AWWA PATH50798

A fluorescence in situ hybridization assay has been developed for the detection of the human-pathogenic microsporidian Encephalitozoon hellem in water samples. This assay employs a species-specific fluorescent-labeled oligonucleotide probe that was designed to be complementary to a highly variable segment of the 16S ribosomal RNA of E. hellem spores. This E. hellem specific oligonucleotide probe was successfully evaluated on tissue culture purified microsporidial spores from all three species of the genus Encephalitozoon. During this evaluation, the fluorescence in situ hybridization assay conditions were optimized to eliminate any non-specific fluorescent signal associated with E. cuniculi and E. intestinalis, while still leaving adequate fluorescent signal intensity in E. hellem spores. Moreover, specificity testing revealed that there was no observable fluorescent signal when the E. hellem specific oligonucleotide probe was tested on Vittaforma cornea, another reported waterborne microsporidial species, and numerous other common waterborne protozoa, bacteria, and algae. Includes 41 references, table, figures.