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determineTaxonomy.pl
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determineTaxonomy.pl
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#!/usr/bin/env perl
use warnings;
use strict;
use threads;
use threads::shared;
use Thread::Queue qw( );
use Thread::Semaphore;
use FAlite;
use English;
use Cpanel::JSON::XS;
use Compress::Zstd qw(compress decompress compress_mt);
use Scalar::Util qw(reftype);
use File::Temp qw/tempdir/;
use RocksDB;
use Sereal;
use Statistics::Basic qw(:all);
my $tmpdir = tempdir( DIR => "/dev/shm", CLEANUP => 1);
my $verb = shift @ARGV;
my $infoFloor = 0;
unless ($verb eq "filter" or $verb eq "classify" or $verb eq "entropy") {
die;
}
my $filter = 0;
if ($verb eq "filter") {
$filter = 1;
}
my $entropy = 0;
if ($verb eq "entropy") {
$entropy = 1;
}
if ($verb eq "classify") {
$infoFloor = shift @ARGV;
}
unless ($infoFloor =~ m/^\d+$/) {
print STDERR "infoFloor must be an integer\n";
die;
}
my $rocksDbAccess = Thread::Semaphore->new(24);
my $finished :shared;
$finished = 0;
my $total :shared;
my $lock :shared = 0;
my $THREADS = $ARGV[4];
my $q = Thread::Queue->new();
my $startQ = Thread::Queue->new();
my $groupQ = Thread::Queue->new();
my $encoder = Sereal::Encoder->new();
my $decoder = Sereal::Decoder->new();
my $lookUpTree = {
"11320" => ["1"],
"95342" => ["2", "1"],
"11983" => ["2", "1"]
};
my $groups = {
"virus" => "10239",
"others" => "28384",
"viroids" => "12884",
"unclassified" => "12908",
"cellularOrganisms" => "131567"
};
my $whiteListFile;
if ($entropy) {
$THREADS = 0;
$whiteListFile = $ARGV[2];
}
my $whiteList = {};
if ($whiteListFile) {
if ($entropy) {
my ($parents, $names, $ranks, $children) = makeTaxonomy($ARGV[0]);
}
open IN, "$whiteListFile";
while (my $line = <IN>) {
chomp $line;
$whiteList->{$line} = 1;
}
close IN;
}
$whiteList = shared_clone($whiteList);
if (not $entropy and $THREADS < 4) {
$THREADS = 4;
} elsif ($entropy) {
$THREADS = 0;
}
my @threads;
for (my $i = 0; $i < $THREADS; $i++) {
$threads[$i] = threads->create(\&worker, $tmpdir, $whiteList, $filter, $infoFloor)
}
my $RocksDBthreads = $THREADS;
if ($RocksDBthreads > 8) {
$RocksDBthreads = 8;
}
my $sortThreads = $THREADS;
if ($sortThreads > 8) {
$sortThreads = 8;
}
open IN, "$ARGV[1]";
my $fasta_file = new FAlite(\*IN);
my $seqCount = 0;
my $dbTAX = RocksDB->new("$tmpdir/TAX", { create_if_missing => 1, db_log_dir => "/dev/null", keep_log_file_num => 1, max_log_file_size => 1, db_stats_log_interval => 9999999, max_bytes_for_level_base => 100000000, target_file_size_base => 20000000});
$dbTAX->put("groups", compress($encoder->encode($groups)));
my $scaffolds = {};
while(my $entry = $fasta_file->nextEntry) {
my $head = $entry->def;
$head =~ s/^>//g;
$seqCount++;
my $seq = $entry->seq;
if ($entropy) {
$head =~ m/taxId=(\d+)/;
my $taxId = $1;
my $print = 1;
my $printErr = "";
if (not defined $taxId) {
$printErr = "WARNING: $head has no taxaId";
} else {
my $seqEnt = entropy($seq);
if ($seqEnt <= 5) {
$print = 0;
$printErr = "FILTERED: $head has entropy of $seqEnt";
}
}
if ($print) {
print ">$head\n";
print "$seq\n";
}
if ($printErr) {
print STDERR "$printErr\n";
}
} else {
$dbTAX->put("scaf.$head", $seq);
$scaffolds->{$head} = 1;
}
}
close IN;
undef($fasta_file);
if ($entropy) {
exit();
}
my $dbAA = RocksDB->new("$tmpdir/AA", { create_if_missing => 1, db_log_dir => "/dev/null", keep_log_file_num => 1, max_log_file_size => 1, db_stats_log_interval => 9999999, max_bytes_for_level_base => 100000000, target_file_size_base => 20000000});
