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L Heyndrickx

Publications and source records attributed to L Heyndrickx.

At least 37 records · Page 2Linked to original sources
PubMed1998

Improved detection of HIV-2 proviral DNA in dually seroreactive individuals by PCR.

OBJECTIVE: To improve the detection rate of HIV-2 proviral DNA in primary uncultured peripheral blood mononuclear cells (PBMC) of HIV-2-seroreactive and HIV-1-HIV-2 dually seroreactive individuals. MATERIALS AND METHODS: Two newly designed HIV-2 PCR primer pairs in the long terminal repeat (LTR) gag and gag-pol regions and a previously described env and LTR HIV-2 PCR primer pairs were tested on samples from 66 confirmed HIV-2-seropositive individuals (The Gambia, 40; Côte d'Ivoire, 17; Guinea-Bissau, nine), 209 dually seroreactive individuals (The Gambia, 82; Côte d'Ivoire, 127), 24 genetically characterized isolated HIV-1 strains (group M subtypes A-H and group O), one simian immunodeficiency virus (SIV) strain cpz, 10 HIV-2 isolates (subtype A, B and unidentified), two SIVsm isolates, and 10 seronegative samples. RESULTS: All HIV-2 primers evaluated showed 100% specificity since there was no amplification observed with 24 HIV-1, one SIVcpz and 10 seronegative samples. One single copy of the HIV-2 genome could be detected with all outer primer pairs as well as all inner primer pairs on one PCR round used. Sensitivity of primers (at least one of the four primer pairs was positive) to HIV-2-seropositive samples was 100% (all nine) in Guinea-Bissau, 71% (12/17) in Côte d'Ivoire, 100% (all 20) in Gambian AIDS patients, and 85% (17/20) in Gambian pregnant women. Doubling the PBMC of dually seroreactive individuals from 7.5 x 10(4) to 1.5 x 10(5) in the PCR revealed the presence of both HIV-1 and 2 proviral DNA in 72% (92/127) in Côte d'Ivoire and 72% (59/82) in The Gambia. By doubling the number of PBMC, HIV-2 detection in dually seroreactive individuals by PCR was increased from 65 to 77% in Côte d'Ivoire and from 67 to 83% in The Gambia. CONCLUSIONS: The use of 1.5 x 10(5) primary uncultured PBMC and the newly designed HIV-2 primer pairs allowed us to document the highest percentage (72%) ever reported of HIV-1-HIV-2 dual infections amongst HIV-1-HIV-2 dually seroreactive individuals in Côte d'Ivoire and The Gambia. Improved detection of HIV-2 proviral DNA, rather than exposure to both viruses, infection with only one virus, or infection with a unique third virus containing epitopes common to both HIV-1 and HIV-2, contributes to a more accurate monitoring of the prevalence of HIV-1-HIV-2 dual infections.

DNA Primers↗
PubMed1998

Evaluation of different V3 peptides in an enzyme immunoassay for specific HIV type 1 group O antibody detection.

Strategies to discriminate group O from group M infections need to be improved. We have developed and evaluated an HIV-1 group O V3 peptide-based enzyme immunoassay (PEIA) for specific HIV-1 group O antibody detection among HIV-1-infected patients. Synthetic peptides, derived from the amino acid sequences of the V3 loop of 15 different group O strains and 7 group O consensus sequences, were evaluated in a PEIA against a panel of genetically confirmed group O (n = 33), group M (n = 90), and HIV-1 antibody-negative sera (n = 17). The best-performing PEIA(s) were then used to screen 134 sera of European and 336 sera of Cameroonian origin for the presence of anti-HIV-1 group O antibodies. The reactivity of reference ("gold standard") sera to individual peptides in the PEIA resulted in the selection of five different peptides with sensitivities (sens), specificities (spec), and test efficiencies (TEs) in the range of 90 to 100%. Improvement of the PEIA was obtained with simultaneous reactivity of at least two different peptides in separate wells of an ELISA plate, together with stringent criteria for positivity. We were able to select seven peptide combinations each with a sens, spec, and TE of 96.9, 100, and 99.2%, respectively. None of the 134 European and 4 (1.2%) of the 336 Cameroonian samples sera were group O positive in the optimized HIV-1 group O PEIA; this was confirmed by the repeated presence of reactives, in agreement with the present knowledge of group O infection distribution. Finally, we were able to develop a strategy with a higher TE (99.2%) than the previously used ANT-70 (98.5%) and ANT-70/MVP5180 (95.7%). Our results show that optimal specificity rather than optimal sensitivity makes the V3 PEIA a sufficiently accurate epidemiological tool to be useful in estimating specifically group O infection among HIV-1-infected patients.

