechiae in the liver at six days pi. Foci of inflammation were visible in the myocardium and brain of animals killed to 64 days pi. This species may serve as an experimental model for sarcocystosis in domestic animals.     

Classical swine fever: virulence and tissue distribution of a 1986 English isolate in pigs

Following the recurrence of classical swine fever in the United Kingdom in 1986, a virus isolated from a single outbreak was studied. A major factor in the spread of this disease is considered to be the presence of infectious virus in tissues taken from animals at certain stages of infection, although their condition may escape detection by routine inspection either before or after slaughter. Intranasal inoculation of the isolate into eight-week-old pigs reproduced the acute form of the disease. The pigs were killed or died between seven and 25 days after inoculation. The virus concentration was determined in a wide range of tissues taken at different stages of infection. Infectious virus was present at high concentrations in all the tissues taken and at all stages of infection. Any porcine tissue is therefore a potential source of infection even when it is taken either before the animal displays detectable signs of disease of after it develops serum neutralising antibodies.     

Studies on the efficacy of intranasal vaccination for the prevention of experimentally induced parainfluenza type 3 virus pneumonia in calves.

The efficacy of intranasal vaccination in preventing or limiting disease of the lower respiratory tract induced by parainfluenza 3 (PI3) virus was evaluated under experimental conditions, using a commercially available live vaccine containing a temperature-sensitive strain of PI3 virus. In a preliminary study four colostrum-deprived calves were vaccinated intranasally at one week and again at two months of age, and two similar calves were given an intranasal placebo. After the second vaccination serum antibodies to PI3 virus were detected in all four vaccinated calves, but not in the control animals. Seventeen days after the second vaccination all six calves were challenged with virulent PI3 virus, and they were killed six days later. The clinical scores and the extent of pulmonary consolidation were reduced in the vaccinated animals; PI3 virus was detected in the upper and lower respiratory tract of the control calves but in none of the vaccinated calves. In a larger scale study with 14 colostrum-fed calves, seven were vaccinated at one week and again at five weeks of age, and seven were given an intranasal placebo. Two weeks after the second vaccination all 14 calves were challenged with virulent PI3 virus. The clinical scores and lung consolidation were significantly reduced in the vaccinated calves in comparison with the controls. Six days after infection, 10 of the 14 calves were killed; PI3 virus was detectable in the nasal secretions of all seven control calves but in only one of the vaccinated animals, and PI3 viral antigen was detected in the lungs of the control calves but not in those of the vaccinated animals. One of the vaccinated calves had developed a severe clinical response after the challenge, but it had only minor lung consolidation when killed.     

Sporadic Bovine Leukosis: A Description of Eight Calves Received at Animal Diseases Research Institute from 1974-1980

Eight calves with sporadic bovine leukosis are described. The common features were generalized lymphadenopathy, visceral involvement and raised total leukocyte and lymphocyte counts. Agar gel immunodiffusion tests for bovine leukemia virus antibodies were negative in eight animals and in all animals from three herds of origin. Lymphocytic nuclear pockets were found in the tissues of one calf but attempts to isolate bovine leukemia virus from two animals were unsuccessful.     

Bluetongue virus serotypes 1 and 3 infection in Poll Dorset sheep

Objective To study the clinical signs following bluetongue virus serotypes 1 and 3 infection in Poll Dorset sheep.
Design A clinical and pathological study.
Procedure Twenty Poll Dorset sheep were inoculated with bluetongue virus serotypes 1 or 3, each inoculum having a different passage history. The sheep were examined daily and their clinical appearance and rectal temperatures recorded. Heparinised and non-heparinised blood samples were taken at intervals for virological and serological study. Gross pathological findings were recorded for several sheep at necropsy and tissue samples were collected from three sheep for virological studies.
Results All inoculated sheep developed clinical disease. The clinical signs and gross pathological changes varied considerably but were consistent with damage to the vascular endothelial system. There was a decline in the titres of infectious bluetongue virus and of antigen in tissues collected between 7 and 12 days after infection.
Conclusions The severity of disease was related to the speed of onset and duration of pyrexia and not the development or titre of viraemia. Generally, those animals with sensitive mouths, depression, coronitis, recumbency and reluctance to move were the most debilitated. Whole blood was the most reliable source of infectious virus from acutely and chronically infected and convalescent animals. However, tissue samples particularly spleen, collected from dead or killed animals suffering from either peracute or acute forms of disease were most appropriate for the rapid confirmation of a clinical diagnosis.     

