Q2-dada2: Error in isBimeraDenovoTable(unqs[[i]], ..., verbose = verbose) : Input must be a valid sequence table

[bassio (Ahmed Bassiouni)]

Error in isBimeraDenovoTable(unqs[[i]], ..., verbose = verbose) : Input must be a valid sequence table.

Hello,
I got the error below at the step of chimera detection.
I wonder if this is that there were no sequences left for chimera detection at the end of that stage.
Running qiime 2 Feb 2018 version.
Let me know if you need any further info

System versions
Python version: 3.5.5
QIIME 2 release: 2018.2
QIIME 2 version: 2018.2.0
q2cli version: 2018.2.0

Installed plugins
alignment 2018.2.0
composition 2018.2.0
cutadapt 2018.2.0
dada2 2018.2.0
deblur 2018.2.0
demux 2018.2.0
diversity 2018.2.0
emperor 2018.2.0
feature-classifier 2018.2.0
feature-table 2018.2.0
gneiss 2018.2.0
longitudinal 2018.2.0
metadata 2018.2.0
phylogeny 2018.2.0
quality-control 2018.2.0
quality-filter 2018.2.0
sample-classifier 2018.2.0
taxa 2018.2.0
types 2018.2.0
vsearch 2018.2.0

See below for error report:

Running external command line application(s). This may print messages to stdout and/or stderr.
The command(s) being run are below. These commands cannot be manually re-run as they will depend on temporary files that no longer exist.

Command: run_dada_paired.R /tmp/tmps4hlcnk7/forward /tmp/tmps4hlcnk7/reverse /tmp/tmps4hlcnk7/output.tsv.biom /tmp/tmps4hlcnk7/filt_f /tmp/tmps4hlcnk7/filt_r 260 260 0 0 2.0 2 consensus 1.0 1 1000000

R version 3.4.1 (2017-06-30)
Loading required package: Rcpp
DADA2 R package version: 1.6.0

Filtering ...

Learning Error Rates
2a) Forward Reads
Initializing error rates to maximum possible estimate.
Sample 1 - 2656 reads in 1987 unique sequences.
Sample 2 - 6060 reads in 2150 unique sequences.
Sample 3 - 11141 reads in 3185 unique sequences.
selfConsist step 2
selfConsist step 3
selfConsist step 4
selfConsist step 5
Convergence after 5 rounds.
2b) Reverse Reads
Initializing error rates to maximum possible estimate.
Sample 1 - 2656 reads in 1987 unique sequences.
Sample 2 - 6060 reads in 2150 unique sequences.
Sample 3 - 11141 reads in 3185 unique sequences.
selfConsist step 2
selfConsist step 3
selfConsist step 4
selfConsist step 5
Convergence after 5 rounds.

Denoise remaining samples

Remove chimeras (method = consensus)
Error in isBimeraDenovoTable(unqs[[i]], ..., verbose = verbose) :
Input must be a valid sequence table.
Calls: removeBimeraDenovo -> isBimeraDenovoTable
In addition: Warning message:
In is.na(colnames(unqs[[i]])) :
is.na() applied to non-(list or vector) of type 'NULL'
Execution halted

[thermokarst]

Hi @bassio!

@bassio

I wonder if this is that there were no sequences left for chimera detection at the end of that stage.

Yep, this is the case! One thing I noticed your "unique sequence" counts are identical between your forward and reverse files - you didn't happen to import forward reads as forward and reverse, did you? That could certainly account for this issue. Otherwise, maybe you could attach a screenshot or QZV of your demux summarize visualization for these data --- the denoising and filtering is sensitive to the quality scores, this plot would help us get at tuning those parameters a bit more. Thanks!

PS - In the future, please try and remember to copy and paste the command or commands that you are trying to run (in addition to the full error message), this helps us help you!

[bassio (Ahmed Bassiouni)]

I looked at the qzvpaired-end-demux.qzv (285.7 KB)

which I obtained using this command

!qiime tools import
--type 'SampleData[PairedEndSequencesWithQuality]'
--input-path './data/fastq/manifest.csv'
--output-path './results/paired-end-demux.qza'
--source-format PairedEndFastqManifestPhred33

The forward and reverse statistics are suspiciously similar, as if the forward are the same as reverse.

Note sure if my fault or bug

[thermokarst]

Thanks @bassio --- can you share your manifest file with us? That will help us figure out exactly what you imported. Thanks!

[bassio (Ahmed Bassiouni)]

Yup my fault .. manifest file duplicating forward and reverse reads.
You can close this issue now!
Thanks for your help!