Analysis of viral tropism.

Tropism of each viral isolate was assessed by infecting U87.CD4 cells expressing either CCR5 or CXCR4, in parallel, with each viral stock and by detection of syncytia on MT2 cell line. To further confirm viral tropism, V3-loop sequence of the gp120 protein of each sample was also evaluated in terms of net amino acid charge (net charge), the presence of positively charged amino acids at codons 11 and/or 25 (bold) of V3-loop (bold italics; HXB2 gp160 positions 306 and 322, respectively) (11/25 rule), and the two major bioinformatics algorithms Position-Specific Scoring Matrix (PSSM) and geno2pheno [coreceptor] (g2p). The 20 CCR5, 4 CXCR4-tropic and 4 dual/mixed-tropic isolates (o) are shown on the table. Sample identifications are represented as follows: subtype B (bold italic), C (normal) and BF (bold). All sequences were obtained from the original PBMC of patients.For net charge, 11/25 rule, PSSM and g2p all possible amino acidic combinations according to amino acid change due to base ambiguities were evaluated. R5-X4: sample B563’s FPR value fell between the two cutoffs, evidencing the presence of X4 variants as well. R5-X4: for sample BF134o three out of four possible amino acidic combinations according to amino acid change due to base ambiguities were R5.