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U Kihm

Publications and source records attributed to U Kihm.

At least 37 records · Page 2Linked to original sources
PubMed1992

Approaches to the identification of non-essential genes of African swine fever virus.

It is poorly understood why vaccines could not be developed for the control and prevention of African swine fever (ASF) virus infection. The aim of our study was to identify genes non-essential for ASF virus replication because there were indications that certain viral gene products, which apparently are non-essential for viral replication, conferred protection from death due to ASF. A cosmid library representing the genome of ASF virus strain France 64 was established and characterized. Then, in order to inactivate viral genes by insertion, the beta-galactosidase (beta-gal) gene was introduced either randomly or at specific locations of selected cloned DNA fragments. These constructions were transfected into cells which had been previously infected with a cell-culture-adapted viral strain in order to allow the generation of recombinant progeny virus. Viable recombinant progeny was identified by at least one of the following means: (1) expression of beta-gal; (2) detection of beta-gal specific DNA by plaque hybridization, and (3) absence of a functional product of the inactivated gene. Presently, we are characterizing a recombinant virus with an insertionally inactivated thymidine kinase gene.

African Swine Fever Virus↗
PubMed1992

Round table on epidemiology and control of fox rabies.

The current epizootic of rabies in Europe has as its main host the fox. Oral vaccination of the fox population has proven to be particularly effective. It is clear that the major components for a successful vaccination programme are a potent and stable vaccine, and an effective baiting system; the latter should attract the target animal but no non-target species. Recently, vaccines of increased stability have been generated; amongst these is a vaccinia recombinant virus which expresses rabies virus glycoprotein. Consequently, both attenuated live virus vaccines and a recombinant vaccine are available for routine field vaccination of the fox population.

Animals↗
PubMed1992

An ELISA for detection of antibodies against porcine epidemic diarrhoea virus (PEDV) based on the specific solubility of the viral surface glycoprotein.

Viral proteins of porcine epidemic diarrhoea virus (PEDV) were extracted from the cytoplasm of infected Vero cells using hypotonic conditions and a non-ionic detergent. Both the pH and the NaCl concentration of the extraction buffer were varied in attempts to increase the solubility of the virion spike glycoproteins (S-protein) and of the nucleocapsid proteins (N-protein). Monoclonal antibodies, hyperimmune sera and convalescent pig sera were used to identify and monitor these proteins by immunoprecipitation and Western blots. The solubility of the S-protein was optimal at pH 4, whereas that of the N-protein was optimal at pH 9. Consequently, it was possible to enrich for either S-protein or N-protein; increases in the NaCl concentration of the buffer were of no advantage in this respect. Enriched preparations of the S-protein and N-protein were used as ELISA antigen for the S-ELISA and N-ELISA, respectively. The S-ELISA proved to be the more effective of the two immunoassays. Antibodies against S-protein remained detectable for longer periods of time than anti-N-protein antibodies in the sera of PEDV-infected pigs. Using this ELISA of increased sensitivity, it was observed that only a small number of farms in Switzerland had been infected with PEDV.

Animals↗
PubMed1992

Relationship between the anti-FMD virus antibody reaction as measured by different assays, and protection in vivo against challenge infection.

The antibody response of cattle after vaccination against foot-and-mouth disease (FMD) virus was monitored using the serum neutralization test (SNT), the sandwich ELISA, liquid-phase ELISA, sandwich competition ELISA, liquid-phase competition ELISA, and the liquid-phase sandwich competition (blocking) ELISA. The competition ELISAs (in particular the "blocking" ELISA) were the most effective at detecting reactivity in these cattle sera. However, 95% of negative sera also competed in the most sensitive ELISA (the "blocking" ELISA) to titres of 1:32 (4% of the sera competed to a titre of 1:128). Comparisons between the different ELISAs, and between these ELISAs and the SNT, demonstrated that the tests were not measuring exactly the same reaction of antibody with FMD virus. With respect to the capacity of animals to resist FMD virus challenge, neither the SNT nor the competition ELISAs were consistently able to identify such animals. The anti-FMD virus antibody titres obtained could be classified into three zones; the "white zone" wherein antibody titres were high and donor animals likely to be protected; the "black zone" wherein antibody titres were low and donor animals likely to be susceptible to infection; the "grey zone" wherein the antibody titres were intermediary and no interpretation could be made with respect to protection. Assays such as ELISA and SNT cannot and do not measure immunological protection; they are a measure of antibody responses and nothing more, and should be interpreted in terms of the "three zone" phenomenon.

Animals↗
PubMed1992

Rapid and biologically safe diagnosis of African swine fever virus infection by using polymerase chain reaction.

