William Horner Andrews (1887-1953)- first professor of physiology at Onderstepoort.

W H Andrews qualified as a veterinarian in London in 1908 and was recruited soon after, in 1909, by Sir Arnold Theiler to join the staff of the newly established veterinary laboratory at Onderstepoort. After initial studies on the treatment of trypanosomosis and on snake venoms he was deployed by Theiler in 1911 to start research on lamsiekte (botulism)at a field station on the farm Kaffraria near Christiana, where he met and married his wife Doris. After a stint as Captain in the SA Veterinary Corps during World War I he succeeded D T Mitchell as head of the Allerton Laboratory in 1918, where he excelled in research on toxic plants, inter alia identifying Matricaria nigellaefolia as the cause of staggers in cattle. When the Faculty of Veterinary Science was established in 1920 he was appointed as the first Professor of Physiology. After the graduation of the first class in 1924, and due to health problems, he returned to the UK, first to the Royal Veterinary College and then to the Weybridge Veterinary Laboratories of which he became Director in 1927. After his retirement in 1947 he returned to South Africa as a guest worker at Onderstepoort where he again became involved in teaching physiology when Prof. Quin unexpectedly died in 1950. Andrews died in Pretoria in 1953 and was buried in the Rebecca Street Cemetery.

Direct vesicular transport of MHC class II molecules from lysosomal structures to the cell surface.

Newly synthesized MHC class II molecules are sorted to lysosomal structures where peptide loading can occur. Beyond this point in biosynthesis, no MHC class II molecules have been detected at locations other than the cell surface. We studied this step in intracellular transport by visualizing MHC class II molecules in living cells. For this purpose we stably expressed a modified HLA-DR1 beta chain with the Green Fluorescent Protein (GFP) coupled to its cytoplasmic tail (beta-GFP) in class II-expressing Mel JuSo cells. This modification of the class II beta chain does not affect assembly, intracellular distribution, and peptide loading of the MHC class II complex. Transport of the class II/ beta-GFP chimera was studied in living cells at 37 degrees C. We visualize rapid movement of acidic class II/beta-GFP containing vesicles from lysosomal compartments to the plasma membrane and show that fusion of these vesicles with the plasma membrane occurs. Furthermore, we show that this transport route does not intersect the earlier endosomal pathway.

Accumulation of HLA-DM, a regulator of antigen presentation, in MHC class II compartments.

The HLA-DM genes encode an unconventional HLA (human leukocyte antigen) class II molecule that is required for appropriate binding of peptide to classical HLA class II products. In the absence of DM, other class II molecules are unstable upon electrophoresis in sodium dodecyl sulfate and are largely associated with a nested set of peptides derived from the invariant chain called CLIP, for class II-associated invariant chain peptides. DMA and DMB associated and accumulated in multilaminar, intracellular compartments with classical class II molecules, but were found infrequently, if at all, at the cell surface. Thus, DM may facilitate peptide binding to class II molecules within these intracellular compartments.

History of bluetongue research at Onderstepoort.

Research on this economically important disease of ruminants, especially sheep, which had been named bluetongue by farmers in the 19th century, has been part and parcel of the activities at Onderstepoort ever since its establishment in 1908 and therefore covers a full century of the OVI's existence. In view of Onderstepoort's centenary celebration a brief overview of this research is given in terms of the historic milestones which influenced and guided global research on this and other viral diseases of animals.

HLA-DO is a negative modulator of HLA-DM-mediated MHC class II peptide loading.

BACKGROUND: Class II molecules of the major histocompatibility complex become loaded with antigenic peptides after dissociation of invariant chainderived peptides (CLIP) from the peptide-binding groove. The human leukocyte antigen (HLA)-DM is a prerequisite for this process, which takes place in specialised intracellular compartments. HLA-DM catalyses the peptide-exchange process, simultaneously functioning as a peptide 'editor', favouring the presentation of stably binding peptides. Recently, HLA-DO, an unconventional class II molecule, has been found associated with HLA-DM in B cells, yet its function has remained elusive. RESULTS: The function of the HLA-DO complex was investigated by expression of both chains of the HLA-DO heterodimer (either alone or fused to green fluorescent protein) in human Mel JuSo cells. Expression of HLA-DO resulted in greatly enhanced surface expression of CLIP via HLA-DR3, the conversion of class II complexes to the SDS-unstable phenotype and reduced antigen presentation to T-cell clones. Analysis of peptides eluted from HLA-DR3 demonstrated that CLIP was the major peptide bound to class II in the HLA-DO transfectants. Peptide exchange assays in vitro revealed that HLA-DO functions directly at the level of class II peptide loading by inhibiting the catalytic action of HLA-DM. CONCLUSIONS: HLA-DO is a negative modulator of HLA-DM. By stably associating with HLA-DM, the catalytic action of HLA-DM on class II peptide loading is inhibited. HLA-DO thus affects the peptide repertoire that is eventually presented to the immune system by MHC class II molecules.

