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test_pick_open_reference_otus.py
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test_pick_open_reference_otus.py
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#!/usr/bin/env python
# File created on 02 Nov 2011
from __future__ import division
__author__ = "Greg Caporaso"
__copyright__ = "Copyright 2011, The QIIME project"
__credits__ = ["Greg Caporaso"]
__license__ = "GPL"
__version__ = "1.9.0-dev"
__maintainer__ = "Greg Caporaso"
__email__ = "gregcaporaso@gmail.com"
from glob import glob
from os import chdir, getcwd
from os.path import exists
from shutil import rmtree
from tempfile import mkdtemp
from unittest import TestCase, main
from skbio.tree import TreeNode
from skbio.util import remove_files
from biom import load_table
from qiime.util import load_qiime_config, count_seqs, get_qiime_temp_dir
from qiime.workflow.util import (call_commands_serially, no_status_updates,
WorkflowError)
from qiime.parse import parse_qiime_parameters, fields_to_dict
from qiime.test import initiate_timeout, disable_timeout, get_test_data_fps
from qiime.workflow.pick_open_reference_otus import (
pick_subsampled_open_reference_otus,
iterative_pick_subsampled_open_reference_otus,
final_repset_from_iteration_repsets,
convergent_pick_subsampled_open_reference_otus)
from bfillings.sortmerna_v2 import build_database_sortmerna
allowed_seconds_per_test = 120
class PickSubsampledReferenceOtusThroughOtuTableTests(TestCase):
""" """
def setUp(self):
"""
"""
self.start_dir = getcwd()
self.qiime_config = load_qiime_config()
self.dirs_to_remove = []
self.files_to_remove = []
self.tmp_dir = get_qiime_temp_dir()
self.test_data = get_test_data_fps()
# override default reference data for reduced runtime and better
# defined tests
self.params = parse_qiime_parameters(
["assign_taxonomy:id_to_taxonomy_fp %s"
% self.test_data['refseqs_tax'][0],
"assign_taxonomy:reference_seqs_fp %s"
% self.test_data['refseqs'][0],
"align_seqs:template_fp %s"
% self.test_data['refseqs_aligned'][0],
"filter_alignment:lane_mask_fp %s"
% self.test_data['refseqs_aligned_lanemask'][0]])
self.wf_out = mkdtemp(dir=self.tmp_dir,
prefix='qiime_wf_out', suffix='')
self.dirs_to_remove.append(self.wf_out)
initiate_timeout(allowed_seconds_per_test)
def tearDown(self):
""" """
# turn off the alarm
disable_timeout()
# change back to the start dir - some workflows change directory
chdir(self.start_dir)
remove_files(self.files_to_remove)
# remove directories last, so we don't get errors
# trying to remove files which may be in the directories
for d in self.dirs_to_remove:
if exists(d):
rmtree(d)
def test_pick_subsampled_open_reference_otus_step_1_and_4(self):
"""pick_subsampled_open_reference_otus functions as expected
"""
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=0.60,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
status_update_callback=no_status_updates,
minimum_failure_threshold=100000)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
prefilter_failures_fp = glob(
'%s/prefilter_otus/*_failures.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(prefilter_failures_fp),
"Prefilter failures file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of seequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# ... and that number is 6
self.assertEqual(len(otu_table.ids(axis='observation')), 6)
# the correct sequences failed the prefilter
prefilter_failure_ids = [s.strip()
for s in open(prefilter_failures_fp, 'U')]
self.assertEqual(len(prefilter_failure_ids), 24)
self.assertTrue('t1_1' in prefilter_failure_ids)
self.assertTrue('p1_2' in prefilter_failure_ids)
self.assertTrue('not16S.1_130' in prefilter_failure_ids)
self.assertTrue('not16S.1_151' in prefilter_failure_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
#
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.CleanUp.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.CleanUp.ReferenceOTU0) is not in "
"the final OTU map.")
