The production of peste des petits ruminants hyperimmune sera in rabbits and their application in virus diagnosis

Hyperimmune sera were produced by serial inoculation of rabbits with Vero cell-adapted, sucrose gradient-purified Nigerian peste des petits ruminants virus (PPRV) isolate. Two antisera produced, neutralized the homologous PPRV but not the heterologous rinderpest Kabette "O" virus. The antisera gave strong precipitin lines with purified PPRV antigens and were used to detect PPRV and rinderpest virus antigens from ante-mortem secretions and post-mortem tissue homogenates from PPR and rinderpest virus infected goats and cattle by the agar gel precipitation tests (AGPT). The hyperimmune sera gave good titration curves with both purified Nigerian goat and the United Arab Emirate wildlife PPRV isolates in the indirect enzyme linked immunosorbent assay (ELISA). Results of indirect ELISA showed that although there were some cross reactions with the rinderpest, canine-distemper and measles viruses, at 1:100 dilution, the antisera would give a positive signal with only the homologous PPR virus.     

Improved isolation of rinderpest virus in transformed bovine T lymphoblast cell lines.

Bovine T lymphoblast cell lines transformed by the protozoan Theileria parva were compared with bovine kidney (BK) and Vero cells for their ability to isolate various strains of rinderpest virus from tissues and infected secretions. All of the strains of rinderpest virus that were tested, including attenuated cell-culture, caprinised and lapinised vaccines, and both mild and virulent pathogenic strains, readily induced syncytial cytopathic effect (cpe) in T lymphoblasts. The cpe could often be detected within one day of inoculation of lymphoblasts, whereas it took three to 14 days to appear in Vero and BK cells. Using lymphoblasts it was possible to reisolate rinderpest virus from nine of 42 swabs collected from three cattle experimentally infected with an isolate from a recent outbreak of mild disease whereas the same swabs yielded only one reisolate on BK cells. It was also possible using the lymphoblasts to detect infectious virus in the ocular, nasal and oral secretions of goats and rabbits infected with caprinised and lapinised virus, respectively. Peste des petits ruminants virus appeared to grow as rapidly as rinderpest virus in the lymphoblasts whereas canine distemper virus readily induced cpe on first passage but less readily on subsequent passage. Measles virus induced relatively little cpe when inoculated into lymphoblasts and did not appear to passage in these cells. The lymphoblasts are easy to maintain in culture and since they rapidly recovered 11 isolates from 37 diagnostic samples could prove useful in laboratories carrying out rinderpest diagnosis.     

The detection of transmissible gastroenteritis viral antibodies by immunodiffusion.

Precipitating antibodies against transmissible gastroenteritis viral antigens were detected by the immunodiffusion test in two transmissible gastroenteritis viral hyperimmune antisera and in antiserum prepared against haemagglutinating encephalomyelitis virus but not in sera from several species of normal animals, in antisera prepared against a variety of othet viruses and bacteria or sera from swine with bacterial enteritis. When the immunodiffusion test was compared with the virus neutralization test for the detection of transmissible gastroeneritis viral antibodies in 20 swine sera certain samples which contained high titres of virus neutralizing antibodies failed to produce precipitation while other sera were positive in the immunodiffusion test although their virus neutralizing antibody titres were relatively low. Precipitating antibodies were also detected by immunodiffusion in several samples of milk whey from a sow which had been vaccinated with inactivated transmissible gastroenteritis virus.     

Effect of Immunization with Plasmid DNA Encoding for Rinderpest Virus Matrix Protein on Systemic Rinderpest Virus Infection in Rabbits

Plasmid vaccine pBK-CMVMP1LC113 expressing the matrix (M) gene of rinderpest virus was assessed for its potential to protect rabbits against a lethal viral challenge. Rabbits immunized with plasmids expressing the M gene were not protected when challenged with lapinized rinderpest virus, despite the production of anti-M antibodies, while rabbits immunized with rinderpest tissue culture vaccine were completely protected from a lethal challenge with lapinized rinderpest virus. The plasmid vaccine also had no significant effect on the lymphopenia in challenged rabbits. The results indicate that rinderpest M protein does not have a protective role in rinderpest infection.     

