Also, this tool along with a few others has recently been changed to work with paired-end data only if that data is arranged as a so-called paired collection. From your screenshot it looks as if you have regular separate datasets for fw and rv reads, but judging from the number of datasets in your history your analysis could profit greatly from using collections. If you’ve never used them, you might want to check out this tutorial although the user interface around collections has changed lately and iss not exactly like shown in this material any more.
If, for now, you prefer to keep on working with separate datasets as you did so far, you can always switch to the previous version of bowtie2 as shown in the screenshot below.
Cheers,
Wolfgang
