High percentage of discordant reads

Question

[Domu]

I am working with Pseudomonas aeruginosa PA14 genomes. I aligned my Illumina reads to the reference genome using bowtie2 and when I look at the stats I get something like the following:

450261 reads; of these:
  450261 (100.00%) were paired; of these:
    191037 (42.43%) aligned concordantly 0 times
    255656 (56.78%) aligned concordantly exactly 1 time
    3568 (0.79%) aligned concordantly >1 times
    ----
    191037 pairs aligned concordantly 0 times; of these:
      183325 (95.96%) aligned discordantly 1 time
    ----
    7712 pairs aligned 0 times concordantly or discordantly; of these:
      15424 mates make up the pairs; of these:
        5469 (35.46%) aligned 0 times
        3976 (25.78%) aligned exactly 1 time
        5979 (38.76%) aligned >1 times
99.39% overall alignment rate

I have read that concordant and discordant alignments are used to detect INDELS, however, 40% reads aligning discordantly seems very high. Can someone tell me what is going on?

Thanks in advance!

[GenoMax]

Did you trim the paired end data files independently? If so they are likely out of sync which will lead to highly discordant alignments.

[Domu]

No, I trimmed them in paired end mode of Trimmomatic

[GenoMax]

If you are willing, try aligning with bbmap.sh from BBMap suite and see what the stats say.

bbmap.sh -Xmx10g in1=file_R1.fq.gz in2=file_R2.fq.gz ref=PA14_genome.fa

should be all you need. Stats are written to STDERR.

[Domu]

Hey, thanks! I will look into this :)