In order to circumvent the need for infectious virus for the diagnosis of African swine fever (ASF), we established the polymerase chain reaction (PCR) technique for the detection of ASF virus (ASFV) DNA. A 740-bp fragment that originated from the conserved region of the viral genome was partially sequenced. From this sequence, four PCR primers and one oligonucleotide probe were designed and synthesized. A specific 640-bp PCR product was amplified by using oligonucleotides 1 and 5 as primers and extracts of the following samples as templates: organs and plasma obtained from ASFV-infected pigs, ASFV-infected cell cultures, and cloned DNA fragments containing the same conserved genomic region as that in the original 740-bp clone. No specific reaction products were observed in the corresponding controls. The identities of the PCR products were confirmed either by a second amplification with nested primers or by hybridization with a specific, biotinylated oligonucleotide probe. PCR proved to be a quicker and more sensitive method than virus isolation followed by the hemadsorption test when spleen and plasma samples from experimentally ASFV-infected pigs were tested. Furthermore, cloned virus DNA could be used as a positive control in the place of a live virus control. This is advantageous whenever the use of live virus is undesirable.

African Swine Fever↗
PubMed1992

Protective immune response against foot-and-mouth disease.

The causative agents of foot-and-mouth disease (FMD) are small icosahedral viruses of the Aphthovirus group within the Picornaviridae family. There is no evidence that these viruses infect cells of the immune system or otherwise interfere detrimentally with their function; additionally, it has not been possible to relate cytotoxicity reactions against virus-infected cells to the efficacy of the immune response against FMD virus infection. In contrast, there is a close association between FMD virus antibody and the protective immune response (10, 14, 15, 20, 24, 25, 29-32). Induction of this antibody is dependent on the structure of the viral antigenic sites (7-9, 11, 18) and on the concomitant presence of Th-lymphocyte epitopes (4, 5, 7, 8), although a Th-lymphocyte-independent response has been reported (2). Recent work by Piatti et al. (26) showed that the immune response induced by FMD virus was only Th-lymphocyte dependent when low doses of antigen were used. This latter work was performed in mice, and it is not certain that a similar situation would be found in cattle. As for the major effector immune defense, this relies on the interaction between antibody-virus complexes and the phagocytic cells of the reticuloendothelial system (17, 19).

Animals↗
PubMed1991

Production of monoclonal antibodies specific for African swine fever virus following in vitro primary immunization of mouse splenocytes in the presence of stimulated T lymphocyte supernatants.

Splenocytes from non-immune mice were stimulated in vitro using a kit of cytokine preparations (obtained from murine MLR and EL-4 cell cultures), and concomitantly immunized with African swine fever (ASF) virus antigen. In addition, fusions were performed at 5 days after primary or secondary stimulation/immunization. The detection of specific antibodies in the culture supernatants was not successful. In contrast, specific antibody-producing hybridomas could be generated, and this was at least comparable to a standard in vivo immunization regime, even though the optimum fusion ratio employed with these in vitro immunized splenocytes was one which is not optimum when in vivo immunized lymphocytes are used. Consequently, it would appear that hybridoma generation is a more sensitive method than the direct measurement of antibody at detecting in vitro primary immune responses. After primary in vitro immunization, the majority of immunoglobulins produced were apparently of the IgG isotype, with only 8-17% clearly IgM. These antibodies were mainly against VP73 (the major viral envelope protein) as expressed on viral antigen extracted from infected cells, although other specificities were also found. This demonstrated, by in vitro means, that the VP73 carried dominant immunogenic epitopes on ASF virus. Such observations show that the in vitro responses were closely related to those which have been detected in vivo.

African Swine Fever Virus↗
PubMed1991

Hybridoma antibody production in vitro in type II serum-free medium using Nutridoma-SP supplements. Comparisons with in vivo methods.

Six hybridoma clones (three producing anti-bovine viral diarrhoea antibodies and three producing anti-hog cholera virus antibodies) were studied with respect to their growth in type II serum-free medium. This was found to be successful in all cases when the serum-free medium was supplemented with 1% v/v Nutridoma, but only after adaptation over a small number of subcultures. Adapted hybridoma cultures could be grown in 250 ml spinner flasks, and the antibody-containing supernatants concentrated by tangential flow filtration. These were compared both with conventional cultures of the clones grown in serum-containing media and with ascites fluids generated using the latter type of culture. Adaptation to growth in the Nutridoma-based serum-free medium did not impair the capacity of the cells to produce and secrete antibodies, although their growth rate was slightly reduced when 250 ml spinner cultures were employed. The titre obtained from the ascites fluids were, as expected, higher than those from the tissue cultures, but contamination by other proteins, a significant problem in ascites fluids, did not present itself with the serum-free cultures. Consequently, 'serum-free' hybridoma antibodies could easily be concentrated in a relatively pure form, with respect to immunoglobulins present, having no need for affinity purification procedures. In addition, the acute in vivo problems of solid tumours and haemorrhage production could be avoided through the use of in vitro methods.

