Provided by: swarm_2.2.2+dfsg-1_amd64 
NAME
swarm — find clusters of nearly-identical nucleotide amplicons
SYNOPSIS
swarm [ options ] filename
DESCRIPTION
Environmental or clinical molecular studies generate large volumes of amplicons (e.g., 16S
or 18S SSU-rRNA sequences) that need to be clustered into molecular operational taxonomic
units (OTUs). Common clustering methods are based on greedy, input-order dependent
algorithms, with arbitrary selection of global cluster size and cluster centroids. To
address that problem, we developed swarm, a fast and robust method that recursively groups
amplicons with d or less differences (i.e. substitutions, insertions or deletions). swarm
produces natural and stable clusters centered on local peaks of abundance, mostly free
from input-order dependency induced by centroid selection.
Exact clustering is impractical on large data sets when using a naïve all-vs-all approach
(more precisely a 2-combination without repetitions), as it implies unrealistic numbers of
pairwise comparisons. swarm is based on a maximum number of differences d between two
amplicons, and focuses only on very close local relationships. For d = 1, the default
value, swarm uses an algorithm of linear complexity that generates all possible single
mutations and performs exact-string matching by comparing hash-values. For d = 2 or
greater, swarm uses an algorithm of quadratic complexity that performs pairwise string
comparisons. An efficient k-mer-based filtering and an astute use of comparisons results
obtained during the clustering process allows swarm to avoid most of the amplicon
comparisons needed in a naïve approach. To speed up the remaining amplicon comparisons,
swarm implements an extremely fast Needleman-Wunsch algorithm making use of the Streaming
SIMD Extensions (SSE2) of modern x86-64 CPUs. If SSE2 instructions are not available,
swarm exits with an error message.
swarm can read nucleotide amplicons in fasta format from a normal file or from the
standard input (using a pipe or a redirection). The amplicon identifier is defined as the
string comprised between the '>' symbol and the first space or the end of the line,
whichever comes first. As swarm outputs lists of amplicon identifiers, amplicon
identifiers must be unique to avoid ambiguity; swarm exits with an error message if
identifiers are not unique. Amplicon identifiers must end with a '_' followed by a
positive integer representing the amplicon copy number (or abundance annotation;
usearch/vsearch users can use the option -z to use the ';size=' annotation). Abundance
annotations play a crucial role in the clustering process, and swarm exits with an error
message if that information is not available. The amplicon sequence is defined as a string
of [ACGT] or [ACGU] symbols (case insensitive, 'U' is replaced with 'T' internally),
starting after the end of the identifier line and ending before the next identifier line
or the file end; swarm silently removes newline symbols ('\n' or '\r') and exits with an
error message if any other symbol is present.
General options
-h, --help
display this help and exit successfully.
-t, --threads positive integer
number of computation threads to use. Values between 1 and 256 are accepted, but
we recommend to use a number of threads lesser or equal to the number of
available CPU cores. Default number of threads is 1.
-v, --version
output version information and exit successfully.
-- delimit the option list. Later arguments, if any, are treated as operands even if
they begin with '-'. For example, 'swarm -- -file.fasta' reads from the file
'-file.fasta'.
Clustering options
-d, --differences zero or positive integer
maximum number of differences allowed between two amplicons, meaning that two
amplicons will be grouped if they have integer (or less) differences. This is
swarm's most important parameter. The number of differences is calculated as the
number of mismatches (substitutions, insertions or deletions) between the two
amplicons once the optimal pairwise global alignment has been found (see
'pairwise alignment advanced options' to influence that step). Any integer from
0 to 255 can be used, but high d values will decrease the taxonomical resolution
of swarm results. Commonly used d values are 1, 2 or 3, rarely higher. When using
d = 0, swarm will output results corresponding to a strict dereplication of the
dataset, i.e. merging identical amplicons. Warning, whatever the d value, swarm
requires fasta entries to present abundance values. Default number of differences
d is 1.
-n, --no-otu-breaking
deactivate the built-in OTU refinement (not recommended). Amplicon abundance
values are used to identify transitions among in-contact OTUs and to separate
them, yielding higher-resolution clustering results. That option prevents that
separation, and in practice, allows the creation of a link between amplicons A
and B, even if the abundance of B is higher than the abundance of A.
Fastidious options
-b, --boundary positive integer
when using the option --fastidious (-f), define the minimum mass of a large OTU.
