Genome-Wide Essentiality Analysis of Mycobacterium abscessus by Saturated Transposon Mutagenesis and Deep Sequencing

• Published in: mBio Vol. 12; no. 3; p. e0104921
• Main Authors: Rifat, Dalin, Chen, Liang, Kreiswirth, Barry N, Nuermberger, Eric L
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 29.06.2021
• Subjects: DNA Transposable Elements; Genes, Essential; Genome, Bacterial; Genomics - methods; High-Throughput Nucleotide Sequencing - methods; Humans; Mutagenesis, Insertional; Mycobacterium abscessus - genetics; Mycobacterium abscessus - pathogenicity; Mycobacterium tuberculosis - genetics; Research Article; transposon; mycobacterium; genetics; deep sequencing; gene disruption; transposon sequencing (Tn-Seq); genomics; Himar1 mutagenesis; essentiality; essential gene; Mycobacterium abscessus
• ISSN: 2150-7511; 2150-7511
• DOI: 10.1128/mBio.01049-21

Mycobacterium abscessus is an emerging opportunistic human pathogen that naturally resists most major classes of antibiotics, making infections difficult to treat. Thus far, little is known about physiology, pathogenesis, and drug resistance. Genome-wide analyses have comprehensively catalogued genes with essential functions in Mycobacterium tuberculosis and Mycobacterium avium subsp. (here, ) but not in . By optimizing transduction conditions, we achieved full saturation of TA insertion sites with Himar1 transposon mutagenesis in the ATCC 19977 genome, as confirmed by deep sequencing prior to essentiality analyses of annotated genes and other genomic features. The overall densities of inserted TA sites (85.7%), unoccupied TA sites (14.3%), and nonpermissive TA sites (8.1%) were similar to results in and . Of the 4,920 annotated genes, 326 were identified as essential, 269 (83%) of which have mutual homology with essential genes, while 39 (12%) are homologous to genes that are not essential in and , and 11 (3.4%) only have homologs in . Interestingly, 7 (2.1%) essential genes have no homologs in either or , two of which were found in phage-like elements. Most essential genes are involved in DNA replication, RNA transcription and translation, and posttranslational events to synthesize important macromolecules. Some essential genes may be involved in pathogenesis and antibiotics response, including certain essential tRNAs and new short open reading frames. Our findings will help to pave the way for better understanding of and benefit development of novel bactericidal drugs against . Limited knowledge regarding pathogenesis and intrinsic resistance to most classes of antibiotics is a major obstacle to developing more effective strategies to prevent and mitigate disease. Using optimized procedures for Himar1 transposon mutagenesis and deep sequencing, we performed a comprehensive analysis to identify genetic elements essential for growth and compare them to similar data sets for and subsp. . Most essential genes have mutual homology with essential genes, providing a foundation for leveraging available knowledge from to develop more effective drugs and other interventions against . A small number of essential genes unique to deserve further attention to gain insights into what makes different from other mycobacteria. The essential genes and other genomic features such as short open reading frames and noncoding RNA identified here will provide useful information for future study of pathogenicity and new drug development.