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A P Mockett

Publications and source records attributed to A P Mockett.

At least 19 recordsLinked to original sources
PubMed1999

Avian encephalomyelitis virus is a picornavirus and is most closely related to hepatitis A virus.

The complete RNA genome of avian encephalomyelitis virus (AEV) has been molecularly cloned and sequenced. This revealed AEV to be a member of the Picornaviridae and consequently it is the first avian picornavirus for which the genome has been sequenced. Excluding the poly(A) tail the genome comprises 7032 nucleotides, which is shorter than that of any mammalian picornavirus sequenced to date. An open reading frame commencing at nucleotide 495 and terminating at position 6896 (6402 nucleotides) potentially encodes a polyprotein of 2134 amino acids. The polyprotein sequence has 39% overall amino acid identity with hepatitis A virus (HAV; genus Hepatovirus), compared to 19 to 21% for viruses from the other five picornavirus genera. Eleven cleavage products were predicted. The highest identity (49%) with HAV was in the P1 region, encoding the capsid proteins. The 5' and 3' untranslated regions (UTRs) comprise 494 and 136 nucleotides, respectively. The 5' UTR is the shortest of any picornavirus sequenced to date and, unlike HAV, it does not contain a long polypyrimidine tract.

Amino Acid Sequence↗
PubMed1993

Avian herpesvirus as a live viral vector for the expression of heterologous antigens.

Control of Marek's disease in the poultry industry has been successfully achieved for several decades by large-scale vaccination of day-old chickens with live herpesvirus of turkeys (HVT) strains. Several features of this virus including lack of pathogenicity and long-term immune protection due to a persistent viraemic infection made us decide to use HVT as a live viral vector for the expression of foreign antigens. Potential sites for the integration of foreign DNA in the unique short region of the HVT genome were identified by the insertion of a beta-galactosidase expression cassette. Vaccination trials with recombinant virus strains indicated that the marker gene was expressed and stably maintained during animal passage. Based on an insertion site mapping in one of the open reading frames of the unique short region, a general recombination vector was designed for the integration of foreign genes into HVT. Recombinant virus-directed expression of individual antigens from Newcastle disease virus was driven by a strong promoter element derived from the lung terminal repeat sequence of Rous sarcoma virus.

Amino Acid Sequence↗
PubMed1991

Fowlpox virus: pathogenicity and vaccination of day-old chickens via the aerosol route.

Day-old chickens were given a single fowlpox virus vaccination (strain HP201) either via the aerosol or wing-web route. Both methods induced protective immunity against a wing-web or intravenous challenge with virulent fowlpox virus at 47 days old, although high titred virus preparations were required for successful aerosol vaccination. However, no clinical signs of infection were observed as a result of aerosol vaccination even if invasive strains of Escherichia coli were administered simultaneously. The use of aerosol fowlpox virus vaccination of day-old chicks has been shown to be a possible means of mass vaccination and could be applied to the use of fowlpox virus in recombinant vaccines.

Aerosols↗
PubMed1991

Molecular cloning and sequence analysis of the genome of chicken anaemia agent.

The replicative form (RF) DNA of chicken anaemia agent (CAA) was isolated and cloned into bacterial plasmids. After religation of the cloned CAA DNA and transfection into MDCC-MSB1 cells, the DNA could induce c.p.e. characteristic of that caused by CAA, and an antigen was produced which gave positive immunofluorescence when detected with an anti-CAA serum. Sanger sequencing of the 2298 bp genome revealed several open reading frames (ORFs); the major ORF encoded a polypeptide of 51.8K. In SDS-PAGE of CAA viral particles a 50K protein has been reported as the only detectable viral protein. The genomic region downstream of the major ORF had several predicted GC-rich inverted repeats, a poly(A) signal and four copies of an 18 bp repeat element. Database searches did not reveal any sequence with homology to the viral genomic DNA, nor to the amino acid sequence of any of the ORFs, apart from the N-terminal 40 amino acids of the major ORF which showed a limited similarity to the structure of protamines.

Amino Acid Sequence↗
PubMed1990

Antibody response to experimental Salmonella typhimurium infection in chickens measured by ELISA.

An indirect ELISA has been developed to detect Salmonella typhimurium antibodies in chicken sera, using whole bacterial cell protein, flagellar protein or lipopolysaccharide as antigens. In experimental infections high concentrations of S typhimurium-specific IgG persisted after the faecal excretion of S typhimurium had ceased, whereas the specific IgM response was transitory. Some uninfected chickens placed in contact with experimentally infected birds developed high IgG titres in the absence of detectable faecal excretion. Other S typhimurium strains, which varied in their invasive abilities, also induced high titres of IgG. The ELISA allowed chickens infected experimentally with S typhimurium to be differentiated from chickens infected with 10 other serotypes, including S enteritidis. The use of whole blood in place of serum in the ELISA reduced the titres slightly. The storage of serum dried on to filter paper strips for four weeks produced little change in ELISA antibody titre, and the treatment of such strips with phenol or chloroform vapour had little or no effect on the antibody titre.