Animals↗
PubMed1990

Eradication of infectious bovine rhinotracheitis in Switzerland: review and prospects.

Ten years after the first outbreak of infectious bovine rhinotracheitis (IBR) in Swiss dairy cows, the national cattle herd is almost free from infection with IBR virus (bovine herpesvirus 1, BHV 1). The national programme for the eradication of IBR was divided into four phases: (1) Prevention of transmission of the infection by restrictions on trade of bovines and assessment of the prevalence of cattle with antibodies to BHV 1. (2) Slaughtering animals with antibodies to BHV 1 in order to eradicate BHV 1 from breeding herds. (3) Detection and eradication of further BHV 1 reservoirs (e.g. fattening cattle). (4) Monitoring programme and legal actions in order to maintain the favourable situation. Approximately 50,000 animals were slaughtered in the course of the eradication of IBR. The total costs amounted to approximately SFr. 110,000,000 over 10 years. The costs for maintaining the situation are estimated at approximately SFr. 5,000,000 per annum.

Animals↗
PubMed1990

Inactivation of foot and mouth disease virus in skimmed milk with propionic acid, citric acid and hydrogen peroxide.

In order to protect farm animals from infections such as foot and mouth disease (FMD) and tuberculosis, the pasteurisation of milk and milk products designated for the feeding of animals is compulsory in Switzerland. Nowadays, milk products are often treated chemically with acids or with hydrogen peroxide in order to keep bacterial contamination low. The capacity of these chemical treatments to inactivate FMD virus in skimmed milk within 6 h at 5 degrees C was tested in this study. The results indicated that the addition of 0.1%-0.3% of consumable acids, such as citric acid or propionic acid, could not guarantee the complete inactivation of FMD virus in skimmed milk. Similar results were obtained both with FMD virus deliberately added to skimmed milk and with skimmed milk obtained from naturally infected cows. Hydrogen peroxide in concentrations of 0.1%-0.3% was also an ineffective means of controlling the risk of FMD virus transmission from contaminated milk.

Animals↗
PubMed1990

[Current information on the subject of African horse sickness (AHS)].

The objective of the present review was to summarize current knowledge of African horse sickness (AHS), based on available literature (which is nonetheless relatively scant) and recent information obtained from the O.I.E. Brief description is made of the biology of AHS virus (an arbovirus, transmitted by Culicoides imicola), isolation of the agent, diagnosis by serotyping procedures, and preventive measures (such as protection of horses from insect infestation, or vaccination programs) which may be taken. The recent outbreaks in Spain, Portugal, and Morocco, have demonstrated that much more research on this disease is required.

Africa↗
PubMed1989

Antibodies to foot-and-mouth disease virus infection associated (VIA) antigen: use of a bioengineered VIA protein as antigen in an ELISA.

An enzyme-linked immunosorbent assay (ELISA) to detect antibodies to foot-and-mouth disease (FMD) virus infection associated (VIA) antigen (viral RNA polymerase) in cattle sera, was developed using a bioengineered VIA (BioVIA) protein antigen. Compared with the classical immunodiffusion test, with viral RNA polymerase purified from infected cell cultures as antigen, this ELISA was more sensitive. However, depending on the cattle population examined, sera with antibodies to viral RNA polymerase, probably due to infection with other picornaviruses, were detected. Despite these observations, the ELISA using BioVIA provided a rapid answer as to whether or not FMD virus circulated in a given herd of cattle. The main advantage of this ELISA is its absolute safety, since in no step of the antigen production was infectious or uninfectious FMD virus involved. The test can therefore be performed under normal laboratory conditions and no isolation units are needed as they are for the immunodiffusion test.

Animals↗
PubMed1989

[Serologic and virologic studies of selected cattle with antibodies to bovine herpesviruses].

