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Biomedical subjects

D Hannant

Publications and source records attributed to D Hannant.

At least 37 records · Page 2Linked to original sources
PubMed2000

Equine immunity to viruses.

The identification of some of the adaptive immune responses to infection with equine viruses has been the first step toward rational immunoprophylactic design. Sufficient knowledge of infection-induced immunity and informed estimates of the requirements for long-term immunity for EIV have now been obtained. Thus, the future for inactivated EIV vaccines is promising now that new adjuvants have been applied to induce cellular immunity and safe methods have been designed to stimulate virus-neutralizing (VN) antibody at mucosal surfaces. Adenoviruses induce circulating VN antibody, the presence of which appears to correlate with protection from reinfection. Therefore, the potential of vaccines to induce VN antibody and protect from challenge is an important next step with this virus. With persistent viruses such as EHV-1, antibody-mediated protection from infection can be achieved only at the site of initial infection, that is, the nasopharynx and upper respiratory tract. Systemic dissemination is very rapid and consequently VN antibody is unlikely to play a major role in prevention of disease once the initial infection event has occurred. Cellular immune responses, particularly CTLs, play a dominant role in protection and recovery and are important in immune surveillance and determination of the outcome of reactivation of latent virus. Therefore, the key to future EHV-1 vaccine design is to focus on stimulation of CTL responses, and this requires the successful presentation of vaccine-derived antigenic peptides to MHC class I molecules that are recognized by specific receptors on CTL. There is some evidence that stimulation of EHV-1-specific CTL precursors may correlate with immunity to this virus. By analogy with gamma herpesviruses in humans, CTL precursor frequency may also function as an immune correlate for EHV-2. Although EAV infection induces strong immunity in females and geldings, persistent infection of the genital tract is an important route of dissemination from stallions. Although inactivated vaccines induce strong immunity (which depends upon VN activity of serum antibody) to first infection, the immunologic control of persistent infection is currently poorly understood; however, analogy with other persistent viruses suggests that CTLs are also likely to play an important role in the control of persistent EAV infections.

Acute-Phase Proteins↗
PubMed1999

Determination of equid herpesvirus 1-specific, CD8+, cytotoxic T lymphocyte precursor frequencies in ponies.

The frequency of antigen-specific, genetically restricted cytotoxic T lymphocyte precursors (CTLp) was measured in peripheral blood mononuclear cells (PBMC) of ponies before and after infection with equid herpesvirus 1 (EHV1). Split-well limiting dilution analysis (LDA) was developed to measure CTLp frequency using EHV1-infected 51Cr-labelled lymphoblasts as targets. Extensive characterisation showed that recombinant human interleukin-2, autologous antigen presenting cells and equine serum containing virus neutralising antibody were necessary for maturation of CTLp into effector CTL in vitro. CTLs were not induced when the equine serum (containing VN antibody) was replaced with either foetal calf serum or foetal equine serum (without VN antibody), or seronegative equine serum. CTLp frequency decreased significantly when CD8+ lymphocytes were depleted from the induction cultures. There was good inter- and intra-assay reproducibility using both fresh and recovered cryopreserved PBMC. Both EHV1 and EHV4 could be used to induce effector CTL which lysed EHV1-infected target cells. CTLp frequencies were measured in 2 groups of ponies: Group 1 consisted of two ponies (approx. 9 years old), which had multiple previous experimental infections with EHV1; Group 2 comprised five young (1-2 years) and two older (7 years) ponies which had presumed natural exposure to EHV1/EHV4 but no previous experimental infections. The results showed that CTLp frequencies were higher in the ponies of Group 1 compared with the others. Moreover, ponies with the higher CTLp frequencies were better protected against re-challenge infection with EHV1, showing reduced or absent clinical and virological signs. Consequently, measurement of EHV1-specific CTLp frequency is a potential in vitro correlate of immunity which may be useful for screening new vaccines in horses before embarking upon challenge protection studies to confirm efficacy.

Animals↗
PubMed1999

Equid herpesvirus-induced immunosuppression is associated with lymphoid cells and not soluble circulating factors.