By default, an OTU with a mass of 3 or more is considered large. Conversely, an
OTU is small if it has a mass of less than 3, meaning that it is composed of
either one amplicon of abundance 2, or two amplicons of abundance 1. Any positive
value greater than 1 can be specified. Using higher boundary values will speed up
the second pass, but also reduce the taxonomical resolution of swarm results.
Default mass of a large OTU is 3.
-c, --ceiling positive integer
when using the option --fastidious (-f), define swarm's maximum memory footprint
(in megabytes). swarm will adjust the --bloom-bits (-y) value of the Bloom filter
to fit within the specified amount of memory. The value must be at least 3.
-f, --fastidious
when working with d = 1, perform a second clustering pass to reduce the number of
small OTUs (recommended option). During the first clustering pass, an
intermediate amplicon can be missing for purely stochastic reasons, interrupting
the aggregation process. The fastidious option will create virtual amplicons,
allowing to graft small OTUs upon bigger ones. By default, an OTU is small if it
has a mass of 2 or less (see the --boundary option to modify that value). To
speed things up, swarm uses a Bloom filter to store intermediate results.
Warning, the second clustering pass can be 2 to 3 times slower than the first
pass and requires much more memory to store the virtual amplicons in Bloom
filters. See the options --bloom-bits (-y) or --ceiling (-c) to control the
memory footprint of the Bloom filter. The fastidious option modifies clustering
results: the output files produced by the options --log (-l), --output-file (-o),
--mothur (-r), --uclust-file, and --seeds (-w) are updated to reflect these
modifications; the file --statistics-file (-s) is partially updated (columns 6
and 7 are not updated); the output file --internal-structure (-i) is partially
updated (column 5 is not updated for amplicons that belonged to the small OTU).
-y, --bloom-bits positive integer
when using the option --fastidious (-f), define the size (in bits) of each entry
in the Bloom filter. That option allows to balance the efficiency (i.e. speed)
and the memory footprint of the Bloom filter. Large values will make the Bloom
filter more efficient but will require more memory. Any value between 2 and 64
can be used. Default value is 16. See the --ceiling (-c) option for an
alternative way to control the memory footprint.
Input/output options
-a, --append-abundance positive integer
set abundance value to use when some or all amplicons in the input file lack
abundance values (_integer, or ;size=integer; when using -z). Warning, it is not
recommended to use swarm on datasets where abundance values are all identical. We
provide that option as a courtesy to advanced users, please use it carefully.
swarm exits with an error message if abundance values are missing and if this
option is not used.
-i, --internal-structure filename
output all pairs of nearly-identical amplicons to filename using a five-columns
tab-delimited format:
1. amplicon A label.
2. amplicon B label.
3. number of differences between amplicons A and B (positive integer).
4. OTU number (positive integer). OTUs are numbered in their order of
delineation, starting from 1. All pairs of amplicons belonging to the
same OTU will receive the same number.
5. cummulated number of steps from the OTU seed to amplicon B (positive
integer). When using the option --fastidious (-f), the actual number
of steps between grafted amplicons and the OTU seed cannot be re-
computed efficiently and is always set to 2 for the amplicon pair
linking the small OTU to the big OTU. Cummulated number of steps in
the small OTU (if any) are left unchanged.
-l, --log filename
output all messages to filename instead of standard error, with the exception of
error messages of course. That option is useful in situations where writing to
standard error is problematic (for example, with certain job schedulers).
-o, --output-file filename
output clustering results to filename. Results consist of a list of OTUs, one OTU
per line. An OTU is a list of amplicon identifiers separated by spaces. That
output format can be modified by the option --mothur (-r). Default is to write to
standard output.
-r, --mothur
output clustering results in a format compatible with Mothur. That option
modifies swarm's default output format.
-s, --statistics-file filename
output statistics to filename. The file is a tab-separated table with one OTU per
row and seven columns of information:
1. number of unique amplicons in the OTU,
2. total abundance of amplicons in the OTU,
3. identifier of the initial seed,
4. initial seed abundance,
5. number of amplicons with an abundance of 1 in the OTU,
6. maximum number of iterations before the OTU reached its natural limit,
7. cummulated number of steps along the path joining the seed and the
furthermost amplicon in the OTU. Please note that the actual number of
differences between the seed and the furthermost amplicon is usually
much smaller. When using the option --fastidious (-f), grafted
amplicons are not taken into account.
