117
persisters related genes by screening the Keio deletion mutant library (Ma et al.
2010). The bacteriostatic antibiotics could induce the bacteria switch to persisters.
By screening the Keio deletion mutant library, 37 and 9 genes were found to have
deficiency in rifampin and tetracycline induced persister formation, respectively.
Those genes were mapped into DNA repair, transcription, transporters, lipopolysaccharides (LPS) biosynthesis, flagella biosynthesis, metabolism, and translation
pathway (Cui et al. 2018). Except the inactivation and deletion mutant library, the
expression library also has been used for persisters related genes identification
(Spoering et al. 2006). By screening the expression library, Lewis group found glpD
(encoding an anaerobic sn-glycerol-3-phosphate dehydrogenase) and plsB (encoding an sn-glycerol-3-phosphate acyltransferase) participate in persister cell formation in E. coli (Spoering et al. 2006).
Transposon based high-throughput screening or sequencing has the potential to
reveal putative functions for most non-essential genes within an organism. However,
this technology missed the essential genes which may play an important role in
persister formation.
5.5 Omics Approaches
5.5.1 Genomics Approaches for Persister Studies
Next generation sequencing (NGS) technologies nowadays can generate large
amounts of whole genome sequences rapidly. The experimental evolution program
has allowed investigators to obtain high persistence mutants. Combination of experimental evolution with NGS is able to identify the mutations that result in high
persistence phenotype. The first study in persister genetics was accomplished by
Moyed and Bertrand. In this study, they identified the first high persistence gene
hipA by mutagenesis-and-selection concept (Moyed and Bertrand 1983). The pathogens in a host usually faces many stresses, such as antibiotics, starvation and
immune system, and these stresses can facilitate pathogens to acquire high persistence mutations in vivo (Didelot et al. 2016). High persister strains have been
reported in clinically isolated Candida albicans (LaFleur et al. 2010), P. aeruginosa
(Mojsoska et al. 2019), S. aureus (McAdam et al. 2011), S. epidermidis (Haunreiter
et al. 2019) and M. tuberculosis (Muller et al. 2013). Genomics approaches have
also enabled identification of the high persistence associated mutation sites of the
clinical isolates. Point mutations of relA (encoding a GTP pyrophosphokinase) and
rlmN (encoding a ribosomal methyltransferase) promote S. aureus persistent infection in host (Gao et al. 2010). Mojsoska and colleague screened 467 longitudinal
clinical isolates of P. aeruginosa from 40 cystic fibrosis patients and classified
25.7% of them as high persister variants. Whole genome sequencing was used to
identify 9 candidate ‘hip’ genes among those variants and the sigma factor encoding
gene rpoN was found to be the top ‘hip’ gene (Mojsoska et al. 2019). Analysis of
5 Molecular and Systems Biology Approaches for Analyzing Drug-Tolerant Bacterial…
persisters related genes by screening the Keio deletion mutant library (Ma et al.
2010). The bacteriostatic antibiotics could induce the bacteria switch to persisters.
By screening the Keio deletion mutant library, 37 and 9 genes were found to have
deficiency in rifampin and tetracycline induced persister formation, respectively.
Those genes were mapped into DNA repair, transcription, transporters, lipopolysaccharides (LPS) biosynthesis, flagella biosynthesis, metabolism, and translation
pathway (Cui et al. 2018). Except the inactivation and deletion mutant library, the
expression library also has been used for persisters related genes identification
(Spoering et al. 2006). By screening the expression library, Lewis group found glpD
(encoding an anaerobic sn-glycerol-3-phosphate dehydrogenase) and plsB (encoding an sn-glycerol-3-phosphate acyltransferase) participate in persister cell formation in E. coli (Spoering et al. 2006).
Transposon based high-throughput screening or sequencing has the potential to
reveal putative functions for most non-essential genes within an organism. However,
this technology missed the essential genes which may play an important role in
persister formation.
5.5 Omics Approaches
5.5.1 Genomics Approaches for Persister Studies
Next generation sequencing (NGS) technologies nowadays can generate large
amounts of whole genome sequences rapidly. The experimental evolution program
has allowed investigators to obtain high persistence mutants. Combination of experimental evolution with NGS is able to identify the mutations that result in high
persistence phenotype. The first study in persister genetics was accomplished by
Moyed and Bertrand. In this study, they identified the first high persistence gene
hipA by mutagenesis-and-selection concept (Moyed and Bertrand 1983). The pathogens in a host usually faces many stresses, such as antibiotics, starvation and
immune system, and these stresses can facilitate pathogens to acquire high persistence mutations in vivo (Didelot et al. 2016). High persister strains have been
reported in clinically isolated Candida albicans (LaFleur et al. 2010), P. aeruginosa
(Mojsoska et al. 2019), S. aureus (McAdam et al. 2011), S. epidermidis (Haunreiter
et al. 2019) and M. tuberculosis (Muller et al. 2013). Genomics approaches have
also enabled identification of the high persistence associated mutation sites of the
clinical isolates. Point mutations of relA (encoding a GTP pyrophosphokinase) and
rlmN (encoding a ribosomal methyltransferase) promote S. aureus persistent infection in host (Gao et al. 2010). Mojsoska and colleague screened 467 longitudinal
clinical isolates of P. aeruginosa from 40 cystic fibrosis patients and classified
25.7% of them as high persister variants. Whole genome sequencing was used to
identify 9 candidate ‘hip’ genes among those variants and the sigma factor encoding
gene rpoN was found to be the top ‘hip’ gene (Mojsoska et al. 2019). Analysis of
5 Molecular and Systems Biology Approaches for Analyzing Drug-Tolerant Bacterial…