Cameroon↗
PubMed1998

Differential diagnosis of HIV type 1 group O and M infection by polymerase chain reaction and PstI restriction analysis of the pol gene fragment.

HIV-1 group O serological screening or confirmation strategies so far have not proved 100% sensitive and specific, indicating a lack of antibody reactivity or cross-reactivity with group O antigens. Therefore, genetic analysis currently represents the only method by which confirm presumed HIV-1 group O or group O/M infections. We have optimized the sensitivity (100%) and specificity (100%) of an HIV-1 group O/M-specific PCR of a pol gene fragment. In addition, we report on a highly sensitive (97.2%) and specific (100%) method for differentiation between HIV-1 group O and group M viruses, using PCR and PstI enzyme restriction fragment analysis of a pol fragment. Compared with sequencing, these methods are fast, inexpensive, and simple.

DNA, Viral↗
PubMed1998

Design and evaluation of an in-house HIV-1 (group M and O), SIVmnd and SIVcpz antigen capture assay.

An enzyme-linked immuno-sorbent assay (ELISA) for the detection of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIVcpz/SIVmnd) antigens was designed using immunoreagents from naturally infected individuals, and compared to the commercially available Vironostika HIV-1 Antigen Microelisa System (Organon Teknika). The in-house assay proved to be specific for HIV-1 isolates belonging to group M (A-H) and group O and for SIVcpz and SIVmnd isolates, but was less sensitive than the Vironostika HIV-1 Antigen Microelisa System, except for SIVmnd. For the strains belonging to HIV-2, SIVmac and SIVagm, the in-house assay could not detect antigen to an appreciable degree. This study shows that a considerably less expensive but sufficiently accurate HIV-1 antigen capture assay can be developed to monitor HIV-1 (group M and O), SIVcpv and SIVmnd antigen in the supernatants of virus cultures.

Antibodies, Monoclonal↗
PubMed1998

Isolation of HIV-1 RNA from plasma: evaluation of seven different methods for extraction (part two).

Some new commercial methods for the extraction of viral RNA have been introduced recently. In addition to the study published previously (Verhofstede, C., Reniers, S., Van Wanzeele. F., Plum J., 1996. AIDS 8, 1421-1427), seven different methods (four newly developed and three reference methods) for extraction of HIV-1 RNA from plasma have been evaluated. The RNA preparation method that gave the best results (acceptable reproducibility, highest sensitivity, reasonable price, fast and easy to perform), was the QIAamp Viral RNA kit from QIAgen. The High Pure Viral RNA Kit (Boehringer Mannheim) as well as the non-commercialised extraction kits were also very sensitive. The non-commercial tests seem less suitable for routine use and for the processing of large number of samples. Two methods, RNA Insta-Pure LS (Eurogentec) and PANext RNA extraction kit 1 (NTL, PANsystems GmbH) are not adapted for HIV plasma extraction. The single step methods using glass fibre or silica column are rapid (from 60 to 75 min depending on the number of wash steps) and although the price is high they are cheaper than the Boom extraction methods: High Pure Viral RNA Kit (Boehringer Mannheim) ($3.3/sample), QIAamp Viral RNA Kit (Qiagen) ($3.6/sample), Boom extraction ($5/sample). The Qiagen kit is the only kit that combines sensitivity with reproducibility, it is commercialised, rapid and affordable in price and can be automated. For most of the methods evaluated the inter-test variability was acceptable (mean variation coefficient between duplicate extractions varied between 26.4 and 48.6%).

