Tissue tropism of African horsesickness virus in the chicken embryo demonstrated with the avidin-biotin complex immunoperoxidase method.

In horses, African horsesickness virus (AHSV) exhibits marked tropism for certain microvascular endothelia and components of the mononuclear phagocyte system. In this study, the tropism of a field isolate of AHSV serotype 5 was studied in 24 chicken embryos. Histopathology on embryonic tissues harvested with 12 hour intervals revealed progressive changes associated with endothelial damage. Immunolabeling demonstrated viral antigens in the microvascular endothelium of the spleen, lungs, and the mesenchymal connective tissue at the base of the neck, from 24 hours post inoculation. Subsequently, specific immunolabeling increased steadily in endothelia of these and other tissues such as skeletal and cardiac muscle, gastrointestinal smooth muscle, mesonephric glomeruli, liver, subcutis and feathers. Positive immunolabeling was also occasionally observed in circulating mononuclear cells and in Kupffer cells in the liver. It was concluded, that this isolate of AHSV displayed similar tissue tropism in the chicken embryo as in the horse.

Aspects of the working life of women in the nursing profession in South Africa: survey results.

This article reports on a survey done among nurses registered with the South African Nursing Council. The survey was carried out in the last quarter of 2003. The purpose of the survey was to investigate aspects of the working life of women in the nursing profession in South Africa and to make recommendations on how their working environment could be improved. The important findings were that pay-related issues dominate as the main problem at work. Improving pay scales and being paid according to extra experience, responsibilities and qualifications could improve the nurses' working environment. Furthermore, training opportunities, medical insurance and equal opportunities should be addressed as a matter of urgency. In general, respondents had a positive attitude towards their job, which leaves the impression that nurses still regard their jobs as something they do for the sake of a service to the community and not only for the money they earn.

Teratogenicity of a mutagenised Rift Valley fever virus (MVP 12) in sheep.

A 5-fluorouracil mutagenised Rift Valley fever virus strain, which was shown to be attenuated and immunogenic in cattle and sheep, was evaluated for its ability to cause teratogenic effects in pregnant sheep. A group of 50 sheep at various stages of pregnancy was inoculated with the virus and the pregnancies followed to term. There were two abortions and 14% of the lambs produced by vaccinated ewes showed teratogenic effects, the most prevalent being spinal hypoplasia, hydranencephaly, brachygnathia inferior and arthrygryposis. The foetal malformations of the central nervous and musculo-skeletal systems were mostly consistent with those observed in sheep vaccinated with the attenuated Smithburn RVF strain. The teratogenic effects of MVP12 were not seen in previous experiments by other authors as immunisation of sheep took place in the second to third trimester of pregnancy, when the foetal brain tissue has completed most of its cell division.

Nursing professionals' views on the workplace. Survey results.

This article reports on a survey done among registered, enrolled and auxiliary nurses registered with the South African Nursing Council. The survey was carried out in the period from the end of December 1997 to the beginning of 1998. The purpose of the survey was to obtain the views of female nurses on various aspects of the workplace. The important findings were the fact that nurses liked working as part of a team and that this contributed the most to their job satisfaction. The item that contributed least to job satisfaction was pay. The most important problems were that they felt that they were not paid enough and that they need better benefits. The majority of nurses were however positive about their jobs and the items the highest on the list of career expectations were job satisfaction, followed by a need for recognition.

Long-lasting protection of sheep against bluetongue challenge after vaccination with virus-like particles: evidence for homologous and partial heterologous protection.

Insect cells co-infected with appropriate recombinant baculoviruses synthesize double-shelled, virus-like particles (VLPs) with bluetongue virus (BTV) VP2 proteins representing serotype 1 (BTV-1), 2 (BTV-2), 10 (BTV-10), 13 (BTV-13) or 17 (BTV-17) as previously reported for BTV-10 (French, T.J., Marshall, J.J.A. and Roy, P. J. Virol. 1990, 64, 5696-5700). The derived particles were purified and used to vaccine sheep, either as single VLP types (BTV-10, BTV-17) or as a combination of all five serotypes. Control sheep received saline. The virus-neutralizing antibody responses were measured. Depending on the experiment, the sheep were challenged with homologous (BTV-10, -13, -17) or selected heterologous (BTV-4, -11, -16) viruses either after 4 months or 14 months, and the disease, viraemias and clinical reactions monitored. The results indicated that two doses of 10 micrograms of VLPs elicited a long-lasting immune response which protected the sheep against challenge with the homologous virulent virus. In certain cases, partial protection was afforded against challenge by heterologous BTV serotypes.

