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/* BaitFisher (version 1.2.8) a program for designing DNA target enrichment baits
* BaitFilter (version 1.0.6) a program for selecting optimal bait regions
* Copyright 2013-2017 by Christoph Mayer
*
* This source file is part of the BaitFisher-package.
*
* This program is free software: you can redistribute it and/or modify
* it under the terms of the GNU General Public License as published by
* the Free Software Foundation, either version 3 of the License, or
* (at your option) 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 General Public License for more details.
*
* You should have received a copy of the GNU General Public License
* along with BaitFisher. If not, see <http://www.gnu.org/licenses/>.
*
*
* For any enquiries send an Email to Christoph Mayer
* c.mayer.zfmk@uni-bonn.de
*
* When publishing work that is based on the results please cite:
* Mayer et al. 2016: BaitFisher: A software package for multi-species target DNA enrichment probe design
*
*/
#include <iostream>
#include <fstream>
#include <ctime>
#include "global-types-and-parameters.h"
#include "CBaitRecord.h"
#include "faststring2.h"
#include "CBlastParser.h"
#include "print_container.h"
#include "Ctriple.h"
#include "climits"
using namespace std;
bool skip_blast = false;
unsigned start_time = 0;
unsigned blast_time = 0;
// Verbosity rules:
// verbosity 0 -> only error messages that lead to exit()
// verbosity 1 -> only warnings
// verbosity 2 -> information on baits, time used
// verbosity 3 -> progress
// verbosity 4 -> more progress
// verbosity
void welcome(std::ostream &os)
{
os << endl << endl;
os << "Welcome to " << PROGNAME << ", version " << VERSION << ".\n\n";
}
void good_bye(std::ostream &os)
{
os << "Your BaitFilter analysis finished successfully.\n";
}
void get_hours_minutes_seconds(unsigned secs, unsigned &h, unsigned &m, unsigned &s)
{
h = secs/3600;
secs %= 3600;
m = secs/60;
s = secs%60;
}
void print_time(unsigned secs, ostream &os)
{
char prev = os.fill('0');
unsigned h, m, s;
get_hours_minutes_seconds(secs, h, m, s);
os << h << ":";
os.width(2);
os << m << ":";
os.width(2);
os << s;
os.fill(prev);
}
void print_stats(const CBaitLoci_collection &blc, const char * header= NULL)
{
if (global_verbosity >= 2)
{
CBaitRegionSelectorSets brss;
if (header)
cout << header << "\n";
blc.fill_locus_selector(brss);
brss.print_num_genes_num_features(cout, " ", blc.has_feature_information());
unsigned NumSeqs, NumBaits;
blc.get_seqs_baits_count(NumSeqs, NumBaits);
// cout << " Number of bait regions: " << blc.get_number_of_loci() << "\n";
cout << " Total number of baits: " << NumBaits << "\n";
cout << " Total number of bait windows fully covered by sequences " << NumSeqs << "\n";
cout << " Proportion of baits saved (1-#baits/#sequences): " << (1.0 -(double)NumBaits/NumSeqs)*100 << "%\n";
cout.precision(6);
cout << " Mean maximum distance of baits to MSAs: " << blc.get_max_dist_mean() << "\n";
cout << " Overall maximum distance of baits to MSA: " << blc.get_overall_max_dist() << "\n";
cout.precision(2);
cout << " Mean CG content of baits: " << blc.get_CG_mean() << "\n";
}
}
void extract_loci_information(const faststring &header, faststring &alignment_name, faststring &gene_name, unsigned &feature_num, unsigned &start)
{
faststring rest1;
faststring rest2;
faststring tmp;
header.divide_at('|', alignment_name, rest1);
rest1.divide_at('|', gene_name, rest2);
rest2.divide_at('|', tmp, rest1);
if (!tmp.isAnUnsigned())
{
cerr << "Error: Header does not obey formatting conventions: " << header << "\n";
cerr << "The third field (separator \"|\") should be an unsigned integer number, but found \"" << tmp << "\" instead.\n";
exit(-3);
}
feature_num = tmp.ToUnsigned();
rest1.divide_at('|', tmp, rest2);
if (!tmp.isAnUnsigned())
{
cerr << "Error: Header does not follow formatting conventions: " << header << "\n";