Animals↗
PubMed1990

A method for the rapid purification of serum IgM for the diagnosis of recent viral infections of chickens.

The rapid purification of chicken IgM from serum was achieved by affinity chromatography. IgM immunoadsorbent gels were prepared using monoclonal antibodies specific to chicken IgM. Five different eluting agents were compared for the dissociation of the adsorbed IgM; the most convenient for routine purposes was 2 M NaCl, Tris-HCl, EDTA (NTE), as this enabled direct assay of eluents by ELISA without requiring the intermediate step of dialysis, which the other eluting agents did. Eluents prepared from sera obtained from infectious bronchitis virus (IBV) and infectious laryngotracheitis (ILTV)-infected chickens, together with samples of the same serum fractionated by gel chromatography, were tested by ELISA for virus-specific antibodies and to confirm the identity of the antibody class. In the case of both IBV and ILTV, similar results were obtained using immunoaffinity and gel chromatography. IBV-specific IgM, as determined by both methods in the ELISA, reached its highest concentration at the 8th day after inoculation and was virtually absent by the 24th day, whilst the highest concentration of ILT-specific IgM was detected at 6 days and no or little IgM was present at 16 days after inoculation. Purification of serum IgM by affinity chromatography followed by ELISA was considered suitable for routine serological diagnosis of IB and ILT, since the time required to complete the assay (3 hours) was considerably less than that for gel chromatography and many samples could be assayed simultaneously.

Animals↗
PubMed1988

Amino acids within hypervariable region 1 of avian coronavirus IBV (Massachusetts serotype) spike glycoprotein are associated with neutralization epitopes.

The spike glycoprotein (S) gene of IBV codes for a precursor protein which is cleaved into the N-terminal S1 and C-terminal S2 glycopolypeptides. The S1 glycopolypeptide, which induces neutralizing antibody, comprises approximately 520 amino acid residues. We have determined the nucleotide sequence of S1 of seven strains of the Massachusetts (Mass) serotype and the first 337 bases of two additional Mass strains. Despite the fact that the strains had been isolated over three decades in Europe and the U.S.A. there was only 4% base and 6% amino acid variation within the group. Nearly one third of the 32 amino acid differences in S1 were in two hypervariable regions (HVRs 1 and 2) comprising residues 38-51 and 99-115, identified by Niesters et al. (1986), showing that HVRs 1 and 2 are a feature of the Mass serotype. Amino acid variation within HVRs 1 and 2 was 29% and 40% respectively. Five vaccine strains could be distinguished from each other by sequencing of the first 337 nucleotides. Variants of M41 which resisted neutralization by two monoclonal antibodies (A13 and A38) had the same, single base change at position 134, resulting in substitution of proline residue 45 by histidine. This indicates that residues within HVR 1 are associated with epitopes which induce neutralizing antibody.

Amino Acid Sequence↗
PubMed1987

Expression of the infectious bronchitis virus spike protein by recombinant vaccinia virus and induction of neutralizing antibodies in vaccinated mice.

A cDNA clone of the infectious bronchitis virus (IBV) spike protein gene has been recombined into vaccinia virus. Cells infected with the recombinant virus synthesized IBV spike antigen which was recognized by antibody raised against purified spike protein. Immunofluorescence showed that the IBV spike antigen was transported to the infected cell surface membrane and immunoprecipitation showed the presence of the glycosylated 180K mol. wt. polypeptide precursor of the two spike subunits S1 and S2 that comigrated with this antigen from IBV-infected cells. Vaccinated mice produced antibody that recognized the IBV spike antigen by ELISA and which neutralized IBV infectivity as shown by ciliostasis tests on tracheal organ cultures.

Antibody Formation↗
PubMed1986

Biochemical characterization of chicken secretory component.

Human dimeric IgA was injected i.v. into chickens whose bile was then collected. The human IgA which had been transported across the hepatocytes of the chicken was subsequently purified from the bile and shown to be associated with a protein of 80 kDa and pI 4.6, which reacted with rabbit antisera to chicken bile proteins but not with an antiserum to human alpha chain. The chicken bile protein thus has functional and biochemical properties similar to those of mammalian secretory component.

Animals↗
PubMed1986

Immune responses to eimeria: quantification of antibody isotypes to Eimeria tenella in chicken serum and bile by means of the ELISA.

