Project description:Leptospirillum ferriphilum is an important acidophilic ferrous iron-oxidizing species for bioleaching or biooxidation to win metals like copper or gold, respectively. L. ferriphilum is inhibited by elevated conentrations of chloride. In the present study, the type strain of L. ferriphilum (i.e. strain DSM 14647) by subcultivation in presence of increasing chloride concentrations was adapted to tolerate higher concentrations of NaCl. The adapted culture was grown in presence of 180 mM NaCl or and the non-adapted culture without NaCl, and both were harvested in the late exponential phase. Total RNA was isolated and checked for quality and integrity. For each of the two conditions three RNA preparations of high quality (integrity number above 7) were pooled. Ribosomal RNA was depleted. DNA libraries for paired-end sequencing on an Illumina MiSeq were generated with a TruSeq stranded mRNA library prep kit (Illumina). Among the genes up-regulated were those coding for proteins likely involved in intracellular pH regulation, response to reactive oxygen species, and Fe-S cluster biosynthesis. Among the genes down-regulated in presence of chloride were those related to lipopolysaccharide and peptidoglykane synthesis, and interestingly also those for (hydroxy) ectoine biosynthesis.
Project description:Pelagic aggregates function as hotspots for microbial activity and biological carbon pumps for exporting OM fixed by photoautotrophs to sediments in lakes and oceans. In iron-rich (ferruginous) lakes, photoferrotrophic or chemolithoautotrophic bacteria appear to contribute to CO2 fixation by oxidizing reduced iron which leads to the formation of iron-rich pelagic aggregates called iron-snow. In acidic lakes, iron snow is colonized mainly by acidophilic iron-cycling microbes that can trigger interspecies aggregation mechanisms. However, the significance of iron oxidizers in carbon fixation, their general role in iron snow functioning, and the flow of carbon within iron snow is still unclear. Here, we combined a two-year metatranscriptome analysis with a 13CO2 metabolic labeling approach to determine general metabolic activities. Protein-based stable isotope probing (protein-SIP) was used to trace the 13CO2 incorporation in iron snow microcosms over time under both oxic and anoxic conditions. Analysis of our mRNA-derived metatranscriptome data identified four key players (Leptospirillum, Ferrovum, Acidithrix, Acidiphilium) with relative abundances (59.6%-85.7%) in iron snow encoding a variety of ecologically relevant pathways, including carbon fixation, polysaccharide biosynthesis, and flagellar-based motility. We did not detect transcriptional activity for carbon fixation from archaea or eukaryotes. The largest numbers of expressed genes (3008, 2991, 2936) matched to the genomes of our previously obtained iron snow isolates (Acidithrix sp. C25, Acidiphilium sp. C61, Acidocella sp. C78) separately. 13CO2 incorporation studies identified Leptospirillum and Ferrovum, as the main active chemolithoautotrophs under oxic conditions, and Ferrovum was the main active organism under anoxic conditions as well. Small amounts of labeled 13C (Relative isotope abundance: 1.0%-5.3%) were found in the heterotrophic Acidiphilium and Acidocella. Overall, our data show that iron oxidizers play an important role in the formation of iron minerals and CO2 fixation, but the majority of fixed C apparently did not reach other iron snow microbes. This finding suggests that most of the fixed C will be directly exported to the sediment without feeding heterotrophs in the water column in acidic ferruginous lakes.