Final Program and Oral Abstracts [International Conference on AIDS (8th: 1992: Amsterdam, Netherlands)]

WeA 1079-WeA 1084

TRACK A: BASIC SCIENCE

WeA 1079 SIMIAN IMMUNODEFICIENCY VIRUSES FROM TANTALUS MONKEYS: EVIDENCE FOR A
NEW SPECIES-SPECIFICSIVAGM-TYPE. Mialer. Michaela C.*; Saksena, N.K.*'";
Chappey, C.**; Herv6, V.***; Nerrienet E.*; Durand, J.-P.""; Georges
A.J.***; Sonigo P.******; Barr6-Sinoussi F.*; *lnstitut Pasteur, Paris;
*/*State University, Syracuse; **URBB263 INSERM, Paris; ""* Institut
Pasteur, Bangui, *"*Institut Pasteur, Dakar, **** ICGM, Paris.
Objective: Simian immunodeficiency viruses (SIV) are widely present in African Green monkeys
(AGM) and may have evolved in parallel with their natural host species. However, information to
date is limited to viruses of only three species: the vervet, the grivet and the sabaeus green
monkeys. We present here a genetic study of lentiviruses of another AGM species, the tantalus. We
also initiated a first study in env of SIV from sabaeus monkeys.
Methods: SIV isolates from tantalus in Central African Republic and from sabaeus in Senegal were
selected. Proviral DNA-fragments in gag-, pol- and env-regions were amplified by PCR. Env
amplified fragments were sequenced directly using SIVagm-specific nested primers. The deduced aasequences were compared to each other and to all other SIVagms reported so far.
Results: 544 bp fragments encoding for the C-terminal half of the EGP of 4 SIV isolates from tantalus
and 4 viral strains from sabaeus were sequenced. The amino acid identity was always greater
between the tantalus isolates themselves (>78 %) than between tantalus SIVs and viruses from
vervet, grivet or sabaeus monkeys (<70%). The greatest variation was observed between one
tantalus and one sabaeus strain (57.1%). In addition, alignment of entire gp120 encoding sequences
revealed that the first and the fourth variable domains of SIV from tantalus are highly divergent in
regard to equivalent sequences of previously published SIVagms.
Conclusion: This analysis confirms the existence of species specific SIVs for vervets, grivets and
sabaeus monkeys and reveals a fourth distinct SIVagm group specific for tantalus. Genetic distances
between these groups are significant and allow to distinguish SIV types according to each host
species. However, enhancing Informations on the phylogeny of SIVagms require additional sequence
analysis. Such informations on genetic diversity may also be relevant for a better understanding of
possible adaptation mechanisms between primate lentiviruses and their natural hosts in Africa, and
their possible consequences on the lack of pathogenicity observed so far in naturally infected
monkeys.
Miiller Michaela C. Institut Pasteur, Unit4 de Biologie des
R4trovirus, 25 Rue du Dr. Roux, 75724 PARIS C4dex 15, France
Tel.: [phone] / Fax: [phone]
WeA 1081 MOLECULAR CLONING AND SEQUENCE ANALYSISOF A VARIANT IMMUNODEFICIENCY VIRUS ISOLATED FROM A WILD CAPTURED CIIlMPANZEE.
Vanden Haesevelde, Marlcen, Peeters M.*, Willems B., Saman E., vandcr Grocn G.*, and
Van Heuverswyn H.
Innogenetics N.V., 9052 Ghent; * Institute of Tropical Medicine, 2000 Antwerp, Belgium.
Objective: The immunodeficiency virus, SIVcp_ anisolated from a wild captured chimpanzee, was molecular cloned
and sequenced. The purpose was to document the genetic diversity exhibited by the two SIVs isolated from
chimpanzees and confirm their position within the HIV/SIV family.
Methods: Primers and nested primers to HIV-1 conserved regions were used in the Polymerase Chain Reaction on
cellular DNA from cultured human PBL's infected with the SIVpzant isolate. The amplified products were cloned
and their information was sequenced using the dideoxy chain termination technique.
Results: The SIV p,_nt, isolate shows considerable sequence variation when compared with the Gabonese isolate
SIVcpz- gab (Huet T. et al., 1990, Nature 345:356-359). The homology level between both chimpanzee isolates varies
