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package bin;
import java.io.File;
import java.io.PrintStream;
import java.util.ArrayList;
import java.util.Arrays;
import java.util.Random;
import java.util.concurrent.atomic.AtomicInteger;
import java.util.concurrent.atomic.AtomicLong;
import dna.AminoAcid;
import fileIO.ByteFile;
import fileIO.FileFormat;
import fileIO.ReadWrite;
import shared.Parse;
import shared.Parser;
import shared.PreParser;
import shared.Shared;
import shared.Timer;
import shared.Tools;
import stream.ConcurrentReadInputStream;
import stream.ConcurrentReadOutputStream;
import stream.FASTQ;
import stream.FastaReadInputStream;
import stream.Read;
import structures.ByteBuilder;
import structures.ListNum;
import tax.TaxNode;
import tax.TaxTree;
import tracker.ReadStats;
/**
* Generates synthetic reads from multiple input fastas.
*
* @author Brian Bushnell
* @date Feb 8, 2025
*
*/
public class RandomReadsMG {
/*--------------------------------------------------------------*/
/*---------------- Initialization ----------------*/
/*--------------------------------------------------------------*/
/**
* Code entrance from the command line.
* @param args Command line arguments
*/
public static void main(String[] args){
//Start a timer immediately upon code entrance.
Timer t=new Timer();
//Create an instance of this class
RandomReadsMG x=new RandomReadsMG(args);
//Run the object
x.process(t);
//Close the print stream if it was redirected
Shared.closeStream(x.outstream);
}
/**
* Constructor.
* @param args Command line arguments
*/
public RandomReadsMG(String[] args){
{//Preparse block for help, config files, and outstream
PreParser pp=new PreParser(args, getClass(), false);
args=pp.args;
outstream=pp.outstream;
}
//Set shared static variables prior to parsing
ReadWrite.USE_PIGZ=ReadWrite.USE_UNPIGZ=true;
ReadWrite.setZipThreads(Shared.threads());
FASTQ.TEST_INTERLEAVED=FASTQ.FORCE_INTERLEAVED=false;
{//Parse the arguments
final Parser parser=parse(args);
Parser.processQuality();
overwrite=ReadStats.overwrite=parser.overwrite;
append=ReadStats.append=parser.append;
out1=parser.out1;
out2=parser.out2;
qfout1=parser.qfout1;
qfout2=parser.qfout2;
extout=parser.extout;
}
validateParams();
doPoundReplacement(); //Replace # with 1 and 2
checkFileExistence(); //Ensure files can be read and written
checkStatics(); //Adjust file-related static fields as needed for this program
//Create output FileFormat objects
ffout1=FileFormat.testOutput(out1, FileFormat.FASTQ, extout, true, overwrite, append, false);
ffout2=FileFormat.testOutput(out2, FileFormat.FASTQ, extout, true, overwrite, append, false);
if("auto".equalsIgnoreCase(taxTreeFile)){taxTreeFile=TaxTree.defaultTreeFile();}
tree=TaxTree.loadTaxTree(taxTreeFile, outstream, true, false);
}
/*--------------------------------------------------------------*/
/*---------------- Initialization Helpers ----------------*/
/*--------------------------------------------------------------*/
/** Parse arguments from the command line */
private Parser parse(String[] args){
//Create a parser object
Parser parser=new Parser();
//Set any necessary Parser defaults here
//parser.foo=bar;
//Parse each argument
for(int i=0; i<args.length; i++){
String arg=args[i];
//Break arguments into their constituent parts, in the form of "a=b"
String[] split=arg.split("=");
String a=split[0].toLowerCase();
String b=split.length>1 ? split[1] : null;
if(b!=null && b.equalsIgnoreCase("null")){b=null;}
if(a.equals("verbose")){
verbose=Parse.parseBoolean(b);
}else if(a.equals("tree")){
taxTreeFile=b;
}else if(a.equals("in") || a.equals("ref")){
Tools.getFileOrFiles(b, inputFiles, true, false, false, false);
}else if(a.equals("depth") || a.equals("cov")){
minDepth=maxDepth=Float.parseFloat(b);
}else if(a.equals("mindepth") || a.equals("mincov")){
minDepth=Float.parseFloat(b);
}else if(a.equals("maxdepth") || a.equals("maxcov")){
maxDepth=Float.parseFloat(b);
}else if(a.equals("depthvariance") || a.equals("variance")){
depthVariance=Float.parseFloat(b);
}else if(a.equals("mode") || a.equals("depthmode")){
depthMode=Tools.find(b.toUpperCase(), modes);
}else if(a.equals("insert") || a.equals("avginsert")){
avgInsert=Float.parseFloat(b);
}else if(a.equals("len") || a.equals("length") || a.equals("readlen") || a.equals("readlength")){
readlen=Integer.parseInt(b);