Experimental sialodacryoadenitis virus infection in athymic CD-1 mice.

The purpose of the study was to investigate the susceptibility of nude mice to sialodacryoadenitis virus. Young adult male CD-1 nude mice were inoculated intranasally with virus, killed at 2, 4, 6, 8, 10 and 20 days postinoculation and examined for virus-induced lesions in tissues including respiratory tract. Inoculated and control mice were examined by virus isolation and serology. In a companion study, male Wistar rats were inoculated intranasally with the same inoculum, and examined by histopathology, immunofluorescence microscopy and serology. In virus-inoculated mice, lesions were minimal in the lower respiratory tract, and were absent in other tissues. Virus was isolated from the lower respiratory tract in animals sampled at six or eight days postinoculation. Antiviral antibody was not detected in sera from inoculated and control mice. Virus-associated lesions and antibodies were readily detected in rats following inoculation. Based on this study, there is no evidence that inadvertent exposure to sialodacryoadenitis virus should pose a threat to CD-1 nude mice, and their susceptibility to the disease appears to be similar to that reported in euthymic CD-1 mice.     

Demonstration of infectious bovine rhinotracheitis virus antigen in paraffin sections

Nine pregnant heifers were inoculated intravenously with infectious bovine rhinotracheitis virus (IBRV) in the sixth month of pregnancy. Tissues were collected from the fetus of a heifer killed 13 days postinoculation (PI), from fetuses of 6 heifers that aborted 16-27 days PI, and from mummified fetuses of 2 heifers that aborted 53 and 85 days PI, respectively. Control tissues were obtained from the fetus of a non-inoculated heifer that was killed in the seventh month of gestation. Tissues were fixed in 10% formalin, embedded in paraffin, and examined for viral antigen by immunohistochemistry, using biotinylated second antibody and alkaline phosphatase-labeled avidin-biotin complex. Antigen was detected in at least 1 tissue from the fetus of each inoculated heifer. Positive tissues included lung, liver, spleen, kidney, adrenal, and placenta. In several fetuses, antigen was identified in tissues from which virus was not isolated in cell culture. This appeared to occur when tissues had only a few small foci of infection or when tissues were severely autolyzed. The observation of viral antigen in tissues from mummified fetuses indicates that this technique may be useful in diagnostic laboratories to detect IBRV infection in tissues that are not suitable for virus isolation or for examination by the cryostat tissue section-fluorescent antibody technique.     

Pathogenesis and Subsequent Cross‐Protection of Influenza Virus Infection in Pigs Sustained by an H1N2 Strain

The H1N1, H3N2 and, more recently, H1N2 subtypes of influenza A virus are presently co‐circulating in swine herds in several countries. The objectives of this study were to investigate the pathogenesis of Sw/Italy/1521/98 (H1N2) influenza virus, isolated from respiratory tissues of pigs from herds in Northern Italy, and to evaluate its potential cross‐protection against the Sw/Fin/2899/82 (H1N1) strain. In the pathogenesis test, eight pigs were intranasally infected with H1N2 virus; at pre‐determined intervals, these animals were killed and necropsied, along with eight uninfected animals. In the cross‐protection test, sixteen pigs were infected by intranasal (i.n.) and intratracheal (i.t.) routes with either H1N2 or H1N1 virus. Twenty days later, all pigs were challenged (by the same route), with either the homologous H1N2 or heterologous H1N1 virus strains. Control group was inoculated with culture medium alone. On post‐challenge days (PCD) 1 and 3, two pigs from each infected group, along with one control pig, were killed. Clinical, virological, serological and histopathological investigations were performed in both the pathogenicity and cross‐protection tests. In the pathogenicity test, mild clinical signs were observed in two pigs during 3 and 4 days, respectively. Virus was isolated from two pigs over 6 days and from lung samples of pigs killed on post‐infection days 2 and 4. Seroconversion was detected in the two infected animals killed 15 days after infection. In the cross‐protection study, mild clinical respiratory signs were detected in all pigs infected with either the H1N2 or H1N1 virus. The virus was isolated from nasal swabs of almost all pigs till 6 days. After the challenge infection, the pigs remained clinically healthy and virus isolation from the nasal secretions or lung samples was sporadic. Antibody titres in H1N1 or H1N2 infected groups were similar, whereas the H1N2 sub‐type induced less protection against re‐infection by homologous and heterologous virus than H1N1 sub‐type. The controls had no signs of the disease. In the H1N2 infected pigs, a reduced number of goblet cells in nasal and tracheal mucosa and small foci of lymphomononuclear cell infiltrates in the submucosa were detected. Furthermore, the goblet cell reduction was related to the time of infection. Diffuse mild interstitial pneumonia was also recorded in pigs infected with the H1N2 virus and challenged with either H1N1or H1N2 pigs. These studies showed the moderate virulence of the H1N2 virus and a partial cross‐protection against heterologous infection.     