cerr << "The fourth field (separator \"|\") should be an unsigned integer number, but found \"" << tmp << "\" instead.\n";
exit(-3);
}
start = tmp.ToUnsigned();
// rest2.divide_at('|', tmp, rest1);
// if (!tmp.isAnUnsigned())
// {
// cerr << "Error: Header does not follow formatting conventions: " << header << "\n";
// cerr << "The fifth field (separator \"|\") should be an unsigned integer number, but found \"" << tmp << "\" instead.\n";
// exit(-3);
// }
// if (!rest1.isAnUnsigned())
// {
// cerr << "Error: Header does not follow formatting conventions: " << header << "\n";
// cerr << "The sixth field (separator \"|\") should be an unsigned integer number, but found \"" << rest1 << "\" instead.\n";
// exit(-3);
// }
// tiling_index = tmp.ToUnsigned();
// counter = rest1.ToUnsigned();
}
int main(int argc, char ** argv)
{
start_time = time(NULL);
cout.setf(ios::fixed);
cout.precision(2);
welcome(cout);
read_and_init_parameters(argc, argv, cerr);
#ifdef SKIPBLAST
skip_blast = true;
global_blast_result_file = "blast_result.txt";
#endif
if (!global_blast_result_file.empty())
{
skip_blast = true;
}
if (global_verbosity >= 2)
{
cerr << '\n';
print_parameters(cerr, "");
print_calling_command_line(cerr, argc, argv);
cerr << '\n';
}
// Parameter tests:
if (0)
{
faststring cmd = global_blast_exe + " -version";
cout << "Blast exe test: " << cmd << "\n";
int err = system(cmd.c_str());
cout << err << "\n";
exit(0);
}
ifstream is;
ofstream os;
is.open(global_bait_filename.c_str());
if (is.fail())
{
cerr << "Error: Input file: " << global_bait_filename << " could not be opened. I guess it does not exist.\n";
exit(-1);
}
if (global_mode != 'S')
{
os.open(global_output_filename.c_str());
if (os.fail())
{
cerr << "\n";
cerr << "Error: Failed to open or create output file. " << global_output_filename << "\n";
cerr << "Exiting.\n";
exit(-1);
}
}
if (global_verbosity >= 3)
{
cout << "PROGRESS: Parsing bait file: ... " << flush;
}
CBaitLoci_collection blc(is);
is.close();
if (global_verbosity >= 3)
{
cout << "Done. Time used since start in seconds: " << (time(NULL)-start_time) << "\n";
}
// Note: Bait loci are sorted in the constructor. Usually, they should be sorted in the bait file,
// if the user has not edited the file.
// DEBUG code:
// blc.print_stats(cout);
// blc.print_all(cout, 's');
// blc.print_all(cout, 'f');
// exit(0);
if (global_mode == 'S')
{
faststring msg;
msg = "Characteristics of the set of bait regions in input file: ";
if (global_verbosity < 2) // Can be the case if (global_mode == 'S')
global_verbosity = 2; // Minimum value required by print_stats() to print something. Could be handled with a parameter!
print_stats(blc, msg.c_str());
exit(0);
}
else if (global_verbosity >= 2)
{
faststring msg;
msg = "Characteristics of the set of bait regions in input file (before filtering): ";
print_stats(blc, msg.c_str());
}
if (global_mode == 'a')
{
if (global_verbosity >= 3)
{
cout << "\n\n";
cout << "PROGRESS: Determining: Best bait region in each alignment/gene with criterion minimum number of baits\n";
}
CBaitLoci_collection blc_best_baitRegions_in_genes_crit_numBaits(blc, 'g', 'b');
blc_best_baitRegions_in_genes_crit_numBaits.print_all(os, 's');
if (global_verbosity >= 2)
{
cout << "\n";
cout << "Characteristics of the set of bait regions after determining the best bait region per alignment/gene under the criterion to minimize the number of baits we have\n";
print_stats(blc_best_baitRegions_in_genes_crit_numBaits);
cout << "\n";
}
}
else if (global_mode == 'A')
{
if (global_verbosity >= 3)
{
cerr << "\n\n";
cerr << "PROGRESS: Determining: Best bait region in each gene with criterion maximum number of sequences.\n";
}
CBaitLoci_collection blc_best_baitRegions_in_genes_crit_numSeqs(blc, 'g', 's');
blc_best_baitRegions_in_genes_crit_numSeqs.print_all(os, 's');
if (global_verbosity >= 2)
{
cout << "\n";
cout << "Characteristics of the set of bait regions after determining the best bait region per alignment/gene under the criterion to maximize the number of sequences we have\n";
