BACKGROUND: Microsporidia are obligate intracellular, eukaryotic pathogens that 
infect a wide range of animals from nematodes to humans, and in some cases, 
protists. The preponderance of evidence as to the origin of the microsporidia 
reveals a close relationship with the fungi, either within the kingdom or as a 
sister group to it. Recent phylogenetic studies and gene order analysis suggest 
that microsporidia share a particularly close evolutionary relationship with the 
zygomycetes.
METHODOLOGY/PRINCIPAL FINDINGS: Here we expanded this analysis and also examined 
a putative sex-locus for variability between microsporidian populations. Whole 
genome inspection reveals a unique syntenic gene pair (RPS9-RPL21) present in 
the vast majority of fungi and the microsporidians but not in other eukaryotic 
lineages. Two other unique gene fusions (glutamyl-prolyl tRNA synthetase and 
ubiquitin-ribosomal subunit S30) that are present in metazoans, 
choanoflagellates, and filasterean opisthokonts are unfused in the fungi and 
microsporidians. One locus previously found to be conserved in many 
microsporidian genomes is similar to the sex locus of zygomycetes in gene order 
and architecture. Both sex-related and sex loci harbor TPT, HMG, and RNA 
helicase genes forming a syntenic gene cluster. We sequenced and analyzed the 
sex-related locus in 11 different Encephalitozoon cuniculi isolates and the 
sibling species E. intestinalis (3 isolates) and E. hellem (1 isolate). There 
was no evidence for an idiomorphic sex-related locus in this Encephalitozoon 
species sample. According to sequence-based phylogenetic analyses, the TPT and 
RNA helicase genes flanking the HMG genes are paralogous rather than orthologous 
between zygomycetes and microsporidians.
CONCLUSION/SIGNIFICANCE: The unique genomic hallmarks between microsporidia and 
fungi are independent of sequence based phylogenetic comparisons and further 
contribute to define the borders of the fungal kingdom and support the 
classification of microsporidia as unusual derived fungi. And the 
sex/sex-related loci appear to have been subject to frequent gene conversion and 
translocations in microsporidia and zygomycetes.