Immunity to canine adenovirus respiratory disease: a comparison of attenuated CAV-1 and CAV-2 vaccines

Four litters of puppies were divided into three groups. One group was vaccinated with a live CAV-1 vaccine and another with a live CAV-2 vaccine. Throat swabs were collected from two dogs in each of these groups to monitor the possible excretion of vaccine virus, but none was found. Both groups, together with the third group of unvaccinated controls, were challenged 17 days later with an aerosol of virulent CAV-2. One dog from each group was killed on the third, fourth, seventh, ninth, 11th and 14th days after challenge. The unvaccinated dogs developed a clinical disease characterised by anorexia, dullness, coughing and tachypnoea. The lungs were consolidated and histological examination revealed the main lesion to be a severe necrotising bronchiolitis. Large amounts of virus were present in the respiratory tissues of these dogs and high titres of virus were isolated from throat swabs. In contrast, both groups of vaccinated dogs remained clinically almost normal with minimal lesions, present for a much shorter period of time. Virus was found on day 4 in the respiratory tissues of one dog vaccinated with CAV-1 but the other vaccinated animals contained little or no virus. In general, the degree of protection afforded by CAV-1 vaccine seemed similar to that provided by CAV-2 vaccine.     

Investigation into avian pneumovirus persistence in poults and chicks using cyclosporin A immunosuppression

One-day-old poults or two-week old chicks were infected oculonasally with avian pneumovirus. Cloacal swabs were collected for virus isolation as were selected tissues (Harderian gland, turbinates, trachea, lungs and kidneys) from birds killed at regular intervals up to 33 days post infection (p.i.) for poults, and up to 40 days p. i. for chicks. In an attempt to induce virus re-excretion, the T-cell-suppressor cyclosporin A (CSA) was given for 12 days starting from three weeks p.i. in poults and from four weeks p.i. in chicks. Birds were sampled for virus isolations up to day 12 post CSA treatment. Virus was recovered only up to day nine p.i. in poults, and day five p.i. in chicks during the acute phase of the infection. Despite T-cell suppression, there was no evidence of re-excretion of the virus, and hence no evidence for the persistence of virus in the tissues examined.     

Experimental Aujeszky's disease in pigs: excretion, survival and transmission of the virus

Airborne Aujeszky's disease virus was recovered from looseboxes containing groups of pigs infected with virus strains from England, Northern Ireland and Denmark from days 1 to 7 after infection. Pigs sampled individually excreted most airborne virus on days 2 and 3 after infection. On a 24 hour basis the maximum amount of airborne virus excreted per pig was log10 5.3 TCID50. Subclinical infection was transmitted from a clinically affected group of pigs to a seronegative group held in separate looseboxes when air was drawn through ducting connecting one box with the other. Tissues taken from pigs killed at varying times after infection showed that the main sites of virus replication were in the head and neck region. Aujeszky's disease virus was detected for up to 40 days in a range of tissues taken from pigs at the acute stage of disease and stored at -20 degrees C.