Journal
FRONTIERS IN MICROBIOLOGY
Volume 6, Issue -, Pages -Publisher
FRONTIERS MEDIA SA
DOI: 10.3389/fmicb.2015.00243
Keywords
FACS; dinitrogen fixation; Logo di Cadagno; green sulfur bacteria; phenotypic noise; phenotypic variability; diversity; single-cell analysis
Categories
Funding
- Leopoldina [LPDS 2009-42]
- Marie-Curie-Intra-European fellowship for career development (FP7-MC-IEF
- Phenofix) [271929]
- Eawag and ETH Zurich
- European Research Council Advanced Grant (BIOCARB) [246749]
- European Research Council (ERC) [246749] Funding Source: European Research Council (ERC)
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Populations of genetically identical microorganisms residing in the same environment can display marked variability in their phenotypic traits; this phenomenon is termed phenotypic heterogeneity. The relevance of such heterogeneity in natural habitats is unknown, because phenotypic characterization of a sufficient number of single cells of the same species in complex microbial communities is technically difficult. We report a procedure that allows to measure phenotypic heterogeneity in bacterial populations from natural environments, and use it to analyze N-2 and 002 fixation of single cells of the green sulfur bacterium Chlorobium phaeobacteroides from the meromictic lake Lago di Cadagno. We incubated lake water with N-15(2) and (CO2)-C-13 under in situ conditions with and without NH4+. Subsequently, we used flow cell sorting with auto-fluorescence gating based on a pure culture isolate to concentrate C. phaeobacteroides from its natural abundance of 0.2% to now 26.5% of total bacteria. C. phaeobacteroides cells were identified using catalyzed-reporter deposition fluorescence in situ hybridization (CARD-FISH) targeting the 16S rRNA in the sorted population with a species-specific probe. In a last step, we used nanometer-scale secondary ion mass spectrometry to measure the incorporation N-15 and 130 stable isotopes in more than 252 cells. We found that C. phaeobacteroides fixes N-2 in the absence of NH4+, but not in the presence of NH4+ as has previously been suggested. N-2 and 002 fixation were heterogeneous among cells and positively correlated indicating that N-2 and CO2 fixation activity interact and positively facilitate each other in individual cells. However, because CARD-FISH identification cannot detect genetic variability among cells of the same species, we cannot exclude genetic variability as a source for phenotypic heterogeneity in this natural population. Our study demonstrates the technical feasibility of measuring phenotypic heterogeneity in a rare bacterial species in its natural habitat, thus opening the door to study the occurrence and relevance of phenotypic heterogeneity in nature.
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