MOLECULAR AND GENETIC CHARACTERIZATION OF THE VIRULENCE PLASMID OF YERSINIA ENTEROCOLITICA.

Persistent Link:
http://hdl.handle.net/10150/187348
Title:
MOLECULAR AND GENETIC CHARACTERIZATION OF THE VIRULENCE PLASMID OF YERSINIA ENTEROCOLITICA.
Author:
DUBEL, JACQUELINE ROBERTA.
Issue Date:
1983
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:
Purified DNA from a nalidixic acid resistant derivative of a virulent serotype 0:3 clinical isolate of Yersinia enterocolitica was subjected to transpositional mutagenesis in an effort to construct avirulent mutants. The resulting transpositional mutants, as well as the wild-type virulent strain and its isogenic derivative that had been cured of the virulence plasmid, were analyzed for plasmid DNA content. The plasmid DNA content of each strain was further characterized by restriction endonuclease digestion and the transposon insertion sites for the mutants were located. All of the strains were then tested for pathogenicity by the following assays: calcium dependence, colonization of the mouse gastrointestinal tract, HEp-2 cell adherence and invasion, HEp-2 cell monolayer detachment, autoagglutination, serum resistance, outer membrane protein production and production of V antigen. In addition, the hydrophobic properties of each strain were examined by a rapid polystyrene plate method and hydrophobic interaction chromatography. The results of the tests were compared to plasmid DNA analyses for each strain in an attempt to identify any plasmid-associated genes that are related to virulence. The wild-type strain was virulent, or positive, by all of the assays employed for evaluation of pathogenicity. In contrast, its isogenic derivative that had been cured of the virulence plasmid was negative, or avirulent, for the same assays with one exception. The avirulent plasmidless strain still retained the ability to adhere to and invade HEp-2 cells, supporting the belief that these properties are probably encoded by the bacterial chromosome. In addition, three transpositional mutants were constructed that were no longer calcium dependent, capable of detaching HEp-2 cell monolayers or able to produce three unique outer membrane proteins. Restriction endonuclease analysis confirmed the presence of the transposon on the Hind III "A" fragment of the virulence plasmid and located the region responsible for the lost virulence properties. The gene or set of genes identified were designated cal and the respective calcium independent mutants Cal⁻. Furthermore, the assays for hydrophobicity indicated that the virulence plasmid, specifically the cal gene(s), codes for hydrophobic properties on the surface of the bacterium. The study demonstrated that a virulence-associated region, cal, is located on the virulence plasmid of Y. entrocolitica and responsible for calcium dependence, HEp-2 cell monolayer detachment, the production of the three plasmid-specified outer membrane proteins and cell-surface hydrophobicity.
Type:
text; Dissertation-Reproduction (electronic)
Keywords:
Plasmids.; Plasmids -- Genetics.; Viruses.; Viral genetics.; Yersinia enterocolitica -- Physiology.
Degree Name:
Ph.D.
Degree Level:
doctoral
Degree Program:
Microbiology; Graduate College
Degree Grantor:
University of Arizona

Full metadata record

DC FieldValue Language
dc.language.isoenen_US
dc.titleMOLECULAR AND GENETIC CHARACTERIZATION OF THE VIRULENCE PLASMID OF YERSINIA ENTEROCOLITICA.en_US
dc.creatorDUBEL, JACQUELINE ROBERTA.en_US
dc.contributor.authorDUBEL, JACQUELINE ROBERTA.en_US
dc.date.issued1983en_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.abstractPurified DNA from a nalidixic acid resistant derivative of a virulent serotype 0:3 clinical isolate of Yersinia enterocolitica was subjected to transpositional mutagenesis in an effort to construct avirulent mutants. The resulting transpositional mutants, as well as the wild-type virulent strain and its isogenic derivative that had been cured of the virulence plasmid, were analyzed for plasmid DNA content. The plasmid DNA content of each strain was further characterized by restriction endonuclease digestion and the transposon insertion sites for the mutants were located. All of the strains were then tested for pathogenicity by the following assays: calcium dependence, colonization of the mouse gastrointestinal tract, HEp-2 cell adherence and invasion, HEp-2 cell monolayer detachment, autoagglutination, serum resistance, outer membrane protein production and production of V antigen. In addition, the hydrophobic properties of each strain were examined by a rapid polystyrene plate method and hydrophobic interaction chromatography. The results of the tests were compared to plasmid DNA analyses for each strain in an attempt to identify any plasmid-associated genes that are related to virulence. The wild-type strain was virulent, or positive, by all of the assays employed for evaluation of pathogenicity. In contrast, its isogenic derivative that had been cured of the virulence plasmid was negative, or avirulent, for the same assays with one exception. The avirulent plasmidless strain still retained the ability to adhere to and invade HEp-2 cells, supporting the belief that these properties are probably encoded by the bacterial chromosome. In addition, three transpositional mutants were constructed that were no longer calcium dependent, capable of detaching HEp-2 cell monolayers or able to produce three unique outer membrane proteins. Restriction endonuclease analysis confirmed the presence of the transposon on the Hind III "A" fragment of the virulence plasmid and located the region responsible for the lost virulence properties. The gene or set of genes identified were designated cal and the respective calcium independent mutants Cal⁻. Furthermore, the assays for hydrophobicity indicated that the virulence plasmid, specifically the cal gene(s), codes for hydrophobic properties on the surface of the bacterium. The study demonstrated that a virulence-associated region, cal, is located on the virulence plasmid of Y. entrocolitica and responsible for calcium dependence, HEp-2 cell monolayer detachment, the production of the three plasmid-specified outer membrane proteins and cell-surface hydrophobicity.en_US
dc.typetexten_US
dc.typeDissertation-Reproduction (electronic)en_US
dc.subjectPlasmids.en_US
dc.subjectPlasmids -- Genetics.en_US
dc.subjectViruses.en_US
dc.subjectViral genetics.en_US
dc.subjectYersinia enterocolitica -- Physiology.en_US
thesis.degree.namePh.D.en_US
thesis.degree.leveldoctoralen_US
thesis.degree.disciplineMicrobiologyen_US
thesis.degree.disciplineGraduate Collegeen_US
thesis.degree.grantorUniversity of Arizonaen_US
dc.identifier.proquest8401260en_US
dc.identifier.oclc690250161en_US
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