print_stats(blc_best_baitRegions_in_genes_crit_numSeqs);
cout << "\n";
}
}
else if (global_mode == 'f')
{
if (global_verbosity >= 3)
{
cerr << "\n\n";
cerr << "PROGRESS: Determining: Best bait region in each feature with criterion minimum number of baits\n";
}
CBaitLoci_collection blc_best_baitRegions_in_features_crit_numBaits(blc, 'f', 'b');
blc_best_baitRegions_in_features_crit_numBaits.print_all(os, 's');
if (global_verbosity >= 2)
{
cout << "\n";
cout << "After determining the best bait region per feature under the criterion to minimize the number of baits we have\n";
print_stats(blc_best_baitRegions_in_features_crit_numBaits);
cout << "\n";
}
}
else if (global_mode == 'F')
{
if (global_verbosity >= 3)
{
cerr << "\n\n";
cerr << "PROGRESS: Determining: Best bait region in each feature with criterion maximum number of sequences.\n";
}
CBaitLoci_collection blc_best_baitRegions_in_features_crit_numSeqs(blc, 'f', 's');
blc_best_baitRegions_in_features_crit_numSeqs.print_all(os, 's');
if (global_verbosity >= 2)
{
cout << "\n";
cout << "Characteristics of the set of bait regions after determining the best bait region per feature under the criterion to maximize the number of sequences we have\n";
print_stats(blc_best_baitRegions_in_features_crit_numSeqs);
cout << "\n";
}
}
else if (global_mode == 't' || global_mode == 'T')
{
unsigned sum_seqs, sum_baits;
unsigned start_count = 0;
unsigned best_index = 1;
unsigned best_num = 0; // This initialisation is only done to silence the compiler warning for using an uninitialised variable.
// This is essential output: Restrict it by verbosity??
if (global_verbosity >= 2)
{
cout << "Running thinning mode:\n";
cout << "\nThe columns are: starting offset, sum of sequences the result is based on, number of required baits.\n";
}
// Run thinning mode:
while (start_count < global_thinning_step_width)
{
blc.count_seqs_baits_for_start_and_offset(start_count, global_thinning_step_width, sum_seqs, sum_baits);
++start_count;
if (start_count == 1) // In this block we will surely initialize best_num:
{
if (global_mode == 't')
best_num = sum_baits;
else // global_mode == 'T'
best_num = sum_seqs;
}
if (global_mode == 't')
{
if (best_num > sum_baits)
{
best_index = start_count;
best_num = sum_baits;
}
}
else // global_mode == 'T'
{
if (best_num < sum_seqs)
{
best_index = start_count;
best_num = sum_seqs;
}
}
if (global_verbosity >= 2)
{
cout << start_count << " " << sum_seqs << " " << sum_baits << "\n";
}
}
cout << "Best starting offset: " << best_index << "\n";
cout << "A bait file that has been thinned out is written to the output file: " << global_output_filename << "\n";
cout << "This file contains each " << global_thinning_step_width << "th bait region starting with bait-region " << best_index << " of the input bait file.\n";
cout << "Numbers correspond to the line numbers in the input file +1, since all bait files have a headline.\n";
blc.print_with_start_stepwidth(os, 's', best_index, global_thinning_step_width);
}
else if (global_mode == 'h' || global_mode == 'H')
{
unsigned sum_seqs, sum_baits;
unsigned start_count;
unsigned best_index = UINT_MAX;
unsigned best_num = 0; // This initialisation is only done to silence the compiler warning for using an uninitialised variable.
unsigned best_penalty;
unsigned penalty_diff;
std::vector<CBaitLocus*>::iterator it_bait_vector, it_next_alignment_start, it_this_alignment_start;
// Loop over all alignments or features stored in this collection.
// Bait loci of all alignments or features are stored this one file.
// We will handle one alignment/feature a time.
// The current alignment or feature ends at it_tmp.
// The file ends at it_end
unsigned alignment_counter = 0;
bool finished = false;
// For each alignment
while (!finished) // The variable finished is being set inside blc.count_seqs_baits_for_start_and_offset_multi_gene_feature_aware
{
++ alignment_counter; // First value in loop is 1.
