Email updates

Keep up to date with the latest news and content from BMC Medical Informatics and Decision Making and BioMed Central.

Open Access Research article

Genotypic tropism testing by massively parallel sequencing: qualitative and quantitative analysis

Martin Däumer1, Rolf Kaiser2, Rolf Klein1, Thomas Lengauer3, Bernhard Thiele1 and Alexander Thielen13*

Author Affiliations

1 Institute of Immunology and Genetics, Pfaffplatz 10, 67655 Kaiserslautern, Germany

2 Institute of Virology, University of Cologne, Fürst-Pückler-Str. 56, 50935 Cologne, Germany

3 Max Planck Institute for Informatics, Stuhlsatzenhausweg E1.4, 66123, Saarbrücken, Germany

For all author emails, please log on.

BMC Medical Informatics and Decision Making 2011, 11:30  doi:10.1186/1472-6947-11-30

Published: 13 May 2011



Inferring viral tropism from genotype is a fast and inexpensive alternative to phenotypic testing. 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.


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[coreceptor].


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[coreceptor] (10%), and defining a minority cutoff of 5%, the results were concordant in all but one isolate.


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.