The enigmatic pathogenesis of malignant catarrhal fever (MCF) involves dysregulated immune responses in susceptible ruminant species. Economically important outbreaks of MCF are due to 2 of the 10 viruses currently comprising the malignant catarrhal fever virus group: ovine herpesvirus 2 (OvHV-2) and alcelaphine herpesvirus 1 (AlHV-1). Attempts to develop effective vaccines for this group of viruses in the 1970s were sufficiently discouraging that they were temporarily abandoned. This review focuses on recent efforts to understand the pathogenesis of MCF, particularly the sheep-associated form of the disease, with the goal of developing rational control methods, including vaccination. The past 2 decades have seen several advances, including recognition of new members of the MCF virus group, better diagnostic assays, induction of disease by a natural route (aerosol), and clearer understanding of OvHV-2’s shedding patterns by domestic sheep. A consistent theme in experimental studies of OvHV-2 in susceptible species is that there are 2 peaks of OvHV-2 gene expression: a preclinical peak involving the respiratory tract and a second in multiple organ systems leading to clinical disease. Latent and lytic gene expression may coexist in tissues during clinical stages in symptomatic animals.

https://journals.sagepub.com/doi/pdf/10.1177/0300985813520435

The pathology of malignant catarrhal fever, with an emphasis on ovine herpesvirus 2.

Malignant catarrhal fever (MCF) is an often lethal infection of many species in the order Artiodactyla. It is caused by members of the MCF virus group within Gammaherpesvirinae. MCF is a worldwide problem and has a significant economic impact on highly disease-susceptible hosts, such as cattle, bison, and deer. Several epidemiologic forms of MCF, defined by the reservoir ruminant species from which the causative virus arises, are recognized. Wildebeest-associated MCF (WA-MCF) and sheep-associated MCF (SA-MCF) are the most prevalent and well-studied forms of the disease. Historical understanding of MCF is largely based on WA-MCF, in which the causative virus can be propagated in vitro. Characterization of SA-MCF has been constrained because the causative agent has never been successfully propagated in vitro. Development of molecular tools has enabled more definitive studies on SA-MCF. The current understanding of MCF, including its etiological agents, epidemiology, pathogenesis, and prevention, is the subj...

Malignant Catarrhal Fever: Inching Toward Understanding

Wildebeest-associated malignant catarrhal fever (WA-MCF), an acute lymphoproliferative disease of cattle caused by alcelaphine herpesvirus 1 (AlHV-1), remains a significant constraint to cattle production in nomadic pastoralist systems in eastern and southern Africa. The transmission of WA-MCF is dependent on the presence of the wildlife reservoir, i.e. wildebeest, belonging to the species Connochaetes taurinus and Connochaetes gnou; hence, the distribution of WA-MCF is largely restricted to Kenya, Tanzania and the Republic of South Africa, where wildebeest are present. WA-MCF is analogous to sheep-associated MCF (SA-MCF) in many aspects, with the latter having sheep as its reservoir host and a more global distribution, mainly in developed countries with intensive livestock production systems. However, unlike SA-MCF, the geographic seclusion of WA-MCF may have contributed to an apparent neglect in research efforts aimed at increased biological understanding and control of the disease. This review aims to highlight the importance of WA-MCF and the need for intensified research towards measures for its integrated control. We discuss current knowledge on transmission and geographical distribution in eastern and southern Africa and the burden of WA-MCF in affected vulnerable pastoral communities in Africa. Recent findings towards vaccine development and pertinent knowledge gaps for future research efforts on WA-MCF are also considered. Finally, integrated control of WA-MCF based on a logical three-pronged framework is proposed, contextualizing vaccine development, next-generation diagnostics, and diversity studies targeted to the viral pathogen and cattle hosts.

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Wildebeest-associated malignant catarrhal fever: perspectives for integrated control of a lymphoproliferative disease of cattle in sub-Saharan Africa

A field vaccine trial in Tanzania demonstrates partial protection against malignant catarrhal fever in cattle.

