Abstract
Our understanding of the biological principles of mycobacterial tolerance to antibiotics is crucial for developing shorter anti-tuberculosis regimens. Various in vitro approaches have been developed to identify the conditions that promote mycobacterial persistence against antibiotics. In our laboratories, we have developed a detergent-free in vitro growth model, in which mycobacteria spontaneously grow at the air–medium interface as self-organized multicellular structures, called biofilms. Mycobacterial biofilms harbor a subpopulation of drug tolerant persisters at a greater frequency than their planktonic counterpart. Importantly, development of these structures is genetically programmed, and defective biofilms of isogenic mutants harbor fewer persisters. Thus, genetic analysis of mycobacterial biofilms in vitro could potentially be a powerful tool to unravel the biology of drug tolerance in mycobacteria. In this chapter we describe a method for screening biofilm-defective mutants of mycobacteria in a 96-well format, which readily yields a clonally pure mutant for further studies.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 215-226 |
| Number of pages | 12 |
| Journal | Methods in Molecular Biology |
| Volume | 1285 |
| DOIs | |
| State | Published - Jan 1 2015 |
| Externally published | Yes |
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Keywords
- Biofilm
- Drug tolerance
- Genetic mutants
- Mycobacteria
ASJC Scopus subject areas
- Molecular Biology
- Genetics
Cite this
Genetic dissection of mycobacterial biofilms. / Ojha, Anil K.; Jacobs, William R.; Hatfull, Graham F.
In: Methods in Molecular Biology, Vol. 1285, 01.01.2015, p. 215-226.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Genetic dissection of mycobacterial biofilms
AU - Ojha, Anil K.
AU - Jacobs, William R.
AU - Hatfull, Graham F.
PY - 2015/1/1
Y1 - 2015/1/1
N2 - Our understanding of the biological principles of mycobacterial tolerance to antibiotics is crucial for developing shorter anti-tuberculosis regimens. Various in vitro approaches have been developed to identify the conditions that promote mycobacterial persistence against antibiotics. In our laboratories, we have developed a detergent-free in vitro growth model, in which mycobacteria spontaneously grow at the air–medium interface as self-organized multicellular structures, called biofilms. Mycobacterial biofilms harbor a subpopulation of drug tolerant persisters at a greater frequency than their planktonic counterpart. Importantly, development of these structures is genetically programmed, and defective biofilms of isogenic mutants harbor fewer persisters. Thus, genetic analysis of mycobacterial biofilms in vitro could potentially be a powerful tool to unravel the biology of drug tolerance in mycobacteria. In this chapter we describe a method for screening biofilm-defective mutants of mycobacteria in a 96-well format, which readily yields a clonally pure mutant for further studies.
AB - Our understanding of the biological principles of mycobacterial tolerance to antibiotics is crucial for developing shorter anti-tuberculosis regimens. Various in vitro approaches have been developed to identify the conditions that promote mycobacterial persistence against antibiotics. In our laboratories, we have developed a detergent-free in vitro growth model, in which mycobacteria spontaneously grow at the air–medium interface as self-organized multicellular structures, called biofilms. Mycobacterial biofilms harbor a subpopulation of drug tolerant persisters at a greater frequency than their planktonic counterpart. Importantly, development of these structures is genetically programmed, and defective biofilms of isogenic mutants harbor fewer persisters. Thus, genetic analysis of mycobacterial biofilms in vitro could potentially be a powerful tool to unravel the biology of drug tolerance in mycobacteria. In this chapter we describe a method for screening biofilm-defective mutants of mycobacteria in a 96-well format, which readily yields a clonally pure mutant for further studies.
KW - Biofilm
KW - Drug tolerance
KW - Genetic mutants
KW - Mycobacteria
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U2 - 10.1007/978-1-4939-2450-9_12
DO - 10.1007/978-1-4939-2450-9_12
M3 - Article
C2 - 25779318
AN - SCOPUS:84925266850
VL - 1285
SP - 215
EP - 226
JO - Methods in Molecular Biology
JF - Methods in Molecular Biology
SN - 1064-3745
ER -