}else if(a.equals("paired") || a.equals("int") || a.equals("interleaved")){
paired=Parse.parseBoolean(b);
}else if(a.equals("seed")){
seed=Long.parseLong(b);
}else if(b==null && Tools.find(arg.toUpperCase(), modes)>=0){
depthMode=Tools.find(arg.toUpperCase(), modes);
}else if(a.equals("parse_flag_goes_here")){
long fake_variable=Parse.parseKMG(b);
//Set a variable here
}else if(parser.parse(arg, a, b)){//Parse standard flags in the parser
//do nothing
}else{
File f=new File(arg);
if(f.exists() && f.canRead()) {
Tools.getFileOrFiles(arg, inputFiles, true, false, false, false);
}else {
outstream.println("Unknown parameter "+args[i]);
assert(false) : "Unknown parameter "+args[i];
}
}
}
return parser;
}
/** Replace # with 1 and 2 in headers */
private void doPoundReplacement(){
//Do output file # replacement
if(out1!=null && out2==null && out1.indexOf('#')>-1){
out2=out1.replace("#", "2");
out1=out1.replace("#", "1");
}
//Ensure out2 is not set without out1
if(out1==null && out2!=null){throw new RuntimeException("Error - cannot define out2 without defining out1.");}
}
/** Ensure files can be read and written */
private void checkFileExistence(){
//Ensure output files can be written
if(!Tools.testOutputFiles(overwrite, append, false, out1, out2)){
outstream.println((out1==null)+", "+(out2==null)+", "+out1+", "+out2);
throw new RuntimeException("\n\noverwrite="+overwrite+"; Can't write to output files "+out1+", "+out2+"\n");
}
//Ensure input files can be read
if(!Tools.testInputFiles(false, true, inputFiles.toArray(new String[0]))){
throw new RuntimeException("\nCan't read some input files.\n");
}
//Ensure that no file was specified multiple times
if(!Tools.testForDuplicateFiles(true, out1, out2)){
throw new RuntimeException("\nSome file names were specified multiple times.\n");
}
}
/** Adjust file-related static fields as needed for this program */
private static void checkStatics(){
//Adjust the number of threads for input file reading
if(!ByteFile.FORCE_MODE_BF1 && !ByteFile.FORCE_MODE_BF2 && Shared.threads()>2){
ByteFile.FORCE_MODE_BF2=true;
}
assert(FastaReadInputStream.settingsOK());
}
/** Ensure parameter ranges are within bounds and required parameters are set */
private boolean validateParams(){
// assert(false) : "TODO";
return true;
}
/*--------------------------------------------------------------*/
/*---------------- Outer Methods ----------------*/
/*--------------------------------------------------------------*/
/** Create read streams and process all data */
void process(Timer t){
//Turn off read validation in the input threads to increase speed
final boolean vic=Read.VALIDATE_IN_CONSTRUCTOR;
Read.VALIDATE_IN_CONSTRUCTOR=true;
//Optionally create a read output stream
final ConcurrentReadOutputStream ros=makeCros();
//Reset counters
readsProcessed=readsOut=0;
basesProcessed=basesOut=0;
//Process the reads in separate threads
spawnThreads(inputFiles, ros);
if(verbose){outstream.println("Finished; closing streams.");}
//Write anything that was accumulated by ReadStats
errorState|=ReadStats.writeAll();
//Close the read streams
errorState|=ReadWrite.closeStream(ros);
//Reset read validation
Read.VALIDATE_IN_CONSTRUCTOR=vic;
//Report timing and results
t.stop();
outstream.println(Tools.timeReadsBasesProcessed(t, readsProcessed, basesProcessed, 8));
outstream.println(Tools.readsBasesOut(t.elapsed, readsOut, basesOut, 8));
//Throw an exception of there was an error in a thread
if(errorState){
throw new RuntimeException(getClass().getName()+" terminated in an error state; the output may be corrupt.");
}
}
/** Create a Read Input Stream */
private ConcurrentReadInputStream makeCris(FileFormat ff){
ConcurrentReadInputStream cris=ConcurrentReadInputStream.getReadInputStream(-1, true, ff, null);
cris.start(); //Start the stream
if(verbose){outstream.println("Started cris");}
return cris;
}
/** Create a Read Output Stream */
private ConcurrentReadOutputStream makeCros(){
if(ffout1==null){return null;}
//Set output buffer size
final int buff=4;
//Notify user of output mode
if(paired && out2==null){
outstream.println("Writing interleaved.");
}
final ConcurrentReadOutputStream ros=ConcurrentReadOutputStream.getStream(
ffout1, ffout2, qfout1, qfout2, buff, null, false);
ros.start(); //Start the stream
return ros;
}
/** Spawn process threads */