Amino acids might be a tool to transform animals from a pro- to an anti-inflammatory phenotype through the downregulation of several genes (TLR-4, NF-κB, TNFa, IL-1β, IL-2, IL-6, IL-8, CCL-5 and CXCL-16) whose expression increases during inflammation. To examine this possibility, each of sixty Chios dairy sheep was assigned to one of the following five dietary treatments: A: basal diet (control group); B: basal diet +6.0 g/head rumen-protected methionine (MetaSmart™ ); C: basal diet +5.0 g/head rumen-protected lysine (LysiGEM™ ); D: basal diet +6.0 g/head MetaSmart™  + 5.0 g/head LysiGEM™ ; and E: basal diet +12.0 g/head MetaSmart™  + 5.0 g/head LysiGEM. The results showed a significant downregulation in the expression of the TLR-4 gene in both the macrophages and the neutrophils of ewes fed rumen-protected amino acids. Significantly lower mRNA transcript accumulation was also observed for the TNFa, IL-1β and CXCL-16 genes in the macrophages and for the IL-1β gene in the neutrophils of ewes supplemented with amino acids. The ewes that received dietary supplementation with rumen-protected lysine alone (C) had significantly lower CCL-5 transcript levels in their macrophages than the ewes fed the other supplemented diets. Diet D enhanced the mRNA expression of the IL-2 gene in ewe neutrophils. Negative correlations were found between: a. TLR-4, TNFa, IL-1β and CXCL-16 gene expression in macrophages and the milk fat and total solids content; b. CCL-5 gene expression in neutrophils and the milk yield and FCM(6%) ; and c. CXCL-16 gene expression and the milk protein content. Moreover, positive correlations were found between the BHBA concentration and the expression of the TLR-4 and CXCL-16 genes in macrophages. In conclusion, the rumen-protected amino acids improved sheep metabolism (as indicated by reduced blood BHBA and urea concentrations), milk chemical composition and immune system function.

The effects of dietary supplementation with rumen‐protected amino acids on the expression of several genes involved in the immune system of dairy sheep

Protection of cattle from alcelaphine herpesvirus-1 (AlHV-1)-induced malignant catarrhal fever (MCF) has been described previously, using an attenuated virus vaccine in an unlicensed adjuvant. The vaccine was hypothesised to induce a protective barrier of virus-neutralising antibody in the oro-nasal region, supported by the observation of high titre neutralising antibodies in nasal secretions of protected animals. Here we describe further analysis of this vaccine strategy, studying the effectiveness of the vaccine formulated with a licensed adjuvant; the duration of immunity induced; and the virus-specific antibody responses in plasma and nasal secretions. The results presented here show that the attenuated AlHV-1 vaccine in a licensed adjuvant protected cattle from fatal intranasal challenge with pathogenic AlHV-1 at three or six months. In addition, animals protected from MCF had significantly higher initial anti-viral antibody titres than animals that succumbed to disease; and these antibody titres remained relatively stable after challenge, while titres in vaccinated animals with MCF increased significantly prior to the onset of clinical disease. These data support the view that a mucosal barrier of neutralising antibody blocks infection of vaccinated animals and suggests that the magnitude of the initial response may correlate with long-term protection. Interestingly, the high titre virus-neutralising antibody responses seen in animals that succumbed to MCF after vaccination were not protective.

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Duration of protective immunity and antibody responses in cattle immunised against alcelaphine herpesvirus-1-induced malignant catarrhal fever.

We wished to determine the effect of of CpG ODN adjuvant on the magnitude and duration of protective immunity against alcelaphine herpesvirus-1 (AlHV-1) malignant catarrhal fever (MCF), a fatal lymphoproliferative disease of cattle. Immunity was associated with a mucosal barrier of virus-neutralising antibody. The results showed that CpG ODN included either with emulsigen adjuvant and attenuated AlHV-1 (atAlHV-1) or alone with atAlHV-1 did not affect the overall protection from clinical disease or duration of immunity achieved using emulsigen and atAlHV-1. This is in contrast to other similar studies in cattle with BoHV-1 or cattle and pigs with various other immunogens. In addition to this, several other novel observations were made, not reported previously. Firstly, we were able to statistically verify that vaccine protection against MCF was associated with virus-neutralising antibodies (nAbs) in nasal secretions but was not associated with antibodies in blood plasma, nor with total virus-specific antibody (tAb) titres in either nasal secretions or blood plasma. Furthermore, CpG ODN alone as adjuvant did not support the generation of virus-neutralising antibodies. Secondly, there was a significant boost in tAb in animals with MCF comparing titres before and after challenge. This was not seen with protected animals. Finally, there was a strong IFN-γ response in animals with emulsigen and atAlHV-1 immunisation, as measured by IFN-γ secreting PBMC in culture (and a lack of IL-4) that was not affected by the inclusion of CpG ODN. This suggests that nAbs at the oro-nasal-pharyngeal region are important in protection against AlHV-1 MCF.

