The altered fatty acylcarnitines, amino acids and organic acids detected in tuberculosis patient urine

Abstract

Mycobacterium tuberculosis is estimated to infect approximately one-third of the world’s population, which can lead to an active, symptomatic disease called tuberculosis (TB), or to asymptomatic states, often referred to as latent TB infection. In 2015 alone, 10.4 million new TB cases were reported, resulting in an estimated 1.8 million deaths. Since the discovery of M. tuberculosis in 1882 by Robert Koch, a vast amount of genomics, proteomics and transcriptomics data have been generated, leading to our current understanding of M. tuberculosis and TB. Most of the data generated from studies used M. tuberculosis cultures; however, it is well-known that this organism’s metabolism and growth in culture differs greatly from growth in the human host, where many different growth mechanisms and energy substrates are preferentially used. Furthermore, very little research to date has focused on the adaptations of M. tuberculosis to the host’s defence mechanisms or growth environment, or for that matter, the host’s adaptations or altered metabolic state in response to the infectious pathogen. This is important since the pathophysiology of M. tuberculosis is directly linked to its metabolism and complex physiology, and to that of the host. Additionally, this pathogen can utilise numerous growth substrates, either by scavenging this from the host or via de novo biosynthesis, in order to ensure its own survival. Metabolomics has served well to expand the current knowledge of the disease and has contributed towards the improved diagnosis and treatment thereof, due to its unique capacity for identifying new disease biomarkers. Metabolomics is defined as the unbiased identification and quantification of the entire metabolome in a specific biological system, with the use of highly advanced analytical instruments, together with various statistical, computational and mathematical analyses. Metabolomics has enabled the identification of new metabolite markers in sputum, blood and urine from TB patients, describing novel M. tuberculosis metabolic pathways and host adaptations. Apart from their possible diagnostic applications, many of these new TB metabolite markers have contributed to the existing knowledge of the biology of the causative pathogen, including various underlying disease mechanisms related to M. tuberculosis drug resistance and virulence, as well as the mechanisms of TB drug action and related side-effects in the host. To date, however, very little data has been published on urine from TB patients, which can be considered an ideal sample matrix to identify markers associated with this host–pathogen interaction. Considering this, in this investigation, a combined semi-targeted liquid and gas chromatography mass spectrometry metabolomics approach was used to compare the urinary fatty acylcarnitines, amino acids and selected organic acids of active TB patients with that of healthy individuals, in order to better characterise the TB-induced alterations to the host metabolome. The generally elevated concentrations of the fatty acylcarnitines and amino acids are most likely due to TB-cachexia. However, the significantly elevated concentrations of arginosuccinate, asparate (and associated asparagine), ornithine (and associated proline and hydroxyproline) and glutamate (and associated glutamine) in particular, indicate a urea cycle abnormality, due to inhibition of N-acetylglutamate synthase by the accumulating propionyl-CoA, isovaleryl-CoA and methylmalonyl-CoA in TB patients. Furthermore, elevated propionylcarnitine, methylmalonate and methylcitrate in the TB patient urine are associated with a vitamin B12 deficiency, which deserves further investigation. Lastly, various metabolites indicative of lactic acidosis, ketoacidosis, oxidative stress and liver damage were identified in the urine of the TB patients