Bridging the Gap: Conference Record [Abstract book, International Conference on AIDS (12th: 1998: Geneva, Switzerland)]

768 Abstracts 41128-41132

12th World AIDS Conference

RNA ~ SD, 5.13 ~ 0.88 vs 4.28 ~ 0.79; p < 0.001). In the one subtype G sample,
the viral load was 1.9 and 1.5 loglo copies per ml higher in the NucliSens assay
than in the standard and modified Amplicor tests, respectively.
Conclusion: These results indicate that commercially available HIV-1 RNA assays vary greatly in their ability to detect HIV-1 subtype A. The modified Amplicor
Monitor assay, which had the best performance in this evaluation could be used
to reliably monitor persons infected with HIV-1 subtype A.

1411281

Comparative evaluation of Amplicor HIV-1 Monitor with
added primer set with quantiplex v2.0 HIV RNA assay and
the nuclisens for quantitation of HIV-1 RNA in a Thai
study population, primarily infected with subtype E

Sasiwimol Ubolyam1, K. Ruxrungtham2, G.J. Weverling3, C. Ungsidhapand4,
J. Lange3, D. Cooper5, P. Phanuphak6. 1Dept. of Microbiology, Chulalongkorn
Hosp., Rama IV Road, Pratumwan Bangkok 10330; 2Dept. of Medicine,
Chulalongkorn Uni., Bangkok; 4HIV-NAT, TRCS; 6 Program on AIDS, TRCS,
Thailand; 3NATEC, Amsterdam, Netherlands; 5NCHECR, Australia
Objective: To comparatively evaluate the reverse transcriptase-coupled polymerase chain reaction (Amplicor HIV-1 Monitor) with added primer set with the
branched DNA method, 2nd version (Quantiplex HIV-1 RNA v2.0) and the nucleic
acid sequence-based assay, 2nd version (NucliSens) for quantitation of HIV-1 in
a Thai study population predominantly infected with HIV-1 subtype E.
Methods: Blinded testing of 319 stored plasma samples taken at sequential study visits from 72 antiretroviral naive participants with CD4+ counts 150 -350/mm3 in a study comparing ddl + d4T in a range of doses with ddl alone. A
subset of 40 samples of 7 randomly selected patients was tested twice with Amplicor with added primer set and twice with NucliSens. All samples were tested
once with Quantiplex. All samples were tested under identical circumstances
according to the manufacturers' recommendations.
Results: HIV subtype was determined from a study baseline serum sample for
all 72 patients: 60 subtype E, 5 E + BMN, 7 could not be subtyped and have been
sent for genotyping.
Out of the 40 samples tested twice with Amplicor, 31 had detectable viral
load in both tests with a median value of 4.2 loglo copies/ml and a difference
of 0.05 loglo copies/mi (std = 0.14, p = 0.05) was found between the two measurements. With the Quantiplex and NucliSens, 5.6% and 4.2% of all samples
had undetectable viral load (cut-offs 500 and 400 copies/ml respectively) before
treatment, compared to 1.4% with Amplicor (cut-off 400 copies/ml). After start
of treatment these percentages were Quantiplex 70.4%, NucliSens 62.3% and
Amplicor 30.0%. Mean results with Quantiplex v2.0 were 0.66 logo1 below those
