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A large-scale whole-genome sequencing analysis reveals false positives of bacterial essential genes

  • Genomics, Transcriptomics, Proteomics
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Essential genes are crucial for bacterial viability and represent attractive targets for novel anti-pathogen drug discovery. However, essential genes determined by the transposon insertion sequencing (Tn-seq) approach often contain many false positives. We hypothesized that some of those false positives are genes that are actually deleted from the genome, so they do not present any transposon insertion in the course of Tn-seq analysis. Based on this assumption, we performed a large-scale whole-genome sequencing analysis for the bacterium of interest. Our analysis revealed that some “essential genes” are indeed removed from the analyzed bacterial genomes. Since these genes were kicked out by bacteria, they should not be defined as essential. Our work showed that gene deletion is one of the false positive sources of essentiality determination, which is apparently underestimated in previous studies. We suggest subtracting the genome backgrounds before the evaluation of Tn-seq, and created a list of false positive gene essentiality as a reference for the downstream application.

Key points

• Discovery of false positives of essential genes defined previously through the analyses of a large scale of whole-genome sequencing data

• These false positives are the results of gene deletions in the studied genomes

• Sequencing the target genome before Tn-seq analysis is of importance while some studies neglected it

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Data availability

SRA data files analyzed in this study are listed in Supplementary Table S1.

Code availability

Used software was listed in the section of “Materials and methods.” Codes can be found at GitHub repository (https://github.com/daiweijun/Whole-genome-sequencing-analysis.git).

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Acknowledgements

Claudine Baraquet (Université de Toulon, France) is acknowledged for helpful discussing the determination of gene deletions basing on read counts. We thank E. Peter Greenberg (University of Washington, USA) for kindly donating the P. aeruginosa PAO1-UW strain. Our lab members are highly appreciable for kind supports in many aspects of this work.

Funding

This work was founded by the National Natural Science Foundation of China (31771341) and by Guangdong Province Science and Technology Innovation Strategy Special Fund (grant no: 2018B020206001).

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Authors and Affiliations

  1. Guangdong Laboratory for Lingnan Modern Agriculture, Guangzhou, 510006, China

    Yuanhao Li, Bo Jiang & Weijun Dai

  2. Guangdong Province Key Laboratory of Microbial Signals and Disease Control, Integrative Microbiology Research Center, South China Agricultural University, Guangzhou, 510642, China

    Yuanhao Li, Bo Jiang & Weijun Dai

Authors
  1. Yuanhao Li
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  2. Bo Jiang
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Contributions

Y.L. and W.D. designed and wrote the manuscript. B.J. conducted the molecular experiments. Y.L. and W.D. performed the bioinformatic analyses. All authors reviewed the manuscript.

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Correspondence to Weijun Dai.

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Li, Y., Jiang, B. & Dai, W. A large-scale whole-genome sequencing analysis reveals false positives of bacterial essential genes. Appl Microbiol Biotechnol 106, 341–347 (2022). https://doi.org/10.1007/s00253-021-11702-3

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  • DOI: https://doi.org/10.1007/s00253-021-11702-3

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