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Biomedical subjects

M J Carter

Publications and source records attributed to M J Carter.

At least 55 records · Page 3Linked to original sources
PubMed1994

Acute arthritis of cats associated with feline calicivirus infection.

Twelve specific pathogen-free cats were infected either by intra-articular inoculation or by contact exposure to one of two strains of feline calicivirus (FCV), either F65, a field strain originating from an outbreak of lameness in a group of cats, or a vaccine strain. Following either route of exposure, both strains induced signs typical of FCV infection including oral and nasal ulceration, conjunctivitis and ocular discharge. These signs were of equal severity for both virus strains, but overall, following either route of infection, F65 induced more severe disease than the vaccine strain, with marked pyrexia, lethargy and lameness. Vaccine virus only induced a relatively mild lameness following intra-articular inoculation. Gross pathological and histopathological lesions were seen in some of the joints, but again changes were more severe in the F65-exposed cats. Virus was isolated from both normal and affected joints from both groups of F65-exposed cats, and from a joint from each cat inoculated intra-articularly with vaccine virus. Mild transient lameness was also seen in one of two control cats inoculated intra-articularly, but no pathological changes were seen or virus isolated from joints. A cDNA probe used in RNA dot blot hybridisation experiments was found to be specific and more sensitive than virus isolation in detecting FCV in selected tissues. This may be useful in future studies on the pathogenesis of FCV disease and in studies on viral persistence in FCV carriers.

Acute Disease↗
PubMed1994

Biological activity, binding site and affinity of monoclonal antibodies to the fusion protein of respiratory syncytial virus.

The neutralizing activity and fusion-inhibition activity per unit weight of immunoglobulin were determined for each of a panel of 20 monoclonal antibodies (MAbs) to the fusion (F) protein of respiratory syncytial (RS) virus. Neutralization did not correlate with fusion-inhibiting activity, suggesting that the F protein plays at least two independent, antibody-sensitive roles in viral infection. Antibodies with the highest biological activity against A2, a subgroup A strain of RS virus, neutralized a subgroup B strain (8/60) poorly, suggesting a degree of antigenic variation that may be important in human infection. All but one fusion-inhibiting MAb bound to protein blots and binding was mapped to two areas on overlapping F protein fragments. One MAb with relatively poor fusion-inhibiting activity bound only to fragments C-terminal of amino acid 384, the remainder bound only to fragments containing residues 253 to 289. MAbs directed to the latter site were heterogeneous in neutralizing activity, subgroup specificity and fusion-inhibiting activity. These variations between MAbs could not be accounted for by differences in their binding avidities. We suggest that this binding site is not the complete antibody epitope which probably includes conformation-dependent elements.

Animals↗
PubMed1994

The complete sequence of a human astrovirus.

We have determined the complete genomic sequence of human astrovirus serotype 1 isolated in Newcastle upon Tyne. The genome is 6813 nucleotides long and contains three sequential open reading frames (ORFs). The two closest to the 5' end are linked by a ribosomal frameshifting motif and contain sequence motifs indicative of non-structural virus proteins: serine protease and RNA-dependent RNA polymerase. A nuclear addressing sequence is also located here. The 3' ORF encodes the virion structural polypeptides as a polyprotein precursor. This genomic organization resembles that of the plant virus family Luteoviridae.

Amino Acid Sequence↗
PubMed1994

The human astrovirus RNA-dependent RNA polymerase coding region is expressed by ribosomal frameshifting.

The genomic RNA of human astrovirus serotype 1 (HAst-1) contains three open reading frames (ORFs), 1a, 1b, and 2. ORF 1b is located downstream of, and overlaps, 1a, and it has been suggested on the basis of sequence analysis that expression of ORF 1b is mediated through -1 ribosomal frameshifting. To examine this possibility, a cDNA fragment containing the 1a-1b overlap region was cloned within a reporter gene and placed under the control of the bacteriophage SP6 promoter in a recombinant plasmid. Synthetic transcripts derived from this plasmid, when translated in the rabbit reticulocyte lysate cell-free system, specified the synthesis of polypeptides whose size and antibody reactivity were consistent with an efficient -1 ribosomal frameshift event at the overlap region. The HAst-1 frameshift signal has two essential components, a heptanucleotide slippery sequence, A6C, and a stem-loop structure in the RNA. The presence of this structure was confirmed by complementary and compensatory mutation analysis and by direct structure probing with single- and double-stranded RNA-specific reagents. The HAst-1 frameshift signal, like that present at the overlap of the gag and pro genes of the retrovirus human T-cell lymphotrophic virus type II, does not involve the formation of an RNA pseudoknot.