obtained with Amplicor before treatment was initiated (4.92 log10-4.26 logio, std
0.39, p = 0.0001), and 0.63 loglo lower (4.20 logio-3.57 loglo, std 0.41, p =
0.0001) after treatment was initiated in samples quantifiable in both assays (n
= 72 before treatment, n = 78 after initiation of treatment). Mean results with
NucliSens were 0.65 loglo below those obtained with Amplicor and 0.010 logo0
below those obtained with Quantiplex for the entire set of samples
Conclusion: Viral load (VL) analysis in this study leads to comparable results
in both drop in VL and % of samples with undetectable VL with Quantiplex and
NucliSens. In contrast, analysis with Amplicor results in higher VL and a smaller %
of samples with undectable VL. The difference impacts on clinical trial outcomes
expressed in % below level of detection.
1343*/41129 High throughput assay for sensitive detection of
HIV-1 RNA of diverse origins, including type O
strains
Cristina Giachetti1, D. Kolk2, J. Dockter2, J.P. Knowlton2, R. Wang2,
S. Hotaling2, S. McDonough2. 110210 Genetic Center Drive, San Diego, CA
92121; 2Gen-Probe Incorporated, San Diego, CA, USA
Background: A major challenge for the development of screening assays for
HIV-1 is its high degree of genetic diversity, which includes two major groups
M and 0, and several subtypes or clades. HIV-1 RNA appears in plasma prior
to antibody response, providing a very sensitive method for early detection of
infection. However, current nucleic acid detection formats are too time consuming
to apply to individual donor screening. To overcome these problems, we have
developed a novel HIV-1 RNA screening assay.
Methods: The assay protocol includes three steps which are automation compatible: (i.) target capture/magnetic microparticle separation of specimen RNA; (ii.)
Transcription-Mediated Amplification (TMA) of viral sequences; and (iii.) amplicon
detection with Hybridization Protection Assay (HPA).
Results: Using a manual mode, one operator can process up to 200 samples
in 5-6 hours. The test detected HIV-1 RNA 11 days -on average- before seroconversion. Analytical sensitivity is at least 100 copies/ml, and sensitive detection
of HIV-1 in clinical isolates and specimens of diverse geographical origin was
demonstrated. Subtype evaluation included more than 250 different strains of
subtypes A-H and 53 different type O variants. Good specificity was observed
in normal plasmas or in plasmas infected with other bloodborne pathogens. No
interference was seen in specimens with autoimmune antibodies, hemoglobin,
bilirubin, or lipid.
Conclusions: The use of target capture, TMA and HPA technologies resulted
in a highly sensitive assay for detection of all known HIV-1 subtypes. The sensitivity, specificity, ease of use, and compatibility with automation of this test will
make it valuable for large volume plasma screening. Implementation of this assay
could have a measurable impact on the safety of donor blood.

