Dada2 Reporting read numbers error

Hi all,

I am denoising 16s PacBio data and running dada2 last night has given me a unexpected error when I run this code:

qiime dada2 denoise-ccs
--i-demultiplexed-seqs Raw_data_archival/archived_reads.qza
--p-min-len 1000
--p-max-len 1800
--p-max-ee 4
--p-front AGRGTTYGATYMTGGCTCAG
--p-adapter RGYTACCTTGTTACGACTT
--p-max-mismatch 3
--p-n-threads 0
--output-dir dada2_output02
--verbose

The error is as follows:

  1. Report read numbers through the pipeline
    Error in cbind(prim, out[, 2], matrix(0, nrow = nrow(out), ncol = 2)) :
    number of rows of matrices must match (see arg 3)
    1: cbind(prim, out[, 2], matrix(0, nrow = nrow(out), ncol = 2))
    Traceback (most recent call last):
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/q2_dada2/_denoise.py", line 467, in denoise_ccs
    run_commands([cmd])
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/q2_dada2/_denoise.py", line 37, in run_commands
    subprocess.run(cmd, check=True)
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/subprocess.py", line 528, in run
    raise CalledProcessError(retcode, process.args,
    subprocess.CalledProcessError: Command '['run_dada.R', '--input_directory', '/var/folders/_3/v5c3hf4j2g34qxfwkqc95rq00000gn/T/qiime2/s.d./data/6c89b129-41b2-4d04-9597-a48fc3cce54f/data', '--output_path', '/var/folders/_3/v5c3hf4j2g34qxfwkqc95rq00000gn/T/tmpw408at7a/output.tsv.biom', '--output_track', '/var/folders/_3/v5c3hf4j2g34qxfwkqc95rq00000gn/T/tmpw408at7a/track.tsv', '--removed_primer_directory', '/var/folders/_3/v5c3hf4j2g34qxfwkqc95rq00000gn/T/tmpw408at7a/nop', '--filtered_directory', '/var/folders/_3/v5c3hf4j2g34qxfwkqc95rq00000gn/T/tmpw408at7a/filt', '--forward_primer', 'AGRGTTYGATYMTGGCTCAG', '--reverse_primer', 'RGYTACCTTGTTACGACTT', '--max_mismatch', '3', '--indels', 'False', '--truncation_length', '0', '--trim_left', '0', '--max_expected_errors', '4', '--truncation_quality_score', '2', '--min_length', '1000', '--max_length', '1800', '--pooling_method', 'independent', '--chimera_method', 'consensus', '--min_parental_fold', '3.5', '--allow_one_off', 'False', '--num_threads', '0', '--learn_min_reads', '1000000', '--homopolymer_gap_penalty', 'NULL', '--band_size', '32']' returned non-zero exit status 1.
    During handling of the above exception, another exception occurred:
    Traceback (most recent call last):
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/q2cli/commands.py", line 520, in call
    results = self._execute_action(
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/q2cli/commands.py", line 581, in _execute_action
    results = action(**arguments)
    File "", line 2, in denoise_ccs
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/qiime2/sdk/action.py", line 342, in bound_callable
    outputs = self.callable_executor(
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/qiime2/sdk/action.py", line 576, in callable_executor
    output_views = self._callable(**view_args)
    File "/Users/s.d./miniconda3/envs/qiime2-amplicon-2024.5/lib/python3.9/site-packages/q2_dada2/_denoise.py", line 476, in denoise_ccs
    raise Exception("An error was encountered while running DADA2"
    Exception: An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more.
    Plugin error from dada2:
    An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more.
    See above for debug info.

As it is filtering verbose tells me that the filtering percent is around 98 which seems correct to me. Is the error telling me there is an issue compiling the data into a table? Thanks for the help with this one.

Hi @tinoracha,

Here's where the root of the issue lies:

This is likely due to some samples with zero reads passing the primer detection. Check out this thread with a similar issue (which also links off to another instance of this error as well, for additional reference).

Cheers :lizard:

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