Hi everyone!!
I am playing with the great q2-sidle to try to analyze five amplicons of five hipervariable regions of the 16S rRNA bacterial gen. However, when I run qiime sidle reconstruct-counts
command, I get the next error message:
qiime sidle reconstruct-counts --p-region AmpliconR1_GG --i-kmer-map /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/green_database/db_ampliconR1/ggenes-13-8-db-Amplicon-R1-100nt-map.qza --i-regional-alignment R1_CCR16_6samples-align-map.qza --i-regional-table /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/v2smurf_11062021-271140879/FASTQ_Generation_2021-06-14_14_52_34Z-427957897/CCR16_6TIPOS/table-R1-TRIM-CA-CCR16-6samples-100nt.qza --p-region AmpliconR2_GG --i-kmer-map /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/green_database/db_ampliconR2/ggenes-13-8-db-Amplicon-R2-100nt-map.qza --i-regional-alignment R2_CCR16_6samples-align-map.qza --i-regional-table /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/v2smurf_11062021-271140879/FASTQ_Generation_2021-06-14_14_52_34Z-427957897/CCR16_6TIPOS/table-R2-TRIM-CA-CCR16-6samples-100nt.qza --p-region AmpliconR4_GG --i-kmer-map /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/green_database/db_ampliconR4/ggenes-13-8-99-db-Amplicon-R4-100nt-map.qza --i-regional-alignment R4_CCR16_6samples-align-map.qza --i-regional-table /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/v2smurf_11062021-271140879/FASTQ_Generation_2021-06-14_14_52_34Z-427957897/CCR16_6TIPOS/table-R4-TRIM-CA-CCR16-6samples-100nt.qza --p-region AmpliconR5_GG --i-kmer-map /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/green_database/db_ampliconR5/ggenes-13-8-99-db-Amplicon-R5-100nt-map.qza --i-regional-alignment R5_CCR16_6samples-align-map.qza --i-regional-table /home/elsa/Documentos/Doctorado/QIIME2/SMURF-CCR/v2smurf_11062021-271140879/FASTQ_Generation_2021-06-14_14_52_34Z-427957897/CCR16_6TIPOS/table-R5-TRIM-CA-CCR16-6samples-100nt.qza --p-n-workers 2 --o-reconstructed-table reconstruction-6samples-noR3/CCR16_6samples-table.qza --o-reconstruction-summary reconstruction-6samples-noR3/CCR16_6samples-summary.qza --o-reconstruction-map reconstruction-6samples-noR3/CCR16_6samples-map.qza --verbose
Traceback (most recent call last):
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2cli/commands.py", line 329, in __call__
results = action(**arguments)
File "<decorator-gen-219>", line 2, in reconstruct_counts
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/qiime2/sdk/action.py", line 244, in bound_callable
outputs = self._callable_executor_(scope, callable_args,
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/qiime2/sdk/action.py", line 390, in _callable_executor_
output_views = self._callable(**view_args)
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2_sidle/_reconstruct.py", line 101, in reconstruct_counts
aligned_kmers = _get_unique_kmers(align_map['kmer'])
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2_sidle/_reconstruct.py", line 525, in _get_unique_kmers
kmers = np.hstack([[a.split("@")[0] for a in kmer.split('|')]
File "<__array_function__ internals>", line 5, in hstack
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/numpy/core/shape_base.py", line 346, in hstack
return _nx.concatenate(arrs, 1)
File "<__array_function__ internals>", line 5, in concatenate
ValueError: need at least one array to concatenate
Plugin error from sidle:
need at least one array to concatenate
See above for debug info.
As you can see, I have indicated that I want to study five regions and I only include four in the command. The reason why I did not add the R3 region it is because I can't denoise it after trimming with dada2 as I did with the others.
I thought that maybe it could be an out of memory problem, but I reanalyzed the data in other server and still got the same error:
qiime dada2 denoise-paired --i-demultiplexed-seqs R3-trimmed-CA-CCR16-6samples.qza --p-trim-left-f 4 --p-trim-left-r 2 --p-trunc-len-f 131 --p-trunc-len-r 136 --o-table table-R3-trimmed-CA-CCR16-6samples.qza --o-representative-sequences rep-seqs-R3-trimmed-CA-CCR16-6samples.qza --o-denoising-stats stats-R3-trimmed-CA-CCR16-6samples.qza
Plugin error from dada2:
An error was encountered while running DADA2 in R (return code 1), please inspect stdout and stderr to learn more.