Phylogenetic analyses of genes from South African LPAI viruses isolated in 2004 from wild aquatic birds suggests introduction by Eurasian migrants.

In 2004, South Africa experienced its first recorded outbreak of a highly pathogenic notifiable avian influenza (HPNAI) viral strain of the H5N2 subtype in ostriches in the Eastern Cape province. The traditional ostrich-farming areas in the Western Cape province report almost yearly outbreaks of low pathogenicity avian influenza (LPAI) in ostriches, which is attributed to introduction by wild birds and certain climatic patterns. During the winter of 2004, LPAI H3N8, H4N8, H5N2 and H5N1 avian influenza viruses were isolated from wild aquatic birds. All eight genes of the H3N8, H4N8 and H5N1 viruses were analysed. The results show that the H5N1 virus does not belong to the HPAI Z/Z+N genotype currently circulating in Asia, but that the most recent common ancestors are Russian H5N2 and H5N3 viruses. The N1 gene lacks the stalk deletion associated with virulence. Internal genes probably originate from a pool containing Chinese, Middle Eastern and Italian viruses. The South African H3N8 and H4N8 viruses appear to have derived their genes from an ecosystem where Asian H5N1, H6N9 and H9N2, Russian H4, and Danish H3N8 viruses have been circulating since 1997. All three viruses share recent nucleoprotein common ancestors with the German and Dutch HPNAI H7N7 viruses from 2003. The diverse pool of genes from which local viruses are derived suggests that reassortment occurred at the Siberian breeding grounds where migratory paths cross, or within the South African ecosystem. This data highlights the importance of surveillance in aquatic migratory birds, particularly members of the Charadriidae, for their potential roles in the introduction of avian diseases to South African poultry and especially ostriches in the case of avian influenza.

Detection of Maedi-Visna virus antibodies using a single fusion transmembrane-core p25 recombinant protein ELISA and a modified receiver-operating characteristic analysis to determine cut-off values.

The core p25 and transmembrane (TM) genes of Maedi-Visna virus (MVV) were cloned individually into the pGEX-2T expression vector. Both proteins were expressed as a combined fusion protein in frame with glutathione S-transferase (GST). The purified recombinant antigens (GST-TM and GST-TM-p25) were used to develop a MVV ELISA. A preliminary assessment of the diagnostic potential of the recombinant antigens (GST-TM and GST-TM-p25) was made by testing the antigens against 46 seropositive and 46 seronegative sheep and comparing the results with a commercial p25 ELISA kit. A two-graph receiver operating characteristic (TG-ROC) analysis program was used to interpret the data. The GST-TM-p25 ELISA was more sensitive than the commercial assay which is based on the p25 antigen alone and more specific than the GST-TM ELISA.

Experimental infection of vaccinated slaughter ostriches in a natural, open-air feedlot facility with virulent Newcastle disease virus.

The presence of virulent Newcastle disease virus (NDV) since the 1993-94 epidemic in southern Africa holds major implications for the export of ostrich products from this region. A challenge experiment with this field strain was conducted in open-air feedlot facilities under strict biosecurity measures. The experiment was designed to follow vaccination and preslaughter quarantine regulations currently enforced in South African export ostrich facilities in order to determine the viremia period and immune response under these specific circumstances. One hundred forty-three slaughter ostriches were allocated into three test groups, according to the time period between pretrial vaccination and challenge (1-2 mo, 2-4 mo, 4-6 mo), and an unchallenged control group. All birds in the test groups were challenged by oral, tracheal, and ocular routes with a field isolate of NDV. They were slaughtered over the next 4 wk on nine separate occasions and bled on 12 occasions. Virus isolation was attempted from seven sets of pooled samples from each bird to determine the viremia period and the serum antibody concentrations were measured by hemagglutination inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) methods to establish an immune response curve. NDV could be back-isolated only up to day 9 postinfection and from only six ostriches with poor immune response titers and corresponding to a rise in antibody levels above an indirect ELISA optical density reading of 0.33. Virus could be recovered only from brain and respiratory tract tissue. The HI test was less sensitive than the ELISA. Immune response curves did not differ significantly between the groups and peaked on day 14 post-infection. From these data, ELISA titers would appear to be a good indicator of the probability that an ostrich will be clinically infected after velogenic NDV challenge. These results also suggest that the current vaccination schedule enforced by the South African Veterinary Authorities results in protective immunity in up to 95% of slaughter ostriches from export approved facilities. The standard 30-day preslaughter quarantine period introduced as part of Crimean-Congo hemorrhagic fever virus control measures also appears sufficient to encompass the determined NDV viremia period of 9-11 days in slaughter ostriches.