Costs and Cost Analysis↗
PubMed1998

Genetic variation of HIV type 1: relevance of interclade variation to vaccine development.

Accumulating data in human immunodeficiency virus (HIV)-infected individuals support the hypothesis that in primary human immunodeficiency virus type 1 (HIV-1) isolates of different clades and phenotype (syncytium inducing [SI] and nonsyncytium inducing [NSI]) common antigenic structures must exist that can stimulate the immune response to produce a broad spectrum (cross-clade and cross SI and NSI) neutralization response. Certain vaccination regimens in chimpanzees and human volunteers with clade B SI type HIV-1 derived candidate vaccines induce neutralizing antibodies against intraclade B SI type primary HIV-1 isolates, but not against intraclade B NSI type of viruses. To be effective against the full antigenic spectrum of primary HIV-1 isolates (cross-clade--SI and NSI) candidate vaccines should contain immunogens of primary isolates representative of the whole antigenic spectrum of HIV-1. There is an urgent need to identify these immunogens and to improve their immunogenicity. As long as we have not yet characterized these cross-HIV-1 spectrum conserved immunogens, candidate vaccines against the more prevalent clades C, A, and E should be developed for evaluation in developing countries. In support of the follow-up and evaluation of the hopefully increasing number of phase 1, 2, and 3 HIV-1 vaccine trials in humans, it is considered a high priority to develop a high throughput neutralization assay, to further expand the use of a limited number of key primary HIV-1 isolates as a surrogate for neutralization of the entire HIV-1 antigenic spectrum (cross-clade--SI and NSI), to develop high throughput subtyping as well as a rapid system to monitor the immunogenic relatedness of different HIV-1 clades.

AIDS Vaccines↗
PubMed1997

The neutralization relationship of HIV type 1, HIV type 2, and SIVcpz is reflected in the genetic diversity that distinguishes them.

Neutralizing antibody (NA) patterns in the sera of individuals naturally infected with human immunodeficiency virus (HIV) type 1, HIV-2, and the simian immunodeficiency virus (SIVcpz) to their homologous and heterologous isolates were determined in a peripheral blood mononuclear cell-based neutralization assay. We examined the role of the V3 loop of HIV-1 and SIVcpz in neutralization and the cross-reactivities among them. Cross-neutralization by sera of humans and chimpanzees naturally infected, respectively, with HIV-1 and SIVcpz isolates was more extensive than the infrequent and low-titer cross-neutralizations observed between HIV-1 and HIV-2. Neutralization of 9 of the 16 HIV-1 isolates by 9 of 10 HIV-2 and all 3 SIVcpz antibody-positive sera were weak and sporadic (titer, 1:10-1:160). Twelve of 15 HIV-1 sera neutralized the 2 SIVcpz isolates with titers of 1:10-1:320 but only sporadically neutralized the 6 HIV-2 isolates (titers: 1:10-1:20). The majority of HIV-1 and SIVcpz sera bound to the V3 peptides although their binding capacity did not readily reflect their neutralizing capacity. The HIV-2 sera did not or only weakly bound to the V3 peptides. These results suggest that HIV-1 and SIVcpz share some structural and functional similarities that set them apart from HIV-2.

Animals↗
PubMed1997

Evaluation of a newly developed HIV antigen test.

