Use of nucleic acid probes and a nonradioactive labeling system for the detection of enteroviruses in water.

Persistent Link:
http://hdl.handle.net/10150/184948
Title:
Use of nucleic acid probes and a nonradioactive labeling system for the detection of enteroviruses in water.
Author:
Richardson, Kenneth James.
Issue Date:
1989
Publisher:
The University of Arizona.
Rights:
Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
Abstract:
Enteroviruses affect a broad segment of the population throughout the world and have been suspected to play a major role in waterborne disease for quite some time. The presence of these viruses in drinking water supplies constitutes a major health risk to the population because of their low infectious dose. The monitoring and study of these viruses in the environment have been limited by the current standard detection methodologies. Nucleic acid probe hybridization is a new and effective approach for the study and detection of these viruses in the environment. An important step in the detection of viruses in concentrated water samples by nucleic acid probes is the isolation of the viral genome from the water sample for hybridization. Previously, a series of time consuming organic extract ions was used to isolate viral RNA. This study reports the development of an alternative method for the isolation and preservation of viral RNA in environmental samples. Briefly, the sample is heated in the presence of an RNase inhibitor, and then applied to a hybridization membrane. This procedure has greatly reduced the time and difficulty of the assay while maintaining sensitivity and increasing consistency. This study reports the development and modification of a nonradioactive labeling system for the detection of viruses in water. Nonradioactive labels such as biotin offer several advantages over radioactive labels including unlimited shelf life, reduced cost and time of assay, and elimination of the radiation hazard. However, radioactive labels are generally the more sensitive method of detection. By combining direct and indirect labeling strategies, the sensitivity of this nonradioactive assay has been increased ten-fold. This assay can detect as little as 100 plaque forming units of poliovirus, only one order of magnitude less sensitive than radiolabeled probes. This assay is also ten-fold less sensitive than radiolabeled probes for the detection of enteroviruses in water samples. Nonradioactive probes offer a safe, inexpensive alternative to radiolabeled probes and tissue culture for the detection of viruses in the environment when ultrasensitivity is not required.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Nucleic acid probes; Enteroviruses; Viral pollution of water -- Measurement; Water -- Pollution -- Measurement; Viruses -- Isolation
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Microbiology and Immunology; Graduate College
Degree Grantor:
University of Arizona

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleUse of nucleic acid probes and a nonradioactive labeling system for the detection of enteroviruses in water.en_US
dc.creatorRichardson, Kenneth James.en_US
dc.contributor.authorRichardson, Kenneth James.en_US
dc.date.issued1989en_US
dc.publisherThe University of Arizona.en_US
dc.rightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.en_US
dc.description.abstractEnteroviruses affect a broad segment of the population throughout the world and have been suspected to play a major role in waterborne disease for quite some time. The presence of these viruses in drinking water supplies constitutes a major health risk to the population because of their low infectious dose. The monitoring and study of these viruses in the environment have been limited by the current standard detection methodologies. Nucleic acid probe hybridization is a new and effective approach for the study and detection of these viruses in the environment. An important step in the detection of viruses in concentrated water samples by nucleic acid probes is the isolation of the viral genome from the water sample for hybridization. Previously, a series of time consuming organic extract ions was used to isolate viral RNA. This study reports the development of an alternative method for the isolation and preservation of viral RNA in environmental samples. Briefly, the sample is heated in the presence of an RNase inhibitor, and then applied to a hybridization membrane. This procedure has greatly reduced the time and difficulty of the assay while maintaining sensitivity and increasing consistency. This study reports the development and modification of a nonradioactive labeling system for the detection of viruses in water. Nonradioactive labels such as biotin offer several advantages over radioactive labels including unlimited shelf life, reduced cost and time of assay, and elimination of the radiation hazard. However, radioactive labels are generally the more sensitive method of detection. By combining direct and indirect labeling strategies, the sensitivity of this nonradioactive assay has been increased ten-fold. This assay can detect as little as 100 plaque forming units of poliovirus, only one order of magnitude less sensitive than radiolabeled probes. This assay is also ten-fold less sensitive than radiolabeled probes for the detection of enteroviruses in water samples. Nonradioactive probes offer a safe, inexpensive alternative to radiolabeled probes and tissue culture for the detection of viruses in the environment when ultrasensitivity is not required.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectNucleic acid probesen_US
dc.subjectEnterovirusesen_US
dc.subjectViral pollution of water -- Measurementen_US
dc.subjectWater -- Pollution -- Measurementen_US
dc.subjectViruses -- Isolationen_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineMicrobiology and Immunologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.contributor.committeememberGerba, Charles P.en_US
dc.contributor.committeememberJoens, Lynn A.en_US
dc.contributor.committeememberSinclair, Norval A.en_US
dc.contributor.committeememberLindell, Thomas J.en_US
dc.contributor.committeememberHowarth, Alan J.en_US
dc.identifier.proquest9014677en_US
dc.identifier.oclc703618663en_US
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