Animals↗
PubMed1994

Cell culture adaptation of astrovirus involves a deletion.

Astroviruses have been adapted to culture by serial blind passage in primary human embryo cells. All viruses thus adapted possess a 45-nucleotide deletion relative to fecal viruses or isolates made in CaCo-2 cells; this deletion may be responsible for the change in host cell range.

Amino Acid Sequence↗
PubMed1993

Identification and sequence determination of the capsid protein gene of human astrovirus serotype 1.

We present the sequence of an open reading frame (ORF) at the 3' end of human astrovirus serotype 1. Primer extension experiments showed that the RNA expressing this gene is shorter than the complete ORF, and could form a protein of M(r) 85,540. The protein was expressed by recombinant baculovirus and was recognized by anti-virion serum, indicating a structural role. Sequence comparison indicates that astrovirus serotypes 1 and 2 differ markedly in the C-terminal half of the protein but are well conserved towards the N-terminus.

Amino Acid Sequence↗
PubMed1993

Detection of Norwalk virus in the UK by the polymerase chain reaction.

We have developed a polymerase chain reaction for the detection of Norwalk virus using the published sequence of the virus RNA dependent RNA polymerase gene and have used this to clone and sequence this region of a virus from a UK outbreak. We have applied this method to a panel of UK Norwalk-like viruses using both Tet-z and Taq DNA polymerases and found that amplification produces a multiplicity of bands from stool samples. However, in combination with Southern blotting, Taq polymerase amplification detected virus in 13 of a panel of 30 clinical samples known to contain these viruses and also detected astroviruses in a mixed infection. Amplification using Tet-z DNA polymerase was less efficient (6/30) and detected predominantly viruses typed as UK type 2 by solid phase immune electron microscopy.

Amino Acid Sequence↗
PubMed1993

Antibiotic prophylaxis in cardiac operations.

This clinical trial, which was composed of 1,031 adults undergoing cardiac operations, compared the efficacy of a single dose of 1 g of ceftriaxone with a 48-our regimen consisting of flucloxacillin and gentamicin. There was no significant difference (p = 0.89) in the overall incidence of major infections: 30 of 515 patients (5.8%; 95% confidence interval, 5.4% to 6.2%) taking ceftriaxone and 29 of 516 patients (5.6%; 95% confidence interval, 5.2% to 6.0%) taking flucloxacillin and gentamicin. Subgroup analyses, with a lower statistical power, failed to show a significant difference between patients who received ceftriaxone and those who received flucloxacillin/gentamicin: major sternal wound infections arose in 2.7% of the patients taking ceftriaxone versus 1.6% in those on the 48-hour regimen (p = 0.20) and major limb wound infections arose in 4.2% and 5.4%, respectively (p = 0.44). Single-dose prophylaxis was associated with fewer intravenous administrations (864 doses versus 9,570 doses) and cost less (A$17,248 versus A$78,510). Although the regimen that included gentamicin was associated with the greatest biochemical impairment of renal function, the overall toxicity for both groups was low. We conclude that a single dose of ceftriaxone provided cost-efficient prophylaxis for adults undergoing cardiac operations when compared with a 48-hour regimen of gentamicin and flucloxacillin. The general principle revealed by our data is that the short-term administration of an appropriate antibiotic regimen represents optimal prophylaxis for patients undergoing cardiac procedures.

Adolescent↗
PubMed1993

Binding of neutralizing monoclonal antibodies to regions of the fusion protein of respiratory syncytial virus expressed in Escherichia coli.

