Joko Imai

Primary infection of Japanese infants with adult T-cell leukaemia-associated retrovirus (ATLV): Evidence for viral transmission from mothers to children

Primary infection with adult T-cell leukemia virus (ATLV) was investigated by follow-up studies on 16 ATLV-seropositive mothers and their breastfed infants in an ATLV-endemic area of Japan. Maternal antibody to ATLV decreased in all the infants, and was detectable in only three of 12 infants tested 6 months after birth. Reappearance of the antibody 9-18 months after birth was observed in only four of the 16 infants. The ATLV-bearing cells in peripheral blood were detected in all 16 mothers after delivery. None of the 16 infants showed ATLV-bearing cells in peripheral or cord blood sampled at birth, or 1, 3 or 6 months after birth. However, virus-bearing cells in the blood became detectable 9-18 months after birth in 13 of the 16 infants. Maternal antibody and virus-bearing cells were never detected in a control group of seven infants of ATLV-seronegative mothers. These findings provide evidence for the high incidence of primary ATLV infection during early infancy among infants born to ATLV-seropositive mothers and suggest maternal viral transmission. Furthermore, samples of breast milk from all 12 seropositive mothers examined contained cell-associated ATLV capable of being transmitted to peripheral leucocytes of neonates. This finding suggests that one of the possible maternal transmission routes of ATLV is via breast milk.

Improvement of Gelatin Particle Agglutination Test for Detection of Anti-HTLV-I Antibody

Partial modifications of antigen components were made to improve the gelatin particle agglutination (PA) test for the detection of antibodies against human T cell leukemia virus type‐I. Envelope glycoproteins prepared by lentil lectin affinity chromatography were further added to the purified viral antigens to be coated on the gelatin particles. Comparative studies with a conventional PA test kit (Serodia ATLA) and indirect immunofluorescence assay showed that the specificity and sensitivity of the new PA test were increased and that abnormal agglutination such as the prozone phenomenon was abolished by this improvement.

Performance Certification of Gelatin Particle Agglutination Assay for Anti-HTLV-1 Antibody: Inconclusive Positive Results

In order to test the performance of particle agglutination assay (PA), 800 preselected PA‐positive sera at 8 blood centers in the Kyushu area were tested in various assays. Most blood centers should improve their PA technique, since a third of the samples were PA‐negative in our hands. A third of our PA high‐titer sera were negative in indirect immunofluorescence and enzyme imiminoassay, the results of which were consistent with each other. Western blots did not detect every positive serum. PA inhibition positivity was not consistent with PA titer. Most IgM antibody‐positive sera also contained IgG antibody. PA should be used in combination with other methods before notifying the results to positive testees.

Rapid, sensitive, specific, and quantitative detection of human T-cell leukemia virus type 1 sequence in peripheral blood mononuclear cells by an improved polymerase chain reaction method with nested primers.

Improving on the nested double polymerase chain reaction (PCR) described previously, we have developed a new two-step PCR (TS-PCR) method for detecting more specifically the human T-cell leukemia virus type 1 (HTLV-1) proviral sequences in peripheral blood mononuclear cells (PBMC). In our TS-PCR method, the point of modification is to use optimal concentrations of primers in the first amplification step in the range of 0.01–0.025 µM. This increases sensitivity and specificity enough to detect from 1 to 105 copies of template DNA without radioisotopes. This method is rapid because of completion in 1 day and is also applicable for quantitative detection of clinical specimens. The data show that the quantitative detection of HTLV-1 proviral sequences by this method correlates with the anti-HTLV-1 antibody titers from serologic analysis of seropositive healthy carriers. Moreover, the TS-PCR method using each specific primer was also attempted for successful detection of other viral genomes; therefore, the principle of this method is widely suitable for routine detection of genomes in the basic and clinical microbiological fields.

Human sera from varicella-zoster virus (VZV) infections cross-react with human T cell leukaemia virus type 1 (HTLV-1): common epitopes in VZV gene 22 protein and HTLV-1 p19 gag protein

Twenty-nine of 100 sera from patients recently infected with varicella-zoster virus (VZV) were found to cross-react with human T cell leukaemia virus type 1 (HTLV-1) antigen in the particle agglutination (PA) assay using HTLV-1 antigen-coated gelatin particles. Anti-VZV IgM antibodies were shown to be responsible for this cross-reactivity. Western blot analysis revealed that PA-positive anti-VZV sera reacted with the HTLV-1 gag p19 protein in HTLV-1-infected cells and recombinant p19 protein produced in Escherichia coli. By using a truncated p19, the cross-reactive region was located to the C-terminal 17 amino acids of p19. One oligopeptide derived from the C terminus, PQIPPPYVEPT (amino acids 115 to 125), was capable of inhibiting PA, suggesting that this peptide carries the cross-reactive epitope. A homologous sequence was found in the VZV gene 22 protein by database analysis, and the oligopeptide TNIPPPLALLR (amino acids 1330 to 1340) had the ability to inhibit PA. These findings suggest that some IgM antibodies against the VZV gene 22 protein produced in the early phase of VZV infection are cross-reactive with the HTLV-1 gag p19 protein because they recognize an antigenic determinant containing an IPPP tetrapeptide.

Sporadic Carriers of Human T-Lymphotropic Virus Type I in Northern Egypt

Sera from 3,158 individuals living in northern Egypt were tested for the presence of antibodies against human T‐lymphotropic virus type I (HTLV‐I) by the newly developed particle agglutination (PA) test. Ten sera gave a positive reaction in the PA test. Eight of these sera were examined further by Western blotting and all of them gave several bands corresponding to HTLV‐I structural proteins. Two of the 8 sera gave positive results in the indirect immunofluorescence test. The results indicate the presence of HTLV‐I carriers in this area, although at very low incidence (0.063%).

Presence of antibodies to p21X and/or p27rex proteins in sera from human T-cell leukemia virus type I-infected individuals

The human T-cell leukemia virus type I (HTLV-I) pX gene encodes three nonstructural proteins, p40tax, p27rex and p21X. So far, natural antibodies to p27rex and/or p21X have not been found in sera from HTLV-I-infected individuals, although antibodies to p40tax have been found. Recently, the viral transcripts specific for these proteins were detected in fresh peripheral blood mononuclear cells from HTLV-I-infected individuals by the polymerase chain reaction coupled to reverse transcription, showing the in vivo expression of these proteins. We detected antibodies to p21X and p27rex by an enzyme-linked immunosorbent assay (ELISA) system using a recombinantly produced p21X protein as a common antigen, because p21X is identical to the C-terminal portion of p27rex. The sensitivity of the ELISA was determined to be approximately 100 times greater than that of Western blotting. From the analyzed sera of 31 ATL patients, 30 asymptomatic carriers, 18 HAM patients and 100 healthy donors, three specimens from one ATL patient and two carriers were found to be positive for anti-p21X/p27rex antibodies. The specificity of the ELISA reaction was confirmed by the competitive ELISA test with the highly purified recombinant p21X protein. As of result, we first determined the presence of anti-p21X/p27rex antibodies in a small percentage (3.8%) of the sera from HTLV-I-infected individuals. Even sera from the ATL patients, whose fresh PBMCs contained the transcripts for these proteins, were not found to contain these antibodies, suggesting that the immune response to these proteins is low in HTLV-I-infected humans.