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

Tuesday, July 9, 1996

Tu.A. 100 - Tu.A.104

Tu.A. 100
FIRST FULL LENGTH SEQUENCE PROTOTYPES OF HIV- I CLADES C, E AND G:
TWO OF THREE ARE MOSAIC GENOMES.
Carr ean K., Salminen, M.O., Leinikki, P2, Johansson, B.3, Burke, D.S.4, tic' rItc an, FE..
' Henry M. Jackson Foundation, Rockville, MD,USA2,National Pubic leaith institute,
Helsinki, Finland,3Huddinge Central Hospital, Huddinge, Sweden rod Valte ir Reed Army
Institute of Research, Rockville, MD, USA.
Objective: To obtain the initial prototypic full length genomic sequences of HIV I isolates of
clades C, E and G.
Methods: DNA from primary virus cultures on donor peripheral i ricirrinrnuliear cells
from HIV- I infections acquired in Ethiopia (C2220, clade C),Thalarnd (CMz'0, rIlde E) and
Kenya (HH8793, clade G) was isolated and used as template for Iong I'R amplification of
virtually full length HIV- I genomes. PCR products were molecularly clr'rI) a f. ully
sequenced using fluorescent dye terminators and an Applied Biosystems rtrrrrrcaled
sequencer. An alignment was built, segmented and the segments analysed usin mraximnum
likelihood and maximum parsimony.
Results: The full length sequence of the Thai and Kenyan viruses revealed i ' to e
mosaics between lade A and lade E, on the one hand, and lade A arI ilde G, on the
otherTheir structure is diagrammed below:
The Thai virus is predominantly clade A(grey) except for the extracellular portion of env
and the U3 region of the LTR.The kenyan virus is predominantly G (white) except for vif
and vpr, and the internal portion of env. It appears that both viruses have con irmon breakpoints at the 3' end of the viral RNA.The clade C virus from Ethiopia was not apparently
recombinant and had the same genomic organization as that of the previors y sequenced
isolates of clades A,B and D. However, the C virus has three NF-kB binding sites in the core
promoter nstead of the usual two, and theThai virus has only one.
Conclusion:The only full length sequences of clades E and G sugg t lit triise 'cslades' are
recombinant.
JK Cart 1600 East Gude Dr., Rockville, MD 20850 USA Telephone: [phone]!9
Fax [phone] email: jcarhiv.hjf.org
Tu.A.101
DISTINCT HIV-I SUBTYPES ASSOCIATED WITH DIFFERENT RISK GROUPS IN
SOUTH AFRICA
Williamson Carolyn***, Engelbrecht S***, van Harmelen J**, van Rensburg EJ**, Bredell
W***,Wood R*". *SAIMR, **University of Cape Town and *"*University orf Sillenbosch
South Africa.
Objectives: To determine HIV I subtypes in different risk groups in Cape Town. South Africa.
Methods: DNA was isolated from blood drawn from 84 patients attending 'oal clinics.
Samples were divided into 4 groups according to presumed mode of transmission: homosexual/bisexual (n=37), heterosexual/vertical (n=42), blood transfusion (n=2), and mode of
transmission unknown (n-3). Proviral DNA was subtyped by HMA based on the 700 bp
V3-V5 region of the env gene (47 samples) and/or by sequence analysis of the vie gene
(500 bp region encoding the p 17 protein) (54 samples). Phylogenetic tier's wet e generated
using both neighbour-joining and maximum parsimony approaches.
Results: Subtypes B, C, D and E were identified. Subtypes were fc'rndl to sere-ate accord
ing to mode of transmission, with subtype B viruses found in 86 % (32/37) of the homosexual/bisexual group and subtype C viruses found in 83% (35/42) of the helerosexual/vertical
transmission group. Subtype B viruses were also found in 5 heterosexual patients, 2 patient
infected by blood transfusion and in 3 patients where the mode of transmission was
unknown. Subtype D viruses were found in 5 homosexual patients anId one heterosexual
patient. A subtype E virus was identified in a heterosexual patients No discrepancy was
found in subtype designation between samples analysed in the gig and env region (n- 19).
Conclusions: The intial epidemic in South Africa in the early 1980s affected r i-t nly the
homosexual population and shifted to a predominantly heterosexual epidemic by the late
1980s. In this study, subtype B viruses were associated with homosexual tranrmi5sion and
subtype C viruses with heterosexual transmission, suggesting two independent epidemics.
C.Williamson, Department of Medical Microbiology University of Cape Town Medical
School, Anzio Road, Observatory 7925, South Africa. Tel: [phone]. Fax: 21 '148 4 110.
email: [email protected]
Tu.A. 102
COMPARISON OF HIV-I SEQUENCES FROM 1981 WITH MORE CURRENT
ISOLATES IN THE UNITED STATES EPIDEMIC
Robbins, Kenneth E. Jaffe HW, Schable CA, Brown TM, Rapier JM, Henes TE, Sch ichetmnan
G, Kalish ML. Centers for Disease Control and Prevention, Atlanta, CA.
Objective: In 1981, specimens were obtained from homosexual men by the.centers for
Disease Control(CDC) task force, formed to investigate increases in opportunistic infections among homosexual men in the U.S.We successfully used seven of these samples to
determine the DNA sequences of two regions of the HIV- I genome.- heI liearly
sequences were compared with other U.S. subtype B sequences to aid i" rhe riderstanding
of the evolving genetic variability of HIV- I in the United States.
Methods: The date of collection of the samples was September to NoviL n'r of 1981,
from patients residing in Georgia, NewYork, New Jersey and California. Both the p1 7 coding region of the gag gene and a 345 b.p. C2-V3 region of the en gene were vequenced
directly from their PCR products by dye primer and dye terminator chenistris sing an
automated sequencer Computer genetic analysis programs (MEGA. PHYLIP GDE) were
used to construct nucleotide and amino acid alignments, genetic distaise mat ices. and phylogenetic trees to compare these with U.S. subtype B sequences fro' the risid 980's (Los
Alamos HIV database) and 1995 (current CDC sequencing project).
Results: Nucleotide genetic distance differences for the C2-V3 reic, r w v' 7% ( 98 I)
9% (mid 1980's), and 13.2% (1995).The pl 7 region also increased n dvergence from 1981
(2.1%) to 1995 (5.3%). All strains in the amino acid alignment have the typorl iorth
American) GPGR tetrapeptide at the tip of the V3 loop and none of the 198 I or 1995
sequences have the genotype associated with the more virulent, syncium inducing phenotype. Phylogenetic analysis demonstrated shorter branch lengths for the 1981 sequences
and no defined group clustering by date of collection.