Antigenic and molecular characterization of host cell-mediated variants of equine H3N8 influenza viruses.

Antigenic differences between three of six equine influenza virus (H3N8) MDCK cell- and egg-derived pairs have been demonstrated using monoclonal and polyclonal antibodies. Sequencing of the haemagglutinin (HA) genes revealed amino acid changes in four of the six virus pairs. These data contrast with those for human isolates of influenza virus in that it was predominantly tissue culture-isolated equine virus and not egg-derived virus which displayed heterogeneity. Some of the molecular changes involved are located within the vicinity of the cell receptor-binding site (positions 156, 158 and 222) whereas others are in the vicinity of the HA1-HA2 cleavage site (positions 18 and 32 of HA1 and position 12 of HA2). Our results indicate that the host cell can play a part in selecting antigenic variants of equine influenza virus and suggest that the egg, and not cell culture as is the case for human isolates, is the preferred host for vaccine and antigenic studies.

Isolation of bluetongue virus serotype 21 from Culicoides spp. in Indonesia.

The isolation of a bluetongue (BLU) virus from Culicoides spp. in Indonesia is reported. BLU serotype 21 was isolated from a mixed pool of C. fulvus and C. orientalis of the Avaritia subgenus.

Encephalitis and chorioretinitis associated with neurotropic African horsesickness virus infection in laboratory workers. Part I. Clinical and neurological observations.

Four laboratory workers from the same vaccine-packing facility developed at different times over an 8-year period an illness characterised by encephalitis (in 3 workers) and uveochorioretinitis (in 4). Low complement fixation titres were detected in all 4 patients to African horsesickness (AHS) virus and enzyme immunoassay and plaque reduction neutralising tests were positive, the latter against both serotypes 1 and 6. Five of 15 laboratory workers from the same facility who were healthy on clinical and ophthalmological examination showed positive plaque reduction neutralising tests but none to both serotypes 1 and 6. It is postulated that the encephalitis with the predominant temporal lobe involvement was caused by an airborne transnasal route of infection of the neurotropic AHS virus released in dried powder form, secondary to the accidental breaking of vaccine bottles. This is possibly the first report of subclinical and probable clinical neurotropic AHS infection in man.

Encephalitis and chorioretinitis associated with neurotropic African horsesickness virus infection in laboratory workers. Part II. Ophthalmological findings.

Four laboratory workers developed uveitis-chorioretinitis, associated with encephalitis in 3 cases. The retinitis was characterised by haemorrhages and areas of retinal oedema, most marked over the posterior polar regions, and was associated with exudative retinal detachments. The lesions progressed over weeks and showed a severe retinal arterial vasculopathy with arteriolar narrowing, ghost vessel formation and the development of optic atrophy. The picture in 2 of the patients resembled that of the acute retinal necrosis syndrome (ARN). Antibodies to African horsesickness (AHS) virus were detected. The serology for AHS virus was positive in all 4 patients as well as in 5 of 15 laboratory workers from the same facility who were clinically and ophthalmologically normal. This is to our knowledge the first description of subclinical and probable clinical neurotropic AHS virus infection in man. AHS is a hitherto-unrecognised possible cause of viral retinitis and the ARN syndrome.

Encephalitis and chorioretinitis associated with neurotropic African horsesickness virus infection in laboratory workers. Part III. Virological and serological investigations.

Four cases of encephalitis with chorioretinitis occurred in the vaccine-packing section of a veterinary research institute: 1 in 1982, 1 in 1985 and 2 in 1989. No viruses were isolated from patients and serological tests failed to reveal significant antibodies to a range of viruses incorporated in veterinary vaccines or to other likely pathogens, except for low titres of complement-fixing antibody to African horsesickness (AHS) virus in all 4 patients. In confirmatory tests, high enzyme immunoassay titres of antibody to AHS virus occurred in the 4 patients and lower titres in 5/58 other workers at the institute. The 4 patients had significant plaque reduction neutralisation antibody titres to some of the strains of virus incorporated in AHS vaccine, particularly to serotypes 1 and 6, which had undergone neuro-adaptation through serial intracerebral passage in mice and which were known to be encephalitogenic following intranasal instillation in horses, guinea pigs and dogs. It is believed that the patients may have acquired aerosol infection with AHS virus as a result of accidental breakage of freeze-dried vaccine bottles.

Encephalitis and chorioretinitis associated with neurotropic African horsesickness virus infection in laboratory workers. Part IV. Experimental infection of the vervet monkey (Cercopithecus pygerythrus).