Antibody response in cattle and rabbits to early antigens of malignant catarrhal fever virus in cultured cells.

Cytosine arabinoside (Ara-C) inhibited the production of late antigens and of infectious virus in monolayers of bovine kidney cells infected with the high-passage, WC-11 strain of malignant catarrhal fever virus. Early antigens were not affected. Using hyperimmune and acute-phase sera from cattle and rabbits in indirect immunofluorescence tests, it was shown that Ara-C treated cultures contained two early antigens; one was diffuse and distributed throughout the cells, the other was particulate and intranuclear. Antibody to early antigens developed later and attained lower titres in infected animals, especially rabbits; only hyperimmune sera reacted with the diffuse early antigen.     

Serological survey concerning the IBR, CHV2, BVD, PI3, BRS and rinderpest viruses in small ruminants in Zaire

More than 400 small ruminant sera from Za?re were screened for antibodies to IBR, CHV2, BVD, bovine and ovine PI3, BRS and rinderpest viruses. Sera from local animals were negative for BVD, PI3 and rinderpest viruses: 8% of sera were positive for IBR virus, all with higher titers to CHV2; 31% of sera were positive to BRS virus.     

Specific fluorescein-labeled antibodies to bovine viral diarrhea virus prepared from sera of rabbits immunized with purified virus.

Specific fluorescein-labeled antibody conjugates to three strains of bovine virus diarrhea virus were prepared from hyperimmune rabbit sera. Viruses used to hyperimmunize the rabbits were purified by four different procedures. Conjugates were comparable in quality and specificity to conjugates prepared from serum of a calf hyperimmunized to bovine virus diarrhea virus in our laboratory. The latter conjugate was tested by Biologics Laboratories, National Veterinary Services, U.S.D.A., Ames, Iowa.     

Antigens and antibodies of malignant catarrhal fever herpesvirus detected by immunodiffusion and counter-immunoelectrophoresis

Using hyperimmune rabbit and cattle sera, immunodiffusion (ID) and counter-immunoelectrophoresis (CIEP) tests detected three or four and two or three malignant catarrhal fever (MCF) virus antigens, respectively, in infected cells. The ID test detected precipitating antibodies to MCF virus in $\text{3}\text{9}$ experimentally infected rabbits, $\text{0}\text{14}$ experimentally infected cattle, $\text{1}\text{13}$ naturally infected cattle, $\text{62}\text{176}$ wildebeest and $\text{3}\text{20}$ hartebeest. The CIEP test detected specific antibodies in $\text{3}\text{9}$ rabbit sera, but non-specific reactions prevented its use with bovine sera.The CIEP test was 2 to 4 times more sensitive than ID for detecting antibodies to MCF virus, but both tests were less sensitive than indirect immunofluorescence.The ID test demonstrated an antigenic relationship between wildebeest and hartebeest strains of MCF virus. Neither ID nor CIEP detected MCFV antigens in tissues infected with MCF virus.     

Serological studies with peste des petits ruminants and rinderpest viruses in Nigeria

One hundred and ninety-five goat and 67 sheep sera collected from various parts of southern Nigeria were screened for neutralising antibodies to both the peste des petits ruminants (PPR) and rinderpest viruses. Neutralising antibodies against both viruses were found in the sheep and goat sera examined. Parallel titration of samples which neutralised both viruses indicated a primary infection with the PPR virus (PPRV). However, some samples which failed to neutralise PPRV neutralised the rinderpest virus (RV) indicating RV activity in sheep and goats in Nigeria. These findings are discussed in relation to the diagnosis of PPRV infection and the recent reappearance of bovine rinderpest in Nigeria.     

Etiology of the stomatitis pneumoenteritis complex in Nigerian dwarf goats.

