Hello! I hope somebody can provide some insight into TopHat and Bowtie2 behavior. I have a data set that I aligned to a reference using Bowtie2 and Tophat, both running default settings. When I used Bowtie, the alignment rates were >90% for all samples. With Tophat, however, the rates were all around 45-55%. Since Tophat uses Bowtie, I'm not sure why the resulting alignment rates are so much lower. My best guess is that when Tophat calls Bowtie, it uses different settings than the default settings for a user using Bowtie directly, but I haven't been able to figure this out from the respective manuals. Could anyone explain why this might be? I'm curious because there's apparently something about my reads that is very sensitive to differences in aligners used, and I want to know what it is.

I'm not sure what additional information is needed to answer this question but I'm ready to provide it. Thank you!

ETA: I'm using Tophat2 and Bowtie2, latest versions of both
Hi everyone, I have an answer to this after consulting with one of the developers. Posting in case anyone here is interested or comes across this when googling.

The difference between how Bowtie2 runs by default and how Tophat2 calls Bowtie2 is in the minimum alignment score required to consider an alignment valid. In Bowtie, this minimum is set to -0.6 - 0.6 * read length. Tophat runs Bowtie with the alignment score minimum set to a constant of -14. So Tophat has a much more stringent criterion for which alignments it considers valid. (When Hisat2 calls Bowtie2, the default minimum threshold is set to 0 - 0.2 * read length. With my samples, this resulted in alignment rates around 75% - right in between the Bowtie and Tophat results.) 

The developer I communicated with was surprised that this parameter made such a drastic difference in alignment results, so it's worth paying attention to. This is likely because I'm dealing with diverse wild populations that might have significantly diverged from the reference genome. 