The isolation and attenuation of a virus causing rhinotracheitis in turkeys in South Africa

In March 1978, a number of turkeys with severe respiratory symptoms affecting over 80% of the flock were investigated for a possible causative agent. With the standard techniques used for the isolation of bacteriae, mycoplasmae and viruses, only Mycoplasma gallisepticum, Mycoplasma meleagridis and Newcastle disease virus were isolated. Tracheal organ cultures were subsequently prepared from 27-day-old turkey embryos and inoculated with sinus exudate from affected turkeys. After an incubation period of 4 days a virus was isolated with which the typical symptoms, as observed in the field, could be reproduced in susceptible turkeys after 3-5 days. Following primary isolation in tracheal organ cultures, the virus grew readily in embryonated eggs and Vero cells. With the electron microscope, virus-like particles, varying in size from 40 nm-500 nm, were observed, having a pleomorphic shape and studded with fine surface projections. The virus seems to fall into the family Paramyxoviridae. A vaccine produced from attenuated virus in embryonated eggs afforded good protection against mortalities due to airsacculitis that normally follows on to turkey rhinotracheitis infection. The serological and clinical effects of the virus on chickens are also reported on.     

Improved immunogenicity of novel baculovirus-derived Theileria parva p67 subunit antigens

East Coast fever (ECF) in cattle is caused by the tick-borne protozoan parasite Theileria parva. The major sporozoite surface antigen of T. parva (p67) is an important candidate for inclusion in a subunit vaccine. Recently, we reported the expression and production of different parts of p67 as fusions to either GFP or to the baculovirus GP64 envelope glycoprotein in insect cells, which resulted in stable proteins recognized by a monoclonal specific for native p67. The immunogenicity of these fusion proteins was examined in out-bred mice and cattle. In mice, the full length p67 molecule without its signal peptide and transmembrane region, but fused to GFP (GFP:p67deltaSS) was the best immunogen followed by the C-terminus of p67 fused to GP64 (GP64:p67C). These two immunogens also provoked a high level of sero-conversion in cattle when formulated in a water-in-oil or saponin-derived adjuvant with only 100 microg of protein and a single booster. The vaccine-elicited antibodies efficiently inhibited the infectivity of T. parva sporozoites in in vitro neutralization assays. This study demonstrated that these new baculovirus-derived p67 vaccines were highly immunogenic, and that in combination with a suitable adjuvant, they have a clear potential to induce protective immunity in cattle.     

Advances in mucosal vaccination

Pathogens that enter the body via mucosal surfaces face unique defense mechanisms that combine the innate barrier provided by the mucus layer with an adaptive response typified by the production and transepithelial secretion of pathogen-specific IgA. Both the measurement and induction of mucosal responses pose significant challenges for experimental and practical application and may need to be adapted to the species under study. In particular, for livestock, immunization procedures developed in small rodent models are not always effective in large animals or compatible with management practices. This paper reviews the latest advances in our understanding of the processes that lead to secretory IgA responses and how this relates to the development of mucosal immunization procedures and adjuvants for veterinary vaccines. In addition, it highlights the complex interactions that can take place between the pathogen and the host's immune response, with specific reference to Chlamydia/Chlamydophila infections in sheep.     

The chicken pituitary expresses an ovoinhibitor-like protein in subpopulations of some, but not all, hormone-producing cell types

Ovoinhibitor is a serine protease-inhibiting protein that was originally purified from egg whites. It is secreted by the oviduct under the control of estrogen and progesterone and it specifically inhibits serine proteinases such as trypsin and chymotrypsin. During recent attempts to raise monoclonal antibodies (Mabs) against chicken bursa of Fabricius proteins, one Mab was produced that specifically recognized chicken ovoinhibitor. This was the first demonstration of ovoinhibitor in an avian immune organ. We presently report on the expression of an ovoinhibitor-like molecule by the pituitary of the chicken as revealed by immunocytochemistry and RT-PCR. Immunofluorescent dual staining experiments using the mouse anti-ovoinhibitor Mab in conjunction with polyclonal antibodies against various hypophysial hormones revealed partial co-localization of an ovoinhibitor-like molecule with growth hormone (GH), luteinizing hormone (LH), and pro-opiomelanocortin (POMC), in a subset of the respective hormone producing cells. By contrast, no co-localization with prolactin (PRL) could be reliably demonstrated. RT-PCR of hypophysial mRNA using ovoinhibitor gene-specific primers yielded an amplicon that was 20% shorter than predicted on the basis of the published ovoinhibitor sequence. Sequencing revealed that of the represented exons only the central portion was expressed in the pituitary and that both 5′ and 3′ ends of each exon had been truncated. While expression of ovalbumin-like serine protease inhibitors (serpins) has been previously reported in the rat pituitary, to our knowledge, this is the first report of a Kazal-type serine protease inhibitor in the vertebrate neuroendocrine system.     

Cellular and molecular characterisation of the ovine rectal mucosal environment

Immunohistological characterisation of ovine rectal tissue has revealed the presence of lymphoid follicles, predominantly in the submucosa, that closely resemble those found in intestinal Peyer's patches (PPs). Distinct T (CD4+, CD8+, gammadelta-TCR+) and B (CD21+, CD45R+) lymphocyte staining patterns were observed within and around follicles of the rectal mucosa. In addition, IgA+ and IgE+ cells were also found at this tissue site, with both phenotypes commonly residing in the lamina propria. RT-PCR examination of the cytokines expressed in the rectal mucosal tissue revealed consistently high levels of TGFbeta and IL-8 mRNA, low levels of IL-2 mRNA and no detectable IL-4 mRNA. The presence of lymphoid follicles, IgA+ plasma cells and IgA-inducing cytokines in rectal tissue of sheep indicate that this may be a suitable route for delivering mucosal vaccines.     

