Abstracts Vol. 1 [International Conference on AIDS (11th: 1996: Vancouver, Canada)]

Tu.A.2056 - Tu.A.2060

Tuesday July 9, 1996

Tu.A.2056
SEQUENCE VARIABILITY OF HIV- I ADCC EPITOPES IN VIVO:V3 LOOP ADCC
EPITOPES APPEAR TO BE THE MAJOR DETERMINANTS FOR ADCC ACTIVITY AT
DIFFERENT STAGES OF THE INFECTION
Merzouki A*, Estable MC * *, Acel A *, Moniz DE SA M **, Montaner J.S. **, Mandy F ***,
Arella M. *. Institut Armand Frappier, Laval, Quebec, * CFE/UBC,Vancouver, B.C.,** Health
and Welfare Canada. Ottawa, Ontario*** Canada.
Objective: To study the relationship between ADCC activity ADCC epitope sequences,
infection stage, CD4+ cell counts, HIV- I mRNA levels in PBMC and the predicted biological
phenotype of the virus.
Methods: CEM NKR cell clones expressing the HIV- I NL4.3 gp 120/I 60 were used as targets to monitor -HIV- I specific ADCC activities in 33 sera from patients at different stages
of the infection. Nested PCR was used to amplify ADCC epitopes in the V3, C4 and C5-C6
env gene regions. Amplicons were cloned and up to 10 from each sample were sequenced.
Phylogenetic analysis and calculations of intra- and inter-patient variability were performed
by using the software PHYLIP version 3.5. Quantitative competitive (Qc)-RT-PCR was used
for measuring the relative abundance of HIV-I regulatory (spliced) mRNA (tat, rev, nef) versus structural (unspliced) mRNA (gag-pol) directly in PBMC.
Results: Sera from healthy seropositive (CDC Class II) subjects mediated substantially higher levels of ADCC activity (25.7 ~ 1I.5%) than sera from individuals in CDC Class III (10.3
~ 8.9%) or IV (6.4 ~ 8.1%). Both intra- and inter-patient V3 loop ADCC epitope sequences
displayed greater variability with infection progression (differences ranging from 5 - 7% at
stage II, 6 - 14% at stage Ill and 12% - 15% at stage IV). In contrast, ADCC epitope
sequences outside the V3 loop (C4 and C5-C6 regions) were highly conserved (differences
ranging from 0 - 4%) irrespective of the degree of infection progression. Sequence analysis
also showed that all patients in stage II and Ill of the infection, who displayed (by Qc-RTPCR) the lowest levels of unspliced HIV- I mRNA, were either exclusively (94%) or predontinantly (6%) infected with variants with a predicted non-syncitium-inducing
(NSI)/rnacrophage tropic phenotype. Importantly all samples with sequences predicting a
syncitiurmn-inducing (51)fT-cell tropic phenotype were from CDC Class IV patients, that also
displayed the highest ratio of spliced to unspliced HIV- I mRNA.
Conclusions: ADCC activity and CD4 T-cell number decline with the progression of the
infection in patients included in this study Secondly,V3 loop ADCC epitopes appear to be
the major determinants for ADCC activity, and the decline or the loss of ADCC activity
could be a consequence of the appearance of variants with predicted SI/T-cell tropic phenotype (rapidly replicating and cytopathic phenotype) that map the same region.
Abderrazzak Merzouki. Institut Armand Frappier, CP 100, 53 I, boul. des Prairies, Laval
Quebec, Canada, H7N 4Z3.Tel: [phone] I 5, Fax: [phone], email:
[email protected]
Tu.A.2057
DNA SEQUENCE OF THE C2-V3 REGION OF THE EXTERNAL GLYCOPROTEIN