// For alignment_counter==1 the variables it_this_alignment_start and it_next_alignment_start are not initialised, but this is intended.
// They will be initialised with the first call to blc.count_seqs_baits_for_start_and_offset_multi_gene_feature_aware.
// If alignment_counter > 1, we start the next iteration.
it_this_alignment_start = it_next_alignment_start;
if (global_verbosity >= 2)
{
faststring original_file_name;
faststring gene_name;
blc.get_original_file_name_gene_name(alignment_counter, it_this_alignment_start, original_file_name, gene_name);
cout << "\n";
cout << "Running thinning mode for next alignment-file/feature: " << original_file_name << "/" << gene_name << "\n";
// cout << "\nThe columns are: starting offset, sum of sequences the result is based on, number of required baits.\n";
}
// Run thinning mode:
start_count = 0;
penalty_diff = 0;
best_penalty = UINT_MAX;
unsigned best_sum_seqs, best_sum_baits;
// Test all starting points in this alignment
while (start_count < global_thinning_step_width)
{
// In the following call, it_this_alignment_start is only changed if alignment_counter==1.
// In this case it is changed to the "beginning of the data set" and therefore to the beginning of the first alignment.
// Otherwise it remains unchanged.
// it_next_alignment_start is set to the start of the next alignment.
blc.count_seqs_baits_for_start_and_offset_multi_gene_feature_aware(alignment_counter,
it_this_alignment_start,
it_next_alignment_start,
finished,
start_count,
global_thinning_step_width,
sum_seqs, sum_baits, penalty_diff);
if (start_count == 0) // In this block we will initialise best_num:
{
if (global_mode == 'h')
{
best_num = sum_baits;
}
else // global_mode == 'H'
{
best_num = sum_seqs;
}
best_penalty = penalty_diff;
best_index = 0;
best_sum_seqs = sum_seqs;
best_sum_baits = sum_baits;
}
else
{
if (global_mode == 'h')
{
// Penalty is the primary criterion to choose the best starting position
if (best_penalty > penalty_diff || (best_penalty == penalty_diff && best_num > sum_baits) )
{
best_num = sum_baits;
best_index = start_count;
best_penalty = penalty_diff;
best_sum_seqs = sum_seqs;
best_sum_baits = sum_baits;
}
}
else // global_mode == 'H'
{
// Penalty is the primary criterion to choose the best starting position
if (best_penalty > penalty_diff || (best_penalty == penalty_diff && best_num < sum_seqs) )
{
best_num = sum_seqs;
best_index = start_count;
best_penalty = penalty_diff;
best_sum_seqs = sum_seqs;
best_sum_baits = sum_baits;
}
}
}
if (global_verbosity >= 5)
{
cout << ": " << start_count << " " << sum_seqs << " " << sum_baits << " " << penalty_diff << "\n";
if (global_verbosity >= 7)
{
cout << "best index, penalty, score: " << best_index << " " << best_penalty << " " << best_num << "\n";
}
}
++start_count;
} // END while (start_count < global_thinning_step_width) // Test all starting points in this alignment
cout << "Best starting offset (add 1 to get the index): " << best_index << "\n";
cout << "Sequences & baits for best starting offset: " << best_sum_seqs << " " << best_sum_baits << "\n";
cout << "Thinned bait loci have been appended to the output file: " << global_output_filename << "\n";
// cout << "This file contains each " << global_thinning_step_width << "th bait region starting with bait-region " << best_index << " of the input bait file." << "\n";
// cout << "Numbers correspond to the line numbers in the input file +1, since all bait files have a headline." << "\n";
// Write result for this alignment file/feature, before we go to the next a/f or finish.
blc.print_with_start_stepwidth_multi_gene_feature_aware(os, 's', best_index, global_thinning_step_width, it_this_alignment_start);
} // END while (!finished)
} // END else if (global_mode == 'h' || global_mode == 'H')
else if (global_mode == 'b' || global_mode == 'B' || global_mode == 'x' || global_mode == 'C')
{
faststring blastfilename = "blast_result.txt";
if (skip_blast)
{
blastfilename = global_blast_result_file;
}
// Conduct the blast search.
if (!skip_blast)
{
// Write baits to fasta file:
ofstream os_tmp("tmp_baits.txt");
blc.print_all(os_tmp, 'f');
os_tmp.close();
// Conduct a blast of the baits sequences against a reference genome:
// In the blast call we use an E-value threshold of:
// global_blast_max_second_hit_evalue*2
// This will report all relevant hits.
faststring e_value_command;
if (!global_blast_evalue_commandline.empty())
{
double tmp = global_blast_evalue_commandline.ToDouble();
if (tmp <= 10 && tmp > 0)
{
e_value_command = global_blast_evalue_commandline;
}
else
{
if (global_blast_min_hit_coverage_of_bait > 0) // For the coverage filter to work, the E-value should not be too small.