# confirm that number of tips in the tree is the same as the number of
# sequences in the alignment
with open(tree_fp) as f:
num_tree_tips = len(list(TreeNode.from_newick(f).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
self.assertEqual(num_tree_tips, 6)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata, and are at least assigned
# to 'bacteria'
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_step_1_through_4(self):
"""pick_subsampled_open_reference_otus functions as expected
"""
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=0.60,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
status_update_callback=no_status_updates,
minimum_failure_threshold=0)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
prefilter_failures_fp = glob(
'%s/prefilter_otus/*_failures.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(prefilter_failures_fp),
"Prefilter failures file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of seequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# ... and that number is 6
self.assertEqual(len(otu_table.ids(axis='observation')), 6)
# the correct sequences failed the prefilter
prefilter_failure_ids = [s.strip()
for s in open(prefilter_failures_fp, 'U')]
self.assertEqual(len(prefilter_failure_ids), 24)
self.assertTrue('t1_1' in prefilter_failure_ids)
self.assertTrue('p1_2' in prefilter_failure_ids)
self.assertTrue('not16S.1_130' in prefilter_failure_ids)
self.assertTrue('not16S.1_151' in prefilter_failure_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
#
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.ReferenceOTU0) is not in "
"the final OTU map.")
# confirm that number of tips in the tree is the same as the number of
# sequences in the alignment
with open(tree_fp) as f:
num_tree_tips = len(list(TreeNode.from_newick(f).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
self.assertEqual(num_tree_tips, 6)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata, and are at least assigned
# to 'bacteria'
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_rdp_tax_assign(self):
"""pick_subsampled_open_reference_otus fns when assigning tax with rdp
"""
self.params.update(
parse_qiime_parameters(
['assign_taxonomy:assignment_method rdp',
'assign_taxonomy:reference_seqs_fp %s'
% self.test_data['refseqs'][0],
'assign_taxonomy:id_to_taxonomy_fp %s'
% self.test_data['refseqs_tax'][0]]))
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=0.60,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
status_update_callback=no_status_updates,
minimum_failure_threshold=0)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
prefilter_failures_fp = glob(
'%s/prefilter_otus/*_failures.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
rdp_tax_fp = ('%s/rdp_assigned_taxonomy/rep_set_tax_assignments.txt'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(prefilter_failures_fp),
"Prefilter failures file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
self.assertTrue(
exists(rdp_tax_fp),
"rdp taxonomy assignment result doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of seequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# ... and that number is 6
self.assertEqual(len(otu_table.ids(axis='observation')), 6)
# the correct sequences failed the prefilter
prefilter_failure_ids = [s.strip()
for s in open(prefilter_failures_fp, 'U')]
self.assertEqual(len(prefilter_failure_ids), 24)
self.assertTrue('t1_1' in prefilter_failure_ids)
self.assertTrue('p1_2' in prefilter_failure_ids)
self.assertTrue('not16S.1_130' in prefilter_failure_ids)
self.assertTrue('not16S.1_151' in prefilter_failure_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
#
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.ReferenceOTU0) is not in "
"the final OTU map.")
# confirm that number of tips in the tree is the same as the number
# of sequences in the alignment
with open(tree_fp) as f:
num_tree_tips = len(list(TreeNode.from_newick(f).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
self.assertEqual(num_tree_tips, 6)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_usearch(self):
"""pick_subsampled_open_reference_otus functions as expected with
usearch
"""
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=0.60,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
denovo_otu_picking_method='usearch61',
reference_otu_picking_method='usearch61_ref',
status_update_callback=no_status_updates,
minimum_failure_threshold=0)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
prefilter_failures_fp = glob(
'%s/prefilter_otus/*_failures.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(prefilter_failures_fp),
"Prefilter failures file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of seequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# ... and that number is 6
self.assertEqual(len(otu_table.ids(axis='observation')), 6)
# the correct sequences failed the prefilter
prefilter_failure_ids = [s.strip()
for s in open(prefilter_failures_fp, 'U')]
self.assertEqual(len(prefilter_failure_ids), 24)
self.assertTrue('t1_1' in prefilter_failure_ids)
self.assertTrue('p1_2' in prefilter_failure_ids)
self.assertTrue('not16S.1_130' in prefilter_failure_ids)
self.assertTrue('not16S.1_151' in prefilter_failure_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
#
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.ReferenceOTU0) is not in "
"the final OTU map.")