private void spawnThreads(final ArrayList<String> files, final ConcurrentReadOutputStream ros){
//Do anything necessary prior to processing
//Determine how many threads may be used
final int threads=Shared.threads();
//Fill a list with ProcessThreads
ArrayList<ProcessThread> alpt=new ArrayList<ProcessThread>(threads);
AtomicInteger atom=new AtomicInteger(0);
for(int i=0; i<threads; i++){
alpt.add(new ProcessThread(files, ros, i, atom));
}
//Start the threads
for(ProcessThread pt : alpt){
pt.start();
}
//Wait for threads to finish
waitForThreads(alpt);
//Do anything necessary after processing
}
/** Wait until all worker threads are finished, then return */
private void waitForThreads(ArrayList<ProcessThread> alpt){
//Wait for completion of all threads
boolean success=true;
for(ProcessThread pt : alpt){
//Wait until this thread has terminated
while(pt.getState()!=Thread.State.TERMINATED){
try {
//Attempt a join operation
pt.join();
} catch (InterruptedException e) {
//Potentially handle this, if it is expected to occur
e.printStackTrace();
}
}
//Accumulate per-thread statistics
readsProcessed+=pt.readsInT;
basesProcessed+=pt.basesInT;
readsOut+=pt.readsOutT;
basesOut+=pt.basesOutT;
success&=pt.success;
}
//Track whether any threads failed
if(!success){errorState=true;}
}
/*--------------------------------------------------------------*/
/*---------------- Inner Methods ----------------*/
/*--------------------------------------------------------------*/
private ConcurrentReadInputStream makeCris(String fname){
FileFormat ff=FileFormat.testInput(fname, FileFormat.FASTA, null, true, true);
ConcurrentReadInputStream cris=ConcurrentReadInputStream.getReadInputStream(-1, false, ff, null);
cris.start(); //Start the stream
if(verbose){outstream.println("Started cris");}
return cris;
}
float chooseDepthForFile(Random randy) {
if(depthMode==MIN4) {return depthMin4(randy);}
else if(depthMode==EXP) {return depthExp(randy);}
else if(depthMode==ROOT) {return depthRoot(randy);}
else if(depthMode==LINEAR) {return depthLinear(randy);}
else {throw new RuntimeException("Unknown mode "+depthMode);}
}
float depthMin4(Random randy) {
float minRoot=(float)Math.sqrt(minDepth);
float range=(float)(Math.sqrt(maxDepth)-minRoot);
final float rootDepth=minRoot+(Tools.min(randy.nextFloat(), randy.nextFloat(),
randy.nextFloat(), randy.nextFloat()))*range;
final float fileDepth=rootDepth*rootDepth;
return fileDepth;
}
float depthLinear(Random randy) {
float range=(float)(maxDepth-minDepth);
return randy.nextFloat()*range+minDepth;
}
float depthRoot(Random randy) {
float range=(float)maxDepth-minDepth;
float root=randy.nextFloat();
return root*root*range+minDepth;
}
float depthExp(Random randy) {
double lambda=1/Math.sqrt(minDepth*maxDepth);
double depth=Tools.exponential(randy, lambda);
while(depth<minDepth || depth>maxDepth) {
depth=Tools.exponential(randy, lambda);
}
return (float)depth;
}
/*--------------------------------------------------------------*/
/*---------------- Inner Classes ----------------*/
/*--------------------------------------------------------------*/
private class ProcessThread extends Thread {
//Constructor
ProcessThread(final ArrayList<String> files_, final ConcurrentReadOutputStream ros_,
int tid_, final AtomicInteger nextFile_){
files=files_;
ros=ros_;
tid=tid_;
nextFile=nextFile_;
}
//Called by start()
@Override
public void run(){
//Do anything necessary prior to processing
randy=Shared.threadLocalRandom(seed>=0 ? seed+tid : -1);
//Process the files
for(int i=nextFile.getAndIncrement(); i<files.size(); i=nextFile.getAndIncrement()) {
String fname=files.get(i);
processFile(fname, i);
}
//Do anything necessary after processing
//Indicate successful exit status
success=true;
}
/** Iterate through the reads */
void processFile(String fname, int fnum){
// System.err.println("Thread "+tid+" processing file "+fnum+"; next="+nextFile);
ConcurrentReadInputStream cris=makeCris(fname);
//Grab the first ListNum of reads
ListNum<Read> ln=cris.nextList();
final float fileDepth=chooseDepthForFile(randy);
int taxID=TaxTree.parseHeaderStatic2(fname, tree);
if(taxID<0 && ln.size()>0) {
Read c0=ln.get(0);
taxID=TaxTree.parseHeaderStatic2(c0.id, tree);
}
System.err.println("File "+fnum+", tid "+taxID+": depth="+String.format("%.2f", fileDepth));
// assert(taxID>0) : "Can't parse taxID from "+fname;
//As long as there is a nonempty read list...
