Genetic and environmental conditions influencing

persistence

Hannes LuidaleppInstitute of Technology

University of TartuAugust 2011

Overview

• Persisters

• Method to follow cell division

• My results

Bacterial “lifecycle” in lab

Resistance

• Including whole population

• Genetically determined• Pumping out antibiotics• Degradation of antibiotics• Altering the target• etc

Persistence

• Part of the population is insensitive – persisters

• Medically important – chronical and persistent infections

• Genetically identical to the mother cells

• Phenotypical property

Persisters

• frequency usually 10-4-10-6 • mainly originate from stationary phase

• common to all bacterial species investigated (also to fungi and cancer cells)

Keren, et al 2004

High persistence (Hip) mutants

• persister frequency usually at least 100x higher

(also some clinical isolates of C. candida, P. aeruginosa)

• most common mutant is hipA7 (point mutations in hipA gene)

• hipA is a toxin from hipBA

antitoxin-toxin pair

Persisters - dormant cells

Balaban, et al 2004

PersistersType I persister: originate from stationary

phase

Type II persisters: originate from growth phase

Balaban, et al 2004

Physiology of persisters

• Gene expression arraysdifferent toxin/antitoxin genes, other unrelated

genes

• Mutant collection screensdifferent unrelated genes

• Over-expression studies

all toxic stuff will increase persistence

Method to follow cell division

Roostalu, et al 2008

Cell division

• LB medium

• 24h stationary phaseculture diluted into fresh media

• Two population dividing

non-dividing

Age of the culture and persistence

Growth resumption dynamics

Length of antibiotic treatment

Persister dynamics in

mutants

Growth resumption of sucB mutant

Other TCA cycle mutants

Summary

• Growth resumption depends on:– media

– age of the culture

• Level of persisters depends on growth resumption

Mutants and persistence

– Mutants show different persister formation dynamics

– Difference from wild type might be observed only in certain conditions

Medical applications

– To make cell to wake up faster

Allison et al., 2011 Kim et al., 2011

– To make cell permanently dormant

Acknowledgments

People from Tanel Tenson’s lab

Specially Arvi Jõers and Niilo Kaldalu