Identification of novel inhibitors of Mycobacterium smegmatis growth through genome-wide overexpression of Cluster P3 mycobacteriophage Xavia genes
Abstract
Bacteriophages encode numerous genes with no known function, many of which can affect essential cellular processes when expressed in the bacterial host. For mycobacteriophages, genome-wide overexpression in Mycobacterium smegmatis can be used to identify proteins that impair growth. To evaluate the cytotoxic potential of the Cluster P3 phage Xavia, we constructed a plasmid library containing 71 predicted Xavia genes under the anhydrotetracycline-inducible promoter pTet and screened this library in a plate-based cytotoxicity assay to measure impacts on M. smegmatis growth. Two genes prevented transformant recovery, consistent with toxicity under basal promoter leakiness, and inducible expression of 18 additional genes impaired growth. These inhibitory proteins include structural components; factors involved in DNA metabolism, lysogeny, and lysis; and several proteins with no known function. These results extend functional screening into a lineage of actinobacteriophages that has not previously been characterized, and identify new proteins that warrant further mechanistic analysis.
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BibTeX
@article{tennakoon2026identification,
title = {{Identification of novel inhibitors of Mycobacterium smegmatis growth through genome-wide overexpression of Cluster P3 mycobacteriophage Xavia genes}},
author = {Anushka Tennakoon and Iresha K Edirisingha and Cole R Jirsa and Ethan Rutledge and Ankita Bhattacharyya and Ella King and Madelyn Moresi and Amara Lovings and Parker R Yarborough and Anna Hoben and Garrett Manns and Toney Ray Gibson and Syed Mohammed Ashfaque Uddin and Yeseul Bae and Natalie Taylor and Taylor L Johnson and Madelyn Futral and Paige Wilson and Jalyn Brown and Zackari L Arbogast and Abigail Brooks and Carson Ward and Courtney Foxworth and Kacey Nguyen and Daphne Fairchild and Deisy Lemus and Hayley White and Holly Craft and Justin Adonis and Kamiya Givan and Sergio Valdivia and Kayci Beth Wallace and Madeline Bent and Shikelya D Chambliss and Dmitri Mavrodi and Danielle M Heller and Ramesh Rijal},
journal = {G3: Genes | Genomes | Genetics},
year = {2026},
month = sep,
doi = {10.1093/g3journal/jkag262},
url = {https://doi.org/10.1093/g3journal/jkag262},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Identification of novel inhibitors of Mycobacterium smegmatis growth through genome-wide overexpression of Cluster P3 mycobacteriophage Xavia genes
AU - Anushka Tennakoon
AU - Iresha K Edirisingha
AU - Cole R Jirsa
AU - Ethan Rutledge
AU - Ankita Bhattacharyya
AU - Ella King
AU - Madelyn Moresi
AU - Amara Lovings
AU - Parker R Yarborough
AU - Anna Hoben
AU - Garrett Manns
AU - Toney Ray Gibson
AU - Syed Mohammed Ashfaque Uddin
AU - Yeseul Bae
AU - Natalie Taylor
AU - Taylor L Johnson
AU - Madelyn Futral
AU - Paige Wilson
AU - Jalyn Brown
AU - Zackari L Arbogast
AU - Abigail Brooks
AU - Carson Ward
AU - Courtney Foxworth
AU - Kacey Nguyen
AU - Daphne Fairchild
AU - Deisy Lemus
AU - Hayley White
AU - Holly Craft
AU - Justin Adonis
AU - Kamiya Givan
AU - Sergio Valdivia
AU - Kayci Beth Wallace
AU - Madeline Bent
AU - Shikelya D Chambliss
AU - Dmitri Mavrodi
AU - Danielle M Heller
AU - Ramesh Rijal
PY - 2026
DA - 2026/09/16
JO - G3: Genes | Genomes | Genetics
DO - 10.1093/g3journal/jkag262
UR - https://doi.org/10.1093/g3journal/jkag262
AB - Bacteriophages encode numerous genes with no known function, many of which can affect essential cellular processes when expressed in the bacterial host. For mycobacteriophages, genome-wide overexpression in Mycobacterium smegmatis can be used to identify proteins that impair growth. To evaluate the cytotoxic potential of the Cluster P3 phage Xavia, we constructed a plasmid library containing 71 predicted Xavia genes under the anhydrotetracycline-inducible promoter pTet and screened this library in a plate-based cytotoxicity assay to measure impacts on M. smegmatis growth. Two genes prevented transformant recovery, consistent with toxicity under basal promoter leakiness, and inducible expression of 18 additional genes impaired growth. These inhibitory proteins include structural components; factors involved in DNA metabolism, lysogeny, and lysis; and several proteins with no known function. These results extend functional screening into a lineage of actinobacteriophages that has not previously been characterized, and identify new proteins that warrant further mechanistic analysis.
SN - 2160-1836
ER -