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The effect of the TLR9 ligand CpG-oligodeoxynucleotide on the protective immune response to alcelaphine herpesvirus-1-mediated malignant catarrhal fever in cattle

In sheep, the A and B loci encoding the α and β chains of the classical class II MHC molecules are DRA and DRB and DQA and DQB. Previous analyses described the duplication of the DQA and DQB genes. The majority of haplotypes include DQA1 and DQA2 loci, however, in a number of haplotypes, DQA1 appears absent and these haplotypes have been described as DQA1 null. In these haplotypes, the DQA2 locus is found in combination with a second locus which appeared more closely related to DQA2 than DQA1, hence the description of this locus as DQA2-like. Here we combine our previous analysis of the DQA transcripts with an analysis of the associated DQB transcripts in ten haplotypes from MHC homozygous animals. This allows the potential for surface expression of different haplotype combinations of DQA and B genes and the functional significance of DQA2-like and its predicted DQB partner to be determined. Atypical DQB transcripts (DQB2-like) were identified in haplotypes classified as DQA1-null and conserved DQB2-like orthologues were identified in other Bovidae indicating trans-species conservation of the allelic lineage. Functional combinations detected by co-transfection of DQ1, DQ2 and DQ2-like genes demonstrates the potential for a wide range of DQ molecules derived from both intra- and inter-haplotype as well as inter-locus combinations. We provide evidence that DQA2-like and B2-like genes form an evolutionary conserved pair which generates structurally distinct class II molecules that are likely to present a distinct range of peptides to CD4+ T cells.

Structural and functional diversity arising from intra- and inter-haplotype combinations of duplicated DQA and B loci within the ovine MHC.

Herpesviruses often contain cryptic, spliced genes that are not obvious from the initial in silico annotation. Alcelaphine herpesvirus 1 (AlHV-1) contains 72 annotated ORFs but there are also a number of gaps between these that may have protein-coding potential. Comparative analysis of coding potential between AlHV-1 and the related ovine herpesvirus 2 (OvHV-2) revealed a putative novel spliced gene that we have termed A9.5. Analysis of cDNA clones from AlHV-1-infected cells revealed three overlapping clones corresponding to A9.5 and the coding sequence was confirmed by reverse transcription PCR of RNA from AlHV-1-infected cattle tissues. The A9.5 gene was predicted to encode a secreted glycoprotein with molecular mass 19 kDa. Empirical analysis showed that a recombinant haemagglutinin-tagged A9.5 fusion protein was secreted from transfected cells and had a molecular mass of 45 kDa, which was reduced to 20 kDa by endoglycosidase F treatment, confirming that A9.5 was a secreted glycoprotein. In situ RNA hybridization showed that A9.5 was expressed in cells associated with malignant catarrhal fever (MCF) lesions in infected cattle. Detailed analysis of the available OvHV-2 sequences revealed an homologous gene (Ov9.5) with conserved splicing signals and predicted amino acid sequence features in both sequenced isolates of this related virus. We have therefore identified a novel spliced gene in two related macaviruses that is expressed in MCF lesions. Future work will determine its importance for the pathogenesis of disease.

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Helen Todd

Papers

Duration of protective immunity and antibody responses in cattle immunised against alcelaphine herpesvirus-1-induced malignant catarrhal fever.

The effect of the TLR9 ligand CpG-oligodeoxynucleotide on the protective immune response to alcelaphine herpesvirus-1-mediated malignant catarrhal fever in cattle

Structural and functional diversity arising from intra- and inter-haplotype combinations of duplicated DQA and B loci within the ovine MHC.

A novel spliced gene in alcelaphine herpesvirus 1 encodes a glycoprotein which is secreted in vitro.

Host gene expression changes in cattle infected with Alcelaphine herpesvirus 1.