Publication Database

Establishment and application of CRISPR/Cas Technology in Non-Tuberculous Mycobacteria (NTM) (2024)

Art

Hochschulschrift

Autor

Akter, Suriya (WE 1)

Quelle

Berlin: Mensch und Buch Verlag, 2024 — XII, 103 Seiten

ISBN: 978-3-96729-237-4

Sprache

Englisch

Verweise

URL (Volltext): https://refubium.fu-berlin.de/handle/fub188/42753

Kontakt

Institut für Veterinär-Anatomie

Koserstr. 20
14195 Berlin
+49 30 838 75784
anatomie@vetmed.fu-berlin.de

Abstract / Zusammenfassung

Non-tuberculous mycobacteria (NTM) are increasingly recognized for their impact on human health, causing infections, especially in individuals with compromised immune systems or underlying lung conditions. Unlike the well-known Mycobacterium tuberculosis (MTB), NTM's intrinsic resistance to common antibiotics poses treatment challenges. Diagnosing NTM infections is complicated due to nonspecific symptoms, often confused with other diseases. Understanding NTM biology is crucial for improved diagnostics and treatments. The adoption of CRISPRi (Clustered Regularly Interspaced Short Palindromic Repeats interference) and CRISPR/Cas9 in NTM represents a significant breakthrough for advancing NTM research and treatment. The study applied the CRISPRi/dCas9 system to NTM species (M. smegmatis, M. abscessus, and M. avium), using a reporter eGFP gene for functional analysis. CRISPRi led to significant downregulation of eGFP in M. smegmatis and M. abscessus. In M. avium, reduced fluorescence, even without sgRNA, suggested the need for further optimization. CRISPRi proved effective for gene knockdown in M. abscessus. CRISPR/Cas9 was employed to knockout the eGFP gene, resulting in loss of fluorescence in M. smegmatis and M. avium but not in M. abscessus. Assessing the system on the mps1 gene in M. abscessus revealed defects in Glycopeptidolipid synthesis, causing morphotype transition. Evaluation on endogenous genes (porin, ppiA, erm-41, and whiB7) showed successful knockout for porin and ppiA. However, targeting erm-41 and whiB7 indicated the need for system optimization. The successful establishment of CRISPRi and CRISPR/Cas9 in NTM signifies a milestone, offering new avenues for research and potential therapies. These technologies enable precise genomic manipulation, enhancing our understanding of NTM biology and facilitating the development of more effective NTM infection treatments.