Evidence for Highly Variable, Region-Specific Patterns of T-Cell Epitope Mutations Accumulating in Mycobacterium tuberculosis Strains

Abstract

Vaccines that confer protection through induction of adaptive T-cell immunity rely on understanding T-cell epitope (TCE) evolution induced by immune escape. This is poorly understood in tuberculosis (TB), an ancient, chronic disease, where CD4 T-cell immunity is of recognized importance. We probed 905 functionally validated, curated human CD4 T cell epitopes in 79 Mycobacterium tuberculosis (Mtb) whole genomes from India. This screen resulted in identifying 64 mutated epitopes in these strains initially using a computational pipeline and subsequently verified by single nucleotide polymorphism (SNP) analysis. SNP based phylogeny revealed the 79 Mtb strains to cluster to East African Indian (EAI), Central Asian Strain (CAS), and Beijing (BEI) lineages. Eighty-nine percent of the mutated T-cell epitopes (mTCEs) identified in the 79 Mtb strains from India has not previously been reported. These mTCEs were encoded by genes with high nucleotide diversity scores including seven mTCEs encoded by six antigens in the top 10% of rapidly divergent Mtb genes encoded by these strains. Using a T cell functional assay readout, we demonstrate 62% of mTCEs tested to significantly alter CD4 T-cell IFNγ and/or IL2 secretion with associated changes in predicted HLA-DR binding affinity: the gain of function mutations displayed higher predicted HLA-DR binding affinity and conversely mutations resulting in loss of function displayed lower predicted HLA-DR binding affinity. Most mutated antigens belonged to the cell wall/cell processes, and, intermediary metabolism and respiration families though all known Mtb proteins encoded mutations. Analysis of the mTCEs in an SNP database of 5,310 global Mtb strains identified 82% mTCEs to be significantly more prevalent in Mtb strains isolated from India, including 36 mTCEs identified exclusively in strains from India. These epitopes had a significantly higher predicted binding affinity to HLA-DR alleles that were highly prevalent in India compared to HLA-DR alleles rare in India, highlighting HLA-DR maybe an important driver of these mutations. This first evidence of region-specific TCE mutations potentially employed by Mtb to escape host immunity has important implications for TB vaccine design.

A Low-Prevalence Single-Nucleotide Polymorphism in the Sensor Kinase PhoR in Mycobacterium tuberculosis Suppresses Its Autophosphatase Activity and Reduces Pathogenic Fitness: Implications in Evolutionary Selection

“…We analyzed the sequence of phoR in 242 mycobacterial strains from BioProject PRJNA235851 in National Center for Biotechnology Information (NCBI) ( Ramaiah et al, 2019 ) with a combination of drug-sensitive/resistant, polydrug- and multidrug-resistant cohort of a South Indian population ( Ramaiah et al, 2019 ) with H37Rv as the reference sequence to identify various SNPs, and we found one that translated into a non-synonymous, missense mutation in the protein. This SNP changed alanine to threonine residue (A400T) at the 400th position, in the HATPase domain ( Figures 1A,B ).…”

“…The sequences of 19 clinical isolates from India, reported as part of a previous study ( Ramaiah et al, 2019 ), along with 223 sequences from BioProject PRJNA235851 ( Manson et al, 2017 ), were analyzed and aligned using T-COFFEE and CLUSTALW ( Madeira et al, 2019 ) using the phoR gene sequence from H37Rv strain as a template. A variation from g cc to a cc at the 1,198 nucleotide of phoR gene was identified and named PhoR’ (A400T).…”

Evolutionary Genetics of Mycobacterium Tuberculosis and HIV-1: “The Tortoise and the Hare”

“…Furthermore, amino acid variants in 10 out of 14 experimentally tested epitopes led to differential immune responses, as measured by IFN-y secretion in the blood of 82 individuals from the Gambia, latently infected with M. tuberculosis [ 61 ]. Other elaborated approaches have been also uncovering relevant CD4 + T cell epitopes, including some that are highly prevalent in certain MTBC lineages [ 62 , 63 , 64 ]. Another important feature of the immune response in TB is the balance between pathogen clearance and immunopathology [ 8 , 10 ].…”

Attenuated but immunostimulatory Mycobacterium tuberculosis variant bovis strain Ravenel shows variation in T cell epitopes

“…Over the course of the infection, immunity develops, targeting of specific antigens arises, the infection is suppressed, and re-infection by the same pathogen later hindered. On the other hand, immune responses to mycobacterial pathogens like M. tuberculosis are well-known to be more nuanced and skewed towards a cell-mediated immune response 4,8,9 . Early research suggested a limited role of B cells or antibodies in protection, and although these subsets do have beneficial roles 3,10,11 , it is known that a successful immune response against MTB infection absolutely requires CD4 + and CD8 + T cell activity 2,3,12 .…”