The causative agent of stomatitis pneumoenteritis complex was isolated in domesticated goats and Vero cell culture. It was identified immunologically and morphologically as identical with the "Peste des Petits Ruminants" virus. There were cross reactions between stomatitis pneumoenteritis complex virus isolate and rinderpest virus by immunodiffusion and complement fixation tests but no cross neutralization. Goats recovered from stomatitis pneumoenteritis complex were protected against a challenge with rinderpest virus that was lethal to control goats. Ultrastructural morphology revealed intracytoplasmic and intranuclear inclusions made up of random arrays of fibrillar strands. Pleomorphic particles budded from the plasma membrane of infected cells and enveloped virions were seen extracellularly. Specific ferritin tagging was demonstrated in the stomatitis pneumoenteritis complex virus infected cells treated with homologous and peste des petits ruminants viral antibody systems but little, if any, tagging in the heterologous rinderpest system.     

Development and distribution of rinderpest virus antigen in experimentally infected goats

Three goats, experimentally infected with rinderpest virus were examined for the development and distribution of precipitating antigens in various tissues and secretions using the agar gel immunodiffusion test. Virus antigens were detected in ocular secretions and lymph node biopsies from the second to the fourth and fifth days of pyrexia, respectively, but were not detected in nasal secretions. Precipitating antigens were demonstrated in various lymphoid organs, the lung and abomasum of a goat killed on the fourth day of pyrexia. These findings are discussed in relation to the epidemiology of rinderpest in goats in Africa.     

Enzyme-linked immunosorbent assay for detection of equine infectious anemia antibody to purified P26 viral protein

An indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of equine infectious anemia (EIA) antibody in horse sera. Purified P26 viral protein was the antigen; alkaline phosphatase linked to rabbit anti-horse immunoglobulin G was the conjugate. The ELISA detected EIA antibodies in horse sera as early as 11 to 14 days after experimental inoculations. There was full agreement between the results of ELISA and the agar-gel immunodiffusion tests on EIA proficiency test sera. The ELISA readily detected EIA antibody in horse sera that had weak positive reactions on agar-gel immunodiffusion.     

Antigenic variation between three strains of rinderpest virus detected by kinetic neutralisation and competition ELISA using early rabbit antisera

Strains of rinderpest virus (RV) with high and low virulence were compared with cell culture-attenuated RV for antigenic differences. Sera collected from rabbits 1 week after inoculation with live virus distinguished between homologous and heterologous strains by kinetic neutralisation and competition ELISA.     

Passive protection of piglets by recombinant baculovirus induced transmissible gastroenteritis virus specific antibodies.

Sera of pigs immunized with parts of the transmissible gastroenteritis virus (TGEV) spike (S) protein expressed by recombinant baculoviruses were tested, together with a TGEV hyperimmune antiserum, for their abilities to protect three-day-old piglets against TGEV infection. The piglets were infected with virulent TGEV and the sera were given orally 3 h before infection, together with the virus, and every 6 h postinfection during the 30 h of the experiment. Virus shedding was monitored by TGEV isolation from rectal swab samples. The sera containing antibodies induced by the complete S protein or the amino terminal half of the S protein showed protective properties, indicated by delayed onset of clinical signs and virus shedding, similar to the TGEV hyperimmune serum. Those immune sera containing antibodies induced by shorter recombinant proteins were not protective.     

Soluble antigens from culture-grown Besnoitia besnoiti endozoites

Soluble antigens from Besnoitia besnoiti cell culture-grown endozoites, obtained either by hypotonic lysis or by freeze-thawing and ultrasonication (FTS) of the organisms, were detected by the agar gel immunodiffusion method. Each antigenic preparation yielded 1-4 precipitin lines when reacted with the corresponding rabbit hyperimmune serum, while no reaction was observed with Besnoitia-positive sera from naturally infected cattle. Soluble exoantigens released by viable Besnoitia endozoites into the supernatant of infected cell cultures formed two precipitin lines with rabbit anti-FTS hyperimmune serum and appeared as positively charged protein in immunoelectrophoresis. The precipitin lines observed were parasite specific since no reaction occurred with either lysates of normal Vero cells or with supernatants from non-infected cell cultures.     

