The use of comparative genomics to unravel metabolic lesions of the Mycobacterium tuberculosis complex: a case study with Mycobacterium caprae.
Romano, Giovanni Emiddio - Master degree student of LaPAM
Pereira, Taiana Tainá Silva - PhD degree student of LaPAM
Zimpel, Cristina Kraemer - PhD degree student of LaPAM
Camargo, Naila Cristina Soler - PhD degree student of LaPAM
Banari, Alexandre Campos - PhD degree student of LaPAM
Guimarães, Ana Marica de Sá - Assistant Professor of Medical Bacteriology and the head of LaPAM

Abstract: The use of comparative genomics to unravel metabolic lesions of the Mycobacterium tuberculosis complex: a case study with Mycobacterium caprae.

The metabolism of intracellular pathogens is central for host adaptation and virulence. Despite highly similar genomes, members of the Mycobacterium tuberculosis complex display distinct phenotypes of host tropism, virulence, and in vitroreplication. In our laboratory, we observed limited in vitro growth of Mycobacterium caprae, an important pathogen of zoonotic tuberculosis, suggesting that this organism presents metabolic alterations when compared to M. tuberculosis. Thus, our aim was to use comparative genomics to detect potential lesions in metabolic pathways of M. caprae and use this information to suggest novel media formulations. Using 72 publicly available M. caprae genomes, we evaluated the presence of common single nucleotide polymorphisms (SNPs) and indels in comparison to M. tuberculosis H37Rv, and the impact of these mutations on proteins and metabolic pathways of M. caprae. Accordingly, 1,577 SNPs and indels common to M. caprae were detected, including 731 proteins carrying non-synonymous SNPs or frameshifts with potential to affect protein function. Based on the PROVEAN software and assuming that proteins with frameshifts have lost their activity, 203/731 (27.77%) proteins presented deleterious mutations for their functions. Our results confirmed the deleterious mutation in the enzyme pyruvate kinase, common to animal strains, and revealed that the enzymes succinyl-CoA synthetase and propionyl-CoA carboxylase (alpha and beta subunits) are also likely altered. These lesions likely influence the production of succinate and reducing molecules, empowered by the inhibition of the isocitrate dehydrogenase through phosphenolpyruvate accumulation. Important changes were also observed on cholesterol (HIP-CoA synthetase) and nitrate (NarG and NarX) metabolisms, while other deleterious enzymes were metabolically redundant. Few virulence factors and proteins of the dormancy metabolism were also affected. Collectively, our findings indicate that M. caprae carry important metabolism deficiencies that result in poor growth in standard 7H9-OADC media with sodium pyruvate, suggesting an increased dependency on the glyoxylate shunt and methylcitrate pathway. We suggest a new media formulation based on strategies to enhance succinate production, and consequently fumarate and malate, to avoid oxidative stress and to increase reducing power. This is the first study to evaluate M. caprae metabolism, refining the understanding of its adaptive traits and in vitro growth, for research and diagnostics purposes.

Keywords: Mycobacterium tuberculosis Complex; Mycobacterium caprae; Comparative genomics; Metabolism.

Giovanni Emiddio Romano BS

University of São Paulo (USP)

Appearances