Deferred Growth Inhibition Assay to Quantify the Effect of Bacteria-derived Antimicrobials on Competition.
Competitive exclusion can occur in microbial communities when, for example, an inhibitor-producing strain outcompetes its competitor for an essential nutrient or produces antimicrobial compounds that its competitor is not resistant to. Here we describe a deferred growth inhibition assay, a method for assessing the ability of one bacterium to inhibit the growth of another through the production of antimicrobial compounds or through competition for nutrients. This technique has been used to investigate the correlation of nasal isolates with the exclusion of particular species from a community. This technique can also be used to screen for lantibiotic producers or potentially novel antimicrobials. The assay is performed by first culturing the test inhibitor-producing strain overnight on an agar plate, then spraying over the test competitor strain and incubating again. After incubation, the extent of inhibition can be measured quantitatively, through the size of the zone of clearing around the inhibitor-producing strain, and qualitatively, by assessing the clarity of the inhibition zone. Here we present the protocol for the deferred inhibition assay, describe ways to minimize variation between experiments, and define a clarity scale that can be used to qualitatively assess the degree of inhibition.
Institute of Integrative Biology, University of Liverpool; moranj@liverpool.ac.uk.
Josephine C
JC
Moran
Institute of Integrative Biology, University of Liverpool.
Emma L
EL
Crank
Institute of Integrative Biology, University of Liverpool.
Hanaa A
HA
Ghabban
Institute of Integrative Biology, University of Liverpool.
Malcolm J
MJ
Horsburgh
D000362:Agar; D001419:Bacteria / Q000254:growth & development* / Q000378:metabolism*; D001431:Bacteriological Techniques / Q000379:methods*; D056265:Microbial Interactions / Q000502:physiology*
D000362:Agar
ISME J. 2010 Jul;4(7):839-51
20182526
Nat Rev Microbiol. 2010 Jan;8(1):15-25
19946288
BMC Genomics. 2016 Feb 29;17:152
26924200
Trends Ecol Evol. 2011 Apr;26(4):183-92
21367482
Eur J Clin Microbiol Infect Dis. 2015 Jan;34(1):123-9
25079512
Evol Appl. 2015 Aug;8(7):738-50
26240609
Lett Appl Microbiol. 2006 Mar;42(3):215-21
16478507
J Hosp Infect. 2000 Feb;44(2):127-33
10662563
PLoS One. 2010 May 17;5(5):e10598
20498722
Infect Immun. 2014 Sep;82(9):3811-5
24980973
PLoS Comput Biol. 2012;8(7):e1002606
22807668
PLoS One. 2011;6(10):e25880
21998712
Sci Adv. 2015 Jun 05;1(5):e1400216
26601194