A new monoclonal antibody-based enzyme immunoassay (Innogenetics) for the detection and quantification of p24 core antigens of HIV-1 (group M and group O) and of HIV-2 was evaluated on 2745 serum samples and 18 culture supernatants and compared with a reference (Coulter) HIV-1 p24 antigen assay. Positive results were confirmed by neutralization with the reagents of the respective tests. As demonstrated by dilution series of HIV cocultures, the new test recognizes p24 antigen of the most common HIV genetic subtypes, including group O and HIV-2. Titres ranged from 729 to 531441. Therefore p24 antigen assay is but very weakly reactive with HIV-2 (titres from 9 to 81). The new test is considerably more sensitive than the reference. In a population of 365 follow-up samples from 86 different patients, representing all stages of infection, the new test detected p24 antigen at least once in 52% (45/86) of these patients, whereas the reference was positive in 31% (27/86). The newly designed test detected antigen in 40% (145/365) of the samples, while the reference was positive in 21% (75/365). In a group of PCR and/or culture positive neonates, 33% (9/27) of the samples were positive with the new test versus 18% (5/27) with the reference. The specificity of the new test, as determined on 2,000 blood donor samples, was 99.65% (initially), 99.80% (after repetition), and 100% (with neutralization). The reference scored 99.95%, 100%, and 100%, respectively. In 300 seronegative samples from persons at risk, the initial specificity of the new test was 98.67% (the reference, 99.00%). With neutralization, both assays were 100% specific.

Antibodies, Monoclonal↗
PubMed1997

Reactivity and amplification efficiency of the NASBA HIV-1 RNA amplification system with regard to different HIV-1 subtypes.

In view of the genetic diversity of the human immunodeficiency virus Type 1, we assessed the sensitivity and quantification efficiency of the HIV-1 RNA NASBA amplification system with respect to different HIV-1 subtypes and recombinants. Twenty cell culture supernatants representing 17 HIV-1 group M and 3 group O strains were tested, and NASBA RNA loads were compared with results obtained with a RT-PCR based HIV-1 RNA quantitation method, with p24-antigen concentrations and with the infective dose. The current HIV-1 RNA NASBA seemed suitable to quantitate representatives of different HIV-1 M subtypes. Differences between NASBA and RT-PCR loads were observed for certain HIV-1 M strains. Significantly lower RT-PCR loads were measured for most gag A, gag B and gag F strains, whereas NASBA detected lower copy numbers in 1 gag H strain and 1 gag H/env G recombinant. NASBA was not able to quantify 1 HIV-1 group M recombinant. Some of these differences could be explained by the presence and position of mismatches with primers. HIV-1 group O strains were not detectable by both RNA amplification methods. A firm correlation was not observed between the measured RNA loads and either the p24-antigen concentration or the infective dose.

HIV-1↗
PubMed1997

Study of the dynamics of neutralization escape mutants in a chimpanzee naturally infected with the simian immunodeficiency virus SIVcpz-ant.

Here we report on the use of spectral map analysis of time-paired sequential neutralization data of 11 serum samples of a chimpanzee naturally infected with a simian immunodeficiency virus (SIVcpz-ant) and 8 primary consecutive SIVcpz-ant isolates, taken at about 4-month intervals. The analysis reveals the existence of three SIVcpz-ant isolate and serum neutralization clusters. Each cluster groups virus isolates and/or sera based on similarities of their neutralization spectra. On average, neutralization escape mutants emerged after 15 months and mounted a neutralization response approximately 8 months later. The entire gp160 regions of eight consecutive isolates were sequenced and analyzed by a new statistical method called polygram, which allowed the deduction of amino acid sequence motifs of gp160 which were specific for SIVcpz-ant isolates belonging to the same isolate neutralization clusters. Changes in specific amino acid quadruplets in V1, V2, C3, V4, V5, and CD4 domains of gp120 and gp40 were seen to correlate with the neutralization clusters with most of the specific changes occurring in the V4 region. This method of analysis may facilitate an understanding of the study of the dynamic interplay between human immunodeficiency virus (HIV) and host neutralization responses as well as providing possible insights into mechanisms of persistence of HIV-1-related lentiviruses in their natural hosts.