while(ln!=null && ln.size()>0){
for(Read c : ln) {
float depthRatio=1f;
if(varyDepthPerContig) {
depthRatio=1f+(depthVariance*(randy.nextFloat()+randy.nextFloat()))-depthVariance;
}
float contigDepth=depthRatio*fileDepth;
// System.err.println("depthRatio = "+depthRatio+"; contigDepth="+contigDepth);
processContig(c, contigDepth, taxID, fnum);
}
//Notify the input stream that the list was used
cris.returnList(ln);
//Fetch a new list
ln=cris.nextList();
}
//Notify the input stream that the final list was used
if(ln!=null){
cris.returnList(ln.id, ln.list==null || ln.list.isEmpty());
}
}
private void processContig(Read contig, float depth, int taxID, int fnum) {
final int basesPerRead=(paired ? 2*readlen : readlen);
readsInT++;
basesInT+=contig.length();
if(contig.length()<basesPerRead+10) {return;}
if(paired && contig.length()<avgInsert) {return;}
long basesToGenerate=(long)(depth*contig.length());
long readsGenerated=0;
long basesGenerated=0;
ArrayList<Read> list=new ArrayList<Read>(200);
float variance=varyDepthPerContig ? 0 : randy.nextFloat()*depthVariance;
// System.err.println("Generating "+basesToGenerate+" for depth-"+depth+" contig "+contig.id);
for(long i=0; basesGenerated<basesToGenerate; i++) {
Read r=generateRead(contig, i, taxID, fnum, contig.numericID, variance);
if(r!=null) {
list.add(r);
readsGenerated+=r.pairCount();
basesGenerated+=r.pairLength();
}
if(list.size()>=200) {
if(ros!=null) {ros.add(list, 0);}
list=new ArrayList<Read>(200);
}
}
if(list.size()>0) {if(ros!=null) {ros.add(list, 0);}}
// System.err.println("Generated "+basesGenerated+" for depth-"+depth+" contig "+contig.id);
readsOutT+=readsGenerated;
basesOutT+=basesGenerated;
}
private Read generateRead(Read contig, long rnum, int taxID, int fnum, long cnum, float variance) {
if(paired) {return generateReadPair(contig, rnum, taxID, fnum, cnum, variance);}
else {return generateReadSingle(contig, rnum, taxID, fnum, cnum, variance);}
}
private Read generateReadSingle(Read contig, long rnum, int taxID, int fnum, long cnum, float variance) {
int insert=readlen;
int start=randy.nextInt(contig.length()-insert);
if(skip((start+insert)/2, contig.length(), variance)){return null;}
int strand=randy.nextInt(2);
byte[] bases=Arrays.copyOfRange(contig.bases, start, start+readlen);
if(strand==1) {AminoAcid.reverseComplementBasesInPlace(bases);}
String header=makeHeader(start, strand, insert, taxID, fnum, cnum, 0);
Read r=new Read(bases, null, header, rnum);
return r;
}
private Read generateReadPair(Read contig, long rnum, int taxID, int fnum, long cnum, float variance) {
double g=randy.nextGaussian()*0.25f;
int insert=(int)((1+g)*avgInsert);
while(insert>=contig.length() || insert<readlen) {
g=randy.nextGaussian()*0.25f;
insert=(int)((1+g)*avgInsert);
}
int start1=randy.nextInt(contig.length()-insert);
int strand=randy.nextInt(2);
int start2=start1+insert-readlen;
if(skip((start1+insert)/2, contig.length(), variance)){return null;}
byte[] bases1=Arrays.copyOfRange(contig.bases, start1, start1+readlen);
byte[] bases2=Arrays.copyOfRange(contig.bases, start2, start2+readlen);
AminoAcid.reverseComplementBasesInPlace(bases2);
if(strand==1) {
byte[] temp=bases1;
bases1=bases2;
bases2=temp;
}
String header1=makeHeader(start1, strand, insert, taxID, fnum, cnum, 0);
String header2=makeHeader(start1, strand, insert, taxID, fnum, cnum, 1);
Read r1=new Read(bases1, null, header1, rnum);
Read r2=new Read(bases2, null, header2, rnum);
r2.setPairnum(1);
r1.mate=r2;
r2.mate=r1;
return r1;
}
//Skip making a read to vary depth within the contig
private boolean skip(int midpoint, int clen, float variance) {
if(variance<=0) {return false;}
//Then we want to vary within contigs
float maxSkipProb=1f-1f/(1f+variance);
float relativePosition=midpoint/(float)clen;
