Background The mating type loci that govern the mating process in fungi are thought to be influenced by negative frequency-dependent selection due to rare allele advantage. In this study we used a mating type linked DNA marker as a proxy to indirectly study the allelic richness and geographic distribution of mating types of one mating type locus (MAT A) in worldwide populations of the dry rot fungus Serpula lacrymans. This fungus, which causes serious destruction to wooden constructions in temperate regions worldwide, has recently expanded its geographic range with a concomitant genetic bottleneck. Results High allelic richness and molecular variation was detected in the mating type linked marker as compared to other presumably neutral markers. Comparable amounts of genetic variation appeared in the mating type linked marker in populations from nature and buildings, which contrast the pattern observed with neutral genetic markers where natural populations were far more variable. Some geographic structuring of the allelic variation in the mating type linked marker appeared, but far less than that observed with neutral markers. In founder populations of S. lacrymans, alleles co-occurring in heterokaryotic individuals were more divergent than expected by chance, which agrees with the expectation for populations where few mating alleles exists. The analyzed DNA marker displays trans-species polymorphism wherein some alleles from the closely related species S. himantoides are more similar to those of S. lacrymans than other alleles from S. himantoides. Conclusions Our results support the idea that strong negative frequency-dependent selection maintains high levels of genetic variation in MAT-linked genomic regions, even in recently bottlenecked populations of S. lacrymans.