AgnosticBamCounter (abc)

The goal was to have an agnostic examination of positions as I found e.g. PacBio HIFI read analysis of bamtools mpileup to be sometimes questionable.

Emanuel Schmid-Siegert
The Agnostic Bam Counter (abc) determines at a given positions the count of observed nucleotides. Simply supply a bed file with positions and obtains a tsv file with counts for each ATCG and reference if provided

abc 0.3.2
Emanuel Schmid-Siegert
The Agnostic Bam Counter (abc) determines at a given positions the count of observed nucleotides.
Simply supply a bed file with positions and obtains a tsv file with counts for each ATCG and reference if provided.
If a reference file is provided it evaluates further the reference and variant allele frequency.
This is though always a sum over all potentially multi-allelic site.

Note: 
 - one needs to supply single nucleotide positions and not a range
 - it will sort both, by chromosome and then position - this might pose problems with funky contig names!
 - it is similar to BED 0 based so a SAM position 1000 would translate to 999-1000


USAGE:
    abc [OPTIONS] --bam <BAM> --outfile <TSV> --positions <BED>

FLAGS:
    -h, --help       Prints help information
    -V, --version    Prints version information

OPTIONS:
    -b, --bam <BAM>            aligned short or long reads
    -o, --outfile <TSV>        the output file which will be in tsv format
    -p, --positions <BED>      A bed file with positions
    -r, --reference <FASTA>    if reference in fasta format provided, reference nucleotide 
                               is provided for each positions, otherwise NA
    -t, --threads <INT>        number of threads used in thread-pool for querying positions  [default: 1]

Limitations

• the bed-file currently accepts only single nucleotide positions, no ranges.
• systematic testing is not complete

Output

Example:

##	abc:	0.3.2											
##	author:	Emanuel Schmid-Siegert											
##	date:	Thu, 03 Aug 2023 06:57:33 +0200										
##	command:	target/debug/abc --bam test/test_mpileup.bam --positions test/test_mpileup.bed --outfile test/test_mpileup.out.tsv --reference test/test_ref.fa									
# chromosome	start	end	reference	A	T	C	G	ambigious	ins	del	depthVAF	RAF
21	10405200	10405201	C	0	2	55	0	0	0	0	57	0.03510.9649
21	10402985	10402986	A	340	1	2	0	0	0	0	343	0.00580.9913
17	1884	1885	C	0	0	20	0	0	0	0	20	0.0000	1.0000
17	1870	1871	C	0	0	18	0	0	0	0	18	0.0000	1.0000
17	1869	1870	C	0	0	18	0	0	0	0	18	0.0000	1.0000
17	303	304	C	0	0	17	0	0	0	0	17	0.0000	1.0000
17	302	303	G	0	0	0	17	0	0	0	17	0.0000	1.0000

The header contains the version and author of the program as well as the execution time and the used command. We have then in the table the following columns:

• contig/chromosome name
• the start (0 based)
• the end (0 based)
• the reference nucleotide if reference was provided, otherwise "NA"
• the count of As
• the count of Ts
• the count of Cs
• the count of Gs
• the count of ambigious based, e.g. N or X
• number of reads with an insertion
• number of reads with a deletion at that position
• the depth at that position
• VAF = variant allele frequency
• RAF = reference allele frequency

Important: VAF and RAF are taking ** all ** modifications into account , as well insertion and deletions. It can happen that a VAF reported is actually the insertion or deletion event.