# confirm that number of tips in the tree is the same as the number of
# sequences in the alignment
num_tree_tips = len(list(TreeNode.from_newick(open(tree_fp)).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
self.assertEqual(num_tree_tips, 6)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata, and are at least assigned
# to 'bacteria'
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_sortmerna_sumaclust(self):
"""pick_subsampled_open_reference_otus functions as expected with
sortmerna and sumaclust (input reference database not indexed),
the minimum_failure_threshold=100000 so steps 2 and 3 are skipped
and all failures.fasta from step 1 are directly passed for de novo
clustering in step 4 """
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=None,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
denovo_otu_picking_method='sumaclust',
reference_otu_picking_method='sortmerna',
status_update_callback=no_status_updates,
minimum_failure_threshold=100000)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
reads_for_denovo_fp = glob(
'%s/step1_otus/*_failures.txt' %
self.wf_out)[0]
reads_for_otumap_fp = glob(
'%s/step1_otus/*_otus.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(reads_for_denovo_fp),
"Denovo reads file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of sequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# the correct sequences failed the prefilter (E-value)
# reads_for_denovo_ids all passed E-value and are considered
# as true rRNA (having <97% id and/or <97% query coverage)
reads_for_denovo_ids = [s.strip()
for s in open(reads_for_denovo_fp, 'U')]
self.assertEqual(len(reads_for_denovo_ids), 59)
self.assertTrue('t1_1' not in reads_for_denovo_ids)
self.assertTrue('p1_2' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_130' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_151' not in reads_for_denovo_ids)
# reads_for_otumap_ids all passed E-value and are considered
# as true rRNA (having >=97% id and >=97% query coverage)
reads_for_otumap_ids = [seq_id for line in
open(reads_for_otumap_fp, 'U')
for seq_id in line.strip().split('\t')]
self.assertEqual(len(reads_for_otumap_ids), 120)
self.assertTrue('t1_1' not in reads_for_otumap_ids)
self.assertTrue('p1_2' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_130' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_151' not in reads_for_otumap_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.CleanUp.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.CleanUp.ReferenceOTU0) is not in "
"the final OTU map.")
self.assertTrue('wf.test.otu.CleanUp.ReferenceOTU5' in otu_map,
"Failure OTU (wf.test.otu.CleanUp.ReferenceOTU5) is not in "
"the final OTU map.")
