Investigating Bacteriophage Genes Involved in the Defense of its Host
Session Title
Science and Technology
College
College of Arts and Sciences
Department
Biology
Abstract
This work is driven by the recent data collected by Winthrop’s SEA-GENES students. We now describe a phenotypic assay that can assess how a phage may express genes that defend its host against further infection. To achieve this, bacterial host Mycobacterium smegmatis was transformed with the pExTra plasmid, a vector that was assembled to contain individual genes of the K6 subcluster bacteriophage; Cain. High titer lysates from a variety of Winthrop’s phage collection were spotted directly onto lawns of transformed smegmatis. Expression of individual Cain genes are induced and observed for a protective effect against host cell lysis during further phage attack. Ideally, Cain’s homotypic defense abilities (superimmunity), can be observed in this way. However, despite repeated attempts to find a suitable temperature for the activity of both transformed host and phage Cain, superimmunity has been very difficult to test. Now, the defense assay focuses on bacteriophages outside of Cain’s subcluster. Phages ExplosiaNervosa (A9), Larva (K5), and Allegro (B2) were used in numerous defense assays to observe heterotypic defense results, or the affect an individual Cain gene may have in protecting its host from unrelated bacteriophages. So far, several of Cain genes have been revealed to have no defensive effect. However, Cain 27 does appear to exhibit defense of the host cell against lysis by phage ExplosiaNervosa. Confirmation is underway to support this discovery with the goal of developing hypotheses on this and other Cain gene characteristics.
Start Date
15-4-2022 12:00 PM
Investigating Bacteriophage Genes Involved in the Defense of its Host
This work is driven by the recent data collected by Winthrop’s SEA-GENES students. We now describe a phenotypic assay that can assess how a phage may express genes that defend its host against further infection. To achieve this, bacterial host Mycobacterium smegmatis was transformed with the pExTra plasmid, a vector that was assembled to contain individual genes of the K6 subcluster bacteriophage; Cain. High titer lysates from a variety of Winthrop’s phage collection were spotted directly onto lawns of transformed smegmatis. Expression of individual Cain genes are induced and observed for a protective effect against host cell lysis during further phage attack. Ideally, Cain’s homotypic defense abilities (superimmunity), can be observed in this way. However, despite repeated attempts to find a suitable temperature for the activity of both transformed host and phage Cain, superimmunity has been very difficult to test. Now, the defense assay focuses on bacteriophages outside of Cain’s subcluster. Phages ExplosiaNervosa (A9), Larva (K5), and Allegro (B2) were used in numerous defense assays to observe heterotypic defense results, or the affect an individual Cain gene may have in protecting its host from unrelated bacteriophages. So far, several of Cain genes have been revealed to have no defensive effect. However, Cain 27 does appear to exhibit defense of the host cell against lysis by phage ExplosiaNervosa. Confirmation is underway to support this discovery with the goal of developing hypotheses on this and other Cain gene characteristics.
