Warnings. None of paired reads aligned properly.

[DanieleDeco]

Description of bug

I run Spades for a few metagenomic samples and in a few of them, I got the following warnings.
=== Error correction and assembling warnings:

• 2:25:51.222 6985M / 14G WARN General (pair_info_count.cpp : 358) Unable to estimate insert size for paired library #0
• 2:25:51.247 6985M / 14G WARN General (pair_info_count.cpp : 364) None of paired reads aligned properly. Please, check orientation of your read pairs.
• 2:25:51.278 6985M / 14G WARN General (repeat_resolving.cpp : 81) Insert size was not estimated for any of the paired libraries, repeat resolution module will not run.
• 4:11:14.894 6192M / 14G WARN General (pair_info_count.cpp : 187) Single reads are not used in metagenomic mode

Do you know what the problem could be?

The paired reads are pre-processed with trimmomatric and checked with fastq-pair.

Thanks

Daniele

warnings.log
params.txt
spades.log

spades.log

spades.log

params.txt

params.txt

SPAdes version

Spades 15.2

Operating System

Debian 5.9.15

Python Version

Python

Method of SPAdes installation

conda

No errors reported in spades.log

• Yes

[asl]

Your input reads are corrupted. E.g. the left ones do not correspond to the right ones. Ensure you're using paired-end aware trimmer.

[DanieleDeco]

I checked that with fastq-pair and It seems that all the reads are paired.

[asl]

fastq-pair only checks read names. So, if your reads were screwed in other way, then it will not help. Alternatively, your assembly is so fragmented that there are no long contigs usable for estimation of fragment length distribution.