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While being highly predictive when performed on clonal samples, sensitivity of predicting CXCR4-using (X4) variants drops substantially in clinical isolates. This is mainly attributed to minor variants not detected by standard bulk-sequencing. Massively parallel sequencing (MPS) detects single clones thereby being much more sensitive. Using this technology we wanted to improve genotypic prediction of coreceptor usage.<\/jats:p><\/jats:sec><jats:sec><jats:title>Methods<\/jats:title><jats:p>Plasma samples from 55 antiretroviral-treated patients tested for coreceptor usage with the Monogram Trofile Assay were sequenced with standard population-based approaches. Fourteen of these samples were selected for further analysis with MPS. Tropism was predicted from each sequence with geno2pheno<jats:sub>[coreceptor]<\/jats:sub>.<\/jats:p><\/jats:sec><jats:sec><jats:title>Results<\/jats:title><jats:p>Prediction based on bulk-sequencing yielded 59.1% sensitivity and 90.9% specificity compared to the trofile assay. With MPS, 7600 reads were generated on average per isolate. Minorities of sequences with high confidence in CXCR4-usage were found in all samples, irrespective of phenotype. When using the default false-positive-rate of geno2pheno<jats:sub>[coreceptor]<\/jats:sub>(10%), and defining a minority cutoff of 5%, the results were concordant in all but one isolate.<\/jats:p><\/jats:sec><jats:sec><jats:title>Conclusions<\/jats:title><jats:p>The combination of MPS and coreceptor usage prediction results in a fast and accurate alternative to phenotypic assays. The detection of X4-viruses in all isolates suggests that coreceptor usage as well as fitness of minorities is important for therapy outcome. The high sensitivity of this technology in combination with a quantitative description of the viral population may allow implementing meaningful cutoffs for predicting response to CCR5-antagonists in the presence of X4-minorities.<\/jats:p><\/jats:sec>","DOI":"10.1186\/1472-6947-11-30","type":"journal-article","created":{"date-parts":[[2011,5,13]],"date-time":"2011-05-13T18:30:01Z","timestamp":1305311401000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":20,"title":["Genotypic tropism testing by massively parallel sequencing: qualitative and quantitative analysis"],"prefix":"10.1186","volume":"11","author":[{"given":"Martin","family":"D\u00e4umer","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rolf","family":"Kaiser","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rolf","family":"Klein","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Thomas","family":"Lengauer","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bernhard","family":"Thiele","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alexander","family":"Thielen","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"297","published-online":{"date-parts":[[2011,5,13]]},"reference":[{"issue":"2","key":"404_CR1","doi-asserted-by":"publisher","first-page":"222","DOI":"10.1001\/jama.288.2.222","volume":"288","author":"PG Yeni","year":"2002","unstructured":"Yeni PG, Hammer SM, Carpenter CC, Cooper DA, Fischl MA, Gatell JM, Gazzard BG, Hirsch MS, Jacobsen DM, Katzenstein DA: Antiretroviral treatment for adult HIV infection in 2002: updated recommendations of the International AIDS Society-USA Panel. 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