(gp 120) FROM CUBAN HIV-I INFECTED INDIVIDUALS
Gon ez Carmen E*, Iglesias E*, Ferntndez J**, Lobaina L**, Noa E**, D'az H**, Rolo F*,
Duarte C.
Objective: To analyze the DNA variability of the C2-V3 region of gpl20 in Cuban HIV-I
infected individuals.
Methods: Total DNA was extracted firom PBMC of six seropositive individuals and PBMC
infected with a primary isolate from other patient.The C2-V3 region of the gpl20 was
amplified using nested PCR with different set of primers, and the products were cloned in a
Tvector using a commercial kit (Amersham). Plasmids were purified firom positive clones
and the DNA sequence determined with Sequenase 2,0.
Results: The sequence of the V3 region from five independent clones of each sample
showed an intrapatient divergence ranging between 0 to 9.8%.The consensus sequences
obtained in all cases clustered with sequences from clade B reported in "Los Alamos" database (3.9- I 8.5% of divergence).The amino acids sequences GPGR, GWGR, GRGR and
T PGR were found in the tip of the V3 loop The last two tetrapeptides are reported here
for the first time.
Conclusions: The seven samples studied, which were taken from persons infected in Cuba
after 1990, represent viruses from the clade B.This suggests that the viral variability that followed an African pattern in the last decade has changed and is closer to the observed in
Europe and America.
CE. G mez.Vaccine Division, CIGB, PO. Box 61I62, Cubanact:n, Havana 10600. Cuba.Terl.:
5 3-7-2 1I8070; Fax: [phone]; Email: [email protected]
Tu.A.2058
SEVERAL GENETIC SUBTYPES OF HIV- I FOUND IN THE AREAS AROUND THE
BALTIC SEA
Liitsola, K.. Laukkanen T., Salminen M.Leinikki Pauli. National Public Health Institute, Helsinki,
Finland
Objective: To determine the genetic subtypes of HIV- I and to analyse phylogenetic relationships of viruses from Finland, the Baltic Countries and Russia.
Methods: Proviral sequences from the nucleocapsid coding region were amplified and
directly sequenced from 95 Finnish, I I Estonian, 10 Latvian, 6 Lithuanian and 13 Russian
patients.The sequences were phylogenetically analysed using maximum likelihood distance
matria and the Fitch-Margoliash methods.
Results: Viruses belonging to six different subtypes were found. In Finland most viruses
(n=70) were classified as subtype B. others belonged to five different subtypes: A (n=6), C
(n-II ), D (n3), F (n- I), G (n=4). In Estonia 10 viruses belonged to subtype B and one to
subtype D. Nine Lithuanian patients harbored subtype B virus and one subtype C virus.
Four Lithuanian sequences were of subtype B and one was of subtype A.Ten Russian
sequences were classified to subtype B and one in subtype G. One Lithuanian and two
Russian sequences, closely related to each other remained unclassified.Thus subtype B was
the most comnion subtype in all of these countries. Homosexual transmissions predominated in the B subtype, and heterosexual in the other subtypes. Non-B infections were mainly
of Afi can origin. Most subtype B viruses in the Baltic countries and Russia were closely
related suggesting the common source of infection, but in Finland more heterogeneity was
found.
Conclusions: Though the spread of HIV- I has been slow in the areas around the Baltic Sea,

