Automatic expansion triggered when load factor was below minimum threshold

[hermeseduardo]

hi there, I got this error message. Any ideas about what could have cause it?

[2018-02-22 08:36:13] INFO: Running Flye 2.3.2-gd46edb7
[2018-02-22 08:36:13] INFO: Assembling reads
[2018-02-22 08:36:13] INFO: Reading sequences
[2018-02-22 08:44:22] INFO: Generating solid k-mer index
[2018-02-22 08:46:32] INFO: Counting kmers (1/2):
0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100%
[2018-02-22 08:54:26] INFO: Counting kmers (2/2):
0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100%
[2018-02-22 09:21:11] INFO: Filling index table
0% 10% 20% 30% 40% 50% 60% 70% 80% 90% 100%
[2018-02-22 10:00:35] INFO: Extending reads
0% [2018-02-22 10:14:55] ERROR: Caught unhandled exception: Automatic expansion triggered when load factor was below minimum threshold
[2018-02-22 10:14:55] ERROR: flye-assemble(_Z16exceptionHandlerv+0x2d) [0x43c73d]
[2018-02-22 10:14:55] ERROR: /usr/lib64/libstdc++.so.6(+0x96706) [0x2aaaab277706]
[2018-02-22 10:14:55] ERROR: /usr/lib64/libstdc++.so.6(+0x96751) [0x2aaaab277751]
[2018-02-22 10:14:55] ERROR: /usr/lib64/libstdc++.so.6(+0xc1708) [0x2aaaab2a2708]
[2018-02-22 10:14:55] ERROR: /lib64/libpthread.so.0(+0x8744) [0x2aaaab789744]
[2018-02-22 10:14:55] ERROR: /lib64/libc.so.6(clone+0x6d) [0x2aaaaba87aad]
[2018-02-22 10:15:15] ERROR: Command '['flye-assemble', '-l', '/flush1/esc003/Flye_cynegetis_assembly/flye/flye.log', '-t', '20', '-v', '5000', '/flush2/esc003/Pacbio_subreads_smartbellremoved.fasta', '/flush1/esc003/Flye_cynegetis_assembly/flye/0-assembly/draft_assembly.fasta', '576716800', '/data/esc003/apps/Flye/flye/resource/asm_raw_reads.cfg']' returned non-zero exit status 1

[mikolmogorov]

Hi,

Thanks for the report. It looks like there is a rare issue with the hash function for sequence ids - it does not provide a good uniform distribution of keys in this case. I incorporated a batter hash function which should solve the problem - please try the updated version from the flye-devel branch. You can either checkout it through the git interface, or download a zip archive: https://github.com/fenderglass/Flye/archive/flye-devel.zip

Let me know if this helps,

Mikhail

[hermeseduardo]

thanks Mikhail, I tried but the problem is still there. Please let me know if you need additional information from my side.
best,

Hermes

[mikolmogorov]

Could you send me the log file and give some information about the dataset: techonology, genome size, coverage etc?

[hermeseduardo]

sure, pacbio subreads (RSII) fasta files, coverage about 40X, genome size around 500MB. Bellow the some info about the sbatch and the whole log file.

HE

#SBATCH --mem=1000GB
#SBATCH --cpus-per-task=20

/data/esc003/apps/Flye-flye-devel/bin/flye --pacbio-raw pacbio.fasta --genome-size 550m --threads 20 -m 5000 -o flye