{
double temp = global_blast_max_second_hit_evalue*2;
if (temp < 0.001)
temp = 0.001;
e_value_command = faststring(temp, 15, true);
}
else
{
e_value_command = faststring(global_blast_max_second_hit_evalue*2, 15, true);
}
cerr << "Warning: The Evalue that can be specified with the blast-evalue-cutoff option has to be greater than 0 and smaller or equal to 10."
<< " The evalue that has been specified is not in this range."
<< " Therefore we are falling back to the default value: " << e_value_command << "\n";
}
}
else
{
if (global_blast_min_hit_coverage_of_bait > 0) // For the coverage filter to work, the E-value should not be too small.
{
double temp = global_blast_max_second_hit_evalue*2;
if (temp < 0.001)
temp = 0.001;
e_value_command = faststring(temp, 15, true);
}
else
{
e_value_command = faststring(global_blast_max_second_hit_evalue*2, 15, true);
}
}
faststring blastcommand = global_blast_exe + " "
+ global_blast_extra_commandline
+ " -outfmt 6 -task blastn -query tmp_baits.txt -db " + global_blast_db
+ " -out " + blastfilename + " -evalue " + e_value_command;
if (global_verbosity >= 2)
{
cout << "Running blast with command:\n";
cout << blastcommand << "\n";
}
blast_time = time(NULL);
// cerr << "Please press return to continue.\n";
// cin.get();
int blast_command_error;
blast_command_error = system(blastcommand.c_str());
if (blast_command_error != 0)
{
cerr << "ERROR: The blast command returned with non-zero error status. This indicates that blast encountered an error. The status blast returned via the system command was: " << blast_command_error << "\n";
cerr << "I suggest to run blast command manually:\n";
cerr << blastcommand << "\n";
cerr << "Exiting.\n";
exit(-1);
}
blast_time = time(NULL)-blast_time;
} // END if (!skip_blast)
if (global_verbosity >= 50)
cout << "PROGRESS: Reading the blast result file: " << blastfilename << " ..." << flush;
CBlast_parser bp(blastfilename.c_str());
if (global_verbosity >= 50)
cout << " Done.\n";
// How to use the CBalstParser to filter hits:
// Mode 0:
// Filter gene if first hit is below global_blast_max_first_hit_evalue and if second hit is below best_e_value_this_query*factor_max_increase
// Mode 1:
// Filter gene if first hit is below global_blast_max_first_hit_evalue and if second hit is below global_blast_max_second_hit_evalue
// double_hits contain only one of the 2 good hits that have been found.
// In case more than 2 good hits are found, it contains all but the first good hit, which is more than what we need.
// But this redundancy is removed when we add the bait information to the locus_selector data structure.
//
CBlast_parser double_hits;
bp.detect_2_good_hits_per_query(global_blast_max_first_hit_evalue, global_blast_max_second_hit_evalue, double_hits, 1);
set<faststring> set_of_names;
double_hits.set_of_query_names(set_of_names);
// The locus_selector contains information about
CBaitRegionSelectorSets locus_selector;
set<faststring>::iterator it, it_end;
it = set_of_names.begin();
it_end = set_of_names.end();
// cout << "Start Time: " << time(NULL) << "\n";
// For all baits
while (it != it_end)
{
faststring alignment_name;
faststring gene_name;
unsigned feature_num;
unsigned start;
// unsigned tiling_index; // Just dummy, will not be used.
// unsigned counter; // Just dummy, will not be used.
