3D-Fluorescence In Situ Hybridization of Intact, Anaerobic Biofilm
FISH (fluorescence in situ hybridization) is a valuable technique to visualize and quantify localization of different microbial species within biofilms. Biofilm conformation can be altered during typical sample preparation for FISH, which can impact observations in multispecies biofilms, including the relative positions of cells. Here, we describe methods to preserve 3-D structure during FISH for visualization of an anaerobic coculture biofilm of Desulfovibrio vulgaris Hildenborough and Methanococcus maripaludis.
Key wordsBiofilm structure Multispecies biofilm Population interactions Sulfate-reducing bacteria Methanogenic archaea
The authors wish to thank Betsey Pitts for microscopy assistance and Dr. Sebastian Lücker for thoughtful discussions. Special thanks to Peg Dirckx for preparing Figs. 1 and 2. This work was supported as a component of ENIGMA, a scientific focus area program supported by the U.S. Department of Energy, Office of Science, Office of Biological and Environmental Research, Genomics: GTL Foundational Science through contract DE-AC02-05CH11231 between Lawrence Berkeley National Laboratory and the U.S. Department of Energy. K.A.B. and L.B.C. were also supported by a NSF-IGERT fellowship in Geobiological Systems at Montana State University (DGE 0654336). The confocal microscopy equipment used was purchased with funding from the NSF-Major Research Instrumentation Program and the M.J. Murdock Charitable Trust.