Unicycler SPAdes error

[MJoseMo]

Hi,

I'm using unicycler and encountering an error. The error message is

Error: SPAdes encountered an error.

In a GitHub discussion, I found that this could be resolved with --spades_options -m 1024. However I encounter a new error:
--spades_options: command not found.

`Starting Unicycler (2025-02-06 17:18:56)

Welcome to Unicycler, an assembly pipeline for bacterial genomes. Since you provided only short reads, Unicycler will essentially function as a SPAdes-optimiser. It will try many k-mer sizes, choose the best based on contig length and graph connectivity, and scaffold the graph using SPAdes repeat resolution.
For more information, please see https://github.com/rrwick/Unicycler

Command: conda/envs/centos/bin/unicycler -1 U2OS_1_L1_1_50.fq.gz -2 U2OS_1_L1_2_50.fq.gz -o U2OS1/

Unicycler version: v0.5.1
Using 8 threads

The output directory already exists and files may be reused or overwritten:
U2OS1

Dependencies:
Program Version Status
spades.py 4.0.0 good
racon not used
makeblastdb 2.16.0+ good
tblastn 2.16.0+ good

Choosing k-mer range for assembly (2025-02-06 17:26:19)

Unicycler chooses a k-mer range for SPAdes based on the length of the input reads. It uses a wide range of many k-mer sizes to maximise the chance of finding an ideal assembly.

SPAdes maximum k-mer: 127
Median read length: 150
K-mer range: 27, 53, 71, 87, 99, 111, 119, 127

SPAdes assemblies (2025-02-06 17:33:23)

Unicycler now uses SPAdes to assemble the short reads. It scores the assembly graph for each k-mer using the number of contigs (fewer is better) and the number of dead ends (fewer is better). The score function is 1/(c*(d+2)), where c is the contig count and d is the dead end count.

spades.py -o U2OS1/spades_assembly -k 27 --threads 8 --gfa11 --isolate -1 U2OS_1_L1_1_50.fq.gz -2 U2OS_1_L1_2_50.fq.gz -m 1024
Error: SPAdes encountered an error:`

[Faiimklin]

@MJoseMo Hi, I get the same error. My problem is the input files. Here is my solution, make sure the paths to your input FASTQ files are correct and that the files themselves are not corrupted. You can use fastqc to check the quality of your reads.

I run #fastqc nsn_forward_paired_downsample.fq.gz but it says "application/gzip Failed to process test_subset/nsn_forward_small.fq.gz java.util.zip.ZipException: Not in GZIP format"
It means that your "gzipped" FASTQ files is not actually gzipped. This is almost certainly why SPAdes is failing.

I use #file nsn_forward_paired_downsample.fq.gz to check the input file.
The output is "nsn_forward_paired_downsample.fq.gz: ASCII text"
The output confirms that the file is not gzipped. It's just plain text, even though it has the .gz extension. This is the core problem.
So, I run these following command lines to make the input file to .gz

#mv nsn_forward_paired_downsample.fq.gz nsn_forward_paired_downsample.fq
#bgzip nsn_forward_paired_downsample.fq > nsn_forward_paired_downsample_original.fq.gz
#file nsn_forward_paired_downsample.fq.gz

The output is "nsn_forward_paired_downsample.fq.gz: Blocked GNU Zip Format (BGZF; gzip compatible), block length 11622"
That means that my file is now correctly gzipped. The file is already in the correct format.

I do both forward and reverse FASTQ files. After that, I can run Unicycler and get results.

I hope I share something helpful to you. Good luck.

[owenburroughs]

I encountered the same error. In my case, it was because my working directory path included spaces in it. Removing the spaces solved the issue. This is an underlying issue with SPAdes as far as I can tell.

[valery-shap]

Hi,

I had the same error. It was solved by creating a new output directory.