Phenotypic Heterogeneity ina PanelofInfectious Molecular HumanImmunodeficiency Virus Type1Clones Derived froma Single Individual

Previously, we andothers havedemonstrated a relation between theclinical courseofhumanimmunodeficiency virus type1(HIV-1) infection andbiological properties ofHIV-1variants suchasreplication rate, syncytium-inducing (SI) capacity, andcytotropism. Forthemolecular analysis ofthebiological variability in these properties, we generated a panel ofphenotypically distinct yetgenetically highly homologous infectious molecular clones. Theseclones were derived fromHIV-1isolates, mostly recovered bydirect clonal isolation, froma single individual inwhoma transition fromnon-SI toSIisolates hadbeenidentified overtime. Of17 molecular clones tested, 8 were infectious. Theclones exhibited differences inSIcapacity andT-cell line tropism. Theirphenotypes corresponded tothose oftheir parental isolates, formally demonstrating that biological variability ofHIV-1isolates can beattributed tosingle molecular clones. Withthese clones we demonstrated thatSIcapacity andtropism fortheH9T-cell line, almost invariably coupled inprimary HIV-1 isolates, arediscernible properties. Alsodifferent requirements appeared toexist forH9andSupTlcell line tropism. Weobtained evidence that T-cell line tropism isnotcaused bydifferences inlevel ofHIV-1expression butmostprobably isrestricted atthelevel ofvirus entry. Restriction mapping offourclones withdivergent phenotypes revealed ahighdegree ofnucleotide sequencehomology (over 96.3%), indicating theusefulness of these clones forthetracking ofgenetic variability critical fordifferences inbiological phenotype. Inrecent years,increasing attention hasbeengiven tothe biological variability ofhumanimmunodeficiency virus type 1 (HIV-1) isolates andthepossible importance ofthis variability forthepathogenesis ofAIDS (2,9,34).In previous longitudinal studies, we demonstrated thatthe majority ofHIV-1isolates recovered fromasymptomatic individuals arenon-syncytium inducing (NSI)inperipheral bloodlymphocyte (PBL)culture anddonotreplicate inthe H9 T-cell line. Instable asymptomatic persons,onlyNSI HIV-1isolates withlowreplication rateswere found. In contrast, theemergence ofsyncytium-inducing (SI), H9tropic viruses invariably heralded a rapid lossofCD4+T cells andprogression toAIDSwithin 1/2to2years(35, 36). Recent reports demonstrating amuchhigher frequency of infected peripheral bloodleukocytes ininfected individuals thanestimated previously havemadeCD4+T-cell depletion causedbydirect viral killing ofCD4+cells more plausible (17,29,32).Indeed, theinvitro properties ofSIvariants (syncytium induction, highinfectivity, andbroadhostrange) allcouldvery wellcontribute toeffective CD4+ T-cell depletion invivo. Ontheother hand, ourfindings thatonly NSIvariants arefoundinstable asymptomatic individuals andthat, compared withSIvariants, these viruses aremuch more monocytotropic indicate thatNSIvariants areimportantforthemaintenance ofHIV-1infection, putatively becauseofan increased capacity toevadetheimmune system(30). Sincebiological variability ofHIV-1isolates thus may well beofdirect pathogenetic relevance, molecular analysis to