Investigator: Kathleen Cornely, Providence College

Scientific Theme: Molecular Toxicology

Abstract: Bacteriophages are viruses that infect bacteria and have shown promise in the treatment of diseases caused by pathogenic bacteria that have become resistant to antibiotics. Phage begin an infection cycle by binding to surface receptors on the bacterial host, inserting the phage DNA into the host and then using host cell machinery to make copies of the phage DNA and structural proteins. Nascent phage particles are assembled, the cell lyses, and the phage are free to infect other host bacteria. This so-called “lytic cycle” (because infection results in host lysis) is the basis of phage therapy in which phage destroy bacterial pathogens. Phage are also capable of forming lysogens in which the phage DNA is incorporated into the bacterial phage DNA. The bacterial cell survives the infection and the phage DNA is replicated along with the bacterial chromosomal DNA when the cell divides. This “lysogenic cycle” also has implications for some pathogenic bacteria (an example is C. botulinum) which require the presence of a temperate phage in order to produce the toxin responsible for the disease. Bacteria harboring phage DNA can “switch” to a lytic cycle; understanding the mechanism of lysogeny will allow scientists to better exploit phage therapy to treat a variety of diseases. Here we propose to study two recently isolated K2 cluster mycobacteriophages, ZoeJ (isolated at Providence College) and Mufasa (isolated at Georgia Gwinett College). These phages infect the non-pathogenic Mycobacterium smegmatis and might also infect other mycobacterial species such as M. tuberculosis and M. leprae. Both phages contain genes coding for integrase, the enzyme that catalyzes the incorporation of phage DNA into the bacterial chromosome and are potentially able to form lysogens. The formation of lysogens will be verified by confirming the ability of the lysogen to release phage, and by PCR analysis of the genome to demonstrate phage DNA incorporation. We will then perform tests with the ZoeJ and Mufasa lysogens to determine if the lysogens are “immune” (that is, resistant to phage infection) to phages of the same (K2) subcluster, other K cluster phages, and phages of other clusters. Time-course experiments will be carried out in which we will visualize the phage infection using electron microscopy.

Human Health Relevance: The discovery a century ago of bacteriophages, viruses that infect bacteria, led scientists to propose the use of phage to treat diseases caused by bacterial pathogens. Cast aside when antibiotics were discovered, phage therapy has re-emerged as a treatment strategy for antibiotic-resistant bacteria. We propose to isolate phage capable of infecting mycobacteria; examples include include M. tuberculosis and M. leprae.