Amino Acid Sequence↗
PubMed1996

Epidemiological and molecular characteristics of HIV infection in Gabon, 1986-1994.

OBJECTIVE: To describe trends in the prevalence of HIV-1 infection in different populations in Gabon, and the molecular characteristics of circulating HIV strains. METHODS: Data were collected on HIV prevalence through sentinel surveillance surveys in different populations in Libreville (the capital) and in Franceville. In Libreville, a total of 7082 individuals (hospitalized patients, tuberculosis patients, pregnant women, asymptomatic adults, prisoners) were recruited between 1986 and 1994. In Franceville, we tested 771 pregnant women and 886 healthy asymptomatic adults (1986-1988). Sera were screened for HIV antibodies by enzyme-linked immunosorbent assay (ELISA) and confirmed by Western blot or line immunoassay (LIA). Reactive samples in ELISA were tested for the presence of antibodies to HIV-1 group O viruses by ELISA using V3 peptides from HIV-1 ANT-70 and HIV-1 MVP-5180 followed by confirmation by LIA and a specific Western blot. Seventeen HIV-1 strains were isolated (1988-1993) and a 900 base-pair fragment encoding the env region containing V3, V4, V5 and beginning of gp41 was sequenced and a phylogenetic tree was constructed. RESULTS: HIV prevalence was relatively low and remained stable (0.7-1.6% in pregnant women, 2.1-2.2% in the general population). The prevalence was also stable among prisoners (2.1-2.6%). Among hospitalized and tuberculosis patients prevalence was higher and increased (1.8-12.7% and 1.5-16.2%, respectively). Only three sera had antibodies to HIV-1 group O. The 17 HIV-1 strains represent six different genetic subtypes including type O. CONCLUSION: Our data from 1986 to 1994 show a stable and low HIV prevalence in Gabon, and a high genetic diversity of HIV-1 strains. This, also observed in Cameroon, is in contrast to that found elsewhere in Africa. Differences in rate of spread of HIV infection are probably explained by interplay between numerous factors. The role of different HIV subtypes in the dynamics of the HIV epidemic should be examined further.

Adult↗
PubMed1996

Phylogenetic analysis of the env gene of HIV-1 isolates taking into account individual nucleotide substitution rates.

OBJECTIVE: To estimate the relative substitution rate of the individual positions in an alignment of HIV-1 env sequences coding for areas V3, V4, V5, and the beginning of gp41, and to study phylogenetic relationships between HIV-1 strains taking into account these substitution rate estimates. DESIGN: Phylogenetic comparison of 145 HIV-1 strains classified in HIV-1 group M, subtypes A-H and isolated from patients of 24 different geographical origins. METHODS: A new method recently developed for measuring the substitution rates of the individual nucleotides in a sequence alignment was applied to an alignment of env gene sequences. From the resulting substitution rate distribution, an equation was derived that describes the relationship between dissimilarity and evolutionary distance better than equations previously available. Phylogenetic trees were then constructed from matrices of distances computed using this new equation. RESULTS: 'Substitution rate calibration' offers detailed information on the relative substitution rate or variability of the nucleotides in the env gene. A large phylogenetic tree of 145 env gene sequences constructed by neighbour-joining and taking into account the substitution rate spectrum for this gene, clearly shows the existence of the eight subtypes A-H, all supported at a bootstrap level of 90% or higher. Intersubtype distances were between 0.25 and 0.38, which is considerably higher than those found in trees not considering differences in substitution rates among different alignment positions. CONCLUSIONS: Evolutionary distances are seriously underestimated when individual substitution rates are not considered in the estimation evolutionary distances. Furthermore, due to the more accurate estimation of evolutionary distances, naturally occurring HIV-1 intersubtype recombinants could be recognized more easily.

Base Sequence↗