Cooperation between food-associated microorganisms
Mareike Katharina Baer, Joana Esslen, André Lipski
Department of Food Microbiology and Hygiene, University of Bonn
Background
Foodstuffs contain complex consortia of microorganisms interacting with each other in a competitive or cooperative way. Cooperative interactions of pathogens and/or spoilage microorganisms with commensal bacteria can threaten consumer’s health or reduce shelf life due to accelerated deterioration. Signal molecules called autoinducers can be secreted to coordinate community-wide functions, e.g. bioluminescence, virulence, or biofilm formation. Autoinducer 2 (AI-2) molecules are produced by both Gram-negative and Gram-positive bacteria, and are thus assumed interspecies quorum sensing (QS) signals.
Foodstuffs used for isolation
We assessed interspecies cooperation of strains isolated from a variety of foodstuffs, e. g. meat, fish, raw milk cheese, and ready-to-eat food.
Strains were identified via 16S rRNA gene sequencing, and tested for AI-2 production using bioluminescent Vibrio campbellii reporter strain ATCC BAA-1121 [1].
Detection of QS molecules
Depending on the foodstuff or origin, about 40-50 % of isolates produced AI-2. Independent of the foodstuff of origin, the genus Pseudomonas did not produce AI-2, while lactic acid bacteria as well as isolates of the genera Kocuria, Bacillus, Listeria, Brochothrix, and the order Enterobacterales were frequently tested positive for AI-2 production.
Growth and biofilm formation of cooperating isolates
Combinations of two isolates displayed enhanced growth or biofilm formation compared to pure cultures. Within the vast majority of tested combinations, we either observed enhanced growth in the planktonic phase or enhanced biofilm formation, rather than a combination of both.
In many cases, potentially cooperating combinations were composed of one AI-2 producer and one non-producer.
Spatial proximity of cooperating cells within biofilms
In some cases, cell-free supernatants of single strains excerted growth- or biofilm-enhancing effects on other strains. However, in most cases the presence of both strains was pivotal for the observed effects. Fluorescence-in-situ-hybridization (FISH) proved spatial proximity of the cooperating species within biofilms formed on stainless steel coupons, indicating beneficial effects of small distances between cooperating partners.
The picture captions the biofilm formed by a co-culture of the isolates Ps. paracarnis 26H (orange), stained with probe PSE-1284-Cy3 specific for Pseudomonas sp., and Cb. maltaromaticum 36H (blue), counterstained with DAPI. Scale bar: 10 µm.
Outlook
We recently established a gas chromatopraphy-mass spectroscopy (GC-MS) method [2] in our lab aiming to detect and quantify AI-2 production under different environmental conditions.
Further research will tackle the questions:
- Are growth or biofilm formation enhanced when cells are grown in separate compartments allowing metabolite and QS-molecule exchange while omitting direct cell contact?
- What is the effect of direct AI-2 administration, and are the effects dose-dependent?
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References
[1] Bassler, BL et al. 1997. Cross-species induction of luminescence in the quorum-sensing bacterium Vibrio harveyi. J Bacteriol 179: 4043-4045
[2] Thiel, V et al. 2009. Identification, quantification, and determination of the absolute configuration of the bacterial quorum-sensing signal autoinducer-2 by gas chromatography-mass spectrometry. ChemBioChem 10: 479-485.
