Construction and characterization of novel Mycobacterium tuberculosis-derived triple and quadruple knockout vaccines against tuberculosis

Authors

• Omar Garnica
• Raja Veerapandian
• Kishore Das
• Abhishek Mishra
• Varsha Rawat
• Areanna Carmona
• Vivek Chauhan
• Rakesh Kumar
• Jessica Chacon
• Shrikanth S. Gadad, The University of Texas Rio Grande ValleyFollow
• Subramanian Dhandayuthapani, The University of Texas Rio Grande ValleyFollow

Document Type

Article

Publication Date

4-21-2026

Abstract

Tuberculosis (TB) is a deadly disease that claims the lives of over a million people each year worldwide. The Bacille Calmette-Guérin vaccine has long been used to protect against TB, but it produces variable effects across different populations and fails to protect against adult pulmonary TB. Therefore, there is an urgent need for alternative vaccines that can offer better protection. We have developed a strategy for the rational deletion of virulence-related genes in Mycobacterium tuberculosis (Mtb) to create hyperattenuation that also enhances immunogenicity. Previously, we generated both single (∆fbpA) and double knockout (DKO) (∆fbpA-∆sapM) mutants of Mtb and assessed their immunogenicity and efficacy using mice. Herein, we have created triple knockout (TKO) and quadruple knockout (QKO) strains to enhance the immunogenicity and safety of the DKO strain by deleting the zmp1 and dosR genes. The resulting TKO strains, TKO-Z (∆fbpA-∆sapM-∆zmp1) and TKO-D (∆fbpA-∆sapM-∆dosR), and the QKO strain (∆fbpA-∆sapM-∆zmp1-∆dosR), were evaluated for their immunogenicity and safety in mice. Whereas TKO-Z and QKO strains exhibited superior immunogenicity compared to the DKO strain, their protective efficacy in mice was comparable. However, survival studies involving SCID mice indicated that the QKO strain was highly attenuated. Therefore, rational deletion of genes in Mtb seems to be an innovative approach for developing safer and more efficacious vaccines against TB.

Comments

Copyright © 2026 Garnica et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International license

Recommended Citation

Garnica, O., Veerapandian, R., Das, K., Mishra, A., Rawat, V., Carmona, A., Chauhan, V., Kumar, R., Chacon, J., Khan, A., Ramos, E. I., Gadad, S. S., Jagannath, C., & Dhandayuthapani, S. (2026). Construction and characterization of novel Mycobacterium tuberculosis-derived triple and quadruple knockout vaccines against tuberculosis. Infection and immunity, 94(5), e0050025. https://doi.org/10.1128/iai.00500-25    

Creative Commons License

This work is licensed under a Creative Commons Attribution 4.0 International License.

DOI

10.1128/iai.00500-25

Academic Level

faculty

Mentor/PI Department

Immunology and Microbiology