from 70% for the more conserved genes (POL) to 55% for the more variable parts of the genome (env). The same
levels of homology were found when the chimpanzee isolates were compared with different members of the HIV-1
family, as well as with very divergent HIV-1 isolate Ant70 (Vanden Haesevelde et al., 1991, VIIth International
Conference on AIDS, Abstracts M.A.1157), whereas the variation within the HIV-1 family is much smaller.
Phylogenetic tree analysis based on the alignment of pol and env nucleotide sequences, situates the SIVcpz within the
HIV-1 family. However, both SIVcpz isolates do not duster together, indicating the SIVepz strains do not form a
separate subgroup within the HIV-1 family.
Conclusion: The present analysis of the SIVc.antisolateindicates that these chimpanzee isolates belong to the HIV-1
family but are more divergent than the isolates described until now in this family. Since the human isolate Ant70 is
also divergent to the same extent within this family, this indicates that the HIIV-1 family might be more variable than
previously appreciated.
VANDEN HAESEVELDE Marleen - Industriepark Zwijnaarde 7 b4 - 9052 GENT -
BELGIUM- (+32) 91/410.711
WeA 1083 THE DEVELOPMENT OF A HUMAN IMMUNODEFICIENCY VIRUS
QUASISPECIES IN VIVO
Cichutek. Klaus*, Merget, H.*, Norley, S.*, Linde, R.#, Kreutz, W.#, Gahr,
M." and Kurth, R.* *Paul-Ehrlich-Institute, D-6070 Langen, Germany,
#University Clinics, Frankfurt/Main,'University Children's Clinic, G6ttingen
Objectives: To analyse the quasispecies of human immunodeficiency virus type 1 (HIV-1) present in
peripheral blood mononuclear cells (PBMC) of hemophiliacs probably infected from the same source.
Methods: HIV-1 was isolated from PBMC by cocultivation with negative donor chord blood
lymphocytes (CBL). Characterization of the env gene regions extending from variable region V3 to V4
was performed by nested PCR (polymerase chain reaction) amplification starting from PBMC or CBL
DNA, molecular cloning and subsequent DNA sequencing of up to 20 subgenomic clones.
Results: During treatment with one specific batch of blood clotting factor IX, a number of
hemophilia B patients were infected in Germany with a clonal HIV-1 substrain belonging to the NorthAmerican/European group. The nucleotide sequences of cloned env gene regions between the variable
V3-loop and variable V4 region analysed from cultured virus and from virus in the peripheral blood cell
DNA of two different patients 5 months post infection and one of the patients 11 months post infection
were shown to be highly homologous. Based on the assumption that the consensus sequence (termed
HIV-1MBK) was identical with the genotype of the initially infecting virus we were able to construct
phylogenetic trees of the quasispecies of both patients. True intermediate genotypes between input and
multiply mutated genotypes were detected by 5 and 11 months post infection and genetic variation was
shown to develop at a faster rate in one of the patients. Except for the HIV-1MBK genotype, the vast
majority of all variants detected 5 months post infection in the blood of the other patient were replaced
with new variants 11 months post infection. Variability was shown to reside in two small regions located
3' of the V3-loop and within the V4-region. Overlapping synthetic peptides corresponding to the entire
sequenced region, including mutants, were produced and the antibody reactivity in patients sera tested by
ELISA. Linear epitopes were found only within the V3 loop and fine mapping showed the sequence
SINIGP to be the immunodominant epitope. The affinity of antibody to the only mutant found in this
epitope (SINTGP) was reduced in comparison to the consensus.
Conclusion: This is the first report of the evolution of an HIV-1 quasispecies starting from a single and
still detectable genotype.
Dr. Klaus Cichutek, Paul-Ehrlich-Institute, Paul-Ehrlich-Str. 51-59, D-6070
Langen, Germany, Tel. x-49-6103-7550, Fax x-49-6103-755-123.

