Following the identification of the C-C chemokines RANTES, MIP-1alpha and MIP-1beta as major human immunodeficiency virus (HIV)-suppressive factors produced by CD8+ T cells, several chemokine receptors were found to serve as membrane co-receptors for primate immunodeficiency lentiretroviruses. The two most widely used co-receptors thus far recognized, CCR5 and CXCR4, are expressed by both activated T lymphocytes and mononuclear phagocytes. CCR5, a specific RANTES, MIP-1alpha and MIP-1 receptor, is used preferentially by non-MT2-tropic HIV-1 and HIV-2 strains and by simian immunodeficiency virus (SIV), whereas CXCR4, a receptor for the C-X-C chemokine SDF-1, is used by MT2-tropic HIV-1 and HIV-2, but not by SIV. Other receptors with a more restricted cellular distribution, such as CCR2b, CCR3 and STRL33, can also function as co-receptors for selected viral isolates. The third variable region (V3) of the gp120 envelope glycoprotein of HIV-1 has been fingered as a critical determinant of the co-receptor choice. Here, we document a consistent pattern of evolution of viral co-receptor usage and sensitivity to chemokine-mediated suppression in a longitudinal follow-up of children with progressive HIV-1 infection. Viral isolates obtained during the asymptomatic stages generally used only CCR5 as a co-receptor and were inhibited by RANTES, MIP-1alpha and MIP-1beta, but not by SDF-1. By contrast, the majority of the isolates derived after the progression of the disease were resistant to C-C chemokines, having acquired the ability to use CXCR4 and, in some cases, CCR3, while gradually losing CCR5 usage. Surprisingly, most of these isolates were also insensitive to SDF-1, even when used in combination with RANTES. An early acquisition of CXCR4 usage predicted a poor prognosis. In children who progressed to AIDS without a shift to CXCR4 usage, all the sequential isolates were CCR5-dependent but showed a reduced sensitivity to C-C chemokines. Discrete changes in the V3 domain of gp120 were associated with the loss of sensitivity to C-C chemokines and the shift in co-receptor usage. These results suggest an adaptive evolution of HIV-1 in vivo, leading to escape from the control of the antiviral C-C chemokines.

The biological phenotype of primary human immunodeficiency virus type 1 (HIV-1) isolates varies according to the severity of the HIV infection. Here we show that the two previously described groups of rapid/high, syncytium-inducing (SI) and slow/low, non-syncytium-inducing (NSI) isolates are distinguished by their ability to utilize different chemokine receptors for entry into target cells. Recent studies have identified the C-X-C chemokine receptor CXCR4 (also named fusin or Lestr) and the CC chemokine receptor CCR5 as the principal entry cofactors for T-cell-line-tropic and non-T-cell-line-tropic HIV-1, respectively. Using U87.CD4 glioma cell lines, stably expressing the chemokine receptor CCR1, CCR2b, CCR3, CCR5, or CXCR4, we have tested chemokine receptor specificity for a panel of genetically diverse envelope glycoprotein genes cloned from primary HIV-1 isolates and have found that receptor usage was closely associated with the biological phenotype of the virus isolate but not the genetic subtype. We have also analyzed a panel of 36 well-characterized primary HIV-1 isolates for syncytium induction and replication in the same series of cell lines. Infection by slow/low viruses was restricted to cells expressing CCR5, whereas rapid/high viruses could use a variety of chemokine receptors. In addition to the regular use of CXCR4, many rapid/high viruses used CCR5 and some also used CCR3 and CCR2b. Progressive HIV-1 infection is characterized by the emergence of viruses resistant to inhibition by ␤-chemokines, which corresponded to changes in coreceptor usage. The broadening of the host range may even enable the use of uncharacterized coreceptors, in that two isolates from immunodeficient patients infected the parental U87.CD4 cell line lacking any engineered coreceptor. Two primary isolates with multiple coreceptor usage were shown to consist of mixed populations, one with a narrow host range using CCR5 only and the other with a broad host range using CCR3, CCR5, or CXCR4, similar to the original population. The results show that all 36 primary HIV-1 isolates induce syncytia, provided that target cells carry the particular coreceptor required by the virus.

