On the use of sequence-quality information in OTU clustering

High-throughput sequencing has become an essential technology in life science research. Despite continuous improvements in technology, the produced sequences are still not entirely accurate. Consequently, the sequences are usually equipped with error probabilities. The quality information is already employed to find better solutions to a number of bioinformatics problems (e.g. read mapping). Data processing pipelines benefit in particular (especially when incorporating the quality information early), since enhanced outcomes of one step can improve all subsequent ones. Preprocessing steps, thus, quite regularly consider the sequence quality to fix errors or discard low-quality data. Other steps, however, like clustering sequences into operational taxonomic units (OTUs), a common task in the analysis of microbial communities, are typically performed without making use of the available quality information.

In this paper, we present quality-aware clustering methods inspired by quality-weighted alignments and model-based denoising, and explore their applicability to OTU clustering. We implemented the quality-aware methods in a revised version of our de novo clustering tool GeFaST and evaluated their clustering quality and performance on mock-community data sets. Quality-weighted alignments were able to improve the clustering quality of GeFaST by up to 10%. The examination of the model-supported methods provided a more diverse picture, hinting at a narrower applicability, but they were able to attain similar improvements. Considering the quality information enlarged both runtime and memory consumption, even though the increase of the former depended heavily on the applied method and clustering threshold.

The quality-aware methods expand the iterative, de novo clustering approach by new clustering and cluster refinement methods. Our results indicate that OTU clustering constitutes yet another analysis step benefiting from the integration of quality information. Beyond the shown potential, the quality-aware methods offer a range of opportunities for fine-tuning and further extensions.