# confirm size of full OTU map
self.assertTrue(len(otu_map), 9)
# confirm that number of tips in the tree is the same as the number of
# sequences in the alignment
num_tree_tips = len(list(TreeNode.from_newick(open(tree_fp)).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata, and are at least assigned
# to 'bacteria'
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_sortmerna_sumaclust_step2_3(self):
"""pick_subsampled_open_reference_otus functions as expected with
sortmerna and sumaclust (input reference database not indexed),
minimum_failure_threshold=0 and percent_subsample=0.7, so a non-empty
subsampled_failures.fasta file is created in step 1 and passed to steps 2/3/4
for analysis. """
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.7,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=None,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
denovo_otu_picking_method='sumaclust',
reference_otu_picking_method='sortmerna',
status_update_callback=no_status_updates,
minimum_failure_threshold=0)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
reads_for_denovo_fp = glob(
'%s/step1_otus/*_failures.txt' %
self.wf_out)[0]
reads_for_otumap_fp = glob(
'%s/step1_otus/*_otus.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(reads_for_denovo_fp),
"Denovo reads file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of sequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# the correct sequences failed the prefilter (E-value)
# reads_for_denovo_ids all passed E-value and are considered
# as true rRNA (having <97% id and/or <97% query coverage)
reads_for_denovo_ids = [s.strip()
for s in open(reads_for_denovo_fp, 'U')]
self.assertEqual(len(reads_for_denovo_ids), 59)
self.assertTrue('t1_1' not in reads_for_denovo_ids)
self.assertTrue('p1_2' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_130' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_151' not in reads_for_denovo_ids)
# reads_for_otumap_ids all passed E-value and are considered
# as true rRNA (having >=97% id and >=97% query coverage)
reads_for_otumap_ids = [seq_id for line in
open(reads_for_otumap_fp, 'U')
for seq_id in line.strip().split('\t')]
self.assertEqual(len(reads_for_otumap_ids), 120)
self.assertTrue('t1_1' not in reads_for_otumap_ids)
self.assertTrue('p1_2' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_130' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_151' not in reads_for_otumap_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus
self.assertEqual(count_seqs(new_refseqs_fp)[0],
count_seqs(self.test_data['refseqs'][0])[0] +
len([o for o in otu_table.ids(axis='observation')
if o.startswith('wf.test.otu')]) +
count_seqs(pynast_failures_fp)[0])
# spot check a few of the otus to confirm that we're getting reference
# and new otus in the final otu map. This is done on the OTU map
# singletons get filtered before building the otu table
otu_map = fields_to_dict(open(otu_map_w_singletons_fp))
self.assertTrue('295053' in otu_map,
"Reference OTU (295053) is not in the final OTU map.")
self.assertTrue('42684' in otu_map,
"Failure OTU (42684) is not in the final OTU map.")
self.assertTrue('wf.test.otu.ReferenceOTU0' in otu_map,
"Failure OTU (wf.test.otu.ReferenceOTU0) is in "
"the final OTU map.")
# confirm size of full OTU map
self.assertTrue(len(otu_map), 11)
# confirm that number of tips in the tree is the same as the number of
# sequences in the alignment
num_tree_tips = len(list(TreeNode.from_newick(open(tree_fp)).tips()))
num_align_seqs = count_seqs(aln_fp)[0]
self.assertEqual(num_tree_tips, num_align_seqs)
# OTU table without singletons or pynast failures has same number of
# otus as there are aligned sequences
otu_table = load_table(otu_table_fp)
self.assertEqual(len(otu_table.ids(axis='observation')),
num_align_seqs)
# Reference OTUs have correct taxonomy assignment (can't confirm the )
obs_idx = otu_table.index('295053', axis='observation')
self.assertEqual(
otu_table.metadata(axis='observation')[obs_idx]['taxonomy'],
["k__Bacteria", "p__Proteobacteria", "c__Gammaproteobacteria",
"o__Enterobacteriales", "f__Enterobacteriaceae", "g__", "s__"])