-u, --uclust-file filename
output clustering results in filename using a tab-separated uclust-like format
with 10 columns and 3 different type of entries (S, H or C). That option does not
modify swarm's default output format. Each fasta sequence in the input file can
be either a cluster centroid (S) or a hit (H) assigned to a cluster. Cluster
records (C) summarize information (size, centroid label) for each cluster. Column
content varies with the type of entry (S, H or C):
1. Record type: S, H, or C.
2. Cluster number (zero-based).
3. Centroid length (S), query length (H), or cluster size (C).
4. Percentage of similarity with the centroid sequence (H), or set to '*'
(S, C).
5. Match orientation + or - (H), or set to '*' (S, C).
6. Not used, always set to '*' (S, C) or to zero (H).
7. Not used, always set to '*' (S, C) or to zero (H).
8. set to '*' (S, C) or, for H, compact representation of the pairwise
alignment using the CIGAR format (Compact Idiosyncratic Gapped
Alignment Report): M (match), D (deletion) and I (insertion). The
equal sign '=' indicates that the query is identical to the centroid
sequence.
9. Label of the query sequence (H), or of the centroid sequence (S, C).
10. Label of the centroid sequence (H), or set to '*' (S, C).
-w, --seeds filename
output OTU representatives to filename in fasta format. The abundance value of
each OTU representative is the sum of the abundances of all the amplicons in the
OTU.
-z, --usearch-abundance
accept amplicon abundance values in usearch/vsearch's style
(>label;size=integer[;]). That option influences the abundance annotation style
used in output files.
Pairwise alignment advanced options
when using d > 1, swarm recognizes advanced command-line options modifying the pairwise
global alignment scoring parameters:
-m, --match-reward positive integer
Default reward for a nucleotide match is 5.
-p, --mismatch-penalty positive integer
Default penalty for a nucleotide mismatch is 4.
-g, --gap-opening-penalty positive integer
Default gap opening penalty is 12.
-e, --gap-extension-penalty positive integer
Default gap extension penalty is 4.
As swarm focuses on close relationships (e.g., d = 2 or 3), clustering results are
resilient to pairwise alignment model parameters modifications. When clustering using a
higher d value, modifying model parameters has a stronger impact.
EXAMPLES
Clusterize the data set myfile.fasta into OTUs with the finest resolution possible (1
difference, built-in breaking, fastidious option) using 4 computation threads. OTUs are
written to the file myfile.swarms, and OTU representatives are written to
myfile.representatives.fasta.
swarm -t 4 -f -w myfile.representatives.fasta < myfile.fasta > myfile.swarms
AUTHORS
Concept by Frédéric Mahé, implementation by Torbjørn Rognes.
CITATION
Mahé F, Rognes T, Quince C, de Vargas C, Dunthorn M. (2014) Swarm: robust and fast
clustering method for amplicon-based studies. PeerJ 2:e593
<https://doi.org/10.7717/peerj.593>
Mahé F, Rognes T, Quince C, de Vargas C, Dunthorn M. (2015) Swarm v2: highly-scalable and
high-resolution amplicon clustering. PeerJ 3:e1420 <https://doi.org/10.7717/peerj.1420>
REPORTING BUGS
Submit suggestions and bug-reports at <https://github.com/torognes/swarm/issues>, send a
pull request on <https://github.com/torognes/swarm>, or compose a friendly or curmudgeonly
e-mail to Frédéric Mahé <mahe@rhrk.uni-kl.de> and Torbjørn Rognes <torognes@ifi.uio.no>.
AVAILABILITY
Source code and binaries are available at <https://github.com/torognes/swarm>
COPYRIGHT
Copyright (C) 2012-2017 Frédéric Mahé & Torbjørn Rognes
This program is free software: you can redistribute it and/or modify it under the terms of
the GNU Affero General Public License as published by the Free Software Foundation, either
version 3 of the License, or any later version.
This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY;
without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE.
See the GNU Affero General Public License for more details.
You should have received a copy of the GNU Affero General Public License along with this
program. If not, see <http://www.gnu.org/licenses/>.
SEE ALSO
swipe, an extremely fast Smith-Waterman database search tool by Torbjørn Rognes (available
from <https://github.com/torognes/swipe>).
vsearch, an open-source re-implementation of the classic uclust clustering method (by
Robert C. Edgar), along with other amplicon filtering and searching tools. vsearch is
implemented by Torbjørn Rognes and documented by Frédéric Mahé, and is available at
<https://github.com/torognes/vsearch>.