the distribution of HIV-I genomic subtypes is probably similar as elsewhere in Europe.
Considerable heterogeneity was found among viruses in Finland, while most Baltic and
Russian subtype B viruses studied seemed to belong to a closely related virus pool.
Background information of Finnish patients confirmed that most infections are imported in
Finland, but homogeneit of Baltic and Russian sequences suggests that the number of infection sources is quite small in these countries probably because of restnricted traveling to the
West during the Soviet era.
P Leinikki, Mannerheinintie 166, 00300 Helsinki, Finland Telephone:[phone], Fax: 358
0 474 446 I, email: [email protected]
Tu.A.2059
EVIDENCE OF MULTIPLE SIVagm-sab SUBTYPES WITHIN AFRICAN GREEN
MONKEYS OF THE SAME TROOP
F Bibollet-Ruche I, C Brengues 1, A Galat-Luong2, G Galat2, X Pourrut2, F Veas l. G Cuny.
I. Laboratoire Retrovirus, ORSTOM, Montpellier. France. 2. Laboratoire de Pimatologie,
ORSTOM, Dakar, Senegal.
Rationale and Objective: Elevated seroprevalence rates have been reported in the four
African green monkey (AGMs) subspecies and each subspecies is infected by SIVagrn strains
exhibiting limited identity. Moreover, genetic diversity within the viruses infecting each AGM
subspecies is much more important to that observed for the other primate lentiviruses.The
objective of this study is to determine genetic diversity of SIVagm-sab from feral AGM of
the sabaeus subspecies (Cercopithecus aethiops sbaeurs).
Methods: AGMs were sampled from two troops living in non overlapping home ranges in
the Fathala forest (Saloum Delta National Park, Senegal). By a sermni nested PCR technique,
two 450bp regions of the genome were amplified and sequenced, spanning the first tat
coding exon and part of the TM glycoprotein.The env fragment allowed to assess the variability of 140 amino acid residues in the gp40, corresponding to the transmembrane
domain and a part of the cytoplasmic region. Combination of en and tat regions allowed
to deduce sequences forTat and Rev regulatory proteins.
Results: Eleven new SIVagm-sab from 2 troops were included in this study Regulatory proteins Tat and Rev showed limited amino acid variability for domains known to be important
for protein functions. However, detailed analysis of these proteins indicate that N terminus
region forTat and C-terminus region for Rev are highly variable; biological relevance of this
variability is not known, but has not yet been reported for regulatory proteins in lentivirus.
Analysis of splicing events for tat also revealed important differences between the different
SIVagm-sab strains. Average homology score between the different viruses for gp40 amino
acid sequences is comparable to homology between SlVagrn-ver strains.lnexpectedly
SIVagm-sab within AGMs from the same troop differ by as ruch as 30% in the portion of
the TM glycoprotein evaluated. Phylogenetic analysis of env and tat regions allowed identification of distinct virus lineages, equidistantly related, referred as SIVagm-sab subtypes.
Identical clustering of these different viruses in both trees indicate that at least two distantly
related subtypes are found in one AGM troop. Moreover these subtypes differ from one
troop to an other.
Conclusions: These results represents the most detailed phylogenetic analysis of SIVagm-sab
within feral animals that has been conducted to date indicating the presence of multiple
subtypes within the same troop.These data strongly suggests ancient introduction of distinct
SIVagm-sab lineages, designated here as SIVagm-sab subtypes.
Bibollet-Ruche, Frederic. Laboratoire Rdtrovirus, 99 I Avenue Agropols. B.P 5045 34032
Montpellier cdx I France.TEL: [phone] FAX; 67 54 78 00
Tu.A.2060
THE IMPACT OF THE HIV-I GENETIC SUBTYPE ON THE NATURAL HISTORY OF
HIV- I INFECTION: PRELIMINARY RESULTS
M.Lasky, JL Perret2, M.Peeters-, F Bibollet- Ruche, D.Dormont3. F.Liegeois, D.Patrel,
C.Gras2, E.Delaporte. I.aboratoire Retrovirus, ORSTOM. Montpellier,; France,. 2. H6pital
Laveran, Marseille, France. 3. CEA, Fontenay, France.
Objective:To evaluate the impact of the genetic subtype on the natural history of HIV-I
infection.
Materials and Methods: Caucasian patients infected oversea, mainly in Africa or Asia
attending the Laveran Hospital in Marseille were included in the stud). For all these patients
the year and the country of infection are known. Every 3 to 6 months the patients undergo
a standardized clinical examination and CD4 counts are determined. A virus isolation is
done for all these patients in order to determine the genetic subtype and the phenotype.
Genotyping was done by HMA (V3--V5 region of gp I20) and by direct sequencing and phylogenetic analysis of the C2-V3 region.Syncytium formation has been determined on the
MT-2 cell line.Viral load was measured for all the patients by a p2-4 ant.gen capturinng test.
Results: A total of 7 I male patients are actually included in the study 2 1 Iave been infected in Europe and 50 in different countries oversea (Djibouti, Cameroon,7chad, Gabon,
RCA, Guyane, C6te d'Ivoire, Mayotte,Cambodge) 20/21(95,2%) patients contamnated in
Europe were infected with genotype B and I with genotype A. Among the 50 patients
infected overseas 9(18%) were infected with subtype A, 14 (28%) subtype B, 14 (28%)subtype C, 3 (6%) subtype D and 4 (8%) with subtype E as determined by HMAThe genetic
subtypes identified by HMA were confirmed by seqience and phylogenetic analysis of the
C2V3 region.The mean follow-up period for the 7 I patients was 40.5 months (7- 10
months), with a mean of 49 months for genotype A, 42.2 05 type B, 36,3m type C, and
24,7m for type E. A regression analysis was done for the different subtypes to measure the
rate of the decline in CD4counts, and fom these preliminary results appears that the rate
of decline in CD4 counts is similar for patients infected with type A, B, C and D. but is significantly faster for patients infected with subtype E (p<0,03) None of the 14 subtype C
strains were Syncytium inducing(SI), whereas 45% of type B, 33,3% of type A and 25% of
type E strains were SI.These differences in phenotypes were not related to the stage of the
disease. 6/14(43%) type C patients were positive for p24 Ag in their plasma versus
1/10(10%) for type A. I I/40(28(%) for type B and I/4 for type E.
Conclusions: From this preliminary results we observe that: non B genetic subtypes of
HIV- I are introduced in Europe by infections occurred overseas: the decline in CD4 counts
is faster for patients infected with subtype E and SI strains seem to be less frequent among
subtype C strains.
Peeters, Martine. Laboratoire Retrovirus, 91 I Avenue Aguopolis B.P 50-45 34032 Montpellie,
France.TEL: [phone] Fax: [phone] cdx I

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