Neurotropic vaccine strains of African horsesickness (AHS) virus types 1 and 6 were implicated as the possible aetiological agents in 4 cases of encephalitis and uveochorioretinitis in laboratory workers accidentally exposed to the freeze-dried vaccine preparations of the virus. To date, AHS virus has not been known to infect man. To ascertain whether or not primates were susceptible to infection with AHS virus, vervet monkeys (Cercopithecus pygerythrus) were inoculated, either transnasally or intraconjunctivally, with vaccine strains of AHS virus types 1 and 6. The course of infection was monitored using parameters such as behavioural changes, febrile reaction, cerebrospinal fluid pleocytosis, serology, magnetic resonance imaging and autopsy. Encephalitis, manifested by varying degrees of fever, behavioural changes and pleocytosis, but no chorioretinitis was detected in all 6 transnasally infected monkeys. This was confirmed by autopsy, where a meningo-encephalitis affecting the medial temporal lobe but no lesions in the eyes was demonstrated. Neither virus appeared to infect the animals after intraconjunctival inoculation. These findings support the theory that the patients were infected by the inhalation of freeze-dried vaccine preparations. The pathogenesis of the eye lesions, however, remains uncertain.

Protective efficacy of virus-like particles for bluetongue disease.

Bluetongue virus-like particles (VLPs) derived from multiple baculovirus expression vectors have been administered in the presence of various adjuvants to sheep, a vertebrate host susceptible to the virus, and the neutralizing antibody responses are measured. Vaccinated sheep are challenged after 4 months of inoculation, and clinical reaction indices and viraemia determined. The results indicate that these multiprotein virus-like particles lacking the genetic material are highly immunogenic and as little as 10 micrograms of VLPs in conjunction with appropriate adjuvant elicit an immune response which protects against infectious virus challenge. The formation of virus-like particles using this new technology offers a novel approach in vaccinology.

Recombinant virus vaccine for bluetongue disease in sheep.

Bluetongue virus proteins derived from baculovirus expression vectors have been administered in different combinations to sheep, a vertebrate host susceptible to bluetongue virus, and the neutralizing antibody responses were measured. Vaccinated sheep were subsequently challenged, and the indices of clinical reaction were calculated. The results indicated that the outer capsid protein VP2 alone in doses of greater than 50 micrograms per sheep elicited protection. A dose of ca. 50 micrograms of VP2 protected some but not all sheep. However, when used in combination with ca. 20 micrograms of the other outer capsid protein, VP5, 50-micrograms quantities of VP2 not only protected all the vaccinated sheep but also elicited a higher neutralizing-antibody response. The addition of viral core proteins VP1, VP3, VP6, and VP7, the nonstructural proteins NS1, NS2, and NS3, and the outer capsid proteins VP2 and VP5 did not enhance this neutralizing-antibody response.

Characterization of Palyam serogroup orbiviruses isolated in South Africa and serologic evidence for their widespread distribution in the country.

The finding that there had been multiple isolations of Palyam serogroup orbiviruses from aborted cattle fetuses in neighbouring Zimbabwe, suggested that there was a need to investigate the possible occurrence of the viruses in South Africa. Unidentified viruses isolated in South Africa, which had been in storage, were examined. Four viruses which had been isolated from Culicoides midges collected at various sites in the years from 1969 to 1977, were identified as three strains of Gweru virus and one of the Nyabira virus (Palyam group serotypes originally described from Zimbabwe). A fifth virus, isolated in 1967 from the blood of a cow with mild fever, was found to be a distinct new member of the Vellore antigenic complex of the Palyam serogroup and was named Apies River virus. Sera from 476 cattle, 150 sheep, 24 goats and 78 humans from 10 farms were tested for neutralizing antibodies to the above three serotypes of virus plus Abadina and Marondera serotypes. Only 1 of 100 cattle sera from two farms in the southern coastal area had antibody, but elsewhere there was a high prevalence of antibody with 254 (53%) of all cattle exhibiting activity for one or more of the five serotypes of virus tested. Only 6 (4%) sheep, 3 (12.5%) goats and 11 (14%) humans had antibody.

Isolation of a capsid protein of bluetongue virus that induces a protective immune response in sheep.