A paresis isolate of equid herpesvirus 1 (EHV1, Ab4/8) and a plaque-purified virus derived from it (EHV1, Ab4/13), induced long-term suppression of both mitogenic and antigen-specific lymphocyte proliferations in adult outbred ponies. Peripheral blood mononuclear cells (PBMC) taken from a pony after EHV1 infection suppressed the in vitro function of normal cells but serum did not. This showed that the observed immune suppression was associated with circulating PBMC and/or their products rather than circulating soluble factors such as antigen or immune complexes. The results suggested that productive infection of lymphocytes by EHV1 was unlikely to result in the observed in vitro effects. Moreover, prostaglandin release from monocytes was not likely to have caused the observed suppression, because lymphocyte responsiveness was not restored in the presence of indomethacin.

Animals↗
PubMed1998

Report of the Second Equine Leucocyte Antigen Workshop, Squaw valley, California, July 1995.

The final assignment of antibody clusters for leucocyte antigens and immunoglobulins, as described in detail in Sections 3 and 4, is summarized in Table 4. Together with other mAbs developed outside of ELAW II (Table 9) this pool of reagents represent a powerful array of tools for the study of equine immunity. The Second Equine Leucocyte Antigen Workshop made considerable advances in pursuing the objectives of establishing the specificities of mAbs and achieving consensus on the nomenclature for equine leucocyte and immunoglobulin molecules. Of equal importance, several productive collaborations were fostered among the participating laboratories and observers. Overall, enormous advances have been made in the past decade since mAbs specific for equine leucocyte antigens and immunoglobulins were first reported. There remains enormous scope and need for further studies of equine leucocyte antigens and immunoglobulins, both for the purposes of comparative immunology and for the good of the horse. In the future novel techniques will be required to develop reagents for specific target antigens such as the orthologues of the CD25 or CD45 isoforms. In studies of equine immunoglobulins the functional role of the IgG isotypes must be better established, reagents for IgE must be developed, and cloning of the immunoglobulin heavy chain genes will be essential if the complexities of the IgG sub-isotypes are to be elucidated. The tasks still facing the currently small group of equine immunologists throughout the world remain formidable, and will only be tackled successfully in a spirit of collaboration.

Animals↗
PubMed1998

Screening of horse polyclonal antibodies with a random peptide library displayed on phage: identification of ligands used as antigens in an ELISA test to detect the presence of antibodies to equine arteritis virus.

A random hexapeptide fusion-phage library was screened to isolate phages that bind to antibodies present in horse sera positive for equine arteritis virus (EAV). Analysis of the peptide sequences displayed by isolated phages identified seven groups. 25% of the isolated phages used as antigens in an ELISA test were specifically recognised by a pool of sera which was positive for EAV in virus neutralisation test (VN). Five of these, when used as antigen in ELISA, detected greater than 50% of sera (n = 30) containing antibodies to EAV as detected by VN. When these five phages were pooled together and used as antigen in ELISA, the detection was improved. The sensitivity and specificity of the ELISA were 99 and 71%, respectively, compared with the EAV neutralisation test (n = 200). This study has shown the potential that phage display libraries have for identifying peptide sequences which could be used as antigen in diagnostic ELISAs.

Amino Acid Sequence↗
PubMed1996

Residence and recruitment of leucocytes to the equine lung after EHV-1 infection.

This study characterised bronchoalveolar leucocytes collected by lavage from five susceptible and two immune ponies before and after nebulised aerosol infection with equid herpesvirus-1 (EHV-1). Leucocyte counts and lymphocyte phenotypic analyses were performed using either differential staining or an indirect immunofluorescence assay with monoclonal antibodies specific for equine (Eq) CD4, CD5, CD8 and B lymphocytes. After EHV-1 infection, significant changes developed: a transient neutrophilia occurred on Day 2, coincident with a reduction in macrophage numbers and an EqCD5+, EqCD4+ and EqCD8+ lymphopaenia. On Day 21, a significant rise in EqCD8+ lymphocytes with an associated fall in the EqCD4+:EqCD4+ ratio and fluctuations in B lymphocyte phenotypes were observed. This study shows that following EHV-1 infection, the bronchoalveolar compartment is subject to dynamic leucocyte migration which may have an important role in immunopathogenesis and recovery from disease.

