I'm using ANI to determine if two bacteria species are related or not. What is the threshold for saying two sequences come from different species ?
I don't think there is such threshold. You will get different identities based on which regions of genome you are looking at. Why not identify both bacterias and take a look on how close they are in evolutionary tree?
I'm using whole genome sequence (chromosomal + plasmids) as comparisons. So it should give an idea if my samples are of the same species or not. These species have not yet been documented previously . By the way, I have found an article saying that ANI values between genomes of the same species are above 95% .
The species boundary is 94-95% depending on the reference you choose. Other boundaries such as genus and family have also been done using AAI (average amino acid identity).
Check out the following references:
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