I need to know how many mutations in DNA resulted in amino acid changes.

I describe the mutation frequencies in DNA as follows:

Reads from Illumina were filtered Aligned the reads with the reference ORF To get the frequencies, I used mpileup

To estimate how many of these mutations resulted in mutations, I have: Translated the filtered reads using umgap for all frames Algined the reads using minimap2

I need a different approach to estimate a low abundance mutation rate that results in a different amino acid, or an alternative to mpileup that I can input translated reads into.