engleski

Visualization and identification of metabolically active microbial populations in hot spring biofilm

The composition and activity of select populations in microbial hot spring biofilms is strongly influenced by fluctuations in temperature and the availability of electron donors and acceptors. To gain further insights into these complex communities, we characterized the microbial communities by 16S rRNA gene amplicons sequencing, and identified active populations under different substrate amendments and incubation conditions. Bioorthogonal noncanonical amino acid tagging (BONCAT) was applied to reveal active cells by tracking the incorporation of synthetic amino acids into newly synthesized proteins, which are later detected by fluorescent staining using azide-alkyne click chemistry. Subsequently, the phylogenetic identity of translationally active cells was determined by combining BONCAT with catalyzed reporter deposition rRNA-targeted fluorescence in situ hybridization (CARD-FISH). Interestingly, the anabolic activity of Gammaproteobacteria and Chloroflexi was increased in almost all incubations regardless if organic or inorganic substrate was amended, but the highest activity was observed in glucose and thiosulfate amended incubations.

hot spring ; biofilm ; 16S rRNA ; Boncat ; CARD-FISH

nije evidentirano