A method to purify the neutralization specific antigen of bluetongue virus P2 in large amounts has been developed. The purified protein is free from virus-specified or cellular contaminants and its immunological specificity has been preserved. The purification is based on the observation that protein P2 can be dissociated from the virion by treatment with monovalent or divalent salts. The salt concentration required to solubilize the outer capsid proteins is pH dependent and in general decreases with a decrease in pH. P2 purified by extraction from polyacrylamide gels does not induce immune-precipitating or neutralizing antibodies. The response against P5, on the other hand, is much less conformational dependent and P5 purified from gels readily induces P5-precipitating antibodies in rabbits. These antibodies do not neutralize the virus. Purified P2, immunoabsorbed with anticore serum to remove trace amounts of P7, was injected into sheep. An initial dose of 50 micrograms of P2 was sufficient to induce P2-precipitating antibodies as well as neutralizing and hemagglutination-inhibiting antibodies. These sheep were fully protected against challenge with a virulent strain of the same BTV serotype. Lower doses of P2 still provided a significant level of protection even though no neutralizing antibodies could be detected.

Rubella--a case for immunising male adolescents.

An outbreak of rubella at a training institution for predominantly adolescent males is described. The cost and inconvenience of the outbreak furnish evidence that immunisation in its own right is worth while, and should not be seen only as a means of protecting susceptible females of child-bearing age.

Comparison of techniques for demonstrating antibodies to Rift Valley fever virus.

Nine serological techniques were compared by monitoring the response to infection with Rift Valley fever (RVF) virus in three sheep. Antibodies were monitored daily for the first 14 days after infection, then weekly and later fortnightly up to week 24. The earliest antibody response was detected in one sheep on day 3 by a plaque reduction neutralization test, and by day 6 antibodies were demonstrable in all three sheep by haemagglutination-inhibition, reversed passive haemagglutination-inhibition, immunodiffusion, indirect immunofluorescence (IF), enzyme-linked immunosorbent assay and neutralization of cytopathic effect in cell cultures. Antibodies were demonstrable by complement fixation on day 8 at the earliest. IF and the two neutralization techniques produced the highest titres, but all tests could be used satisfactorily for the serological diagnosis of RVF. Inactivated antigen could be used for all except the neutralization tests. A radioimmunoassay technique using 125I-labelled staphylococcal protein A detected antibodies on day 8 at the earliest and produced lower mean titres than some of the other techniques. This was probably because sheep immunoglobulins bind protein A poorly.

Comparative pathogenicity and antigenic cross-reactivity of Rift Valley fever and other African phleboviruses in sheep.

Homologous and heterologous haemagglutination-inhibition (HAI), complement-fixation (CF), immunodiffusion (ID) and mouse neutralization tests were performed with the Lunyo (LUN) and a Zimbabwean strain of Rift Valley fever (RVF) virus, the prototype and a South African strain of Arumowot (AMT) virus and prototype strains of Gordil (GOR), Saint-Floris (SAF) and Gabek Forest (GF) viruses, using immune mouse ascitic fluids prepared against these viruses. Reactions of identity occurred in all tests between LUN and the Zimbabwean strains of RVF and between the two strains of AMT virus. Otherwise, cross-reactions occurred between all the phleboviruses in HAI tests, while reactions in CF, ID and neutralization tests were monospecific for virus serotypes, except that weak cross-reaction occurred between GOR and SAF viruses in CF and ID tests. Four sheep infected subcutaneously with the Zimbabwean strain of RVF virus developed transient fever, viraemia, leucopaenia, relative thrombocytopaenia, haemoconcentration and raised serum enzyme levels, which indicated that the sheep had developed necrotic hepatitis. Disseminated focal necrotic hepatitis was confirmed in a sheep killed for examination on day 4 post-infection. The other three sheep recovered uneventfully after only mild depression and anorexia. Groups of three sheep infected with SAF, GOR, AMT and GF viruses had no demonstrable viraemia or other sign of infection or illness, except that the sheep infected with AMT developed mild fever lasting less than 24 h. Antibody responses were monitored at intervals over a period of 24 weeks in all sheep by homologous and heterologous HAI, CF and cell culture neutralization (CPENT) tests. Homologous antibody responses were marked in the RVF-infected sheep and their sera cross-reacted strongly in HAI tests with antigens of the other viruses. The sera of the RVF-infected sheep cross-reacted less markedly in CF and CPENT tests. Homologous antibody responses were poor in all the sheep infected with phleboviruses other than RVF, and the cross-reactivity of their sera for RVF antigen or virus was negligible. All sheep were challenged with RVF virus 48 weeks after their initial infection. The sheep which had originally been infected with RVF virus were immune and developed neither fever nor viraemia. All other sheep developed fever, viraemia and antibodies to RVF virus. It was concluded that the African phleboviruses, other than RVF, are unlikely to cause disease in livestock or to induce antibodies which could cause confusion in the diagnosis of RVF.