An 8-year study on the prevalence and drug resistance of mycobacteria in clinical specimens (2011-2018)
CLINICAL EPIDEMIOLOGY AND GLOBAL HEALTH
Authors: Aghajani, Jafar; Saif, Shima; Farnia, Parissa; Farnia, Poopak; Ghanavi, Jalaledin; Velayati, Ali Akbar
Non-tuberculous mycobacteria cause a wide range of clinical disorders. Isoniazid (INH) and rifampin (RIF) are the most effective first-line antibiotics against for Mycobacterium tuberculosis. The aim of this study was to investigate the prevalence of mycobacteria in clinical samples collected during 8 years (2011-2018) in the National Research Institute of Tuberculosis and Lung Disease, Tehran, Iran and to determine the tuberculosis drug resistance to first-line antibiotics, INH and RIF. In this study, 15829 different clinical specimens were collected at the NRITLD National Tuberculosis Center. A multiplex allele specific polymerase chain reaction (MAS-PCR) was used to identify mutations related to RIF and INH resistance. The genes involved were katG315, inhA for INH and rpoB516, rpoB526 and rpoB531 for RIF. In total (7528/15829, 47.56%), mycobacterial isolates including 6937 MTBC (43.82%) and 591 NTM (3.73%) were obtained. The frequency of MTBC isolates decreased from 65.17% (2015/3092) in 2011 to 47.06% (1224/2601) in 2018. Among NTM isolates, M. simiae was the most prevalent with 55.33% (327/591). The average INH resistance ratio between 7528 MTB and NTM isolates was 21%, from 15.98% in 2011 to 18.76% in 2018. In the case of RIF, the same resistance trend has been gradually increasing from 12.45% in 2011 to 14.55% in 2018. The prevalence of MDR TB has increased during the study period, from 6.49% (134/2065) in 2011 to 12.58% (174/1354) in 2018. The results of this study indicate that early detection of mycobacterial strains and determination of their drug resistance are necessary.
Integrating High-Resolution MALDI Imaging into the Development Pipeline of Anti-Tuberculosis Drugs
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
Authors: Treu, Axel; Kokesch-Himmelreich, Julia; Walter, Kerstin; Hoelscher, Christoph; Roempp, Andreas
Successful treatment of tuberculosis (TB) requires antibiotics to reach their intended point of action, i.e., necrotizing granulomas in the lung. MALDI mass spectrometry imaging (MSI) is able to visualize the distribution of antibiotics in tissue, but resolving the small histological structures in mice, which are most commonly used in preclinical trials, requires high spatial resolution. We developed a MALDI MSI method to image antibiotics in the mouse lung with high mass resolution (240k @ m/z 200 fwhm) and high spatial resolution (10 mu m pixel size). A crucial step was to develop a cryosectioning protocol that retains the distribution of water-soluble drugs in small and fragile murine lung lobes without inflation or embedding. Choice and application of matrices were optimized to detect human-equivalent drug concentrations in tissue, and measurement parameters were optimized to detect multiple drugs in a single tissue section. We succeeded in visualizing the distribution of all current first-line anti-TB drugs (pyrazinamide, rifampicin, ethambutol, isoniazid) and the second-line drugs moxifloxacin and clofazimine. Four of these compounds were imaged for the first time in the mouse lung. Accurate mass identification was confirmed by on-tissue MS/MS. Evaluation of fragmentation pathways revealed the structure of the double-protonated molecular ion of pyrazinamide. Clofazimine was imaged for the first time with 10 mu m pixel size revealing clofazimine accumulation in lipid deposits around airways. In summary, we developed a platform to resolve the detailed histology in the murine lung and to reliably detect a range of anti-TB drugs at human-equivalent doses. Our workflow is currently being employed in preclinical mouse studies to evaluate the efficacy of novel anti-TB drugs.