[2018-02-24 01:57:36] root: DEBUG: Genome size: 576716800
[2018-02-24 01:57:36] root: INFO: Running Flye 2.3.2-release
[2018-02-24 01:57:36] root: DEBUG: Cmd: /data/esc003/apps/Flye-flye-devel/bin/flye --pacbio-raw /flush2/esc003/Pacbio_subreads_smartbellremoved_headers_removed.fasta --genome-size 550m --threads 20 -m 5000 -o flye
[2018-02-24 01:57:36] root: INFO: Assembling reads
[2018-02-24 01:57:36] root: DEBUG: -----Begin assembly log------
[2018-02-24 01:57:36] root: DEBUG: Running: flye-assemble -l /flush1/esc003/Flye_cynegetis_assembly/flye/flye.log -t 20 -v 5000 /flush2/esc003/Pacbio_subreads_smartbellremoved_headers_removed.fasta /flush1/esc003/Flye_cynegetis_assembly/flye/0-assembly/draft_assembly.fasta 576716800 /data/esc003/apps/Flye-flye-devel/flye/resource/asm_raw_reads.cfg
[2018-02-24 01:57:36] DEBUG: Build date: Feb 23 2018 21:50:23
[2018-02-24 01:57:36] DEBUG: Parameters:
[2018-02-24 01:57:36] DEBUG: kmer_size=15
[2018-02-24 01:57:36] DEBUG: kmer_size_big=17
[2018-02-24 01:57:36] DEBUG: big_genome_threshold=50000000
[2018-02-24 01:57:36] DEBUG: maximum_jump=1500
[2018-02-24 01:57:36] DEBUG: maximum_overhang=1500
[2018-02-24 01:57:36] DEBUG: hard_min_coverage_rate=10
[2018-02-24 01:57:36] DEBUG: repeat_coverage_rate=10
[2018-02-24 01:57:36] DEBUG: jump_divergence_rate=2
[2018-02-24 01:57:36] DEBUG: overlap_divergence_rate=5
[2018-02-24 01:57:36] DEBUG: penalty_window=100
[2018-02-24 01:57:36] DEBUG: max_coverage_drop_rate=5
[2018-02-24 01:57:36] DEBUG: chimera_window=100
[2018-02-24 01:57:36] DEBUG: min_reads_in_contig=4
[2018-02-24 01:57:36] DEBUG: max_inner_reads=10
[2018-02-24 01:57:36] DEBUG: max_inner_fraction=0.25
[2018-02-24 01:57:36] DEBUG: max_separation=500
[2018-02-24 01:57:36] DEBUG: tip_length_threshold=20000
[2018-02-24 01:57:36] DEBUG: unique_edge_length=50000
[2018-02-24 01:57:36] DEBUG: min_repeat_res_support=0.5
[2018-02-24 01:57:36] DEBUG: out_paths_ratio=5
[2018-02-24 01:57:36] DEBUG: graph_cov_drop_rate=10
[2018-02-24 01:57:36] DEBUG: coverage_estimate_window=100
[2018-02-24 01:57:36] DEBUG: low_cutoff_warning=1
[2018-02-24 01:57:36] DEBUG: assemble_kmer_sample=1
[2018-02-24 01:57:36] DEBUG: assemble_gap=500
[2018-02-24 01:57:36] DEBUG: repeat_graph_kmer_sample=5
[2018-02-24 01:57:36] DEBUG: repeat_graph_gap=100
[2018-02-24 01:57:36] DEBUG: repeat_graph_max_kmer=500
[2018-02-24 01:57:36] DEBUG: read_align_kmer_sample=1
[2018-02-24 01:57:36] DEBUG: read_align_gap=500
[2018-02-24 01:57:36] DEBUG: read_align_max_kmer=500
[2018-02-24 01:57:36] DEBUG: Running with k-mer size: 17
[2018-02-24 01:57:36] INFO: Reading sequences
[2018-02-24 02:05:51] DEBUG: Mean read length: 8958
[2018-02-24 02:05:51] DEBUG: Estimated coverage: 46
[2018-02-24 02:05:51] INFO: Generating solid k-mer index
[2018-02-24 02:05:51] DEBUG: Hard threshold set to 4
[2018-02-24 02:05:51] DEBUG: Started kmer counting
[2018-02-24 02:08:01] INFO: Counting kmers (1/2):
[2018-02-24 02:15:22] INFO: Counting kmers (2/2):
[2018-02-24 02:33:14] DEBUG: Filtered 1247354 repetitive kmers
[2018-02-24 02:33:14] DEBUG: Estimated minimum kmer coverage: 8, 537665765 unique kmers selected
[2018-02-24 02:33:14] INFO: Filling index table
[2018-02-24 02:34:25] DEBUG: Solid kmers: 537665765
[2018-02-24 02:34:25] DEBUG: Kmer index size: 11555644070
[2018-02-24 02:56:33] INFO: Extending reads
[2018-02-24 03:07:29] DEBUG: Mean read coverage: 9
[2018-02-24 03:07:39] DEBUG: Assembled contig 1
With 7 reads
Start read: +m170808_50191
At position: 2
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 4
Inner reads: 0
Length: 31157
[2018-02-24 03:07:39] DEBUG: Inner: 16 covered: 88 total: 6005312
[2018-02-24 03:07:42] DEBUG: Assembled contig 2
With 13 reads
Start read: -m170713_180044
At position: 1
leftTip: 1 rightTip: 1
Suspicios: 1
Mean extensions: 3
Inner reads: 0
Length: 79404
[2018-02-24 03:07:42] DEBUG: Inner: 82 covered: 200 total: 6005312
[2018-02-24 03:07:44] DEBUG: Assembled contig 3
With 12 reads
Start read: +m170515_163220
At position: 9