WeA 1 080 GENETICALLY-DIVERSE. SIV,,-RELATED HIV-2 IN
WEST AFRICA: EVIDENCE FOR ZOONOTIC INFECTION. Gao, F., Yue, L', White,
A.', Pappas, P., Barchue, J., Hanson, A., Sharp, P.", Shaw, G." and Hahn, Beatrice'.
University of Alabama at Birmingham, Birmingham, Alabama, USA, "Liberian Institute
for Biomedical Research, "Trinity College, Dublin, Ireland.
Objectives: Current understanding of the natural history and phylogeny of Human Immunodeficiency Virus
Type 2 (HIV-2) derives from studies of culture-amplified viruses from urban populations experiencing epidemic
spread of infection and disease. Since tissue culture is highly selective for viral strains with an in vitro growth
advantage, we hypothesized that such isolates may represent only a subset of a much larger and genetically
more diverse group of viruses.
Methods: Blood samples from two HIV-2 seropositive, healthy Liberian agricultural workers (F0784, 2238) and
a symptomatic urban dweller from Cote dlvoire (7312A) were processed for virus culture and PCR analysis.
Peripheral blood mononuclear cells were cultured alone, in combination with normal donor lymphocytes and
macrophages, and with immortalized T-cell lines. HIV-2 sequences were amplified directly from uncultured
PBMC DNA using nested PCR. Evolutionary trees were constructed using the neighbor-joining method.
Results: Using nested PCR, HIV-2 sequences were identified in all three individuals. Viral isolates, however,
were obtained only from subject 7312A but not from subjects F0784 and 2238, despite several attempts.
Sequence analysis of PCR derived pol, env, and LTR regions revealed an unexpected degree of genotypic
variation, with up to 23% sequence differences in regions where most HIV-2 viruses differ by less than 10%.
Subsequent phylogenetic analysis identified one virus (HIV-2,0o.) to be significantly more closely related to
simian immunodeficiency viruses infecting sooty mangabeys and rhesus macaques than to any virus of human
derivation, a second virus (HIV-2,) to be most closely related to the previously reported highly divergent
HIV-2,. strain, and a third virus (HIV-2,,) to represent a likely intergenic recombinant. In addition, one
subject (F0784) was found to harbor multiply-defective viral genomes that resulted from G to A hypermutation.
Conclusions: These results indicate that HIV-2 in man and SIV in mangabeys and captive macaques
represent members of a single, genetically-diverse group of viruses which cannot be separated into distinct
phylogenetic lineages according to species of origin. Although the evolutionary origins and transmission
patterns of this virus group remain to be defined, there is mounting evidence that the sooty mangabey is a
natural reservoir and that human infections may represent a zoonosis.
Hahn, Beatrice. University of Alabama at Birmingham, 701 S 19th Street,
Birmingham, AL 35294, USA Telephone: [phone], FAX [phone].
WeA 1082 PHENOTYPE ASSOCIATED SEQUENCE VARIATION IN THE THIRD VARIABLE DOMAIN
OF THE HIV-1 gpl20 MOLECULE. Fouchier. Ron A.M.; Groenink, M.; Kootstra, N.A.;
Tersmette, M.; Huisman, H.G.; Miedema, F.; Schuitemaker H. Central Laboratory of the
Netherlands Red Cross Blood Transfusion Service and Laboratory of Experimental and Clinical
Immunology of the University of Amsterdam, Amsterdam, The Netherlands.
Objectives: The third variable (V3) domain of HIV-1 gpl20 elicits type specific neutralizing antibodies,
and contains a type specific T-cell epitope. V3 is critical for gpl20 function, and has been implied in
determining the virus phenotype including fusion capacity and monocytotropism. Fusion capacity of virus
isolates has been shown usefull as a prognostic marker for disease progression. Comparison of small
numbers of isolates have not provided insight as how the high degree of sequence variability may give
rise to large subgroups of HIV-1 strains.
Methods: PCR amplification of V3 domains from a large panel (60 isolates from 30 infected persons)
of primary isolates with well defined syncytium inducing capacity and monocytotropism was performed,
followed by direct sequencing of the PCR products.
Results: Sequence analysis of V3 domains from virus isolates with well defined biological phenotypes
revealed that fast-replicating, syncytium-inducing (SI)/non-monocytotropic isolates contained V3
sequences with a significantly higher positive charge than those of slow-replicating, non-syncytiuminducing (NSI)/monocytotropic isolates. On either side of the loop, midway the cysteine and the central
GPG motif, a highly variable amino acid residue is located that was negatively charged or uncharged in
NSI/monocytotropic isolates whereas in SI/non-monocytotropic isolates either one or both were positively
charged. These substitutions result in changes in predicted secondary structure of the V3 domain. Site
directed mutagenesis experiments of the critical amino acid residues are now in progress to formerly
prove the functional relevance of our findings.
Conclusions: Our data indicate that the V3 domain is an important determinant for the biological
phenotype of HIV-1 isolates and further suggest that an efficacious AIDS vaccine conferring protection
to the relevant HIV-1 variants should include V3 sequences specific for NSI/monocytotropic in addition
to the now generally used SI/non-monocytotropic isolates.
Fouchier, Ron A. M., Central Lab. of the Neth. Red Cross Blood Transfusion Service and Lab.
of Experimental and Clinical Immunology of the University of Amsterdam. Plesmanlaan 125, 1066
CX Amsterdam, The Netherlands. Telephone [phone]; FAX [phone].
VWeA 1084 SIMPLLIFIED A'PPROACHES FOR DIAGNOSIS AND TREATMENT OF
ST[) IN AFRICA.
Ndoye__*, Sakho ML*, Tardi M**, Diaw I**, Mbacke KD**,
Sarr LC*, Mboup S*. *: Comite SIDA Senegal, **: Centre
MST d(l 1'IHS, Dakar, Senegal.
Objectifs: To improve the accessibility of diagnosis and treatment
of STD.
Methods: 1. Study on the prevalence of STD, including the
importance of antimicrobial resistance.
2. Study on the knowledge and feasibility of algorithms
for diagnosis of STD.
3. Cost-efficacy study on the treatment of STD using
drugs from the essential drug programme.
Results: 1. High STD prevalences were found among the general
population and among high risk groups, such as prostitutes.
2. lorithms performed best for genital discharge and
genital ulcer.
3. Treatment with essential drugs was successful in
most cases.
Clinical algorithms on 1) genital discharge in men andy
women and 2) on genital ulcer in men and women have been developed and
are now used at the primary and secondary health care level.
Conclusions: Such a study is necessary and must be conducted in all
developing countries experiencing difficulties in establishing a
proper diagnosis and an affordable treatment for STD. Using clinical
algorithms, the health worker at the primary level can cope with more
than 60% of the STD in developing countries.

Dr NDOYE IBRA
BP: 3435 - Dakar/SENEGAL
Tel: [phone]

Fax: [phone]

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