# All observations have 'taxonomy' metadata, and are at least assigned
# to 'bacteria'
for o in otu_table.iter(axis='observation'):
self.assertTrue(
o[2]['taxonomy'][0] in ['k__Bacteria', 'Unassigned'])
def test_pick_subsampled_open_reference_otus_sortmerna_sumaclust_db(self):
"""pick_subsampled_open_reference_otus functions as expected with
sortmerna and sumaclust (input reference database indexed) """
# Create sortmerna indexed database
sortmerna_db, files_to_remove = \
build_database_sortmerna(self.test_data['refseqs'][0],
max_pos=10000,
output_dir=self.tmp_dir)
self.files_to_remove.extend(files_to_remove)
self.params['pick_otus']['sortmerna_db'] = sortmerna_db
pick_subsampled_open_reference_otus(
input_fp=self.test_data['seqs'][0],
refseqs_fp=self.test_data['refseqs'][0],
output_dir=self.wf_out,
percent_subsample=0.5,
new_ref_set_id='wf.test.otu',
command_handler=call_commands_serially,
params=self.params,
prefilter_refseqs_fp=None,
prefilter_percent_id=None,
qiime_config=self.qiime_config,
step1_otu_map_fp=None,
step1_failures_fasta_fp=None,
parallel=False,
suppress_step4=False,
logger=None,
denovo_otu_picking_method='sumaclust',
reference_otu_picking_method='sortmerna',
status_update_callback=no_status_updates,
minimum_failure_threshold=0)
otu_map_w_singletons_fp = '%s/final_otu_map.txt' % self.wf_out
final_failure_fp = '%s/final_failures.txt' % self.wf_out
final_repset_fp = '%s/rep_set.fna' % self.wf_out
new_refseqs_fp = '%s/new_refseqs.fna' % self.wf_out
reads_for_denovo_fp = glob(
'%s/step1_otus/*_failures.txt' %
self.wf_out)[0]
reads_for_otumap_fp = glob(
'%s/step1_otus/*_otus.txt' %
self.wf_out)[0]
tree_fp = '%s/rep_set.tre' % self.wf_out
aln_fp = '%s/pynast_aligned_seqs/rep_set_aligned.fasta' % self.wf_out
otu_table_fp = ('%s/otu_table_mc2_w_tax_no_pynast_failures.biom'
% self.wf_out)
pynast_failures_fp = ('%s/pynast_aligned_seqs/rep_set_failures.fasta'
% self.wf_out)
self.assertTrue(
exists(otu_map_w_singletons_fp),
"OTU map doesn't exist")
self.assertFalse(exists(final_failure_fp),
"Final failures file shouldn't exist, but it does")
self.assertTrue(
exists(final_repset_fp),
"Final representative set doesn't exist")
self.assertTrue(
exists(new_refseqs_fp),
"New refseqs file doesn't exist")
self.assertTrue(
exists(reads_for_denovo_fp),
"Denovo reads file doesn't exist")
self.assertTrue(exists(tree_fp), "Final tree doesn't exist")
self.assertTrue(exists(aln_fp), "Final alignment doesn't exist")
self.assertTrue(exists(otu_table_fp), "Final BIOM table doesn't exist")
self.assertTrue(
exists(pynast_failures_fp),
"PyNAST failures file doesn't exist")
# all OTUs in final OTU table occur more than once
otu_table = load_table(otu_table_fp)
for row in otu_table.iter_data(axis='observation'):
self.assertTrue(sum(row) >= 2,
"Singleton OTU detected in OTU table.")
# number of OTUs in final OTU table equals the number of sequences in
# the alignment...
self.assertEqual(len(otu_table.ids(axis='observation')),
count_seqs(aln_fp)[0])
# the correct sequences failed the prefilter (E-value)
# reads_for_denovo_ids all passed E-value and are considered
# as true rRNA (having <97% id and/or <97% query coverage)
reads_for_denovo_ids = [s.strip()
for s in open(reads_for_denovo_fp, 'U')]
self.assertEqual(len(reads_for_denovo_ids), 59)
self.assertTrue('t1_1' not in reads_for_denovo_ids)
self.assertTrue('p1_2' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_130' not in reads_for_denovo_ids)
self.assertTrue('not16S.1_151' not in reads_for_denovo_ids)
# reads_for_otumap_ids all passed E-value and are considered
# as true rRNA (having >=97% id and >=97% query coverage)
reads_for_otumap_ids = [seq_id for line in
open(reads_for_otumap_fp, 'U')
for seq_id in line.strip().split('\t')]
self.assertEqual(len(reads_for_otumap_ids), 120)
self.assertTrue('t1_1' not in reads_for_otumap_ids)
self.assertTrue('p1_2' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_130' not in reads_for_otumap_ids)
self.assertTrue('not16S.1_151' not in reads_for_otumap_ids)
# confirm that the new reference sequences is the same length as the
# input reference sequences plus the number of new non-singleton otus