Person: Ford, Christopher Burton
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Ford
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Christopher Burton
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Ford, Christopher Burton
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Publication The Evolution of Drug Resistant Mycobacterium Tuberculosis(2012-10-29) Ford, Christopher Burton; Fortune, Sarah Merritt; Lipsitch, Marc; Rubin, Eric; Bloom, Barry; Waldor, MatthewMycobacterium tuberculosis (Mtb) poses a global health catastrophe that has been compounded by the emergence of highly drug resistant Mtb strains. We used whole genome sequencing (WGS) to directly compare the accumulation of mutations in Mtb isolated from cynomolgus macaques with active, latent and early reactivation disease. Based on the distribution of single nucleotide polymorphisms (SNPs) observed, we calculated the mutation rates for these disease states. Our data suggest that during latency, Mtb acquires a similar number of chromosomal mutations as would be expected to emerge in a logarithmically growing culture over the same period of time despite reduced bacterial replication during latent infection. The pattern of polymorphisms suggests that the mutational burden in vivo is due to oxidative DNA damage. We next sought to determine why some strains of Mtb are preferentially associated with high-level drug resistance. We demonstrate that Mtb strains from the East Asian lineage acquire drug resistances in vitro more quickly than Mtb strains from the Euro-American lineage. Their higher drug resistance rate in vitro reflects a higher basal mutation. Moreover, the in vitro mutation rate correlates well with the bacterial mutation rate in humans as determined by whole genome sequencing of clinical isolates. Finally, using an agent-based model, we show that the observed differences in mutation rate predict a significantly higher probability of multi-drug resistance in patients infected with East Asian lineage strains of Mtb. Lastly, we sought to determine the mechanisms Mtb uses to proofread nascently polymerized DNA. Through fluctuation analysis of deletion mutants of two potential \(polIII\epsilon\) homologs, we demonstrate that neither is responsible for the maintenance of DNA replication fidelity. To explore the possibility that one of these homologs, Rv3711c, participates in an unknown redundant pathway, we used transposon capture and sequence (TraCS) to identify genes conditionally essential in an Rv3711c deletion mutant. Our analysis suggests that while Rv3711c does not participate in proofreading, it may act in an alternative novel DNA repair pathway. Taken together, our fluctuation analysis and TraCS data suggest that mycobacteria do not use canonical methods of proofreading to maintain genomic fidelity.Publication Mycobacterium tuberculosis mutation rate estimates from different lineages predict substantial differences in the emergence of drug resistant tuberculosis(2013) Ford, Christopher Burton; Shah, Rupal R.; Maeda, Midori Kato; Gagneux, Sebastien; Murray, Megan; Cohen, Ted; Johnston, James C.; Gardy, Jennifer; Lipsitch, Marc; Fortune, SarahA critical question in tuberculosis control is why some strains of Mycobacterium tuberculosis are preferentially associated with multiple drug resistances. We demonstrate that M. tuberculosis strains from Lineage 2 (East Asian lineage and Beijing sublineage) acquire drug resistances in vitro more rapidly than M. tuberculosis strains from Lineage 4 (Euro-American lineage) and that this higher rate can be attributed to a higher mutation rate. Moreover, the in vitro mutation rate correlates well with the bacterial mutation rate in humans as determined by whole genome sequencing of clinical isolates. Finally, using a stochastic mathematical model, we demonstrate that the observed differences in mutation rate predict a substantially higher probability that patients infected with a drug susceptible Lineage 2 strain will harbor multidrug resistant bacteria at the time of diagnosis. These data suggest that interventions to prevent the emergence of drug resistant tuberculosis should target bacterial as well as treatment-related risk factors.Publication The mutation rate of mycobacterial repetitive unit loci in strains of M. tuberculosis from cynomolgus macaque infection(BioMed Central, 2013) Ragheb, Mark N; Ford, Christopher Burton; Chase, Michael; Lin, Philana Ling; Flynn, JoAnne L; Fortune, SarahBackground: Mycobacterial interspersed repetitive units (MIRUs) are minisatellites within the Mycobacterium tuberculosis (Mtb) genome. Copy number variation (CNV) in MIRU loci is used for epidemiological typing, making the rate of variation important for tracking the transmission of Mtb strains. In this study, we developed and assessed a whole-genome sequencing (WGS) approach to detect MIRU CNV in Mtb. We applied this methodology to a panel of Mtb strains isolated from the macaque model of tuberculosis (TB), the animal model that best mimics human disease. From these data, we have estimated the rate of MIRU variation in the host environment, providing a benchmark rate for future epidemiologic work. Results: We assessed variation at the 24 MIRU loci used for typing in a set of Mtb strains isolated from infected cynomolgus macaques. We previously performed WGS of these strains and here have applied both read depth (RD) and paired-end mapping (PEM) metrics to identify putative copy number variants. To assess the relative power of these approaches, all MIRU loci were resequenced using Sanger sequencing. We detected two insertion/deletion events both of which could be identified as candidates by PEM criteria. With these data, we estimate a MIRU mutation rate of 2.70 × 10-03 (95% CI: 3.30 × 10-04- 9.80 × 10-03) per locus, per year. Conclusion: Our results represent the first experimental estimate of the MIRU mutation rate in Mtb. This rate is comparable to the highest previous estimates gathered from epidemiologic data and meta-analyses. Our findings allow for a more rigorous interpretation of data gathered from MIRU typing.Publication Distinct Effects on Diversifying Selection by Two Mechanisms of Immunity Against Streptococcus pneumoniae(Public Library of Science, 2012) Li, Yuan; Gierahn, Todd; Thompson, Claudette; Trzciński, Krzysztof; Ford, Christopher Burton; Croucher, Nicholas J; Gouveia, Paulo; Flechtner, Jessica B.; Malley, Richard; Lipsitch, MarcAntigenic variation to evade host immunity has long been assumed to be a driving force of diversifying selection in pathogens. Colonization by Streptococcus pneumoniae, which is central to the organism's transmission and therefore evolution, is limited by two arms of the immune system: antibody- and T cell- mediated immunity. In particular, the effector activity of CD4+ TH17 cell mediated immunity has been shown to act in trans, clearing co-colonizing pneumococci that do not bear the relevant antigen. It is thus unclear whether TH17 cell immunity allows benefit of antigenic variation and contributes to diversifying selection. Here we show that antigen-specific CD4+ TH17 cell immunity almost equally reduces colonization by both an antigen-positive strain and a co-colonized, antigen-negative strain in a mouse model of pneumococcal carriage, thus potentially minimizing the advantage of escape from this type of immunity. Using a proteomic screening approach, we identified a list of candidate human CD4+ TH17 cell antigens. Using this list and a previously published list of pneumococcal Antibody antigens, we bioinformatically assessed the signals of diversifying selection among the identified antigens compared to non-antigens. We found that Antibody antigen genes were significantly more likely to be under diversifying selection than the TH17 cell antigen genes, which were indistinguishable from non-antigens. Within the Antibody antigens, epitopes recognized by human antibodies showed stronger evidence of diversifying selection. Taken together, the data suggest that TH17 cell-mediated immunity, one form of T cell immunity that is important to limit carriage of antigen-positive pneumococcus, favors little diversifying selection in the targeted antigen. The results could provide new insight into pneumococcal vaccine design.