my $dbNUC = RocksDB->new("$tmpdir/NUC", { create_if_missing => 1, db_log_dir => "/dev/null", keep_log_file_num => 1, max_log_file_size => 1, db_stats_log_interval => 9999999, max_bytes_for_level_base => 100000000, target_file_size_base => 20000000});
my ($parents, $names, $ranks, $children) = makeTaxonomy($ARGV[0]);
sub readBlastLines {
my $reference = $_[0];
my $db = $_[1];
my $topPerPos = $_[2];
my $topPerPosPerTaxa = $_[3];
my $parents = $_[4];
my $ranks = $_[5];
my $names = $_[6];
my $corrections = $_[7];
my $ok = $_[8];
my $localScore = $_[9];
my $overallScores = $_[10];
my $maxLocal = $_[11];
my $maxPoss = $_[12];
my $groups = $_[13];
my $selfMax = $_[14];
my $localMax = $_[15];
my $scale = $_[16];
my $codons = $_[17];
my $repPerPosPerTaxa = $_[18];
my $decoder = $_[19];
my $otherCont = $_[20];
my $tmpdir = $_[21];
my $scaffold = $_[22];
my $blosum62 = $_[23];
my $singleMode = $_[24];
my $lambda = $_[25];
my $k = $_[26];
my $expectedBitPerNuc = $_[27];
my $localDepth = {};
my $tid = threads->tid();
my $hit = 0;
my $goodAfterCycle = {};
$rocksDbAccess->down();
my $lines = $decoder->decode(decompress($db->get($reference)));
$rocksDbAccess->up();
my $maxPerAlignPos = {};
my $isProt = 0;
my $totalHitPerPos = {};
my $taxaPerPos = {};
my $prevTopPerPos = {};
if (scalar(keys %$codons)) {
$isProt = 1;
}
for (my $cycle = 0; $cycle < 2; $cycle++) {
foreach my $line (@$lines) {
my @parts = split /\t/, $line;
my $head = $parts[0];
my $coordAdjust = 0;
if ($head =~ m/;coords=(\d+)/) {
$coordAdjust = $1;
$head =~ s/;coords.*//g;
}
my $evalue = $parts[10];
if ($evalue > .1) {
next;
}
my $refHit = $parts[1];
$refHit =~ m/;taxId=(\d+)/;
my $refTaxa = $1;
unless (defined $refTaxa) {
next;
} elsif ($refTaxa == 0) {
next;
}
if (exists $corrections->{$refTaxa}) {
$refTaxa = $corrections->{$refTaxa};
}
$hit = 1;
if ($cycle > 0) {
unless (exists $goodAfterCycle->{$refTaxa}) {
next;
}
}
my $queryStart = $parts[6] + $coordAdjust;
my $queryStop = $parts[7] + $coordAdjust;
my $frame = 1;
if ($queryStop < $queryStart) {
my $temp = $queryStart;
$frame = -1;
$queryStart = $queryStop;
$queryStop = $temp;
}
my $bitScore = $parts[11];
my $bitScorePerAlign = $bitScore / (($queryStop - $queryStart) + 1);
if (not $isProt and $bitScorePerAlign > $expectedBitPerNuc) {
$bitScorePerAlign = $expectedBitPerNuc;
}
my $gotTop = 0;
for (my $i = $queryStart; $i <= $queryStop; $i++) {
unless (exists $totalHitPerPos->{$i}) {
$totalHitPerPos->{$i} = 0;
}
if ($cycle == 0) {
unless (exists $topPerPos->{$i}) {
$topPerPosPerTaxa->{$i}->{$refTaxa} = $bitScorePerAlign;
$topPerPos->{$i} = $bitScorePerAlign;
$gotTop = 1;
} elsif ($bitScorePerAlign > $topPerPos->{$i}) {
$topPerPosPerTaxa->{$i}->{$refTaxa} = $bitScorePerAlign;
$topPerPos->{$i} = $bitScorePerAlign;
$gotTop = 1;
foreach my $checkTaxa (keys %{$topPerPosPerTaxa->{$i}}) {
if ($topPerPosPerTaxa->{$i}->{$checkTaxa} < (.90 * $bitScorePerAlign) ) {
delete $topPerPosPerTaxa->{$i}->{$checkTaxa};
}
}
} elsif ((not exists $topPerPosPerTaxa->{$i}->{$refTaxa} and $bitScorePerAlign > (.90 * $topPerPos->{$i})) or (exists $topPerPosPerTaxa->{$i}->{$refTaxa} and $bitScorePerAlign > $topPerPosPerTaxa->{$i}->{$refTaxa})) {
$topPerPosPerTaxa->{$i}->{$refTaxa} = $bitScorePerAlign;
}
} else {
if (exists $topPerPos->{$i}) {
my $ratio = ($bitScorePerAlign / $topPerPos->{$i});
if ($ratio > 1) {
lock($lock);
lockPrintStderr("top per pos $i = $topPerPos->{$i}, bitscoreperslign = $bitScorePerAlign (scaled), ratio = $ratio, should never happen, taxa = $refTaxa");
}
$taxaPerPos->{$i}->{$refTaxa}++;
if (not exists $topPerPosPerTaxa->{$i}->{$refTaxa} or $topPerPosPerTaxa->{$i}->{$refTaxa} < $bitScorePerAlign) {
$topPerPosPerTaxa->{$i}->{$refTaxa} = $bitScorePerAlign;
}
my $localScale = $scale;
