Keywords:
MICROSPORIDIA, ENCEPHALITOZOON HELLEM, E. INTESTINALIS, E. CUNICULI, PROTOZOA, WATER,
Project Information:
Progress
:A fluorescent in situ hybridization (FISH) assay has been developed which targets a variable region of the rRNA of Encephalitozoon hellem with a fluorescent-labeled oligonucleotide probe. The specificity and reliability afforded by targeting unique genetic sequences within the rRNA far exceeds those requiring the biochemical expression of specific markers that may not be constant over time. In situ hybridization has the benefit of molecular information at the level of nucleic acid sequences, similar to PCR, but with the added advantage of morphological observation that PCR lacks. This assay appears to be highly sensitive and specific to an individual species, and has potential for being performed at relatively low cost. Furthermore, the FISH assay appears to work in conjunction with a fluorescently labeled antibody directed against the spore surface. Preliminary studies also have indicated that the FISH assay may be able to detect E. hellem spores seeded into source water.
Reports:
Hester, J.D., H.D.A. Lindquist, A.M. Bobst, and F.W. Schaefer, III. 2000. Fluorescent in situ detection of Encephalitozoon hellem spores with a 6-carboxyfluorescein-labeled ribosomal-RNA targeted oligonucleotide probe. Journal of Eukaryotic Microbiology 47:299-308.
Dufour, A.P., M. Feige, H.D.A. Lindquist, Editor, M. Messner, S. Regli, C. Rodgers, Frank W. Schaefer, III, S. Schaub, J. Sinclair, and L.J. Wymer. 1999. Criteria for evaluation of proposed protozoan detection methods. U.S. Environmental Protection Agency, Office of Research and Development, Cincinnati, OH. EPA 815-K-99-02.
Lindquist, H.D.A., A.P. Dufour, L.J. Wymer, F.W. Schaefer, III. 1999. Criteria for evaluation of proposed protozoan detection methods. J. Microbiol. Method. 37: 33-43.
Presentations:
Hester, J.D. H.D.A. Lindquist, A.M. Bobst, and F.W. Schaefer, III. 1999. In Situ Detetion of Encephalitozoon hellem Spores in Seeded Water Samples with a Fluorescent Labeled Ribosomal RNA-Targeted Oligonucleotide Probe. Ohio River Base Consortium for Research and Education; Cincinnati, OH.
Hester, J.D. 1999. Fluorescent In Situ Detection of Encephalitozoon hellem Spores in Environmental Water Samples. Ecology Division Seminar. Miami University, Oxford, Ohio.
Hester, J.D. H.D.A. Lindquist, A. M. Bobst and F.W. Schaefer III. 1999. Fluorescent In Situ Detection of Encephalitozoon hellem Spores in Environmental Water Samples. 2000 Joint Meeting of the American Society of Parasitologists and the Society of Protozologists, San Juan Puerto Rico. 24-28 June 2000.
Relevance
:The Office of Research and Development and the Office of Water jointly published the Research Plan for Microbial Pathogens and Disinfection By-Products in Drinking Water in 1997. The support functions and methods being developed under this task relate to the plan's priority areas, "methods for detecting and enumerating Cryptosporidium and Giardia (including identifying the viability potential) in source and finished drinking water, and methods for other emerging protozoa." Specifically, research is being performed in exposure task EX.M.7, "new protozoan agents."
The Safe Drinking Water Act (SDWA), as amended in 1996, requires the Agency to establish criteria for a monitoring program for unregulated contaminants and to publish a list of contaminants to be monitored. The Office of Water has established a Contaminant Candidate List (CCL) that identifies agents being considered for monitoring under this rule. This list includes microsporidia.
Clients
:Office of Ground Water and Drinking Water (Dr. Paul Berger)
Research Component
:M/DBP (MICROBIAL)
Risk Paradigm
:EXPOSURE
Progress
:A fluorescent in situ hybridization (FISH) assay has been developed which targets a variable region of the rRNA of Encephalitozoon hellem with a fluorescent-labeled oligonucleotide probe. The specificity and reliability afforded by targeting unique genetic sequences within the rRNA far exceeds those requiring the biochemical expression of specific markers that may not be constant over time. In situ hybridization has the benefit of molecular information at the level of nucleic acid sequences, similar to PCR, but with the added advantage of morphological observation that PCR lacks. This assay appears to be highly sensitive and specific to an individual species, and has potential for being performed at relatively low cost. Furthermore, the FISH assay appears to work in conjunction with a fluorescently labeled antibody directed against the spore surface. Preliminary studies also have indicated that the FISH assay may be able to detect E. hellem spores seeded into source water.
Reports:
Hester, J.D., H.D.A. Lindquist, A.M. Bobst, and F.W. Schaefer, III. 2000. Fluorescent in situ detection of Encephalitozoon hellem spores with a 6-carboxyfluorescein-labeled ribosomal-RNA targeted oligonucleotide probe. Journal of Eukaryotic Microbiology 47:299-308.
Dufour, A.P., M. Feige, H.D.A. Lindquist, Editor, M. Messner, S. Regli, C. Rodgers, Frank W. Schaefer, III, S. Schaub, J. Sinclair, and L.J. Wymer. 1999. Criteria for evaluation of proposed protozoan detection methods. U.S. Environmental Protection Agency, Office of Research and Development, Cincinnati, OH. EPA 815-K-99-02.
Lindquist, H.D.A., A.P. Dufour, L.J. Wymer, F.W. Schaefer, III. 1999. Criteria for evaluation of proposed protozoan detection methods. J. Microbiol. Method. 37: 33-43.
Presentations:
Hester, J.D. H.D.A. Lindquist, A.M. Bobst, and F.W. Schaefer, III. 1999. In Situ Detetion of Encephalitozoon hellem Spores in Seeded Water Samples with a Fluorescent Labeled Ribosomal RNA-Targeted Oligonucleotide Probe. Ohio River Base Consortium for Research and Education; Cincinnati, OH.
Hester, J.D. 1999. Fluorescent In Situ Detection of Encephalitozoon hellem Spores in Environmental Water Samples. Ecology Division Seminar. Miami University, Oxford, Ohio.
Hester, J.D. H.D.A. Lindquist, A. M. Bobst and F.W. Schaefer III. 1999. Fluorescent In Situ Detection of Encephalitozoon hellem Spores in Environmental Water Samples. 2000 Joint Meeting of the American Society of Parasitologists and the Society of Protozologists, San Juan Puerto Rico. 24-28 June 2000.
Relevance
:The Office of Research and Development and the Office of Water jointly published the Research Plan for Microbial Pathogens and Disinfection By-Products in Drinking Water in 1997. The support functions and methods being developed under this task relate to the plan's priority areas, "methods for detecting and enumerating Cryptosporidium and Giardia (including identifying the viability potential) in source and finished drinking water, and methods for other emerging protozoa." Specifically, research is being performed in exposure task EX.M.7, "new protozoan agents."
The Safe Drinking Water Act (SDWA), as amended in 1996, requires the Agency to establish criteria for a monitoring program for unregulated contaminants and to publish a list of contaminants to be monitored. The Office of Water has established a Contaminant Candidate List (CCL) that identifies agents being considered for monitoring under this rule. This list includes microsporidia.
Clients
:Office of Ground Water and Drinking Water (Dr. Paul Berger)
Research Component
:CCL (MICROBIAL)
Risk Paradigm
:EXPOSURE
Project IDs:
ID Code
:EX.M.7
Project type
:ORD-DW Plan
ID Code
:1974
Project type
:OMIS