Biomedical subjects
M Aymard
Publications and source records attributed to M Aymard.
Contribution to the study of the parainfluenza antigens.
From epidemiological data it appeared that an epidemic of parainfluenza type 1 occurred in Lyons in October-November 1967. Thereafter a few sporadic cases were detected. Then in 1970 strains were isolated which showed a change in the hemagglutinin antigen. The persistence of antibodies in the population is shorter than that of the other parainfluenza viruses (i. e. parainfluenza 3); it is not known if reinfections with parainfluenza 1 occur since there are cross-reactions by CF test between parainfluenza 1 and parainfluenza 3 which are known to cause reinfections. The study of antigens of parainfluenza strains was made by HI, Nt, CF and immunodiffusion tests with various antisera prepared in various animal species. This study showed antigenic differences and similarities between parainfluenza 1 and parainfluenza 3.
[cRNA probes (riboprobes) synthesized in vitro for detecting enterovirus by molecular hybridization].
Radioactively labelled RNA transcripts made in vitro of various fragments from cDNA clones of poliovirus type 1 and of hepatitis A virus under the control of bacteriophage T7 or SP6 promoters have been evaluated for diagnostic purposes. The RNA transcripts were 2 orders of magnitude more sensitive as hybridization probes than corresponding cDNA preparations labelled by the nick translation procedure. A combination of hybridization analysis and sequence comparison showed that some regions of the genome of a number of enteroviruses are highly conserved, while others show very little homology; the general order of conservation is: 5'-non-coding greater than 3'-terminal greater than central (2C) greater than VP3 greater than VP1. The 350 bases of the poliovirus VP1 region were highly specific for that virus, while the 450-bases of the 5'NC region showed extensive cross-reaction with other enteroviruses. However, these probes did not hybridize with HAV, which was detected only by HAV-specific riboprobes. The transcripts have been successfully applied as hybridization probes in diagnostic tests on supernatants of infected cell culture lysates and in clinical samples, mainly in stool extracts.
Use of magnetic beads versus guanidium thiocyanate-phenol-chloroform RNA extraction followed by polymerase chain reaction for the rapid, sensitive detection of enterovirus RNA.
The current study compares the sensitivity of RNA extraction using magnetic beads versus that of a standard extraction method. Streptavadin-coated magnetic beads were labelled with a biotinylated, enterovirus-specific oligonucleotide. RNA was extracted using labelled beads or guanidium thiocyanate-phenol-chloroform from 1, 0.1 and 0.01 TCID50/100 microliters of stock coxsackievirus types A9 and B3, echovirus type 11, enterovirus type 70 and poliovirus type 1. Each strain was tested three times. RNA extraction using magnetic beads was > 50% faster than the standard method. The RNA was amplified using RT-PCR, and the products were detected using agarose gel electrophoresis; 6/15 and 7/15 samples at an initial concentration of 0.01 TCID50/100 microliters were detected using magnetic beads or standard extraction, respectively. Negative-stain electron microscopy was used to determine that 0.01 TCID50/100 microliters of coxsackievirus B3 contained approximately 3 genomes. Thus, use of magnetic beads labelled with an enterovirus-specific oligonucleotide was less toxic, more rapid and as sensitive as the current standard RNA extraction method.
Comparison of procedures for the detection of enteroviruses in murine heart samples by in situ polymerase chain reaction.
A protocol for the in situ polymerase chain reaction (IS-PCR) detection of viral nucleic acid in the heart tissue of four-to-five-week-old CD1 mice infected with coxsackievirus B3 (CBV3) Nancy strain is described. To compare the effects of formalin concentration on the IS-PCR process, two different concentrations (10 and 37%) were employed. Using 37% formalin, 25 PCR cycles were sufficient and a permeabilization step could be omitted. However, postfixation of tissues with 4% paraformaldehyde and 100% ethanol after the deparaffinization, reverse transcriptase and amplification steps was required in order to minimize artefacts. When the tissues were fixed in 10% formalin, postfixation with 4% paraformaldehyde was not required, but a permeabilization step had to be employed and 40 cycles of PCR amplification were needed. To detect the PCR product in the 10% formalin-fixed samples, incubation with 0.3 U/ml of an anti-digoxigenin antibody conjugated to alkaline phosphatase was performed for 90 min. When 37% formalin-fixed samples were used, the concentration of the antibody conjugate had to be increased to 3 U/ml and the exposure time was decreased to 30 min. Enterovirus (EV) nucleic acid was detected in the cytoplasm of myocytes. Thus, IS-PCR was successful in localizing EV nucleic acid in the cytoplasm of myocytes in mice infected with a cardiotropic strain of CBV3. Using this technique, 10% formalin-fixed tissues gave better results than 37% formalin-fixed tissues.
[Comparative value of temperature marker and neutralization test for subtyping of herpes simplex viruses (author's transl)].
Two methods for the differentiation of herpes simplex virus (HSV)--rct and neutralization by specific antisera--have been applied to 8 prototype strains and 105 strains isolated from various clinical cases. The evaluation of the II/I index allows to choose the most specific rabbit antisera. The rct 40 degrees 3 C marker test gives comparable results for HSV grown in RK13 and human fibroblasts as well; it is in agreement with the immunological test in 48% of the cases. The delta tc values used to express the immunological typing allow to differentiate 76% of the strains (types 1 and 2). The others (24%) are equally neutralized by both specific antisera; they represent "intermediate" strains. The infection of man and animal by HSV (type 1, 2 or "intermediate") also produces heterospecific antibodies in variable amount. From clinical and immunological data rises the hypothesis that "intermediate" strains derive from HSV1 strains by antigenic variation during a persistent infection.
[Gastroenteritis caused by rotavirus. An epidemiologic survey in Lyons. Diagnostic value of the ELISA method].
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[Necrotizing enterocolitis and cytomegalovirus infection (author's transl)].
During an 18-month period, 11 cases of necrotizing enterocolitis (NEC) were diagnosed in infants treated in Neonate Department. This represents 1% of total admissions. Six cases occurred during hospitalization. Clinically, the disease took three different forms: surgical form, medical form responding well to exclusive parenteral feeding and isolated haemorrhagic form, which deserves discussion. Seven of these infants had cytomegalovirus (CMV) infection. NEC accounted for 22% of clinical symptoms in CMV infection. These data suggest that the cytomegalovirus may act as aggravating factor in NEC.
[Evaluation methodology of viral vaccines].
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