if (not $isProt and not exists $otherCont->{$i} and not $singleMode) {
$localScale = 3;
}
my $delta = (($bitScorePerAlign / $topPerPos->{$i}) ** (3 ** $localScale)) / $localScale;
if ($delta > 1) {
lockPrintStderr("delta = $delta, should never happen, on taxa $refTaxa");
}
#### DB Rep per base per accepted alignmet per taxa ####
$repPerPosPerTaxa->{$i}->{$refTaxa} += (($bitScorePerAlign / $topPerPos->{$i}) ** (3 ** $localScale)) / $localScale;
$totalHitPerPos->{$i}++;
} else {
lockPrintStderr("Top hit not in second round: $i");
$topPerPos->{$i} = 0;
$totalHitPerPos->{$i} = 0;
}
}
}
if ($gotTop) {
my $segment = uc(substr($scaffold, $queryStart - 1, ($queryStop - $queryStart) + 1));
my $score = 0;
my $segLength = length($segment);
if ($isProt) {
if ($frame == -1) {
$segment = reverse($segment);
$segment =~ tr/ACGT/TGCA/;
}
my @codons = $segment =~ /(...)/g;
foreach my $codon (@codons) {
if (exists $codons->{$codon} and exists $blosum62->{$codons->{$codon}}) {
$score += $blosum62->{$codons->{$codon}};
} else {
$score--;
}
}
} else {
my $nCount = $segment =~ tr/Nn//;
$score = (2 * ($segLength - $nCount)) - (2 * $nCount);
}
my $bestBitPerBase = raw2bit($score, $lambda, $k) / $segLength;
if (not $isProt and $bestBitPerBase > $expectedBitPerNuc) {
$bestBitPerBase = $expectedBitPerNuc;
} elsif ($isProt and $bestBitPerBase < $bitScorePerAlign) {
#rounding errors
$bestBitPerBase = $bitScorePerAlign;
}
for (my $i = $queryStart; $i <= $queryStop; $i++) {
if (not exists $maxPerAlignPos->{$i} or $maxPerAlignPos->{$i} < $bestBitPerBase) {
$maxPerAlignPos->{$i} = $bestBitPerBase;
}
}
}
}
if ($cycle == 0) {
foreach my $pos (keys %$topPerPosPerTaxa) {
foreach my $putTaxa (keys %{$topPerPosPerTaxa->{$pos}}) {
$goodAfterCycle->{$putTaxa} = 1;
}
}
}
}
my @reps = values %$totalHitPerPos;
my $minHitProt = "inf";;
my $avgHit = 1;
if (scalar @reps) {
$avgHit = mean(\@reps);
$minHitProt = .1 * $avgHit;
}
my $totalBitScore = 0;
my $totalDenom = 0;
my $kept = {};
foreach my $pos (sort {$a <=> $b} keys %$topPerPos) {
my $add = $maxPerAlignPos->{$pos};
unless ($totalHitPerPos->{$pos}) {
next;
}
$kept->{$pos} = 0;
if (not $isProt or ($avgHit < 10) or ($totalHitPerPos->{$pos} > $minHitProt)) {
$totalBitScore += $add;
$totalDenom++;
$kept->{$pos} = 1;
${$selfMax} += $add;
${$localMax} += $add;
} else {
if (exists $topPerPosPerTaxa->{$pos}) {
delete($topPerPosPerTaxa->{$pos});
}
if (exists $topPerPos->{$pos}) {
delete($topPerPos->{$pos});
}
if (exists $totalHitPerPos->{$pos}) {
delete($totalHitPerPos->{$pos});
}
if (exists $repPerPosPerTaxa->{$pos}) {
delete($repPerPosPerTaxa->{$pos});
}
next;
}
foreach my $taxa (keys %{$topPerPosPerTaxa->{$pos}}) {
$maxLocal->{$taxa} += $add;
$maxPoss->{$taxa} += $add;
$localScore->{$taxa} += $topPerPosPerTaxa->{$pos}->{$taxa};
$overallScores->{$taxa} += $topPerPosPerTaxa->{$pos}->{$taxa};
}
}
my $avgBestBit = 0;
if ($totalDenom) {
$avgBestBit = $totalBitScore / $totalDenom;
}
return($hit, $avgBestBit);
}
sub checkWhiteList {
my $whiteList = $_[0];
my $checkTaxa = $_[1];
my $parents = $_[2];
my $escape = 0;
while ($checkTaxa != 1) {
$escape++;
if ($escape == 100) {
return(0);
}
if (exists $whiteList->{$checkTaxa}) {
return(1);
}
if (exists $parents->{$checkTaxa}) {
$checkTaxa= $parents->{$checkTaxa};
}
}
return(0);
}
sub worker {
use RocksDB;
select(STDERR);
$| = 1;
my $tid = threads->tid();
my $tmpdir = $_[0];
my $whiteList = $_[1];
my $filter = $_[2];
my $infoFloor = $_[3];
my $return = {};
my $ok = {};
my $nucLambda = 0.625;
my $nucK = 0.410;
my $protLambda = 0.267;
my $protK = 0.041;
my $maxAddAA = 0;
my $maxAddNucl = 0;
my $aaScale = 3;
my $nucScale = 1;
my $expectedBitPerNuc = (2 * ($nucLambda)) / log(2);