41130_ Sensitivity, linear range and performance of HIV-1
subtypes of the COBASAMPLICOR HIV-1 monitor test
Cynthia Pletcher, D. De Bonville, J. Wang, K. Lavallee, A. Chutoransky, K. Arch,
A. Wang2. Roche Molecular Systems Somerville, 1080 US Highway 202 South
Somerville New Jersey 08876, USA
Objectives: Determine the sensitivity, linear range, precision and performance on
Group M subtypes of HIV-1 of the AMPLICOR HIV-1 MONITOR Test, version 1.5,
and an automated version of the test, the COBAS AMPLICOR HIV-1 MONITOR
Test. The HIV-1 MONITOR version 1.5 is an upgraded test with new PCR primers
designed to yield equivalent performance on all Group M subtypes of HIV-1, and
a new specimen preparation method to increase the sensitivity of the test to 50
HIV RNA copies/mL
Methods: Sample panels for determination of sensitivity, linear range and precision were prepared by serial dilution of cultured HIV-1 in normal human plasma.
For analysis of different subtypes a panel of 30 HIV-1 isolates of subtypes A-G
was prepared from cultured virus stocks that were quantified by electron microscopic pare tice counting. Each isolate was tested in triplicate at 20,000 and 2,000
virus particle/mL. To determine the correlation between version 1.5, the COBAS
test and the current test (version 1.0) a panel of 100 North American plasma
samples (subtype B) was tested.
Results: Analysis of the subtype panel with version 1.0 yielded results consistent with the particle counts for subtypes B, C and D. Subtypes A, E and F
yielded results at least 10-fold less than the expected value. Within the precision
of the study version 1.5 and COBAS yielded equivalent results for all subtypes.
Version 1.5 and COBAS had equivalent sensitivity to version 1.0 of approximately
400 HIV RNA copies/mL, and were linear to approximately 500,000 HIV RNA
copies/mL. Using the UltraSensitive Specimen Preparation Method, version 1.5
and COBAS had sensitivity of approximately 50 HIV RNA copies/mL.
Conclusions: The version 1.5 and COBAS AMPLICOR HIV-1 MONITOR yield
equivalent results on Group M subtypes A-G. The sensitivity, linearity and precision of the COBAS and version 1.5 tests, with the standard specimen preparation
method are equivalent to version 1.0. With the UltraSensitive specimen preparation method, version 1.5 and COBAS had a sensitivity of approximately 50 HIV
RNA copies/mL.
411311 Phase 1/11 study of single dose nevirapine in HIV infected
pregnant Ugandan women and their infants
P. Musoke Mudido1, L. Guay2, D. Bagenda1, F. Mmiro1, C. Nakabiito1,
M. Mirochnick3, J.B. Jackson2. 1Makerere University, Kampala, Uganda; 2Johns
Hopkins University, Baltimore, MD; 3Boston University Boston, MA, USA
Objectives: To determine the pharmacokinetics, safety and tolerance of single
dose nevirapine (NVP) in HIV-1 infected pregnant women in labor and their
infants.
Methods: Twenty one HIV infected pregnant women received a single 200
mg dose of nevirapine during active labor. Eight infants received no NVP (cohort
1) and 13 infants received a single dose of NVP suspension at 2 mg/kg at 72
hours of life (cohort 2). Mothers and infants were monitored to determine safety,
tolerance, NVP, and HIV-1 RNA levels.
Results: The median maternal CD4 cell count at enrollment was 595 cells/ul
with a median plasma HIV RNA level of 22,746 (neg - 1,796,670) copies/ml. The
median time to maternal peak plasma NVP level was 8.1 hours (2-26). Only 3
mothers received NVP < 1 hour before delivery. The median maternal NVP concentration at delivery was 1623 ng/ml (238-2356) with a median cord/maternal
blood ratio of 0.75 (0.37-0.93). The median half life in mothers was 58 hours
(27.4-90 hrs) and in infants was 54 hours (39.6-85.1). There was a median 1.3
log reduction in maternal plasma HIV RNA levels 7 days after dosing. Nine of 20
maternal RNA levels had returned to baseline levels by 6 weeks post delivery. Of
infants born more than one hour after maternal dosing, 6 of 6 evaluable infants in
cohort 1 had NVP levels >100 ng/ml (10 x IC50) and 10 of 10 evaluable infants in
cohort 2 had NVP levels consistently >150 ng/ml (177-757) 7 days after delivery.
Two of 21 infants have confirmed evidence of HIV infection at 6 weeks of age by
plasma HIV RNA detection. NVP was well tolerated in both mothers and infants
with no serious adverse events that could be directly related to NVP.
Conclusion: A single maternal labor dose of NVP was safe, and well tolerated
with marked reduction in viral load. This maternal dose yielded significant infant
NVP levels through the first week of life. The additional infant dosing in cohort 2
yielded higher infant NVP levels through the first week of life.
41132 Comparative evaluation of a new test for HIV-RNA
quantitation in plasma
Giancarlo Cardi, Filippo Ansaldi, V. Venturino, M.T. de Martini, L. Bagnasco,
A. Pasini, A. Demetri, G.C. Icardi. Institute of Hygiene-Univ. of Genoa Via
Pastore L Genoa, Italy
The aim of the study was the evaluation of a new test for the quantitative dosage

of HIV1-RNA named HYBRID CAPTURE II (HC-II) HIV-1 RNA TEST (Digene
Corp., USA). In this test, the nucleic acid is measured directly, its quantity being proportional to the resulting chemiluminescent signal. Biotin-labelled DNA
probes cover 92% of the genomic sequence. Results range between 500 and
106 copies/ml and are obtained in approximately 6 hours. Data from the new
test were compared with those from 3 reference tests: Reverse Transcriptase
Polimerase Chain Reaction (RT-PCR) Amplicor HIV Monitor (Roche Diagnostics); Nucleic Acid Sequence-Based Amplification (NASBA) Nuclisens (Organon