# Possible bug in qiime cutadapt demux paired (2024.10)

**URL:** https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928
**Category:** Technical Support
**Tags:** demux, cutadapt
**Created:** [January 23, 2026, 6:02pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928 "2026-01-23T18:02:57Z")
**Posts on this page:** 7
**Page:** 1

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### Author: ![ppflrs](https://forum.qiime2.org/letter_avatar_proxy/v4/letter/p/cc9497/32.png) [@ppflrs](https://forum.qiime2.org/u/ppflrs)
#### Post date: [January 23, 2026, 6:02pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/1 "2026-01-23T18:02:57Z")

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Dear QIIME2 team,

troubleshooting a big library we encountered a behaviour we think could be a bug in qiime cutadapt demux paired.

We are using QIIME2 version 2024.10 and this is the command that’s being run via snakemake

```bash
qiime cutadapt demux-paired \
       --i-seqs {input.q2_import} \
       --p-error-rate 0.17 \
       --m-forward-barcodes-file {input.mapping_file} \
       --m-reverse-barcodes-file {input.mapping_file} \
       --m-forward-barcodes-column Forward_Barcode \
       --m-reverse-barcodes-column Reverse_Barcode \
       --o-per-sample-sequences {output.q2_demux_file} \
       --p-mixed-orientation TRUE \
       --p-forward-cut 2 \
       --p-reverse-cut 2 \
       --p-anchor-forward-barcode TRUE \
       --p-anchor-reverse-barcode TRUE \
       --p-batch-size 24 \
       --p-cores {threads} \
       --verbose \
       --o-untrimmed-sequences {output.q2_demux_untrimmed}

```

The problematic part is the combination of ‘--p-forward-cut/--p-reverse-cut’ with ‘--p-batch-size’.

Our amplicons are designed with an AC spacer at the beginning of the reads, therefore we chose to use the ‘cut’ parameters. By examining the demux log file I noticed that when qiime invokes cutadapt it will work ok for the first round. The problem is that the first batch of cutadapt will trim the bases and pass the trimmed version of the reads to the second round of cutadapt where it will trim once again. In our case from the 3rd round of cutadapt it started reporting mostly 0 adaptor trimmings for all the samples.

I think the correct behaviour could be:

1st round: trim, pass trimmed version to 2nd round

2nd round: don’t trim, pass input to 3rd round  
3rd round: don’t trim, pass input to next round  
and so on

For us removing ‘--p-forward-cut’ and ‘--p-revese-cut’ fixes the issue.

Attached the log file.

Thanks!

[Novogene\_11Gb\_v8\_q2\_demux.txt](https://forum.qiime2.org/uploads/short-url/y141BHSx37wKVJzCJIDX5jbem9Z.txt) (100.8 KB)

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### Author: ![cherman2](https://forum.qiime2.org/user_avatar/forum.qiime2.org/cherman2/32/17129_2.png) [@cherman2](https://forum.qiime2.org/u/cherman2)
#### Post date: [January 23, 2026, 7:13pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/2 "2026-01-23T19:13:49Z")

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### Author: ![cherman2](https://forum.qiime2.org/user_avatar/forum.qiime2.org/cherman2/32/17129_2.png) [@cherman2](https://forum.qiime2.org/u/cherman2)
#### Post date: [January 24, 2026, 12:15am UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/3 "2026-01-24T00:15:35Z")

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Hi @ppflrs,  
Thank you for pointing this out. You are 100% right about the behavior. We have opened a [PR on GitHub](https://github.com/qiime2/q2-cutadapt/pull/74), if you are interested.

I will come back and inform you once this fix is merged.

Again, thank you for noting this.

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### Author: ![lizgehret](https://forum.qiime2.org/user_avatar/forum.qiime2.org/lizgehret/32/17128_2.png) [@lizgehret](https://forum.qiime2.org/u/lizgehret)
#### Post date: [January 26, 2026, 3:57pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/4 "2026-01-26T15:57:29Z")

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Hi @ppflrs,

I just wanted to qiime in for @cherman2 and let you know that the fix for this has been merged- thanks again for reporting this! 🙌🏼

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### Author: ![lizgehret](https://forum.qiime2.org/user_avatar/forum.qiime2.org/lizgehret/32/17128_2.png) [@lizgehret](https://forum.qiime2.org/u/lizgehret)
#### Post date: [January 26, 2026, 3:57pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/5 "2026-01-26T15:57:35Z")

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<div class="post-metadata">

### Author: ![ppflrs](https://forum.qiime2.org/letter_avatar_proxy/v4/letter/p/cc9497/32.png) [@ppflrs](https://forum.qiime2.org/u/ppflrs)
#### Post date: [January 28, 2026, 1:43pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/6 "2026-01-28T13:43:14Z")

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Hi @lizgehret @cherman2,

thanks a lot for fixing the issue!

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### Author: ![system](https://forum-qiime2-org.s3.dualstack.us-west-2.amazonaws.com/original/3X/2/1/21af5fe23cb6f4579467c66a9ed94e55274ca7bd.svg) [@system](https://forum.qiime2.org/u/system)
#### Post date: [February 28, 2026, 7:43pm UTC](https://forum.qiime2.org/t/possible-bug-in-qiime-cutadapt-demux-paired-2024-10/33928/7 "2026-02-28T19:43:38Z")

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