Animals↗
PubMed1994

Duration of protective efficacy of equine influenza immunostimulating complex/tetanus vaccines.

Seven previously untreated five-month-old New Forest ponies received two doses of equine influenza immunostimulating complex vaccines, one with and one without an immunopurified tetanus toxoid component, given by deep intramuscular injection six weeks apart, followed by a booster dose without tetanus toxoid five months later. Fifteen months after the third dose of vaccine, the ponies were challenged by exposure to an aerosol of influenza A/Equine 2/Sussex/89 (H3N8), a virus isolated from a recent outbreak of influenza A/equine 2 in Britain. The challenge produced severe clinical signs of influenza (pyrexia and coughing) in five unvaccinated control ponies. Four of the vaccinated ponies were completely protected against clinical disease, and two of these were also protected against infection as demonstrated by their lack of an antibody response after challenge. No coughing was recorded among the vaccinated ponies, and only three of the seven vaccinated ponies experienced a transient mild pyrexia. The mean duration and severity of the pyrexia among the vaccinated ponies was significantly less (P < 0.01) than among the controls, and the excretion of virus was almost eliminated, thus demonstrating the protective efficacy of the vaccines 15 months after vaccination. Monitoring of tetanus antitoxin antibodies showed that protective levels (> or = 0.01/iu/ml) were maintained for at least 20 months after vaccination.

Animals↗
PubMed1994

Antigenicity and immunogenicity of experimental equine influenza ISCOM vaccines.

A comparison of the antigenicity and immunogenicity of ISCOM vaccines prepared from equine influenza viruses H3N8 and H7N7 was made with inactivated whole-virus vaccines containing equivalent amounts of virus haemagglutinin. ISCOMs stimulated superior antibody responses in terms of both amount and duration. As with conventional whole-virus vaccines, the levels of antibody to virus haemagglutinin induced by ISCOMs correlated with protection.

Animals↗
PubMed1994

Antigenicity and immunogenicity of equine influenza vaccines containing a Carbomer adjuvant.

Equine influenza vaccines containing inactivated whole virus and Carbomer adjuvant stimulated higher levels and longer lasting antibody to haemagglutinin in ponies than vaccines of equivalent antigenic content containing aluminium phosphate adjuvants. Five months after the third dose of vaccine containing Carbomer adjuvant, ponies were protected against clinical disease induced by an aerosol of virulent influenza virus (A/equine/Newmarket/79, H3N8). In contrast ponies which received vaccine containing aluminium phosphate adjuvant were susceptible to infection and disease. There was an inverse correlation between prechallenge levels of antibody detected by single radial haemolysis (SRH) and duration of virus excretion, pyrexia and coughing. All ponies with antibody levels equivalent to SRH zones of > or = 154 mm2 were protected against infection and all those with levels > or = 85 mm2 were protected from disease.

Acrylic Resins↗
PubMed1994

Distribution of equid herpesvirus-1 (EHV-1) in the respiratory tract of ponies: implications for vaccination strategies.

Twelve adult ponies and 2 conventional foals were exposed to 10(6.6) TCID50 of Equid herpesvirus-1 (EHV-1), strain Ab4 and samples of respiratory tract tissues were recovered. Infectious virus in tissue homogenates was detected using susceptible cell monolayers and expression of viral antigens was monitored using indirect immunoperoxidase histochemistry of paraffin sections. The results illustrated the rapid dissemination of EHV-1 throughout the respiratory tract, with early replication in the lungs one day after exposure. Endothelial cell infection was prominent in all areas of the nasopharynx by Day 4 emphasising the role of endotheliotropism and viraemia in dissemination of this virus to sites of secondary replication. Clinical disease in the adult ponies was mild.

Animals↗
PubMed1994

Distribution of equid herpesvirus-1 (EHV-1) in respiratory tract associated lymphoid tissue: implications for cellular immunity.