my $e = 2.7182818284590452353602874713527;
my $pi = 3.1415926535897932384626433832795;
my $sqrt2 = sqrt(2);
my $cubrt2 = 2 ** (1/3);
my $taxaAccuracyBase = 10;
my $entropyLimit = 5.5;
my $filterRatio = .99;
my $rescueRatio = .98;
my $startNow = $startQ->dequeue();
my $encoder = Sereal::Encoder->new();
my $decoder = Sereal::Decoder->new();
$rocksDbAccess->down();
my $dbAA = RocksDB->new("$tmpdir/AA", { read_only => 1, db_log_dir => '/dev/null', keep_log_file_num => 1}) or die "can't open DB\n";
my $dbNUC = RocksDB->new("$tmpdir/NUC", { read_only => 1, db_log_dir => '/dev/null', keep_log_file_num => 1}) or die "can't open DB\n";
my $dbTAX = RocksDB->new("$tmpdir/TAX", { read_only => 1, db_log_dir => '/dev/null', keep_log_file_num => 1}) or die "can't open DB\n";
my $groups = $decoder->decode(decompress($dbTAX->get("groups")));
my $singleMode = $dbTAX->get("singleMode");
$rocksDbAccess->up();
my $suppressors = {
"others" => 1,
"unclassified" => 1
};
my $codons = {
TTT => "F", TTC => "F", TTA => "L", TTG => "L",
TCT => "S", TCC => "S", TCA => "S", TCG => "S",
TAT => "Y", TAC => "Y", TAA => "*", TAG => "*",
TGT => "C", TGC => "C", TGA => "*", TGG => "W",
CTT => "L", CTC => "L", CTA => "L", CTG => "L",
CCT => "P", CCC => "P", CCA => "P", CCG => "P",
CAT => "H", CAC => "H", CAA => "Q", CAG => "Q",
CGT => "R", CGC => "R", CGA => "R", CGG => "R",
ATT => "I", ATC => "I", ATA => "I", ATG => "M",
ACT => "T", ACC => "T", ACA => "T", ACG => "T",
AAT => "N", AAC => "N", AAA => "K", AAG => "K",
AGT => "S", AGC => "S", AGA => "R", AGG => "R",
GTT => "V", GTC => "V", GTA => "V", GTG => "V",
GCT => "A", GCC => "A", GCA => "A", GCG => "A",
GAT => "D", GAC => "D", GAA => "E", GAG => "E",
GGT => "G", GGC => "G", GGA => "G", GGG => "G",
};
my $blosum62 = {
A => 4,
R => 5,
N => 6,
D => 6,
C => 9,
Q => 5,
E => 5,
G => 6,
H => 8,
I => 4,
L => 4,
K => 5,
M => 5,
F => 6,
P => 7,
S => 4,
T => 5,
W => 11,
Y => 7,
V => 4,
X => -1,
"*" => 1
};
while (1) {
my $reference = $q->dequeue();
unless (defined $reference) {
my $peek = $q->peek();
if (not defined $peek) {
last;
} else {
redo;
}
}
lockPrintStderr("GOT $reference");
my $isContig = 0;
if ($reference =~ m/^contig/) {
$isContig = 1;
}
my $fromProt = 0;
if ($reference =~ m/NoNucHits/) {
$fromProt = 1;
}
my $mixedSources = 0;
$reference =~ m/taxId=(\d+)/;
my $refTaxId = $1;
unless ($refTaxId) {
lockPrintStderr("$reference has no detectable taxId");
}
$rocksDbAccess->down();
my $parents = $decoder->decode(decompress($dbTAX->get("par.$reference")));;
my $ranks = $decoder->decode(decompress($dbTAX->get("ran.$reference")));;
my $names = $decoder->decode(decompress($dbTAX->get("nam.$reference")));
my $taxa2Group = $decoder->decode(decompress($dbTAX->get("t2g.$reference")));
my $corrections = $decoder->decode(decompress($dbTAX->get("cor.$reference")));
my $whiteListed = checkWhiteList($whiteList, $refTaxId, $parents);
my $scaffold = $dbTAX->get("scaf.$reference");
$rocksDbAccess->up();
my $scafEnt = entropy($scaffold);
if ($filter and $scafEnt <= 5) {
lockPrintStderr("FILTERED: $reference is low complexity $scafEnt bit per triplet");
next;
}
my $lcCount = () = $scaffold =~ /[acgt]/g;
if ($lcCount > .2 * length($scaffold)) {
$mixedSources = 1;
}
my $topPerPosAA = {};
my $topPerPosNucl = {};
my $topPerPosPerTaxaAA = {};
my $topPerPosPerTaxaNucl = {};
my $repPerPosPerTaxaAA = {};
my $repPerPosPerTaxaNucl = {};
my $repPerPosPerTaxa = {};
my $overallScoresAA = {};
my $overallScoresNucl = {};
my $overallScores = {};
my $maxPossAA = {};
my $maxPossNucl = {};
my $maxPoss = {};
my $selfMaxAA = 0;
my $selfMaxNucl = 0;
my $selfMax = 0;
my $nonFilterAddMax = 0;
