And reading the file as binary. There's a lesson about the overhead of unicode strings here ;)

Your first example takes 3.1 seconds, my previous comment takes 2.3 seconds, this one takes 1.4 seconds.

    start = time.perf_counter()

    with open("orthocoronavirinae.fasta", "rb") as f:
    total = sum(map(lambda s: 0 if s[0]==65 else s.count(b"G") + s.count(b"C"), f.readlines()))
    
    end = time.perf_counter()
    
    print(total, " total")
    print(end-start, " seconds")