VERSION HISTORY
New features and important modifications of swarm (short lived or minor bug releases are
not mentioned):
v2.2.2 released December 12, 2017
Version 2.2.2 fixes a bug that would cause Swarm to wait forever in very
rare cases when multiple threads were used.
v2.2.1 released October 27, 2017
Version 2.2.1 fixes a memory allocation bug for d = 1 and duplicated
sequences.
v2.2.0 released October 17, 2017
Version 2.2.0 fixes several problems and improves usability. Corrected
output to structure and uclust files when using fastidious mode. Corrected
abundance output in some cases. Added check for duplicated sequences and
fixed check for duplicated sequence IDs. Checks for empty sequences. Sorts
sequences by additional fields to improve stability. Improves compatibility
with compilers and operating systems. Outputs sequences in upper case.
Allows 64-bit abundances. Shows message when waiting for input from stdin.
Improves error messages and warnings. Improves checking of command line
options. Fixes remaining errors reported by test suite. Updates
documentation.
v2.1.13 released March 8, 2017
Version 2.1.13 removes a bug with the progress bar when writing seeds.
v2.1.12 released January 16, 2017
Version 2.1.12 removes a debugging message.
v2.1.11 released January 16, 2017
Version 2.1.11 fixes two bugs related to the SIMD implementation of
alignment that might result in incorrect alignments and scores. The bug
only applies when d > 1.
v2.1.10 released December 22, 2016
Version 2.1.10 fixes two bugs related to gap penalties of alignments. The
first bug may lead to wrong aligments and similarity percentages reported in
UCLUST (.uc) files. The second bug makes Swarm use a slightly higher gap
extension penalty than specified. The default gap extension penalty used
have actually been 4.5 instead of 4.
v2.1.9 released July 6, 2016
Version 2.1.9 fixes errors when compiling with GCC version 6.
v2.1.8 released March 11, 2016
Version 2.1.8 fixes a rare bug triggered when clustering extremely short
undereplicated sequences. Also, alignment parameters are not shown when d =
1.
v2.1.7 released February 24, 2016
Version 2.1.7 fixes a bug in the output of seeds with the -w option when d >
1 that was not properly fixed in version 2.1.6. It also handles ascii
character #13 (CR) in FASTA files better. Swarm will now exit with status 0
if the -h or the -v option is specified. The help text and some error
messages have been improved.
v2.1.6 released December 14, 2015
Version 2.1.6 fixes problems with older compilers that do not have the
x86intrin.h header file. It also fixes a bug in the output of seeds with the
-w option when d > 1.
v2.1.5 released September 8, 2015
Version 2.1.5 fixes minor bugs.
v2.1.4 released September 4, 2015
Version 2.1.4 fixes minor bugs in the swarm algorithm used for d = 1.
v2.1.3 released August 28, 2015
Version 2.1.3 adds checks of numeric option arguments.
v2.1.1 released March 31, 2015
Version 2.1.1 fixes a bug with the fastidious option that caused it to
ignore some connections between large and small OTUs.
v2.1.0 released March 24, 2015
Version 2.1.0 marks the first official release of swarm v2.
v2.0.7 released March 18, 2015
Version 2.0.7 writes abundance information in usearch style when using
options -w (--seeds) in combination with -z (--usearch-abundance).
v2.0.6 released March 13, 2015
Version 2.0.6 fixes a minor bug.
v2.0.5 released March 13, 2015
Version 2.0.5 improves the implementation of the fastidious option and adds
options to control memory usage of the Bloom filter (-y and -c). In
addition, an option (-w) allows to output OTU representatives sequences with
updated abundances (sum of all abundances inside each OTU). This version
also enables swarm to run with d = 0.
v2.0.4 released March 6, 2015
Version 2.0.4 includes a fully parallelised implementation of the fastidious
option.
v2.0.3 released March 4, 2015
Version 2.0.3 includes a working implementation of the fastidious option,
but only the initial clustering is parallelized.
v2.0.2 released February 26, 2015
Version 2.0.2 fixes SSSE3 problems.
v2.0.1 released February 26, 2015
Version 2.0.1 is a development version that contains a partial
implementation of the fastidious option, but it is not usable yet.