Newcastle disease and avian influenza A virus in wild waterfowl in South Africa.

In an intensive ostrich farming area in South Africa with a history of ostrich influenza outbreaks, we conducted a survey of avian influenza virus (AIV) and Newcastle disease virus (NDV) in wild aquatic birds. During late autumn and winter 1998, the time of year when outbreaks in ostriches typically start to occur, 262 aquatic birds comprising 14 species were sampled and tested for both virus infections. From eight samples, AIV, serotype H10N9, could be isolated. All isolates were apathogenic as determined by the intravenous pathogenicity index (0.00). Conversely, none of 33 sera of these wild birds showed antibodies against H10. However, one bird was found serologically positive for H6 AIV. This AIV serotype was later isolated from ostriches during an avian influenza outbreak in this area. No NDV was isolated although 34 of 46 serum samples contained NDV-specific antibodies. This is the first H10N9 isolate to be reported from Africa. In addition, our data support the notion that wild aquatic birds may function as a reservoir for AIV and NDV in South Africa.

Detection of antibodies to Newcastle disease virus in ostriches (Struthio camelus) by an indirect ELISA.

A two-graph receiver operating characteristic analysis, performed on the hemagglutination-inhibition (HI) and enzyme-linked immunosorbent assay (ELISA) test results of a Newcastle disease virus (NDV)-positive and NDV-negative control group of ostrich sera, proved that the ELISA was superior to the HI in both sensitivity and specificity. Comparison of results of the two assays performed on a panel of simulated positive sera ranging from very weak to very strong showed that the ELISA was at least 10 times more sensitive than the HI in detecting low levels of ostrich antibodies to NDV. The ELISA also has the advantage of using untreated serum in a single dilution as opposed to the HI test, which uses pretreated serum in titration.

Mannose receptor mediated uptake of antigens strongly enhances HLA-class II restricted antigen presentation by cultured dendritic cells.

Dendritic cells (DCs) use macropinocytosis and mannose receptor mediated endocytosis for the uptake of exogenous antigens. Here we show that the endocytosis of the mannose receptor and mannosylated antigen is distinct from that of a non-mannosylated antigen. Shortly after internalization, however, both mannosylated and non-mannosylated antigen are found in an MIIC like compartment. The mannose receptor itself does not reach this compartment, and probably releases its ligand in an earlier endosomal structure. Finally, we found that mannosylation of peptides strongly enhanced their potency to stimulate HLA class II-restricted peptide-specific T cell clones. Our results indicate that mannosylation of antigen leads to selective targeting and subsequent superior presentation by DCs which may be useful for vaccine design.

Ostrich diseases.

Scientific knowledge of ostrich diseases is incomplete and very fragmented, with specific details on technical aspects of diagnostic and/or screening tests completely absent in most cases. Salmonella Typhimurium is common in multispecies collections and causes mortality in chicks younger than three months on commercial farms, but is rarely found in chicks older than six months, or slaughter birds of twelve to fourteen months in southern Africa. Campylobacter jejuni and Chlamydia psittaci are occasionally reported, mainly in young ostriches, but both remain a diagnostic challenge. Crimean-Congo haemorrhagic fever is transmitted to domestic animals including ostriches, principally by ticks of the genus Hyalomma. In the ostrich, the disease causes no clinical symptoms during a viraemia of approximately four days. Spongiform encephalopathy has not been reliably reported in ostriches, while anthrax has occurred rarely in modern times but was reportedly an important cause of death approximately 100 years ago in South Africa. Salmonella Gallinarum and S. Pullorum are unknown in ostriches. Pasteurella multocida occurs but is easily contained with antibiotics. Mycoplasma spp. are regularly found in an upper respiratory disease syndrome complicated by opportunistic bacterial pathogens. Ostriches of all ages are susceptible to challenge by velogenic Newcastle disease virus (NDV), but standard inactivated La Sota poultry vaccines can stimulate protective immunity lasting over six months. The viraemic period in vaccinated slaughter ostriches is between nine and eleven days and there are no indications of a carrier state or presence of the virus in the meat or any other tissues after this period, with peak immunoglobulin G response reached on day fourteen post infection. Haemagglutination inhibition tests are significantly less sensitive and less specific than enzyme-linked immunosorbent assays. Cloacal and choanal swabs used for direct virological screening in clinically affected cases (field and experimental) could not detect NDV. All avian influenza isolates reported from ostriches have been non-pathogenic to poultry, even the H5 and H7 subtypes. Some of the latter have been associated with mortality of ostrich chicks in localised outbreaks during periods of inclement weather and with significant wild bird (waterfowl) contact. Borna disease causes a nervous syndrome in ostrich chicks, but to date, has only been reported in Israel. Eastern and Western equine encephalomyelitides cause fatal disease in ostriches and other ratites, with mortality ranging from less than 20% to over 80% in affected flocks. These diseases are present in North, Central and South America where the associated ornithophilic mosquito vectors occur. Equine and human vaccines are apparently safe and efficacious in ratites. Wesselsbron disease, infectious bursal disease (type 2), adenovirus and coronavirus infections have been reported from ostriches but the significance of these diseases is unclear. Due to the paucity of data regarding ostrich diseases and the unvalidated state of most poultry tests in this unique group of birds, strict observation of a pre-slaughter quarantine of thirty days is strongly advised, whilst live exports and fertile eggs should be screened through the additional use of sentinel chickens and/or young ostriches.