Conclusions: I he Los Alamos National Labrtoratory has estimated a 0.5- I%/y evolutionary
rate of cihange for the V3 region of the env gene of I lV-I.The low inter person nucleotide
divergence (5.2%) of these I198 1 sequences is consistent with this rate and the recent
nature ofi the HIV epidemic. The shorter brarnch lengths in the tree analysis indicate an earlier evrlutionary or igin frorn contemporariy chide B sequences, yet the lack of early strain
cluster ing needs to be explored further with additional sequences from this time period.
Kenneth E. Robbins CDC Mail stop D-12 Alainta, GA 30333, USA.
email: Ker2n ciddas I.er.cdc.gov Telephone: [phone] FAX: [phone]
Tu.A. 103
HIV- I DIVERSITY IN PATIENTS FROM RIO DE JANEIRO, BRAZIL
Mo gdoaiza GI, Guirnaries, MI, Grilpp, CBG., Costa, CII,Santos,VGV, LinharesCarvalho, MlBastos, FI2, Galvio Castro, B, Castilhof EA2/5, Bongertz,V. Inst Oswaldo
Cruz I & DIS/CICt2, FIOCRUZ, RJ; Amb Bco da Pr evidencia. RI; L ASP/CPqGM4, FIOCRUZ,
B; S1TD/AIDS/BI Ministry o f I Iealth5, Braslia, BR.
Objective: It'eeH IV Isubtypes, B, F 1io C have bee fund in Br azil up now, in addition
to a recombinant B/ I yenouie, with a clear predortinarce of subtype B isolates. Moreover:
if so, stir ies i sI'' also shovi tiat many BranIi-in subtype B isolates present a typical
armino eacid cc position (GWGR) at the conserved crown of the gp 120 V3 loop, suggest
ing tiat subtype B isolates could be split in two main groups, one corresponding to the
IU SA/Eur pe. consensus B sequence and a se icond one, called B', typically found in Brazil.
The airm of this work is to follow IlIV I diversity ii Rio de Janeiro City anrid try to correlate
the subtype dItr ibution with exposure categories.
Methods: 11V!sramples analyzed up to niow were obtai ined frot positive in divi duals fromr
two cohorts followed at medical centers in Rio de Janeiro, BR (Evandro Chagas HospFICt 1 a d A bulatn corsensus B sequence and a second one, called B', typically found
in Brazt center s or in the streets. I)NA sramples were PR ampified by a nested protocol
using an outer primer set whicht amplifies a 2.0Kb fragmnent spanning from the first exon of
RIV p t tifIe ti ansnerembrane protein gpi4 cciding region of ENV. For the 2nd round
PR, the inner prinner sets covering respectively the VI -V5 or C2- C3 regions of the FHIV I
FNV were used. I1IIV I subtyping was determinate by heteroduplex mobility assay as
described elsewhere (Delwart et al.1993, Scienc e 262:1257 61).
Results: We have aiialyzed 1102 individuals, 57 men and 45 women, which were distributed
by the exp r n) i'ategories as follows: I/ nhomo sexual & 10 bisexual men; 9 heterosexuals
(8 inen), 16 I)Us (one woman) and 10 without available information. From the eighty one
H IV I sanples typed until now, 73 (90%) wetre included as subtype B and 8 (I 0%) as subtype F. to sinifcrant statistical association (Fisher's exact test) was identified cross coi par
ing IIV I sultypes with exposure categories, genres and transmission routes. Discriminatioi