cDNA containing the entire coding sequence of the respiratory syncytial (RS) virus fusion (F) protein gene (574 amino acids) and two large PstI restriction fragments, encoding amino acids 18 to 212 and 214 to 574, were expressed in Escherichia coli as C-terminal chimeras with beta-galactosidase (beta-gal) in the pEX expression vector system. A further cDNA fragment, overlapping the PstI restriction site and encoding amino acids 190 to 289, was derived by PCR and expressed in a similar manner. Polyclonal rabbit serum raised against RS virus bound to all four chimeric proteins but most strongly to those containing C-terminal sequences. Two monoclonal antibodies (MAbs), 1E3 and RS348, capable of neutralizing the virus and inhibiting the viral fusion function, bound to all chimeras except that derived from the N-terminal PstI fragment, suggesting that their binding sites were located between amino acids 214 and 289. Further analysis of binding to expressed fragments from restriction enzyme digests and PCR amplification demonstrated that both antibodies bound to amino acids 253 to 289. MAb RS348 bound to 12-mer overlapping synthetic peptides containing the sequence 265 to 272 (PITNDQKK) but MAb 1E3 failed to bind to any 12-mer peptide derived from the F protein sequence. Immunization of mice with chimeric proteins containing the whole F protein coding sequence or amino acids 253 to 384, which includes the binding site of the two MAbs identified here, failed to induce antibodies that recognized the native RS virus F protein or could neutralize the virus. This suggests that either the beta-gal partner inhibits the immune response to the protein or that elements missing from the protein expressed in E. coli, perhaps conformational or added post-translation, contribute to the neutralizing antibody epitope.

Amino Acid Sequence↗
PubMed1992

Genomic 3' terminal sequence comparison of three isolates of rabbit haemorrhagic disease virus.

Comparison of sequence data is necessary in older to investigate virus origins, identify features common to virulent strains, and characterize genomic organization within virus families. A virulent caliciviral disease of rabbits recently emerged in China. We have sequenced 1100 bases from the 3' ends of two independent European isolates of this virus, and compared these with previously determined calicivirus sequences. Rabbit caliciviruses were closely related, despite the different countries in which isolation was made. This supports the rapid spread of a new virus across Europe. The capsid protein sequences of these rabbit viruses differ markedly from those determined for feline calicivirus, but a hypothetical 3' open reading frame is relatively well conserved between the caliciviruses of these two different hosts and argues for a functional role.

Amino Acid Sequence↗
PubMed1992

The 3' terminal sequence of a human astrovirus.

We have determined the sequence for 1,000 bases from the 3' terminus of a human astrovirus serotype 1 isolated in Newcastle. This is the first sequence reported for a representative of this virus family. We find one open reading frame which terminates 83 bases from a poly A tail. The 3' non-coding-region has similarities to some picornavirus termini. However the amino acids specified by the coding region have no significant homology to the picornavirus protein 3D, encoded at the 3' terminus of these viruses. Northern blot analysis of intracellular virus-specific RNAs revealed one size of transcript which corresponded to full-length virus RNA. Available data thus indicate that astroviruses may resemble picornaviruses in replication strategy.

Amino Acid Sequence↗
PubMed1992

Identification and sequence determination of the capsid protein gene of feline calicivirus.

We have determined 4380 bases of the sequence from a cDNA clone containing the 3' end of feline calicivirus strain F9. We find four candidate open reading frames of which three are complete and comprise 245, 317 and 2012 nucleotides. The fourth continues toward the 5' end. We have expressed the largest complete open reading frame in E. coli. Sera raised to this antigen react specifically with the capsid protein and its intracellular precursor molecule. N-terminal sequence analysis of purified, mature capsid protein confirms this assignment and has identified the position at which precursor is cleaved.

Amino Acid Sequence↗
PubMed1992

The complete nucleotide sequence of a feline calicivirus.

We have determined the complete sequence of a feline calicivirus. The virus genome is 7690 bases long and contains two large open reading frames. Proteins specified by these have similarity to those encoded in the corresponding regions of a candidate calicivirus rabbit hemorrhagic disease virus, but are distinctly different from those specified by another such virus, hepatitis E virus. A third, small open reading frame at the 3' end of the genome is present in both feline and rabbit viruses but is absent from hepatitis E. These findings suggest that the calicivirus family, which consists of a single genus, may require subdivision.

Amino Acid Sequence↗
PubMed1992

Location of monoclonal antibody binding sites in the capsid protein of feline calicivirus.

We report the localization of three monoclonal antibody (MAb) binding sites in the capsid protein of feline calicivirus. Gene fragments were generated by restriction enzyme digestion or the polymerase chain reaction, and expressed as beta-galactosidase fusion proteins in Escherichia coli. These chimeric molecules were screened using three MAbs. A non-neutralizing MAb recognized a region within 36 amino acids of the C terminus. Two neutralizing MAbs bound to a different region of 37 amino acids in the centre of the protein. Comparative sequence analysis shows this area to be the major variable region of the capsid protein.

Amino Acid Sequence↗