my $protHits = 0;
my $nucHits = 0;
my $posPerGroupHits = {};
($protHits, $maxAddAA) = readBlastLines($reference, $dbAA, $topPerPosAA, $topPerPosPerTaxaAA, $parents, $ranks, $names, $corrections, $ok, $overallScoresAA, $overallScores, $maxPossAA, $maxPoss, $groups, \$selfMax, \$selfMaxAA, $aaScale, $codons, $repPerPosPerTaxaAA, $decoder, {}, $tmpdir, $scaffold, $blosum62, $singleMode, $protLambda, $protK, 0);
($nucHits, $maxAddNucl) = readBlastLines($reference, $dbNUC, $topPerPosNucl, $topPerPosPerTaxaNucl, $parents, $ranks, $names, $corrections, $ok, $overallScoresNucl, $overallScores, $maxPossNucl, $maxPoss, $groups, \$selfMax, \$selfMaxNucl, $nucScale, {}, $repPerPosPerTaxaNucl, $decoder, $topPerPosAA, $tmpdir, $scaffold, {}, $singleMode, $nucLambda, $nucK, $expectedBitPerNuc);
my @letters = split //, $scaffold;
unshift @letters, "N";
my $total = 0;
my $unal = 0;
my $badTop = 0;
my $virusDist = 0;
my $badTopAA = 0;
my $badTopNucl = 0;
my $virusLost = 0;
my $suppressRescue = 0;
my $filterUnal = 0;
my $rescueDist = 0;
my $groupScorePerPosAA = {};
my $groupScorePerPosNucl = {};
my $groupScorePerPos = {};
my $avgFail = {};
my $avgFailOverlap = {};
my $suppressCount = {};
my $repPerGroup = {};
my $avgRepPerPos = 0;
my $unalVirusRepAdd = 0;
my $taxaRepAA = {};
my $taxaRepNucl = {};
my $taxaRep = {};
my $posHit = 0;
my $highUnknown = 0;
my $denovoUnal = 0;
my $nonFilterAddAA = 0;
my $nonFilterAddNucl = 0;
my $avgBitDensity = 0;
my $highHitLowDensity = 0;
if (not $protHits and not $nucHits and $filter) {
lockPrintStderr("KEPT: NO hits bypass on filter for $reference");
$return->{$reference} = $scaffold;
next;
}
for (my $pos = 0; $pos < scalar(@letters); $pos++) {
my $hitGroups = {};
if (not defined $letters[$pos]) {
next;
}
if ($letters[$pos] eq "N" or $letters[$pos] eq "n") {
next;
}
$total++;
if (not exists $topPerPosAA->{$pos} and not exists $topPerPosNucl->{$pos}) {
unless ($letters[$pos] =~ m/[acgt]/) {
$unal++;
} else {
$denovoUnal++;
}
if ($filter) {
$filterUnal++;
}
$nonFilterAddNucl += $maxAddNucl;
$nonFilterAddAA += $maxAddAA;
next;
}
$posHit++;
my $localGroupRep = {};
if (exists $repPerPosPerTaxaAA->{$pos}) {
foreach my $taxa (keys %{$repPerPosPerTaxaAA->{$pos}}) {
$repPerGroup->{$taxa2Group->{$taxa}} += $repPerPosPerTaxaAA->{$pos}->{$taxa};
$hitGroups->{$taxa2Group->{$taxa}} = 1;
$localGroupRep->{$taxa2Group->{$taxa}} += $repPerPosPerTaxaAA->{$pos}->{$taxa};
$taxaRepAA->{$taxa} += $repPerPosPerTaxaAA->{$pos}->{$taxa};
$taxaRep->{$taxa} += $repPerPosPerTaxaAA->{$pos}->{$taxa};
$avgRepPerPos += $repPerPosPerTaxaAA->{$pos}->{$taxa};
}
}
if (exists $repPerPosPerTaxaNucl->{$pos}) {
foreach my $taxa (keys %{$repPerPosPerTaxaNucl->{$pos}}) {
$repPerGroup->{$taxa2Group->{$taxa}} += $repPerPosPerTaxaNucl->{$pos}->{$taxa};
$hitGroups->{$taxa2Group->{$taxa}} = 1;
$localGroupRep->{$taxa2Group->{$taxa}} += $repPerPosPerTaxaNucl->{$pos}->{$taxa};
$taxaRep->{$taxa} += $repPerPosPerTaxaNucl->{$pos}->{$taxa};
$taxaRepNucl->{$taxa} += $repPerPosPerTaxaNucl->{$pos}->{$taxa};
$avgRepPerPos += $repPerPosPerTaxaNucl->{$pos}->{$taxa};
}
}
if (exists $topPerPosPerTaxaAA->{$pos}) {
foreach my $taxa (keys %{$topPerPosPerTaxaAA->{$pos}}) {
if (not exists $groupScorePerPosAA->{$pos}->{$taxa2Group->{$taxa}} or $groupScorePerPosAA->{$pos}->{$taxa2Group->{$taxa}} < $topPerPosPerTaxaAA->{$pos}->{$taxa}) {
$groupScorePerPosAA->{$pos}->{$taxa2Group->{$taxa}} = $topPerPosPerTaxaAA->{$pos}->{$taxa};
}
}
}
if (exists $topPerPosPerTaxaNucl->{$pos}) {
foreach my $taxa (keys %{$topPerPosPerTaxaNucl->{$pos}}) {
if (not exists $groupScorePerPosNucl->{$pos}->{$taxa2Group->{$taxa}} or $groupScorePerPosNucl->{$pos}->{$taxa2Group->{$taxa}} < $topPerPosPerTaxaNucl->{$pos}->{$taxa}) {