HIV-1 uses chemokine coreceptors for cell entry. CXCR4 is the major coreceptor for T-cell-line-adapted isolates and CCR5 for non-T-cell-line-adapted isolates. This study investigated if coreceptor usage differs between genetic subtypes of HIV-1. Eighty-one primary isolates representing nine different genetic subtypes (A-J, except I) were tested on U87.CD4 glioma cells stably expressing chemokine receptor CCR1, CCR2b, CCR3, CCR5, or CXCR4. Coreceptor usage was compared to biological phenotype of the isolates (rapid/high, syncytium-inducing or slow/low, non-syncytium-inducing) and to clinical and immunological status of the study subjects. CXCR4 usage was perfectly correlated to the biological phenotype for all subtypes; all of 26 isolates with rapid/high phenotype and none of 55 isolates with slow/low phenotype could infect the CXCR4 expressing cell line. Importantly, the CXCR4-positive, rapid/high phenotype was underrepresented among subtype C isolates. Furthermore, dual tropism for CXCR4 and CCR5 was not found among subtype D isolates. Uni- and multivariate analyses indicated that these subtype-specific differences in coreceptor usage were not due to differences in clinical status, CD4 counts, or treatment. This study shows that CXCR4 usage determines the biological phenotype for all subtypes, but that there appear to exist subtype-dependent differences in frequency of usage of certain coreceptors. This opens up the possibility that genetic subtypes may differ in important biological properties such as virulence, tissue tropism, and transmissibility.

Coreceptor usage of primary human immunodeficiency virus type 1 (HIV-1) isolates varies according to biological phenotype. The chemokine receptors CCR5 and CXCR4 are the major coreceptors that, together with CD4, govern HIV-1 entry into cells. Since CXCR4 usage determines the biological phenotype for HIV-1 isolates and is more frequent in patients with immunodeficiency, it may serve as a marker for viral virulence. This possibility prompted us to study coreceptor usage by HIV-2, known to be less pathogenic than HIV-1. We tested 11 primary HIV-2 isolates for coreceptor usage in human cell lines: U87 glioma cells, stably expressing CD4 and the chemokine receptor CCR1, CCR2b, CCR3, CCR5, or CXCR4, and GHOST(3) osteosarcoma cells, coexpressing CD4 and CCR5, CXCR4, or the orphan receptor Bonzo or BOB. The indicator cells were infected by cocultivation with virus-producing peripheral blood mononuclear cells and by cell-free virus. Our results show that 10 of 11 HIV-2 isolates were able to efficiently use CCR5. In contrast, only two isolates, both from patients with advanced disease, used CXCR4 efficiently. These two isolates also promptly induced syncytia in MT-2 cells, a pattern described for HIV-1 isolates that use CXCR4. Unlike HIV-1, many of the HIV-2 isolates were promiscuous in their coreceptor usage in that they were able to use, apart from CCR5, one or more of the CCR1, CCR2b, CCR3, and BOB coreceptors. Another difference between HIV-1 and HIV-2 was that the ability to replicate in MT-2 cells appeared to be a general property of HIV-2 isolates. Based on BOB mRNA expression in MT-2 cells and the ability of our panel of HIV-2 isolates to use BOB, we suggest that HIV-2 can use BOB when entering MT-2 cells. The results indicate no obvious link between viral virulence and the ability to use a multitude of coreceptors.