Debug info has been saved to /tmp/qiime2-q2cli-err-a8ocf3ae.log
(qiime2-2021.4) elsa@compostela:~/MICROBIOTA/v2smurf_11062021-271140879/FASTQ_Generation_2021-06-14_14_52_34Z-427957897/CCR16_6TIPOS$ cat /tmp/qiime2-q2cli-err-a8ocf3ae.log
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/tmp6pzax_r6/forward /tmp/tmp6pzax_r6/reverse /tmp/tmp6pzax_r6/output.tsv.biom /tmp/tmp6pzax_r6/track.tsv /tmp/tmp6pzax_r6/filt_f /tmp/tmp6pzax_r6/filt_r 131 136 4 2 2.0 2.0 2 12 independent consensus 1.0 1 1000000
R version 4.0.3 (2020-10-10)
Loading required package: Rcpp
DADA2: 1.18.0 / Rcpp: 1.0.6 / RcppParallel: 5.1.2
1) Filtering The filter removed all reads: /tmp/tmp6pzax_r6/filt_f/CCR-16-1-FFPE-Gmet_S56_L001_R1_001.fastq.gz and /tmp/tmp6pzax_r6/filt_r/CCR-16-1-FFPE-Gmet_S56_L001_R2_001.fastq.gz not written.
The filter removed all reads: /tmp/tmp6pzax_r6/filt_f/CCR-16-1-FFPE-GsinMet_S57_L001_R1_001.fastq.gz and /tmp/tmp6pzax_r6/filt_r/CCR-16-1-FFPE-GsinMet_S57_L001_R2_001.fastq.gz not written.
The filter removed all reads: /tmp/tmp6pzax_r6/filt_f/CCR-16-1-FFPE-TumGrasa_S55_L001_R1_001.fastq.gz and /tmp/tmp6pzax_r6/filt_r/CCR-16-1-FFPE-TumGrasa_S55_L001_R2_001.fastq.gz not written.
The filter removed all reads: /tmp/tmp6pzax_r6/filt_f/CCR-16-1-FFPE-Tumor-A_S53_L001_R1_001.fastq.gz and /tmp/tmp6pzax_r6/filt_r/CCR-16-1-FFPE-Tumor-A_S53_L001_R2_001.fastq.gz not written.
The filter removed all reads: /tmp/tmp6pzax_r6/filt_f/CCR-16-1-FFPE-Tumor-B_S54_L001_R1_001.fastq.gz and /tmp/tmp6pzax_r6/filt_r/CCR-16-1-FFPE-Tumor-B_S54_L001_R2_001.fastq.gz not written.
Some input samples had no reads pass the filter.
xx.xxx
2) Learning Error Rates
127 total bases in 1 reads from 1 samples will be used for learning the error rates.
134 total bases in 1 reads from 1 samples will be used for learning the error rates.
3) Denoise samples .
.
4) Remove chimeras (method = consensus)
Error in isBimeraDenovoTable(unqs[[i]], ..., verbose = verbose) :
Input must be a valid sequence table.
Calls: removeBimeraDenovo -> isBimeraDenovoTable
Ejecución interrumpida
Traceback (most recent call last):
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2_dada2/_denoise.py", line 266, in denoise_paired
run_commands([cmd])
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2_dada2/_denoise.py", line 36, in run_commands
subprocess.run(cmd, check=True)
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/subprocess.py", line 516, in run
raise CalledProcessError(retcode, process.args,
subprocess.CalledProcessError: Command '['run_dada_paired.R', '/tmp/tmp6pzax_r6/forward', '/tmp/tmp6pzax_r6/reverse', '/tmp/tmp6pzax_r6/output.tsv.biom', '/tmp/tmp6pzax_r6/track.tsv', '/tmp/tmp6pzax_r6/filt_f', '/tmp/tmp6pzax_r6/filt_r', '131', '136', '4', '2', '2.0', '2.0', '2', '12', 'independent', 'consensus', '1.0', '1', '1000000']' returned non-zero exit status 1.
During handling of the above exception, another exception occurred:
Traceback (most recent call last):
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2cli/commands.py", line 329, in __call__
results = action(**arguments)
File "<decorator-gen-616>", line 2, in denoise_paired
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/qiime2/sdk/action.py", line 244, in bound_callable
outputs = self._callable_executor_(scope, callable_args,
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/qiime2/sdk/action.py", line 390, in _callable_executor_
output_views = self._callable(**view_args)
File "/home/elsa/miniconda3/envs/qiime2-2021.4/lib/python3.8/site-packages/q2_dada2/_denoise.py", line 279, in denoise_paired
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.
So, I think that, perphas, these two errors are interrelated.
Thank you for your help!
Elsa