Hello, I’ve been playing around with some data that could add to my research that’s been uploaded to the NCBI Geo repository (GSE216685).
The issue I’m having is that these samples are all technically positive for Bovine Herpesvirus, with most of them being in the early stages of reactivation of the virus. However, I have not been able to align anything to BoHV on my own. I’m talking, absolutely 0% of reads align using Hisat2, Bowtie2, Star, and BWA. Neither on my personal computer or using Galaxy.
I’ve aligned these reads to the Bos taurus genome using Hisat2 and Bowtie2 with Galaxy, which had no problem.
For example, here are the outputs for the same files when aligning to bos tau and to BoHV:
Bos taurus
52806106 reads; of these:
52806106 (100.00%) were paired; of these:
18781691 (35.57%) aligned concordantly 0 times
23049091 (43.65%) aligned concordantly exactly 1 time
10975324 (20.78%) aligned concordantly >1 times
BoHV
52806106 reads; of these:
52806106 (100.00%) were paired; of these:
52806106 (100.00%) aligned concordantly 0 times
0 (0.00%) aligned concordantly exactly 1 time
0 (0.00%) aligned concordantly >1 times
I’ve also taken the unaligned reads resulting from the Bos taurus alignments and used those and still received:
6241348 reads; of these:
6241348 (100.00%) were paired; of these:
6241348 (100.00%) aligned concordantly 0 times
0 (0.00%) aligned concordantly exactly 1 time
0 (0.00%) aligned concordantly >1 times
----
6241348 pairs aligned concordantly 0 times; of these:
0 (0.00%) aligned discordantly 1 time
----
6241348 pairs aligned 0 times concordantly or discordantly; of these:
12482696 mates make up the pairs; of these:
12482696 (100.00%) aligned 0 times
0 (0.00%) aligned exactly 1 time
0 (0.00%) aligned >1 times
0.00% overall alignment rate
However, the paper associated with this data was able to somehow detect BoHV-1 sequences (I am contacting the authors as well). These are paired-end reads, and I’ve tried running them individually as single-end (nothing). I’ve tried using the untrimmed files (I used trimmomatic for trimming) - nothing. I use JX898220.1 as my reference BoHV-1 genome, but there is also the older outdated reference genome that’s now suppressed on NCBI. Using either still gives me 0% alignment.
Do you have any idea what could be happening here? Is there maybe something obvious that I’m missing?
Thank you for any advice!
You probably should see some kind of an alignment. Took the BICP4 gene and used it to blast against one of the SRA samples at 30 min. Web blast allows blasting against SRA samples ( a unique offering that is not available elsewhere).
I see bits of alignments like this (this search should stay valid for a day or so): https://blast.ncbi.nlm.nih.gov/Blast.cgi?CMD=Get&RID=TVK4F2KK016
Data processing section here https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSM6685970 does not mention using Bovine Herpesvirus genome at all. Is this virus integrated in Bos genome?
Blasting is a great idea I'll try if all else fails. It isn't integrated into the Bos tau genome, it exists as an episome, only transcribing a couple genes until reactivation (hence this very study). They mention using the BoHV-1 reference to guide the alignment along with bos tau with Tophat2.