A three-step method for analysing bacterial biofilm formation under continuous medium flow.
For the investigation and comparison of microbial biofilms, a variety of analytical methods have been established, all focusing on different growth stages and application areas of biofilms. In this study, a novel quantitative assay for analysing biofilm maturation under the influence of continuous flow conditions was developed using the interesting biocatalyst Pseudomonas taiwanensis VLB120. In contrast to other tubular-based assay systems, this novel assay format delivers three readouts using a single setup in a total assay time of 40 h. It combines morphotype analysis of biofilm colonies with the direct quantification of biofilm biomass and pellicle formation on an air/liquid interphase. Applying the Tube-Assay, the impact of the second messenger cyclic diguanylate on biofilm formation of P. taiwanensis VLB120 was investigated. To this end, 41 deletions of genes encoding for protein homologues to diguanylate cyclase and phosphodiesterase were generated in the genome of P. taiwanensis VLB120. Subsequently, the biofilm formation of the resulting mutants was analysed using the Tube-Assay. In more than 60 % of the mutants, a significantly altered biofilm formation as compared to the parent strain was detected. Furthermore, the potential of the proposed Tube-Assay was validated by investigating the biofilms of several other bacterial species.
Laboratory of Chemical Biotechnology, Department of Biochemical and Chemical Engineering, TU Dortmund University, Emil-Figge-Strasse 66, 44227, Dortmund, Germany.
Karolin
K
Schmutzler
Andreas
A
Schmid
Katja
K
Buehler
D001431:Bacteriological Techniques / Q000295:instrumentation / Q000379:methods*; D018441:Biofilms / Q000187:drug effects / Q000254:growth & development*; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000378:metabolism; D011549:Pseudomonas / Q000187:drug effects / Q000502:physiology*
C062025:bis(3',5')-cyclic diguanylic acid; D006152:Cyclic GMP