v2.0.0 released December 3, 2014
Version 2.0.0 is faster and easier to use, providing new output options
(--internal-structure and --log), new control options (--boundary,
--fastidious, --no-otu-breaking), and built-in OTU refinement (no need to
use the python script anymore). When using default parameters, a novel and
considerably faster algorithmic approach is used, guaranteeing swarm's
scalability.
v1.2.21 released February 26, 2015
Version 1.2.21 is supposed to fix some problems related to the use of the
SSSE3 CPU instructions which are not always available.
v1.2.20 released November 6, 2014
Version 1.2.20 presents a production-ready version of the alternative
algorithm (option -a), with optional built-in OTU breaking (option -n). That
alternative algorithmic approach (usable only with d = 1) is considerably
faster than currently used clustering algorithms, and can deal with datasets
of 100 million unique amplicons or more in a few hours. Of course, results
are rigourously identical to the results previously produced with swarm.
That release also introduces new options to control swarm output (options -i
and -l).
v1.2.19 released October 3, 2014
Version 1.2.19 fixes a problem related to abundance information when the
sequence identifier includes multiple underscore characters.
v1.2.18 released September 29, 2014
Version 1.2.18 reenables the possibility of reading sequences from stdin if
no file name is specified on the command line. It also fixes a bug related
to CPU features detection.
v1.2.17 released September 28, 2014
Version 1.2.17 fixes a memory allocation bug introduced in version 1.2.15.
v1.2.16 released September 27, 2014
Version 1.2.16 fixes a bug in the abundance sort introduced in version
1.2.15.
v1.2.15 released September 27, 2014
Version 1.2.15 sorts the input sequences in order of decreasing abundance
unless they are detected to be sorted already. When using the alternative
algorithm for d = 1 it also sorts all subseeds in order of decreasing
abundance.
v1.2.14 released September 27, 2014
Version 1.2.14 fixes a bug in the output with the --swarm_breaker option
(-b) when using the alternative algorithm (-a).
v1.2.12 released August 18, 2014
Version 1.2.12 introduces an option --alternative-algorithm to use an
extremely fast, experimental clustering algorithm for the special case d =
1. Multithreading scalability of the default algorithm has been noticeably
improved.
v1.2.10 released August 8, 2014
Version 1.2.10 allows amplicon abundances to be specified using the usearch
style in the sequence header (e.g. '>id;size=1') when the -z option is
chosen.
v1.2.8 released August 5, 2014
Version 1.2.8 fixes an error with the gap extension penalty. Previous
versions used a gap penalty twice as large as intended. That bug correction
induces small changes in clustering results.
v1.2.6 released May 23, 2014
Version 1.2.6 introduces an option --mothur to output clustering results in
a format compatible with the microbial ecology community analysis software
suite Mothur (<http://www.mothur.org/>).
v1.2.5 released April 11, 2014
Version 1.2.5 removes the need for a POPCNT hardware instruction to be
present. swarm now automatically checks whether POPCNT is available and uses
a slightly slower software implementation if not. Only basic SSE2
instructions are now required to run swarm.
v1.2.4 released January 30, 2014
Version 1.2.4 introduces an option --break-swarms to output all pairs of
amplicons with d differences to standard error. That option is used by the
companion script `swarm_breaker.py` to refine swarm results. The syntax of
the inline assembly code is changed for compatibility with more compilers.
v1.2 released May 16, 2013
Version 1.2 greatly improves speed by using alignment-free comparisons of
amplicons based on k-mer word content. For each amplicon, the presence-
absence of all possible 5-mers is computed and recorded in a 1024-bits
vector. Vector comparisons are extremely fast and drastically reduce the
number of costly pairwise alignments performed by swarm. While remaining
exact, swarm 1.2 can be more than 100-times faster than swarm 1.1, when
using a single thread with a large set of sequences. The minor version
1.1.1, published just before, adds compatibility with Apple computers, and
corrects an issue in the pairwise global alignment step that could lead to
sub-optimal alignments.
v1.1 released February 26, 2013
Version 1.1 introduces two new important options: the possibility to output
clustering results using the uclust output format, and the possibility to
output detailed statistics on each OTU. swarm 1.1 is also faster: new
filterings based on pairwise amplicon sequence lengths and composition
comparisons reduce the number of pairwise alignments needed and speed up the
clustering.
v1.0 released November 10, 2012
First public release.