I am not sure why you are using so much indexing.

Here is an example that demonstrates finding the two letter combinations at any point in the string.

I didn't want to have to test all the substring stuff, but this should give you an idea of how to avoid all the needless indexing.

#!/usr/bin/perl
use warnings;
use strict;
use Data::Dumper;

my @strings = ("AB", "AC", "AD", "AE", "FG");
my $counts;
for my $file (glob ('File/*')) {
    open my $fh, "<", $file || die "Could not open $!!";
    for my $line (<$fh>) {
        for my $string (@strings) {
            $counts->{$string}++ for $line =~ m/$string/g;
        }
    }
    close $fh;
}
print Dumper($counts);
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