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Cryptosporidium parvum oocysts recovered from water by the membrane filter dissolution method retain their
T K Graczyk1, R Fayer, M R Cranfield
1Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.
Abstract:
Cryptosporidium parvum oocysts infectious to neonatal BALB/c mice were processed by the cellulose-acetate membrane (CAM) filter dissolution method to determine if the procedure that utilizes acetone incubation and alcohol centrifugations alters their viability (determined by in vitro excystation) or infectivity (determined by infectivity bioassay). In addition, most oocysts with altered viability by desiccation, heat inactivation, and snap freezing that were processed by the CAM filter dissolution method were nonrefractile, unstained oocyst ghosts. The remaining organisms, oocyst shells, were lightly stained with the acid-fast stain. Infectious oocysts retained their infectivity and nonviable oocysts (oocyst shells) retained their morphology when processed by the CAM dissolution method. Infectious oocysts, oocyst shells, and oocyst ghosts produced positive reactions of similar intensity in direct immunofluorescence antibody staining, utilizing the MERIFLUOR Cryptosporidium/Giardia test kit. Cryptosporidium oocysts recovered from finished drinking water by the CAM dissolution method can be subjected to testing for their viability and infectivity.
Insights
The cellulose-acetate membrane (CAM) filter dissolution method preserves Cryptosporidium parvum oocyst infectivity and morphology. This method allows for accurate viability and infectivity testing of oocysts recovered from drinking water.
Area of Science:
- Environmental microbiology
- Parasitology
- Water quality analysis
Background:
- Cryptosporidium parvum is a significant waterborne pathogen.
- Accurate detection of viable oocysts in water is crucial for public health.
- Existing methods for oocyst recovery and viability assessment can be challenging.
Purpose of the Study:
- To evaluate the impact of the cellulose-acetate membrane (CAM) filter dissolution method on Cryptosporidium parvum oocyst viability and infectivity.
- To determine if the CAM filter dissolution method is suitable for recovering oocysts from finished drinking water for subsequent viability and infectivity testing.
Main Methods:
- Infectious Cryptosporidium parvum oocysts were processed using the CAM filter dissolution method.
- Oocyst viability was assessed by in vitro excystation.
- Oocyst infectivity was determined by infectivity bioassay in neonatal BALB/c mice.
- Oocyst morphology and staining characteristics were examined using acid-fast stain and direct immunofluorescence antibody staining (MERIFLUOR Cryptosporidium/Giardia test kit).
Main Results:
- The CAM filter dissolution method did not alter the infectivity of viable oocysts.
- Nonviable oocysts (oocyst shells and ghosts) retained their morphology after processing.
- Both infectious oocysts and oocyst shells showed similar positive reactions in immunofluorescence antibody staining.
- The method successfully recovered Cryptosporidium oocysts from finished drinking water.
Conclusions:
- The CAM filter dissolution method is a viable technique for recovering Cryptosporidium oocysts from drinking water.
- This method preserves oocyst infectivity and allows for subsequent accurate assessment of viability and infectivity.
- The findings support the use of the CAM method for routine monitoring of Cryptosporidium in water supplies.