The role of veterinary research laboratories in the provision of veterinary services.

Veterinary research laboratories play an essential role in the provision of veterinary services in most countries. These laboratories are the source of new knowledge, innovative ideas and improved technology for the surveillance, prevention and control of animal diseases. In addition, many laboratories provide diagnostic and other services. To ensure the optimal integration of various veterinary activities, administrators must understand the functions and constraints of research laboratories. Therefore, a brief discussion is presented of the following: organisational structures methods for developing research programmes outputs of research scientists and how these are measured the management of quality assurance funding of research. Optimal collaboration can only be attained by understanding the environment in which a research scientist functions and the motivational issues at stake.

Biotechnology, viral oncogenesis and jaagsiekte.

A brief description is given of the discovery of retroviral and cellular oncogenes and of their putative role in oncogenesis. Attempts to apply the biotechnological techniques that were so successful in the study of other retroviruses to the newly-discovered jaagsiekte retrovirus are briefly reviewed.

A scanning and transmission electron microscopy study of jaagsiekte lesions.

A scanning electron microscopy (SEM) and transmission electron microscopy (TEM) study was made of lesions from acute, experimentally induced cases of jaagsiekte. In the SEM study tumour cells were easily identified by the abundant microvilli on their peripheral surface. The SEM study gave further insight into the development of lesions and the spatial relationship of cells involved in jaagsiekte. TEM revealed that the tumour cells were in a state of rapid protein synthesis and had many characteristics in common with other malignant cells.

Transmission of jaagsiekte (ovine pulmonary adenomatosis) by means of a permanent epithelial cell line established from affected lungs.

An epithelial cell line, designated JS-15,4, has been established in culture from jaagiekte lesions and subcultured in vitro for almost 2 years. It exhibits morphological and other features of transformed cells and has been shown by electron microscopy to consist of type B ovine alveolar epithelial cells. Jaagiekte was successfully transmitted to 3 new-born lambs by the intratracheal injection of cells following immunosuppressive treatment with either anti-thymocyte immunoglobulin alone or combined with anti-macrophage immunoglobulin. Incubation periods as short as 10 weeks were recorded. Evidence was also obtained that natural transmission may result from the inhalation of viable cells.

Immunosuppression in new-born lambs.

A novel technique, based on cytotoxicity-neutralization, was developed for the in vitro titration of anti-sheep lymphocyte and anti-sheep macrophage sera. The titres obtained for a number of antisera were compared with those found in an agglutination assay. Anti-lymphocyte sera with a high cytotoxicity-neutralization titre very effectively suppressed the number of circulating lymphocytes in the peripheral blood of treated new-born lambs, thus indicating in vivo immunosuppressive activity.

Presence of Herpesvirus ovis DNA sequences in cellular DNA from sheep lungs affected with jaagsiekte (pulmonary adenomatosis).

To investigate further the possible involvement of Herpesvirus ovis in the aetiology of jaagsiekte, the kinetics of reassociation of viral DNA and DNA isolated from tumour tissue as well as from cell cultures derived from it were studied. Although DNA-DNA hybridization could be demonstrated in 2 cases of jaagsiekte, no correlation was found between the presence of Herpesvirus ovis genome sequences and the occurrence of the disease.