A wind density model to quantify the airborne spread of Culicoides species during north-western Europe bluetongue epidemic, 2006

Increased transport and trade as well as climate shifts play an important role in the introduction, establishment and spread of new pathogens. Arguably, the introduction of bluetongue virus (BTV) serotype 8 in Benelux, Germany and France in 2006 is such an example. After its establishment in receptive local vector and host populations the continued spread of such a disease in a suitable environment will mainly depend on movement of infected vectors and animals. In this paper we explore how wind models can contribute to explain the spread of BTV in a temperate eco-climatic setting. Based on previous work in Greece and Bulgaria filtered wind density maps were computed using data from the European Centre for Medium-Range Weather Forecasts (ECMWF). Six hourly forward wind trajectories were computed at pressure levels of 850 hPa for each infected farm as from the recorded onset of symptoms. The trajectories were filtered to remove wind events that do not contribute to possible spread of the vector. The suitable wind events were rastered and aggregated on a weekly basis to obtain weekly wind density maps. Next to this, cumulated wind density maps were also calculated to assess the overall impact of wind dispersal of vectors. A strong positive correlation was established between wind density data and the horizontal asymmetrical spread pattern of the 2006 BTV8 epidemic. It was shown that short (<5 km), medium (5–31 km) and long (>31 km) distance spread had a different impact on disease spread. Computed wind densities were linked to the medium/long-distance spread whilst short range spread was mainly driven by active Culicoides flight. Whilst previous work in the Mediterranean basin showed that wind driven spread of Culicoides over sea occurred over distances of up to 700 km, this phenomenon was not observed over land. Long-distance spread over land followed a hopping pattern, i.e. with intermediary stops and establishment of local virus circulation clusters at distances of 35–85 km. Despite suitable wind densities, no long range spread was recorded over distances of 300–400 km. Factors preventing spread Eastwards to the UK and Northwards to Denmark during the 2006 epidemic are discussed. Towards the east both elevation and terrain roughness, causing air turbulences and drop down of Culicoides, were major factors restricting spread. It is concluded that the proposed approach opens new avenues for understanding the spread of vector-borne viruses in Europe. Future developments should take into consideration both physical and biological factors affecting spread.     

Low impact of present and historical landscape configuration on the genetics of fragmented Anthyllis vulneraria populations

Decreasing habitat fragment area and increasing isolation may cause loss of plant population genetic diversity and increased genetic differentiation between populations. We studied the relation between the historical and the present landscape configuration (i.e., patch area and patch connectivity), and the present management of calcareous grassland fragments on the one hand, and the within and between population genetic structure of 18 Anthyllis vulneraria populations on the other hand. Despite the long-time fragmentation history and the mainly selfing breeding system of the species, we detected very low genetic differentiation (Φ_st = 0.056) among habitat fragments and no significant isolation-by-distance relation. Average within fragment genetic diversity measured as molecular variance and expected heterozygosity, were relatively high (16.46 and 0.28, respectively), and weakly positively correlated with the current fragment area, most likely because larger fragments contained larger populations. We found no effects of the historical landscape configuration on the genetic diversity of the populations. Our data suggest that the consequences of habitat fragmentation for genetic differentiation and genetic diversity of A. vulneraria are relatively minor which is very likely due to the historical high levels of seed exchange among fragments through grazing and roaming livestock. This study provides indirect evidence that nature management by grazing not only positively affects habitat quality but that it might also mitigate the genetic consequences of habitat fragmentation. From the conservation point of view, this study illustrates the importance of grazing and of the regular transport of livestock between fragments to prevent the long-term effects of fragmentation on the genetic diversity of the populations studied.     

Red clover varieties of Mattenklee type have higher production,protein yield and persistence than Ackerklee types in grass–clover mixtures

Dairy farmers have increased interest in multiyear red clover–ryegrass ley pasture swards because of their high productivity and protein concentration but the adoption of such mixtures has been restricted because of poor persistency of many red clover cultivars currently used. We aimed (i) to assess dry‐matter yield (DMY), persistence and nutritive value of four Mattenklee and four Ackerklee cultivars and (ii) to investigate the underlying plant morphological traits. We conducted a 4‐year field experiment with red clover–ryegrass mixtures, in combination with a short‐term pot experiment in which the red clover cultivars were grown both in monoculture and in mixture with Lolium perenne L. In the field, Mattenklees showed higher production (on average 42% higher) and clover content (on average 34% higher) in the third and fourth year of production compared to Ackerklees. In the fourth year, Mattenklees had slightly lower digestibility and lower protein concentration compared to Ackerklees cultivars; however, the digestible DMY and the protein yield were higher for Mattenklees. Both persistence and nutritive value of the different red clover cultivars were closely correlated with plant morphological traits. For stem length, there was close correlation (r > .77) between the results from the pot experiment and the field experiment, suggesting that stem length may be an important trait for plant breeding.