Mathematical modeling suggests heterogeneous replication of Mycobacterium tuberculosis in rabbits

Tuberculosis (TB), the disease caused by Mycobacterium tuberculosis (Mtb), remains a major health problem with 10.6 million cases of the disease and 1.6 million deaths in 2021. It is well understood that pulmonary TB is due to Mtb growth in the lung but quantitative estimates of rates of Mtb replication and death in lungs of patients or animals such as monkeys or rabbits remain largely unknown. We performed experiments with rabbits infected with a novel, virulent clinical Mtb isolate of the Beijing lineage, HN878, carrying an unstable plasmid pBP10. In our in vitro experiments we found that pBP10 is more stable in HN878 strain than in a more commonly used laboratory-adapted Mtb strain H37Rv (the segregation coefficient being s = 0.10 in HN878 vs. s = 0.18 in H37Rv). Interestingly, the kinetics of plasmid-bearing bacteria in lungs of Mtb-infected rabbits did not follow an expected monotonic decline; the percent of plasmid-bearing cells increased between 28 and 56 days post-infection and remained stable between 84 and 112 days post-infection despite a large increase in bacterial numbers in the lung at late time points. Mathematical modeling suggested that such a non-monotonic change in the percent of plasmid-bearing cells can be explained if the lung Mtb population consists of several (at least 2) sub-populations with different replication/death kinetics: one major population expanding early and being controlled/eliminated, while another, a smaller population expanding at later times causing a counterintuitive increase in the percent of plasmid-bearing cells. Importantly, a model with one kinetically homogeneous Mtb population could not explain the data including when the model was run stochastically. Given that in rabbits HN878 strain forms well circumscribed granulomas, our results suggest independent bacterial dynamics in subsets of such granulomas. Our model predictions can be tested in future experiments in which HN878-pBP10 dynamics in individual granulomas is followed over time. Taken together, our new data and mathematical modeling-based analyses illustrate differences in Mtb dynamics in mice and rabbits confirming a perhaps somewhat obvious observation that "rabbits are not mice".