The enzyme-linked immunosorbent assay (ELISA) has been used to study the serum IgM and IgG response and the bile IgA and IgM response of chickens to a primary, secondary and tertiary inoculation of Eimeria tenella. In the serum there was a rapid and transient IgM response to the primary infection; second and third inoculations of oocysts had comparatively little effect on the concentrations of this antibody isotype. This differed from the specific IgG response which was later and of similar magnitude after each inoculum. In the bile, specific IgA reached its highest concentrations 9-10 days after the primary inoculation and then declined rapidly. The second and third inoculations each induced low concentrations of this isotype. The specific biliary IgM response to the primary inoculation was similar in profile to that of the IgA, but IgM was detected only on day 14-15 after the second and not at all after the third inoculation. The findings are discussed and are compared with the results obtained using other host species, principally the rat, and with other gut organisms. The relevance of the antibody response in resistance to eimerian infections is also discussed.

Animals↗
PubMed1985

Envelope proteins of avian infectious bronchitis virus: purification and biological properties.

Immunoadsorbents, made with monoclonal antibodies, were used to purify the spike and membrane proteins of infectious bronchitis virus (IBV). The purified proteins were inoculated into rabbits to produce antisera. The rabbit anti-spike sera neutralized the infectivity of the virus whereas the anti-membrane sera did not. IBV-infected chickens produced antibodies to both the spike and membrane proteins. Both these antibodies were at their highest concentration about 9-11 days after inoculation, whereas neutralizing antibodies were present only at very low concentrations at that time. Neutralizing antibodies were at their highest concentration 21 days after inoculation. A second inoculation of virus at 42 days induced an anamnestic antibody response to the spike and membrane proteins and also for the neutralizing antibodies. The neutralizing, anti-spike and anti-membrane antibodies all reached highest concentrations 7-11 days after this inoculation. The advantages of purifying viral proteins using affinity chromatography with monoclonal antibodies are discussed.

Animals↗
PubMed1984

Monoclonal antibodies to the S1 spike and membrane proteins of avian infectious bronchitis coronavirus strain Massachusetts M41.

We have established four murine hybridoma cell lines which secrete specific antibody to avian infectious bronchitis virus (IBV) strain Massachusetts M41. Two monoclonal antibodies reacted with the spike protein and two reacted with the membrane protein. The specificity of the monoclonal antibodies for the external structural proteins was detected by immunoprecipitations using radiolabelled virus. The reactions of the monoclonal antibodies showed that one (S1) of the two glycopolypeptides associated with the spike protein has a strain-specific region involved in neutralization and haemagglutination, and the membrane protein has antigenic determinants which are present on the three strains of IBV tested (M41, Beaudette and D41).

Antibodies, Monoclonal↗
PubMed1983

The use of an enzyme-linked immunosorbent assay to detect IgG antibodies to serotype-specific and group-specific antigens of fowl adenovirus serotypes 2, 3 and 4.

A sensitive enzyme-linked immunosorbent assay (ELISA) has been developed which can detect serotype and group-specific antibodies (IgG) to fowl adenovirus serotypes 2, 3 and 4. The chickens produced principally type-specific antibodies after a single oral inoculation of virus which enabled that strain to be identified by the ELISA. However, inoculation of an heterologous serotype, although inducing strain-specific antibodies to itself, also induced high levels of antibody to the group-specific antigens. This masked the serotype-specific antibodies in the ELISA to the first serotype. The ELISA, which has similar sensitivity to the serum neutralisation test, could be used as a rapid, easily performed test to identify fowl adenovirus-specific antibodies.

Adenoviridae↗
PubMed1983

Antibodies to coccidia: detection by the enzyme-linked immunosorbent assay (ELISA).

The ELISA test was used for the detection of antibodies to coccidia in the serum and/or egg yolk of chickens infected with Eimeria acervulina, E. maxima or E. tenella and in the serum of rats infected with E. nieschulzi. Antigens prepared from different developmental stages of the parasite were tested and the cross-reaction between different species of Eimeria were examined. The variability in cross-reactivity of different species and the advantages and possible applications of the test are discussed.

Animals↗
PubMedDate not supplied

IgM responses in chicken serum to live and inactivated infectious bronchitis virus vaccines.

Intramuscular (i.m.) administration of infectious bronchitis virus (IBV) oil-emulsion vaccine (OEV) to IBV-primed or unprimed chickens resulted in the production of zero or minimal concentrations of IBV-specific IgM in the serum, as measured by enzyme-linked immunosorbent assay of gel chromatography fractions. Live-attenuated infectious bronchitis (IB) vaccine given i.m. or by eyedrop stimulated the production of IBV-specific IgM in similar amounts following inoculation by both routes. These levels were comparable to those found in earlier studies following intranasal inoculation with a virulent strain of IBV and confirm that the detection of IBV-specific IgM is a valuable aid to the diagnosis of recent infection. As expected, administration of live-attenuated IB vaccines i.m. or by eyedrop protected the respiratory tract against challenge with virulent virus 24 days later; however, OEV given i.m. did not.

Animals↗