In order to investigate the specificity of low titer antibodies to BHV 1, twelve cattle were subjected to stress and dexamethasone treatment. They were monitored virologically by inoculating cell cultures with naso-pharyngeal-, ocular- and vaginal- or preputial swabs and serologically by assessing the prevalence and incidence of antibodies to bovine, caprine-, porcine-, and equine herpesviruses and to bovine leukemia virus. Antibodies were classified as specific for BHV 1 if the animals excreted IBR virus, or if the antibodies neutralized BHV 1 and reacted with BHV 1 antigens, or if they reacted additionally with CapHV antigens. Animals whose sera recognized BHV 1 and BHV 2 but not other herpesviruses, were judged to have experienced both infections. Nine of the twelve animals had specific BHV 1 antibodies. With three animals the question for specificity of their antibodies remains open. Two animals experienced several herpesvirus infections. Therefore, the induction of crossreacting antibodies, directed against epitopes common to herpesviruses, could not be ruled out. The sera of one animal reacted with BHV 1 and BHV 4 antigens in ELISA tests. They did, however, not neutralize BHV 1.

Animals↗
PubMed1989

[The effectiveness of foot-and-mouth disease vaccines in Switzerland. I. Screening tests and herd immunity].

Protection of the Swiss national cattle herd against foot-and mouth disease is attempted by annual vaccination with inactivated trivalent (O, A, C) vaccines. With the serotype A5 as an example, this paper demonstrates the procedure of potency testing. Serological data obtained with two vaccines in primovaccinated feeder bulls showed that neutralizing antibodies developed within 7 to 14 days post vaccination. There was no statistically significant difference in the anti-serotype antibody titers induced by each of the vaccines; differences were seen between anti-O serotype and anti-A serotype antibody titers, regardless of which vaccine was used. Epidemiological analyses with about 3000 cattle demonstrated that single-vaccinated, and to a lesser degree twice-vaccinated, animals often had short lived immunity; that is, the antibody titers waned within a few months. In contrast, the majority of thrice and multiple vaccinated animals maintained relatively high antibody titers throughout the twelve month period of observation.

Animals↗
PubMed1989

[The effectiveness of foot-and-mouth disease vaccines in Switzerland. II. Stability problems].

One of the vaccines that were used in 1988 to immunize the Swiss national cattle population against foot-and mouth disease (FMD) was apparently not stable. Data, provided by the manufacturer, indicated a high initial antigenic content for serotype O. Protection experiments at the end of the vaccination campaign, however, indicated a substantial loss of serotype O antigen in the vaccine. Serological data, obtained during the campaign indicated that only 12% of the primovaccinated animals and 63% of previously vaccinated animals received an amount of FMD viral antigen sufficient to induce protective immunity. The primovaccinated animals were revaccinated in fall 1988 with a new batch provided by the same manufacturer. The new vaccine induced high titers of neutralizing antibodies in primo- and an anamnestic response in revaccinated cattle.

Animals↗
PubMed1989

[The control of infectious bovine rhinotracheitis (IBR) in Switzerland from 1978 to 1988].

The eradication of Infectious bovine rhinotracheitis/Infectious pustular vulvovaginitis (IBR/IPV) in Switzerland is reviewed. In 1978 IBR was reported in dairy cattle in the Eastern part of Switzerland. No preexisting eradication program was available at that time. In 1983, following a period of hesitation, the legal basis for the eradication of IBR was issued. This aim was achieved by: i) Controls and restrictions of the traffic of susceptible animals in order to prevent further transmission of IBR. ii) Slaughter of seropositive cattle, based on the assumption that animals with antibodies to BHV 1 were virus carriers and therefore an IBR-virus-reservoir. iii) The fact that besides the cattle population no BHV 1 reservoir existed in Switzerland. iv) Never licensing IBR-vaccines because they were not able to prevent the infection and the establishment of latency. The costs of the eradication program amounted to approx. SFr. 114,000,000. A total of 51,911 animals were slaughtered in order to eradicate IBR. An amount of SFr. 5,000,000 per annum is estimated to be necessary in order to maintain the favourable situation concerning IBR. In the future, the experience concerning IBR is applied for the prevention and control of other infectious diseases in the Swiss cattle population.

Animals↗
PubMed1988

Reduction of the number of mice used for potency testing of human and animal rabies vaccines.

Eight different rabies vaccines were tested for their potency in the standard mouse potency test using 3-, 5- and 7-week-old mice. 5-week-old mice seem to be best suited for this purpose, variability from test to test could be reduced considerably. An ELISA was used in parallel for the evaluation of the rabies glycoprotein content of rabies vaccines. Results of the mouse potency test correlated well with those of the ELISA if highly purified human vaccines were tested. Unspecific reactions in the ELISA caused by adjuvanted veterinary vaccines could not be blocked. Further experiments will be needed in order to evaluate the potency of inactivated veterinary rabies vaccines by a in vitro test.

Aging↗