compare.py

import sys
from Bio import SeqIO


def mismatch(seq1, seq2):
    index = []

    for i, (s1, s2) in enumerate(zip(seq1, seq2)):
        if s1 != s2:
            index.append(i)

    return index


sequences = SeqIO.index(sys.argv[1], "fasta")
result = []

for i in range(0, len(sequences['species1'].seq), 3):
    codon_s1 = sequences['species1'].seq[i:i+3]
    codon_s2 = sequences['species2'].seq[i:i+3]
    codon_s3 = sequences['species3'].seq[i:i+3]

    if codon_s1 != codon_s2:
        effect = "."
        mismatches = mismatch(codon_s1, codon_s2)

        if len(mismatches) == 1:
            if codon_s1.translate() == codon_s3.translate():
                effect = "synonymous"
            else:
                effect = "non-synonymous"

        for m in mismatches:
            result.append((
                i+m+1,
                codon_s1[m],
                codon_s2[m],
                codon_s3[m],
                effect
            ))

print("#POS", "species1", "species2", "species3", "effect", sep="\t")

for row in result:
    print("\t".join(map(str, row)))

Answer on biostars question: https://www.biostars.org/p/320742