interleaved mode bug

[jianshu93]

Dear bowtie2 team,

For version 2.5.0, there are bugs with interleaved mode but not with R1 and R2 mode, see details below;

Jianshus-MBP-2:test_data jianshuzhao$ time bowtie2 -x all_mag.fasta -f -1 T4AerOil_sbsmpl5.R1.fa.gz -2 T4AerOil_sbsmpl5.R2.fa.gz -S try.sam --threads 8 1157425 reads; of these: 1157425 (100.00%) were paired; of these: 1012592 (87.49%) aligned concordantly 0 times 144601 (12.49%) aligned concordantly exactly 1 time 232 (0.02%) aligned concordantly >1 times

1012592 pairs aligned concordantly 0 times; of these:
  18872 (1.86%) aligned discordantly 1 time
----
993720 pairs aligned 0 times concordantly or discordantly; of these:
  1987440 mates make up the pairs; of these:
    1982525 (99.75%) aligned 0 times
    4731 (0.24%) aligned exactly 1 time
    184 (0.01%) aligned >1 times

14.36% overall alignment rate

real 0m6.884s user 0m51.170s sys 0m0.767s

Jianshus-MBP-2:test_data jianshuzhao$ time bowtie2 -x all_mag.fasta -f --interleaved T4AerOil_sbsmpl5.fa -S try.sam --threads 8 (ERR): bowtie2-align died with signal 11 (SEGV)

interleaved file was created by seqtk mergepe R1.fa.gz R2.fa.gz. This bug has been there for at least several versions. Can you please check why, the sam output is identical though.

Thanks,

Jianshu

[jianshu93]

By the way, interleaved reads file looks like this:

readPair_20168658_read1 GGGCAGCAGATGGAGGTCGATGGCCCATATCTCTATCCGCCGCTGATCAGTTCGGTGGCGCCACGCAGCGCGGCGATGGATGCGGGGTTTCGCCCCGGCGATGTGATCGTTGCCGTCGATGGCGACCCGATCTTTGCCTTCGACCAGTTGA readPair_20168658_read2 CGTCGGTGCGCCGCGGCGCCAGCGCGAATTGCAGGTTTTCACCCTCACGCCAGACCGACACTTCCAGAACCCGTCCATCCGACGCCTCGACCCGGCTTTTCAACTGGTCGAAGGCAAAGATCGGGTCGCCATCGACGGCAACGATCACAT readPair_3900954_read1 AGGTGGTAGTGATTTTTTCGGGCTCGAGTTCGAGTACCAGGTAACCATTGTTTGTGGTGTTGCTGTAATGCACGTGCGGGTTGGTCAGTGGCGCGAGTTCCTGGCCAGGGATGCCAGGCATGGAAATAGCCGGGGTGGCGAATTCCACAG readPair_3900954_read2 ACATCATTGAGGGCCAGCAACTGGATAACGTCGTCATTCTCACCGGTGATGTACATTCGTCCTGGGCCATAGATGTCCCGCGTGACCCCTACGACCCGAC

Thanks,

Jianshu

[ch4rr0]

Hello @Jianshu93

Thank you for reporting this issue. I have pushed a change to the bug_fixes branch that should resolve this.

./bowtie2-align-s-debug -x example/index/lambda_virus -f  --interleaved foo.fa
Warning: Running in debug mode.  Please use debug mode only for diagnosing errors, and not for typical use of Bowtie 2.
2 reads; of these:
  2 (100.00%) were paired; of these:
    2 (100.00%) aligned concordantly 0 times
    0 (0.00%) aligned concordantly exactly 1 time
    0 (0.00%) aligned concordantly >1 times
    ----
    2 pairs aligned concordantly 0 times; of these:
      0 (0.00%) aligned discordantly 1 time
    ----
    2 pairs aligned 0 times concordantly or discordantly; of these:
      4 mates make up the pairs; of these:
        4 (100.00%) aligned 0 times
        0 (0.00%) aligned exactly 1 time
        0 (0.00%) aligned >1 times
0.00% overall alignment rate
@HD VN:1.5  SO:unsorted GO:query
@SQ SN:gi|9626243|ref|NC_001416.1|  LN:48502
@PG ID:bowtie2  PN:bowtie2  VN:2.5.1    CL:"/home/bowtie2/bowtie2-align-s-debug -x example/index/lambda_virus -f --interleaved foo.fa"
readPair_20168658_read1 77  *   0   0   *   *   0   0   GGGCAGCAGATGGAGGTCGATGGCCCATATCTCTATCCGCCGCTGATCAGTTCGGTGGCGCCACGCAGCGCGGCGATGGATGCGGGGTTTCGCCCCGGCGATGTGATCGTTGCCGTCGATGGCGACCCGATCTTTGCCTTCGACCAGTTGA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII YT:Z:UP
readPair_20168658_read2 141 *   0   0   *   *   0   0   CGTCGGTGCGCCGCGGCGCCAGCGCGAATTGCAGGTTTTCACCCTCACGCCAGACCGACACTTCCAGAACCCGTCCATCCGACGCCTCGACCCGGCTTTTCAACTGGTCGAAGGCAAAGATCGGGTCGCCATCGACGGCAACGATCACAT  IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII  YT:Z:UP
readPair_3900954_read1  77  *   0   0   *   *   0   0   AGGTGGTAGTGATTTTTTCGGGCTCGAGTTCGAGTACCAGGTAACCATTGTTTGTGGTGTTGCTGTAATGCACGTGCGGGTTGGTCAGTGGCGCGAGTTCCTGGCCAGGGATGCCAGGCATGGAAATAGCCGGGGTGGCGAATTCCACAG  IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII  YT:Z:UP
readPair_3900954_read2  141 *   0   0   *   *   0   0   ACATCATTGAGGGCCAGCAACTGGATAACGTCGTCATTCTCACCGGTGATGTACATTCGTCCTGGGCCATAGATGTCCCGCGTGACCCCTACGACCCGA IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII YT:Z:UP

[jianshu93]

Hello @ch4rr0 ,

I can confirm that it is fixed:

Jianshus-MBP-2:test_data jianshuzhao$ time bowtie2 -x all_mag.fasta -f --interleaved T4AerOil_sbsmpl5.fa -S try.sam --threads 8 1157425 reads; of these: 1157425 (100.00%) were paired; of these: 1011517 (87.39%) aligned concordantly 0 times 145616 (12.58%) aligned concordantly exactly 1 time 292 (0.03%) aligned concordantly >1 times

1011517 pairs aligned concordantly 0 times; of these:
  17903 (1.77%) aligned discordantly 1 time
----
993614 pairs aligned 0 times concordantly or discordantly; of these:
  1987228 mates make up the pairs; of these:
    1982475 (99.76%) aligned 0 times
    4640 (0.23%) aligned exactly 1 time
    113 (0.01%) aligned >1 times

14.36% overall alignment rate

real 0m5.379s user 0m40.471s sys 0m0.789s

Please update bio conda channel soon if possible. This mode was used by a many teams or pipelines.

Thanks,

Jianshu

[ch4rr0]

This has been fixed in v2.5.2. Thanks for reporting.