$groupScorePerPosNucl->{$pos}->{$taxa2Group->{$taxa}} = $topPerPosPerTaxaNucl->{$pos}->{$taxa};
}
}
}
foreach my $group (keys %$groups) {
if (exists $groupScorePerPosNucl->{$pos}->{$group}) {
$groupScorePerPos->{$pos}->{$group} += $groupScorePerPosNucl->{$pos}->{$group};
}
if (exists $groupScorePerPosAA->{$pos}->{$group}) {
$groupScorePerPos->{$pos}->{$group} += $groupScorePerPosAA->{$pos}->{$group};
}
}
my $topSuppress = {};
my $topSuppressAA = {};
my $topSuppressNucl = {};
my $suppress = 0;
foreach my $suppressor (keys %$suppressors) {
if (exists $groupScorePerPosAA->{$pos}->{$suppressor}) {
$topSuppressAA->{$suppressor} = $groupScorePerPosAA->{$pos}->{$suppressor};
$topSuppress->{$suppressor} += $groupScorePerPosAA->{$pos}->{$suppressor};
$suppress = 1;
}
if (exists $groupScorePerPosNucl->{$pos}->{$suppressor}) {
$topSuppressNucl->{$suppressor} = $groupScorePerPosNucl->{$pos}->{$suppressor};
$topSuppress->{$suppressor} += $groupScorePerPosNucl->{$pos}->{$suppressor};
$suppress = 1;
}
}
my @bestScores = sort {$groupScorePerPos->{$pos}->{$b} <=> $groupScorePerPos->{$pos}->{$a}} keys %{$groupScorePerPos->{$pos}};
if (exists $groupScorePerPos->{$pos}->{'virus'}) {
my $ratios = {};
foreach my $group (@bestScores) {
$ratios->{$group} = $groupScorePerPos->{$pos}->{'virus'} / $groupScorePerPos->{$pos}->{$group};
}
if ($ratios->{$bestScores[0]} < $filterRatio) {
if ($suppress) {
my @bestSuppress = sort {$topSuppress->{$b} <=> $topSuppress->{$a}} keys %$topSuppress;
if ($ratios->{$bestSuppress[0]} >= $rescueRatio and $ratios->{$bestSuppress[0]} < 1) {
foreach my $suppressor (@bestSuppress) {
$repPerGroup->{$suppressor} -= $localGroupRep->{$suppressor};
}
$suppressRescue += $ratios->{$bestSuppress[0]};
$rescueDist++;
$suppressCount->{$bestSuppress[0]}++;
} else {
$suppress = 0;
}
}
foreach my $group (keys %$ratios) {
if ($group eq 'virus') {
next;
}
$avgFail->{$group} += $ratios->{$group};
$avgFailOverlap->{$group}++;
}
unless ($suppress) {
$virusLost++;
$badTop++;
}
}
} else {
$badTop++;
}
foreach my $group (keys %$hitGroups) {
$posPerGroupHits->{$group}++;
}
}
foreach my $group (keys %$repPerGroup) {
if ($repPerGroup->{$group} > 0) {
$repPerGroup->{$group} = log10($repPerGroup->{$group}) * $posPerGroupHits->{$group};
} else {
$repPerGroup->{$group} = 0;
}
}
my $totalScore = 0;
foreach my $val (values %$topPerPosAA) {
$totalScore += $val
}
foreach my $val (values %$topPerPosNucl) {
$totalScore += $val
}
if ($posHit) {
$avgBitDensity = $totalScore / $posHit;
$avgRepPerPos = $avgRepPerPos / $posHit;
}
$unalVirusRepAdd = $avgRepPerPos * $filterUnal;
my $unalLimit = .8;
$highUnknown = (($isContig or $mixedSources) and (($unal + $denovoUnal) / $total > $unalLimit) and $total > 500);
my $freePass = ($highUnknown or $whiteListed);
if (not $freePass and not $filter and not $posHit and not $isContig) {
lockPrintStderr("FILTERED: $reference no hits on pseudo constructed contig");
next;
}
if (not $freePass and not $filter and ($unal / ($total - $denovoUnal)) > $unalLimit and not ($isContig or $mixedSources)) {
lockPrintStderr("FILTERED: Contig has <20% of pseudoconstructed segments hitting the database: $reference");
next;
}
if ($unalVirusRepAdd) {
$repPerGroup->{'virus'} += $unalVirusRepAdd;
}
foreach my $group (keys %$repPerGroup) {
$repPerGroup->{$group} = $repPerGroup->{$group} / $total;
}
unless (exists $repPerGroup->{'virus'}) {
$repPerGroup->{'virus'} = 0;
}
my $avgRescue = 0;
my $rescueStr = "";
if ($rescueDist) {
$avgRescue = $suppressRescue / $rescueDist;
my @bestSuppress = sort {$suppressCount->{$b} <=> $suppressCount->{$a}} keys %$suppressCount;