Detection of turkey enteric coronavirus by enzyme-linked immunosorbent assay and differentiation from other coronaviruses

A double-antibody ELISA for the detection of coronaviruses in intestinal contents from turkey poults with diarrhea was developed. Antibodies were raised in rabbits and guinea pigs against a Minnesota isolate of turkey enteric coronavirus (TCV) propagated in embryonating turkey eggs and were purified by density-gradient centrifugation. The specificity of antisera was confirmed by hemagglutination-inhibition and immunoelectron microscopy. Absorption of anti-TCV hyperimmune sera with egg extracts or egg ovalbumin and the use of different dilution and blocking buffers influenced the sensitivity and specificity of the ELISA. Reciprocal cross-reactivity was detected among turkey, chicken, bovine, and murine coronaviruses. Antisera to the transmissible gastroenteritis virus of swine, the rabbit enteric coronavirus, or the human coronavirus strain 299E failed to react with TCV. The TCV cross-reacted only moderately with the avian infectious bronchitis virus and the hemagglutinating encephalomyelitis virus of swine. Investigations with samples from 47 commercial turkey flocks in Quebec with episodes of transmissible enteritis revealed that the ELISA was more sensitive than was electron microscopy for detection of TCV.     

凝集试验检测兰氏D群链球菌血清抗体方法的建立

建立凝集试验,用于检测兰氏D群链球菌血清抗体.采用兰氏D群链球菌C55914株制成凝集试验抗原,制备的抗原与兰氏D群链球菌兔高免阳性血清、猪正常免疫阳性血清发生凝集反应,不与健康兔阴性血清及健康猪阴性血清发生凝集反应,与链球菌兰氏C群、兰氏E群、巴氏杆菌A群、B群阳性血清呈阴性反应.试验证明,凝集试验能够快速地检验出兰...     

Re-infection of wildlife populations with rinderpest virus on the periphery of the Somali ecosystem in East Africa

We report surveillance for rinderpest virus in wildlife populations in three major ecosystems of East Africa: Great Rift Valley, Somali and Tsavo from 1994 to 2003. Three hundred and eighty wild animals were sampled for detection of rinderpest virus, antigen or genome and 1133 sampled for antibody in sera from Kenya, Uganda, Ethiopia and Tanzania from 20 species. This was done modifying for wildlife the internationally recommended standards for rinderpest investigation and diagnosis in livestock. The animals were selected according to susceptibility and preference given to gregarious species, and populations were selected according to abundance, availability and association with livestock. Rinderpest virus, antigen and/or genome were detected in Kenya; within Tsavo, Nairobi and Meru National Parks. Serological results from 864 animals (of which 65% were buffalo) from the region were selected as unequivocal; showing the temporal and spatial aspects of past epidemics. Recent infection has been only in or peripheral to the Somali ecosystem (in Kenya). Our evidence supports the hypothesis that wildlife is not important in the long-term maintenance of rinderpest and that wildlife are infected sporadically most likely from a cattle source, although this needs to be proven in the Somali ecosystem. Wildlife will continue to be a key to monitoring the remaining virus circulation in Africa.     

The effect of antibody and complement on the expression of herpesvirus of bovine malignant catarrhal fever in cultured rabbit lymphocytes

Using indirect immunofluorescence with a hyperimmune calf serum, a virus-induced antigen was demonstrated on the surface of lymphocytes expressing intracellular malignant catarrhal fever virus antigens. Antibody to the antigen was also detected in terminal sera of both cattle and rabbits. Antisera did not restrict virus expression in explanted lymph nodes unless they were supplemented with two to four units of lytic complement per ml culture. While human, bovine and guinea pig complements caused immune lysis of infected lymphocytes, rabbit complement was ineffective. The relevance of the findings in the pathogenesis of the lymphoid proliferation caused by MCFV is discussed.     

A recent outbreak of rinderpest in East Africa

Rinderpest was brought under control in Kenya in 1976 but in April 1986 an outbreak of the disease occurred in cattle in Western Kenya, five kilometres from the Kenya-Uganda border. This was the first confirmed field outbreak of the disease in Kenya after a lull of over 10 years. Clinical disease was confined to unvaccinated zebu calves aged six to eight months from which rinderpest virus was isolated. High titres of antibodies to rinderpest virus were demonstrated in sera collected from sheep and goats that were grazing together with the affected cattle herds; there was, however, no evidence of clinical disease in these small ruminants and wildlife species in the affected area. The disease outbreak was rapidly stamped out by quarantine and vaccination.