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 2
Inner reads: 0
Length: 83051
[2018-02-24 03:07:44] DEBUG: Inner: 124 covered: 306 total: 6005312
[2018-02-24 03:07:45] DEBUG: Assembled contig 4
With 11 reads
Start read: +m170712_152947
At position: 2
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 4
Inner reads: 0
Length: 75947
[2018-02-24 03:07:45] DEBUG: Inner: 196 covered: 429 total: 6005312
[2018-02-24 03:07:49] DEBUG: Assembled contig 5
With 20 reads
Start read: +m170516_157558
At position: 18
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 4
Inner reads: 0
Length: 116272
[2018-02-24 03:07:49] DEBUG: Inner: 312 covered: 658 total: 6005312
[2018-02-24 03:07:58] DEBUG: Assembled contig 6
With 30 reads
Start read: -m170713_72622
At position: 16
leftTip: 1 rightTip: 1
Suspicios: 5
Mean extensions: 3
Inner reads: 0
Length: 157831
[2018-02-24 03:07:58] DEBUG: Inner: 444 covered: 972 total: 6005312
[2018-02-24 03:08:00] DEBUG: Assembled contig 7
With 25 reads
Start read: +m170516_207012
At position: 12
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 5
Inner reads: 0
Length: 170570
[2018-02-24 03:08:00] DEBUG: Inner: 600 covered: 1393 total: 6005312
[2018-02-24 03:08:01] DEBUG: Assembled contig 8
With 22 reads
Start read: -m170713_59169
At position: 13
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 4
Inner reads: 0
Length: 139444
[2018-02-24 03:08:01] DEBUG: Inner: 734 covered: 1720 total: 6005312
[2018-02-24 03:08:04] DEBUG: Assembled contig 9
With 23 reads
Start read: -m170809_48452
At position: 10
leftTip: 1 rightTip: 0
Suspicios: 2
Mean extensions: 4
Inner reads: 0
Length: 147044
[2018-02-24 03:08:04] DEBUG: Inner: 852 covered: 1976 total: 6005312
[2018-02-24 03:08:04] DEBUG: Assembled contig 10
With 22 reads
Start read: -m170712_111658
At position: 8
leftTip: 1 rightTip: 1
Suspicios: 2
Mean extensions: 6
Inner reads: 0
Length: 120833
[2018-02-24 03:08:04] DEBUG: Inner: 980 covered: 2388 total: 6005312
[2018-02-24 03:08:08] DEBUG: Assembled contig 11
With 14 reads
Start read: -m170609_42878
At position: 8
leftTip: 1 rightTip: 1
Suspicios: 3
Mean extensions: 5
Inner reads: 0
Length: 81458
[2018-02-24 03:08:08] ERROR: Caught unhandled exception: Automatic expansion triggered when load factor was below minimum threshold
[2018-02-24 03:08:08] ERROR: flye-assemble(_Z16exceptionHandlerv+0x2d) [0x43cc8d]
[2018-02-24 03:08:08] ERROR: /usr/lib64/libstdc++.so.6(+0x96706) [0x2aaaab277706]
[2018-02-24 03:08:08] ERROR: /usr/lib64/libstdc++.so.6(+0x96751) [0x2aaaab277751]
[2018-02-24 03:08:08] ERROR: /usr/lib64/libstdc++.so.6(+0xc1708) [0x2aaaab2a2708]
[2018-02-24 03:08:08] ERROR: /lib64/libpthread.so.0(+0x8744) [0x2aaaab789744]
[2018-02-24 03:08:08] ERROR: /lib64/libc.so.6(clone+0x6d) [0x2aaaaba87aad]
-----------End assembly log------------
[2018-02-24 03:08:24] root: ERROR: Command '['flye-assemble', '-l', '/flush1/esc003/Flye_cynegetis_assembly/flye/flye.log', '-t', '20', '-v', '5000', '/flush2/esc003/Pacbio_subreads_smartbellremoved_headers_removed.fasta', '/flush1/esc003/Flye_cynegetis_assembly/flye/0-assembly/draft_assembly.fasta', '576716800', '/data/esc003/apps/Flye-flye-devel/flye/resource/asm_raw_reads.cfg']' returned non-zero exit status 1

[mikolmogorov]

Thanks,

It seems strange, we have never encountered issues with the newer hash function before. Potentially, it could be machine/OS specific. Could you provide "uname -a" output? Do you also have a possibility to run the dataset on a different machine?

[hermeseduardo]

sure:
uname -a
Linux XXX 4.4.59-92.17-default #1 SMP Thu Apr 6 14:16:09 UTC 2017 (7bc489d) x86_64 x86_64 x86_64 GNU/Linux

[mikolmogorov]

Thanks,

I was not able to reproduce this problem on our datasets so far, so it makes the problem hard to debug. Is it possible for you to share the data - that would make debugging much easier? If so, you can write me to fenderglass@gmail.com

[hermeseduardo]

looks like it was something to do with the installation in the cluster, it seems to be working now (v 2.3.2).

thanks