$rescueStr = "best suppressor: $bestSuppress[0] with $suppressCount->{$bestSuppress[0]} based recovered";
}
foreach my $group (keys %$avgFail) {
$avgFail->{$group} = $avgFail->{$group} / $avgFailOverlap->{$group};
}
my $remove = 0;
my $status = "";
my @repOrder = sort {$repPerGroup->{$b} <=> $repPerGroup->{$a}} keys %$repPerGroup;
my $bestRepGroup = $repPerGroup->{$repOrder[0]};
if (not $freePass and ($badTop / $total > .5 or $bestRepGroup == 0 or $repPerGroup->{'virus'} < $bestRepGroup)) {
if (exists $suppressors->{$repOrder[0]} and $repPerGroup->{'virus'} > (.5 * $bestRepGroup)) {
$status = "KEPT";
lockPrintStderr("RECOVERED: $repOrder[0] is a suppressed category, virus is > 50% is it's score: $repPerGroup->{'virus'} vs $bestRepGroup");
} else {
$remove = 1;
$status = "FILTERED";
}
if ($repPerGroup->{'virus'} < $bestRepGroup and $status ne "KEPT") {
lockPrintStderr("FILTERED: $repOrder[0] had a higher score than virus");
}
} else {
$status = "KEPT";
}
unless ($filter) {
$filterUnal = $unal;
}
{lock($lock);
lockPrintStderr("$status: $reference ($badTop bp/$total bp) bad bases; $virusLost bp virus lost; $filterUnal bp unaligned; $rescueDist bp rescued; avg rescue: $avgRescue; entropy: $scafEnt");
lockPrintStderr("$status: avgRepPerPos = $avgRepPerPos");
}
if ($filter) {
unless ($remove) {
$return->{$reference} = $scaffold;
}
next;
} elsif ($remove) {
next;
}
#### END FILTER MODE ####
#### TAXA ENGINE ####
my $groupScores = {};
my $groupScoresAA = {};
my $groupSciresNucl = {};
my $maxAA = 0;
my $maxNucl = 0;
my $maxNuclTaxa = 0;
my $maxAATaxa = 0;
my $max = 0;
foreach my $taxa (keys %$overallScores) {
$groupScores->{$taxa2Group->{$taxa}}->{$taxa} = $overallScores->{$taxa};
if ($overallScores->{$taxa} > $max) {
$max = $overallScores->{$taxa};
}
if (exists $overallScoresAA->{$taxa} and $overallScoresAA->{$taxa} > $maxAA) {
$maxAA = $overallScoresAA->{$taxa};
$maxAATaxa = $taxa;
}
if (exists $overallScoresNucl->{$taxa} and $overallScoresNucl->{$taxa} > $maxNucl) {
$maxNucl = $overallScoresNucl->{$taxa};
$maxNuclTaxa = $taxa;
}
}
my $groupScoresSortedOrder = {};
my $maxes = {};
foreach my $group (keys %$groupScores) {
my @sorted = sort {$groupScores->{$group}->{$b} <=> $groupScores->{$group}->{$a}} keys %{$groupScores->{$group}};
$groupScoresSortedOrder->{$group} = \@sorted;
$maxes->{$group} = $groupScores->{$group}->{$sorted[0]};
if ($maxes->{$group} > $selfMax) {
$selfMax = $maxes->{$group};
}
}
unless (exists $maxes->{'virus'}) {
$maxes->{'virus'} = 0;
}
lockPrintStderr("BITDENSITY: $reference = $avgBitDensity");
if (not $freePass and $maxes->{'virus'} < $infoFloor and ($selfMax < 1.25 * $infoFloor) and not $highUnknown) {
lockPrintStderr("FILTERED: $reference has a max virus score of $maxes->{'virus'} and a selfMax of $selfMax which is less than the limit of $infoFloor and 125% of $infoFloor");
next;
}
foreach my $taxa (keys %$taxaRep) {
unless (exists $taxaRep->{$taxa}) {
lockPrintStderr("WARNING: taxa id: $taxa not present in $reference db calculations");
$taxaRep->{$taxa} = 0;
}
if (exists $taxaRepAA->{$taxa}) {
$taxaRepAA->{$taxa} = $taxaRepAA->{$taxa} / $total;
} else {
$taxaRepAA->{$taxa} = 0;
}
if (exists $taxaRepNucl->{$taxa}) {
$taxaRepNucl->{$taxa} = $taxaRepNucl->{$taxa} / $total;
} else {
$taxaRepNucl->{$taxa} = 0;
}
$taxaRep->{$taxa} = $taxaRep->{$taxa} / $total;
}
my $minPresence = (.90 * $max);
if (not $freePass and $maxes->{'virus'} < $minPresence) {
lockPrintStderr("lower limit of scoring: $minPresence");
lockPrintStderr("FILTERED: virus below $minPresence threshold $reference");
lockBrowseErr("Max scores of groups:", $maxes);
next;
}
{lock($lock);