// Extract locus information from bait sequence name:
extract_loci_information(*it, alignment_name, gene_name, feature_num, start);
if (0)
{
if (!locus_selector.exists(alignment_name) )
cout << "New alignmentname: " << alignment_name << "\n";
else
cout << "Existing alignmentname: " << alignment_name << "\n";
if (!locus_selector.exists(alignment_name, feature_num) )
cout << "New alignmentname feature_num: " << alignment_name << " " << feature_num << "\n";
else
cout << "Existing alignmentname feature_num: " << alignment_name << " " << feature_num << "\n";
if (!locus_selector.exists(alignment_name, feature_num, start) )
cout << "New alignmentname feature_num start: " << alignment_name << " " << feature_num << " " << start << "\n";
else
cout << "Existing alignmentname feature_num start: " << alignment_name << " " << feature_num << " " << start << "\n";
}
locus_selector.add(alignment_name, feature_num, start);
++it;
} // END For all baits: while (it != it_end)
if (0)
{
cout << "\n";
cout << "Print locus selectors:\n";
locus_selector.print_all(cout);
cout << "\n";
}
// cout << "End Time: " << time(NULL) << "\n";
// Filter bait loci according to double_hits:
if (global_verbosity >= 100)
{
cout << "START: List of double hits:\n";
double_hits.print_all(cout);
cout << "END: List of double hits:\n\n";
}
// The following code was moved to the top.
// if (global_verbosity >= 2)
// {
// // cout << "Number of bait regions before searching/filtering for baits with multiple blast hits: " << blc.get_number_of_loci() << "\n";
// cout << "Characteristics of input file (before filtering): ";
// print_stats(blc);
// cout << "\n";
// }
// Allowed modes in this constructor:
// Filter modes: b, B, x:
// No filter: C. Means that filtered_blc obtains a copy of blc.
CBaitLoci_collection filtered_blc(blc, locus_selector, global_mode);
// Print stats with multiple hits results: (Do not print stats in coverage filter only mode. This case is handled below.)
if (global_verbosity >= 2 && global_mode != 'C')
{
// cout << "Number of bait regions after searching/filtering for baits with multiple blast hits: " << filtered_blc.get_number_of_loci() << "\n";
cout << "\n";
cout << "Characteristics of set of bait regions after filtering for baits with multiple blast hits to reference genome:\n";
print_stats(filtered_blc);
cout << "\n";
}
if (global_blast_min_hit_coverage_of_bait > 0)
{
CBaitLoci_collection coverage_filter(filtered_blc, bp, global_blast_min_hit_coverage_of_bait);
// if (global_verbosity >= 2)
// {
// cout << "Number of bait regions after hit coverage filtering: " << coverage_filter.get_number_of_loci() << "\n";
// }
coverage_filter.print_all(os, 's');
if (global_verbosity >= 2)
{
if (global_blast_min_hit_coverage_of_bait > 0)
{
cout << "\n";
cout << "Characteristics of set of bait regions after filtering for bait regions without sufficient hit coverage values in blast against a reference genome:\n";
print_stats(coverage_filter);
cout << "\n";
}
}
}
else
{
// Report what we have found:
filtered_blc.print_all(os, 's');
}
} // END if (global_mode == 'b' || global_mode == 'B' || global_mode == 'x' || global_mode == 'C')
else if (global_mode == 'c')
{
if (global_conversion_mode == 1)
{
// Write baits to file in Agilent-Germany format
blc.print_all(os, 'A', global_ProbeID_prefix.c_str());
if (global_verbosity >= 2)
{
cout << "Bait statistics for the converted bait file:\n";
print_stats(blc);
cout << "\n";
}
}
else
{
cerr << "INTERNAL ERROR: Unknown conversion mode : " << global_conversion_mode << "\n";
cerr << "Please report this problem to the developer.\n";
exit(-13);
}
} // END if (global_mode == 'c')
else
{
cerr << "INTERNAL ERROR: Unknown internal mode: " << global_mode << "\n";
cerr << "Please report this problem to the developer.\n";
exit(-13);
}
unsigned total_seconds = time(NULL)-start_time;
if (global_verbosity >= 2)
{
cout << "Total time used: ";
print_time(total_seconds, cout);
cout << "\n";
}
if (global_mode == 'b' || global_mode == 'B' || global_mode == 'x' || global_mode == 'C')
{
if (global_verbosity >= 2)
{
cout << "Time used excluding the Blast search: ";
print_time(total_seconds-blast_time, cout);
cout << "\n";
}
}
os.close();
good_bye(cout);
}
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