It has been estimated that, to date, about 48% of all HIV-infected people in the world carry HIV-1 subtype C virus. Therefore, it is of great importance to gain better knowledge about the genetic and biological characteristics of this virus subtype. In the present study, the biological properties of HIV-1 isolates obtained from nine Ethiopian patients with AIDS were studied. DNA sequencing of the V3 loop of gp120 classified the isolates as subtype C. In primary isolation cultures, virus infection was accompanied by syncytium formation and cell lysis. Interestingly, when examining the growth in primary monocyte-macrophage cultures, initial low-level virus replication was followed by a nonproductive state, from which virus could be rescued by cocultivation with Jurkat(tat) cells. Furthermore, none of the isolates replicated in T cell lines (CEM, MT-2, HuT-78, and H9) or in the promonocytic cell line U937 clone 2. All isolates could use CCR5 as coreceptor, whereas no isolates could use CCR2b, CCR3, CCR5, CXCR4, Bonzo/STRL33, or BOB/GPR15. The genotype of the V3 region correlated with the MT-2 negative/non-syncytium-inducing (NSI) phenotype. Comparative studies revealed that the scarcity of CXCR4 usage as well as other phenotypic characteristics of subtype C isolates distinguish this subtype. On the basis of these data, we suggest that in addition, factors other than viral phenotype may govern the pathogenic potential of subtype C isolates.

The V3 region of the human immunodeficiency virus type 1 envelope protein gp120 constitutes a potential neutralization target, but the oligosaccharide of one conserved N-glycosylation site in this region protects it from neutralizing antibodies. Here, we determined whether N-linked glycans of other gp120 domains were also involved in protection of V3 neutralization epitopes. Two molecular clones of HIV-1, one lacking three N-linked glycans of the V1 region (HIV-1(3N/V1)) and another lacking three N-linked glycans of the C2 region (HIV-1(3N/C2)), were created and characterized. gp120 from both mutated viral clones had higher electrophoretic mobilities than gp120 from wild-type virus, confirming loss of N-linked glycans. Wild-type virus and both mutant clones replicated equally well in established T cell lines and all three viruses were able to utilize CXCR4 but not CCR5 as a coreceptor. The induced mutations increased gp120 affinity for CXCR4 but caused no corresponding increase in viral ability to replicate in T cell lines. HIV-1(3N/V1) was neutralized at about 25 times lower concentrations of an antibody to the V3 region than were wild-type virus and HIV-1(3N/C2). Soluble, monomeric gp120 from HIV-1(3N/V1) and wild type virus had identical avidity for the V3 antibody, indicating that the V1 glycans were able to shield V3 only in oligomeric but not monomeric gp120. In conclusion, one or more N-linked glycans of gp120 V1 is engaged in protection of the V3 region from potential neutralizing antibodies, and this effect is dependent on the oligomeric organization of gp120/gp41.

Neutralizing activity against primary human immunodeficiency virus type 1 (HIV-1) isolates from 17 persons who were long-term disease nonprogressors (LTNPs) and 13 persons who were fast progressors (FPs) was compared. Sera from LTNPs showed higher neutralizing activity both in titer and in host spectrum than did sera from FPs. However, LTNP sera had limited neutralizing activity against HIV-1 subtypes from different geographic areas. Sera collected 6 years earlier from both groups had limited neutralizing activity, indicating that early responses are not predictive for disease progression. LTNPs had very low virus loads, as reflected by only one positive isolation, which was an MT-2-negative phenotype. Virus was isolated from all FPs, and the isolates showed a phenotype switch from MT-2 negative to MT-2 positive. Development of high-titer, broadly cross-reactive neutralizing antibodies is associated with control of virus replication and low virus load in HIV-1-infected LTNPs.

We evaluated the performance of 11 SARS-CoV-2 antibody tests using a reference set of heat-inactivated samples from 278 unexposed persons and 258 COVID-19 patients, some of whom contributed serial samples. The reference set included samples with a variation in SARS-CoV-2 IgG antibody titers, as determined by an in-house immunofluorescence assay (IFA). The five evaluated rapid diagnostic tests had a specificity of 99.0% and a sensitivity that ranged from 56.3 to 81.6% and decreased with low IFA IgG titers. The specificity was > 99% for five out of six platform-based tests, and when assessed using samples collected ≥ 22 days after symptom onset, two assays had a sensitivity of > 96%. These two assays also detected samples with low IFA titers more frequently than the other assays. In conclusion, the evaluated antibody tests showed a heterogeneity in their performances and only a few tests performed well with samples having low IFA IgG titers, an important aspect for diagnostics and epidemiological investigations.