foreach my $group (keys %$groupScores) {
my $checkLimit = 10;
my @keys = @{$groupScoresSortedOrder->{$group}};
my $groupMax = $overallScores->{$keys[0]};
my $groupMemberCount = scalar(@keys);
if ($groupMemberCount < $checkLimit) {
$checkLimit = $groupMemberCount;
}
unless ($checkLimit) {
next;
}
lockPrintStderr("Top $checkLimit scoring taxa IDs in group $group");
for (my $i = 0; $i < $checkLimit; $i++) {
my $printNucl = 0;
my $printAA = 0;
unless (exists $overallScoresAA->{$keys[$i]}) {
$overallScoresAA->{$keys[$i]} = 0;
} else {
$overallScoresAA->{$keys[$i]} = $overallScoresAA->{$keys[$i]};
}
unless (exists $overallScoresNucl->{$keys[$i]}) {
$overallScoresNucl->{$keys[$i]} = 0;
} else {
$overallScoresNucl->{$keys[$i]} = $overallScoresNucl->{$keys[$i]};
}
if ($i < $checkLimit) {
my $printScore = int($overallScores->{$keys[$i]});
my $printRep = int($taxaRep->{$keys[$i]});
my $printTaxaRepNucl = int($taxaRepNucl->{$keys[$i]});
my $printTaxaRepAA = int($taxaRepAA->{$keys[$i]});
my $printScoreAA = int($overallScoresAA->{$keys[$i]});
my $printScoreNucl = int($overallScoresNucl->{$keys[$i]});
lockPrintStderr("SCORE: $printScore\tprot: $printScoreAA\tnuc: $printScoreNucl\tdbRep: $printRep\tAA: $printTaxaRepAA\tNucl: $printTaxaRepNucl\t\tTaxID: $keys[$i] - $names->{$keys[$i]}");
}
}
lockPrintStderr("");
}
}
if ($maxAA > $selfMaxAA) {
$selfMaxAA = $maxAA;
}
if ($maxNucl > $selfMaxNucl) {
$selfMaxNucl = $maxNucl;
}
my $baseNucl = ($e / ($pi * $sqrt2)) / 10;
my $baseAA = ($cubrt2 / 4) / 10;
my $baseBlurNucl = 0;
my $baseBlurAA = 0;
my $baseRadiusNucl = 0;
my $accuracyNucl = 0;
my $accuracyAdjustNucl = 0;
my $maxNuclFrac = 0;
my $approxIdNucl = 0;
#### LCA radius calculated here ####
if ($maxNucl) {
$accuracyNucl = $maxNucl / $selfMaxNucl;
$approxIdNucl = (((($accuracyNucl * $expectedBitPerNuc) * log(2)) / $nucLambda) + 3) / 5;
my $denom = log($maxNucl) * log10($maxNucl);
$baseBlurNucl = (-1 * log($approxIdNucl) / $denom) + $baseNucl;
$baseRadiusNucl = ($e/log10($maxNucl)) * $baseBlurNucl;
$accuracyAdjustNucl = (-1 * log10($approxIdNucl) / log10(100 * $baseBlurNucl)) * $baseBlurNucl;
$maxNuclFrac = $maxNucl / ($maxNucl + $maxAA);
}
my $baseRadiusAA = 0;
my $accuracyAA = 0;
my $accuracyAdjustAA = 0;
my $maxAAFrac = 0;
my $approxIdAA = 0;
if ($maxAA) {
$accuracyAA = $maxAA / $selfMaxAA;
my $denom = log($maxAA) * log10($maxAA);
$baseBlurAA = (-1 * log($accuracyAA) / $denom) + $baseAA;
$baseRadiusAA = ($e/log10($maxAA)) * $baseBlurAA;
$accuracyAdjustAA = (-1 * log10($accuracyAA) / log10(100 * $baseBlurAA)) * $baseBlurAA;
$maxAAFrac = $maxAA / ($maxNucl + $maxAA);
}
my $minUnit = 50;
my $bitRadiusNucl = ($baseRadiusNucl + $accuracyAdjustNucl) * $maxNucl;
my $bitRadiusAA = ($baseRadiusAA + $accuracyAdjustAA) * $maxAA;
my $radNucl = ($baseRadiusNucl + $accuracyAdjustNucl);
my $radAA = ($baseRadiusAA + $accuracyAdjustAA);
my $bitRadius = $bitRadiusNucl + $bitRadiusAA;
if ($bitRadius < $minUnit) {
$bitRadius = $minUnit;
$bitRadiusNucl = $maxNuclFrac * $minUnit;
$bitRadiusAA = $maxAAFrac * $minUnit;
}
my $lowerLimit = $max - $bitRadius;
my $lowerLimitNucl = $maxNucl - $bitRadiusNucl;
my $lowerLimitAA = $maxAA - $bitRadiusAA;
my $printLimit = int($lowerLimit);
my $printLimitNucl = int($lowerLimitNucl);
my $printLimitAA = int($lowerLimitAA);
my $worryLimit = $max - (.14 * $max);
my $printWorry = int($worryLimit);
lockPrintStderr("OrigTaxId:$refTaxId");
my $bestAA = {};
my $bestNucl = {};
my $pool = {};
my $bestOverall = {};
my $poolScores = {};
my $poolRep = {};
sub scaleForAccuracy {
my $rep = $_[0];
my $accuracy = $_[1];
my $base = $_[2];
my $minRep = $_[3];
if ($accuracy == 1 or $rep == 0) {
return($rep);