between B and B' patterns is under evaluation.
Conclusions: No segregation between HIV I subtypes and exposure categories was identi
fied in Rio di Jin areiro. A tendency of increasing the frequency of the HIV-I F subtype was
observed when compared to our previous results (Morgado et ail. 1994 AIDS Res Hum
Rer, 10:569 576), where only one F subtype was identified among 18 DNA samples
obtained f1r an,yrnptomatic individuals from 1')990 to 1992, although with no sistatistical significance, possibly due to the small samnple size evaluated up now (SupIportnd by
W -O/GPAIA/VI )Ul, PIAF/FIO(CRUZ and UNDP/World BankPN-DST/AIDS, BR.)
Mariza.oclves M"lorgado-IDept. of Immunrology Institutlo Oswaldo Cruz FIOCRUZ Av.
Brasil 43n,5 Mvin 'nIios RJ CFP210t5 Brazil 1tel1:55 21 2801t46
Tu.A. 104
ENVELOPE SEQUENCES AND BIOLOGICAL CHARACTERIZATION OF
HIV-I SUBTYPE E ISOLATES FROM THAILAND
Auewarak i aent*, McI ane MF' Wasii -*,1ssex M'. *'IIarvard AIDS Institute, Boston,
USA;: MPrahidolUniversity, Bangkok, Thailarrd.
Objective:. o btain HIV I subtype E isolates fromn heterosexually infected individuals in
Thailand and to analyze their envelope sequences and biological properties.
Methods: s I I were isolated by co cultivation of peripheral blood mnononuclear cell
(PBM(-) fino nrinfected pregnant womern with donor PBMC.1he infected individuals had no
history of intiraven ous drug use or blood transflusion. DNAs were prepared from the cultur ed PBMC, and n ested PCR were performed to obtain 2.8 kb fragment covering the
whole op160 ensn'lope gene. Ihe fiagrments were cloned into PCR 11 vector and sequenced.
(crowth kinetics in PBMC, syncytium in duction in M 1-2 cell, and neutralization or enhancing
activity of the isolates by homologous plasma weret udied.
Results: Four HIVSI subtype E isolates and their comnplete envelope sequences were arnavzed. Of these, two isolates grew to high liter in PBMC (#106, '149), one of which induced
syrcytia in MT 2 (#449). Infectivity of this isolate in PBIC was enhanced, whereas that of
the other thmree were neutralized by homologous plasma. [he three isolates (#401, 406,
1 8) with non-syrncytia inducing (NSI) phenotype had GPGQ amino acid sequence at the
tip ofV3 icp, iwhereas the one with syncyltia inducing (SI) fphenotype had GPGR.The isolates that crew only to low titer in PBMC (#401,4 18) carried an extra N-linked glycosylaioi.ite in th' V3 oop. he V3 loop of the isolates with high tiler growth (#406, 4,t9)
showed hign nt positive charte. 1w isolates (06, 4 18) carried an extra disulfide bridgne
in Vi.1e enelopes are being subcloned intoa IIXB2 background to confirm the phenotype of the loes.
Conclusion: cPR motif, lac of extra N-linked glycosylation site and net positive charge in
Vi oo o,-, relsl"d with igh titer growth aind SI phenotype irn the subtype E envelope
clones anayzed.
P Auewarakui Cancer Biology i Harvard School of Public I-Health, 665 Huntington Ave,
Boton, MA )i o, USA.Telephone: 617. 21302 Fax: 617 7d9818 email.
ivew l I i.invoild.edu

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