No abstract

Change of HIV-1 coreceptor use has been connected to progression of disease in children infected with HIV-1, presumably subtype B. It has not been possible to discern whether the appearance of new viral phenotypes precedes disease development or comes as a consequence of it. We studied the evolution of coreceptor use in HIV-1 isolates from 24 vertically infected children. Their clinical, virological, and immunological status was recorded and the env V3 subtype was determined by DNA sequencing. Coreceptor use was tested on human cell lines, expressing CD4 together with CCR5, CXCR4, and other chemokine receptors. The children carried five different env subtypes (nine A, five B, four C, three D, and one G) and one circulating recombinant form, CRF01_AE (n = 2). Of the 143 isolates, 86 originated from peripheral blood mononuclear cells (PBMCs) and 57 originated from plasma, received at 90 time points. In 52 of 54 paired plasma and PBMC isolates the coreceptor use was concordant. All 74 isolates obtained at 41 time points during the first year of life used CCR5. A change from use of CCR5 to use of CXCR4 occurred in four children infected with subtype A, D, or CRF01_AE after they had reached 1.5 to 5.8 years of age. There was a significant association with decreased CD4+ cell levels and severity of disease but, interestingly, the coreceptor change appeared months or even years after the beginning of the immunological deterioration. Thus CXCR4-using virus may emerge as a possible consequence of immune deficiency. The results provide new insights into AIDS development in children.

Human immunodeficiency virus type 1 (HIV-1) enters cells through the chemokine receptors CCR5 (R5 virus) and/or CXCR4 (X4 virus). Loss of N-linked glycans and increased net charge of the third variable loop (V3) of the gp120 envelope glycoprotein have been observed to be important steps towards CXCR4 use. All reported sequences using CCR5 or CXCR4 exclusively, or using both, were gathered from the Los Alamos HIV Database and analysed with regard to the V3 N-linked glycosylation motifs (sequons) and charge. The V3 loop glycan had a sensitivity of 0?98 and a 0?92 positive predictive value in the context of CCR5 use. The difference from X4 was remarkable (P<10 "12). Especially, the sequon motif NNT within the V3 loop was conserved in 99?2 % of the major clades. The results suggest a close association between the V3 loop glycan and CCR5 use and may provide new insight into HIV-1 tropism and help to improve phenotype-prediction models.To enter cells, human immunodeficiency virus type 1 (HIV-1) uses the coreceptors CCR5 and/or CXCR4, together with the T-cell differentiation antigen CD4. Virus using the CCR5 receptor (R5 virus;Berger et al., 1998) or of the non-syncytium-tropic phenotype is suggested to be more transmissible (Zhang et al., 1993), whilst virus using the CXCR4 receptor (X4 virus;Berger et al., 1998) has been associated with disease progression (Connor et al., 1997; Scarlatti et al., 1997).The third variable (V3) loop of the HIV-1 gp120 envelope glycoprotein was recognized early on to play an important role in governing the choice of target cells (Hwang et al., 1991;De Jong et al., 1992;Fouchier et al., 1992;Shioda et al., 1992). Few amino acid substitutions and an increasing net charge of the V3 loop were sufficient to confer a change in cellular tropism in vitro (De Jong et al., 1992;De Wolf et al., 1994). A decreased number of N-linked glycosylation sites (sequons) in gp120, especially within and around the V3 region, has been demonstrated during evolution from the R5 to the X4 phenotype (Pollakis et al., 2001;Polzer et al., 2001Polzer et al., , 2002.Computer-based models have been developed to predict the biological phenotype of HIV sequences (Briggs et al., 2000;Resch et al., 2001;Jensen et al., 2003;Pillai et al., 2003), based on multiple linear regression (Briggs et al., 2000), specific amino acids within V3 and overall charge among subtype B viruses (Resch et al., 2001) and other subtypes (Pillai et al., 2003). Machine learning has been used to develop phenotype classifiers (Resch et al., 2001) and position-specific scoring matrices have also been used (Jensen et al., 2003).These methods relate to the assumption or possibility that the R5 and X4 phenotypes may be evaluated or classified, based on the properties of their specific amino acids, from the same scale or sum of scales. If the R5-and X4-specific properties relate to different characteristics, however, such programs may overlook essential differences in the R5 versus X4 phenotypes.In a previous study (Clevestig et al., 2005), a phylogenet...

Coreceptor use was determined for human immunodeficiency virus type 1 (HIV-1) isolates of various subtypes from 11 women during pregnancy and their infected children. Isolates from peripheral blood mononuclear cells (n=79) and from plasma (n=59) were available. The clinical and immunological stages of HIV-1 infection were recorded. Coreceptor use was tested on human cell lines expressing CD4 and different chemokine receptors. The R5 virus predominated, and only 9 isolates from 2 mothers used CXC chemokine receptor 4. All children carried the R5 virus at the time of diagnosis of HIV-1 infection. In 2 children of mothers carrying the X4 virus, the virus switched from R5 to X4 or to R5X4 by age 18 months (child no. 9) and age 48 months (child no. 10), whereas no children followed up to a similar age whose mothers were carrying the R5 virus experienced such a switch (P=.048). This points to a link between the presence of X4 virus in the mother and the emergence of X4 virus in her child.

Previously, we found that emergence of the X4 viral phenotype in HIV-1-infected children was related to the presence of X4 in their mothers (C.H. Casper et al., J Infect Dis 2002; 186:914-921). Here, we investigated the origin of the X4 phenotype in the child, analyzing two mother-child pairs (Ma-Ca, Mb-Cb) where the mothers carried X4 and their children developed X4 after an initial presence of R5. We used nested polymerase chain reaction of the env V3 region to generate 203 HIV-1 clones for sequencing (Ma, n = 44; Ca, n = 73; Mb, n = 61; Cb, n = 25) from DNA of peripheral blood mononuclear cell (PBMC) lysates, altogether 167 clones, or from cDNA of plasma RNA, 36 clones. PBMC and plasma isolate sequences from each time point enabled us to assign the probable phenotype to clone sequences in a phylogenetic tree. The transmission and evolution were reconstructed using the maximum likelihood method. In mother-child pair Ma-Ca, one maternal R5 isolate clustered with the child's R5 sequences, at the earliest time when R5 was isolated in the child, confirming this as a likely source of the transmitted R5 phenotype. At age 3, an X4 population was present in the child that had evolved from the child's own R5-associated population, clearly distinct from the maternal X4 sequences. The second mother-child pair (Mb-Cb) displayed a similar pattern. Amino acid substitution patterns corroborated the conclusions from the phylogenetic tree. Thus, in both children, the X4 virus developed from their own R5 population, and was not caused by transmission of X4.