float skipProb=relativePosition*maxSkipProb;
return randy.nextFloat()<skipProb;
}
private String makeHeader(int start, int strand, int insert, int taxID, int fnum, long cnum, int pnum) {
bb.clear().append('f').under().append(fnum).under().append('c').under().append(cnum);
bb.under().append('s').under().append(strand).under().append('p').under().append(start);
bb.under().append('i').under().append(insert).under().append("tid");
bb.under().append(taxID).space().append(pnum+1).colon();
return bb.toString();
}
/** Number of reads retained by this thread */
protected long readsOutT=0;
/** Number of bases retained by this thread */
protected long basesOutT=0;
/** Number of input contigs processed by this thread */
protected long readsInT=0;
/** Number of input bases processed by this thread */
protected long basesInT=0;
/** True only if this thread has completed successfully */
boolean success=false;
/** Shared output stream */
private final ConcurrentReadOutputStream ros;
/** Thread ID */
final int tid;
final AtomicInteger nextFile;
private final ArrayList<String> files;
private Random randy;
private ByteBuilder bb=new ByteBuilder(128);
}
private class Genome {
public Genome(String fname_) {
fname=fname_;
}
private void load() {
FileFormat ff=FileFormat.testInput(fname, FileFormat.FASTA, null, true, true);
contigs=ConcurrentReadInputStream.getReads(-1, false, ff, null, null, null);
taxID=TaxTree.parseHeaderStatic2(fname, tree);
if(taxID<0) {taxID=TaxTree.parseHeaderStatic2(contigs.get(0).id, tree);}
}
ArrayList<Read> contigs;
String fname;
int taxID=-1;
}
/*--------------------------------------------------------------*/
/*---------------- Fields ----------------*/
/*--------------------------------------------------------------*/
/** Primary input file path */
private ArrayList<String> inputFiles=new ArrayList<String>();
/** Primary output file path */
private String out1=null;
/** Secondary output file path */
private String out2=null;
private String qfout1=null;
private String qfout2=null;
/** Override output file extension */
private String extout=null;
private String taxTreeFile=null;
/*--------------------------------------------------------------*/
/** Number of reads processed */
protected long readsProcessed=0;
/** Number of bases processed */
protected long basesProcessed=0;
/** Number of reads retained */
protected long readsOut=0;
/** Number of bases retained */
protected long basesOut=0;
private AtomicLong nextReadID=new AtomicLong(0);
private float minDepth=1;
private float maxDepth=256;
private float depthVariance=0.5f;
private float avgInsert=300;
private int readlen=150;
private boolean paired=true;
private long seed=-1;
private boolean varyDepthPerContig=false;
static final String[] modes={"MIN4", "EXP", "ROOT", "LINEAR"};
static final int MIN4=0, EXP=1, ROOT=2, LINEAR=3;
int depthMode=MIN4;
/*--------------------------------------------------------------*/
/*---------------- Final Fields ----------------*/
/*--------------------------------------------------------------*/
/** Primary output file */
private final FileFormat ffout1;
/** Secondary output file */
private final FileFormat ffout2;
private final TaxTree tree;
/*--------------------------------------------------------------*/
/*---------------- Common Fields ----------------*/
/*--------------------------------------------------------------*/
/** Print status messages to this output stream */
private PrintStream outstream=System.err;
/** Print verbose messages */
public static boolean verbose=false;
/** True if an error was encountered */
public boolean errorState=false;
/** Overwrite existing output files */
private boolean overwrite=true;
/** Append to existing output files */
private boolean append=false;
}
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