cutadapt / trim-paired / option "front" and "adapter"

Hello,

I don't know if it's too late but I found this topic " Fungal ITS analysis tutorial"

In fact there is a section in this tutorial which is exactly my problem in this topic:
"One issue with ITS (and other marker genes with vast length variability) is readthrough , which occurs when read lengths are longer than the amplicon itself!"

There is the command used like me but only in one step:

qiime cutadapt trim-paired \
  --i-demultiplexed-sequences demux.qza \
  --p-adapter-f AYTTAAGCATATCAATAAGCGGAGGCT \
  --p-front-f AACTTTYRRCAAYGGATCWCT \
  --p-adapter-r AGWGATCCRTTGYYRAAAGTT \
  --p-front-r AGCCTCCGCTTATTGATATGCTTAART \
  --o-trimmed-sequences demux-trimmed.qza

I tried downloading the data from this tutorial to see if my problem is with these data too, but the forward primers have already been trimmed in the raw reads.

So I think the command in the Fungal ITS analysis tutorial has the same problem that I had, I think we need to use the 2 step cutadapt solution.


Moreover, I detected an error in this tutorial: the forward primer and the reverse primer are swapped.

So, Instead of the command seen in the tutorial above the real command is (with the wildcard enabled!):

qiime cutadapt trim-paired \
  --i-demultiplexed-sequences demux.qza \
  --p-adapter-r AYTTAAGCATATCAATAAGCGGAGGCT \
  --p-front-r AACTTTYRRCAAYGGATCWCT \
  --p-adapter-f AGWGATCCRTTGYYRAAAGTT \
  --p-front-f AGCCTCCGCTTATTGATATGCTTAART \
  --p-match-read-wildcards
  --p-match-adapter-wildcards
  --o-trimmed-sequences demux-trimmed_swapped.qza

=> So to have the expected results: the 2 steps solution:

qiime cutadapt trim-paired
--i-demultiplexed-sequences demux-paired-end.qza
--p-adapter-f AGWGATCCRTTGYYRAAAGTT
--p-adapter-r AYTTAAGCATATCAATAAGCGGAGGCT
--p-match-read-wildcards
--p-match-adapter-wildcards
--verbose
--o-trimmed-sequences trimmed_remove_primers_wild.qza

qiime cutadapt trim-paired
--i-demultiplexed-sequences trimmed_remove_primers_wild.qza
--p-front-f AGCCTCCGCTTATTGATATGCTTAART
--p-front-r AACTTTYRRCAAYGGATCWCT
--p-match-read-wildcards
--p-match-adapter-wildcards
--p-discard-untrimmed
--verbose
--o-trimmed-sequences trimmed_remove_primers_wild_2.qza

How can I report this error on the tutorial Fungal ITS analysis tutorial?