Twelve adult ponies and 2 conventional foals were exposed intranasally to EHV-1, strain Ab4 (TCID50 10(-6.6) and samples of respiratory tract associated lymphoid tissues were recovered between 12 h and 13 days after infection. Infectious virus was detected in tissue homogenates using susceptible cell monolayers and expression of viral antigens was monitored using indirect immunoperoxidase histochemistry on paraffin sections. The results showed both infectious EHV-1 and viral antigens in respiratory tract associated lymph nodes 12 h after exposure. Infected leucocytes were identified morphologically as lymphocytes, monocytes, macrophages and plasma cells. The rapid intracellular localisation of EHV-1 in lymph nodes implies that cell mediated immunity is an important aspect of the equine response to this virus.

Animals↗
PubMed1994

Susceptibility of ponies to infection with Streptococcus pneumoniae (capsular type 3)

Welsh Mountain ponies were inoculated with an isolate of Streptococcus pneumoniae, SPE 1618 (capsular type 3) recovered from the equine respiratory tract: 10 ml of a suspension of 10(8) or 10(9) cfu/ml were instilled intratracheally. Fever was observed after either dose but the greater concentration also produced coughing, ocular and nasal discharge, depression and enlargement of submandibular lymph nodes. Cytological evidence of infection was also observed in tracheal washings during the first week after inoculation and corresponded with isolation of S. pneumoniae from the washes. Morbid anatomical and histopathological examinations of selected animals revealed focal pneumonia affecting the ventral lung, especially the cardiac area and accessory lobe, with a propensity to affect the right lung. S. pneumoniae was isolated directly in pure culture from these lesions or was demonstrable by immunostaining of macrophages bearing specific capsular type 3 antigen. By 10 days after inoculation, the ponies were healthy and had developed antibodies to S. pneumoniae. S. pneumoniae was therefore a primary pathogen in the horse under the conditions of the challenge.

Animals↗
PubMed1993

The outbreak of equine influenza (H3N8) in the United Kingdom in 1989: diagnostic use of an antigen capture ELISA.

In July 1989 influenza A/equine-2 (H3N8) was isolated from a nasopharyngeal swab taken from a non-thoroughbred horse exhibiting acute clinical respiratory disease. This was the first isolation of equine influenza virus in the United Kingdom since 1981. Subsequent investigations of acute respiratory disease in horses indicated that the infection was dispersed throughout the UK. However, unlike the previous epidemic of 1979, the first horses from which the virus was isolated had been vaccinated. This outbreak of influenza provided an opportunity to evaluate an antigen capture ELISA, directed against the influenza virus nucleoprotein, as a rapid method for detecting virus in the nasopharyngeal secretions of naturally infected horses.

Animals↗
PubMed1993

Modulation of the serological response of specific pathogen-free (EHV-free) foals to EHV-1 by previous infection with EHV-4 or a TK-deletion mutant of EHV-1.

EHV-1 was inoculated into specific pathogen-free (SPF) foals in order to study uncomplicated primary responses. Infection resulted in a strong serological response recognizing EHV-1-specific antigens; this contrasts with a previous publication where a weak response was recorded in SPF animals. Antibodies to EHV-1 were readily detected by four techniques (virus neutralization, complement fixation, Western blots and immune precipitation), yet there was comparatively little cross-reaction to EHV-4 target antigen. Re-inoculation with the same virus strain stimulated antibodies to EHV-1 but no additional antigens were recognized and antibodies cross-reacting with EHV-4 antigens were not enhanced. Having characterized the uncomplicated primary response to EHV-1 in SPF foals, further animals were exposed to either EHV-4 or a thymidine kinase-deficient mutant of EHV-1 prior to challenge with w/t EHV-1 to investigate how these infections might modulate the immune responses to EHV-1 or 4. Primary inoculation with EHV-4 or with a thymidine kinase-deficient mutant of EHV-1 produced productive infections as evidenced by virus shedding and pyrexia. In both these cases, however, in contrast to that with w/t EHV-1, the serological response was very weak. Re-infection of foals primed with either EHV-4 or TK-deficient EHV-1 with w/t EHV-1 resulted in a strong response to EHV-1 antigens detected by all four methods. In addition, in the foals given a primary inoculation with EHV-4, superinfection with EHV-1 resulted in a strong cross-reactive response to EHV-4 target antigens. The relevance of these observations to the interpretation of previously reported serological responses to EHVs in SPF and naturally reared animals is discussed.

Animals↗