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. 2022 Sep 16;18(1):112.
doi: 10.1186/s13007-022-00944-5.

Protocol: a simple method for biosensor visualization of bacterial quorum sensing and quorum quenching interaction on Medicago roots

Amanda Rosier  1 , Harsh P Bais  2
Affiliations

Protocol: a simple method for biosensor visualization of bacterial quorum sensing and quorum quenching interaction on Medicago roots

Amanda Rosier et al. Plant Methods. .

Abstract

Background: Defining interactions of bacteria in the rhizosphere (encompassing the area near and on the plant root) is important to understand how they affect plant health. Some rhizosphere bacteria, including plant growth promoting rhizobacteria (PGPR) engage in the intraspecies communication known as quorum sensing (QS). Many species of Gram-negative bacteria use extracellular autoinducer signal molecules called N-acyl homoserine lactones (AHLs) for QS. Other rhizobacteria species, including PGPRs, can interfere with or disrupt QS through quorum quenching (QQ). Current AHL biosensor assays used for screening and identifying QS and QQ bacteria interactions fail to account for the role of the plant root.

Methods: Medicago spp. seedlings germinated on Lullien agar were transferred to soft-agar plates containing the broad-range AHL biosensor Agrobacterium tumefaciens KYC55 and X-gal substrate. Cultures of QS and QQ bacteria as well as pure AHLs and a QQ enzyme were applied to the plant roots and incubated for 3 days.

Results: We show that this expanded use of an AHL biosensor successfully allowed for visualization of QS/QQ interactions localized at the plant root. KYC55 detected pure AHLs as well as AHLs from live bacteria cultures grown directly on the media. We also showed clear detection of QQ interactions occurring in the presence of the plant root.

Conclusions: Our novel tri-trophic system using an AHL biosensor is useful to study QS interspecies interactions in the rhizosphere.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Fig. 1
Fig. 1
Three day after germination (DAG) M. truncatula and bacteria on KYC55 biosensor plates co-incubated with treatment for 3 days. Seedlings are 6 DAG in figures. a Control plants without bacteria b Rm8530 applied to root zone and KYC55 response to Rm8530 AHLs c KYC55 response indicating quorum quenching of Rm8530 by UD1022
Fig. 2
Fig. 2
Six day after germination (DAG) M. truncatula seedlings and YtnP lactonase on KYC55 plates. a Rm8530 and 100 µg ml−1 heat killed YtnP lactonase b Rm8530 and 100 µg ml−1 YtnP lactonase
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