Dynamics of Methane-Consuming Biomes from Wieliczka Formation: Environmental and Enrichment Studies

Abstract

The rocks surrounding Wieliczka salt deposits are an extreme, deep subsurface ecosystem that as we studied previously harbors many microorganisms, including methanotrophs. In the presented research bacterial community structure of the Wieliczka Salt Mine was determined as well as the methanotrophic activity of the natural microbiome. Finally, an enrichment culture of methane-consuming methanotrophs was obtained. The research material used in this study consisted of rocks surrounding salt deposits in the Wieliczka Salt Mine. DNA was extracted directly from the pristine rock material, as well as from rocks incubated in an atmosphere containing methane and mineral medium, and from a methanotrophic enrichment culture from this ecosystem. As a result, the study describes the composition of the microbiome in the rocks surrounding the salt deposits, while also explaining how biodiversity changes during the enrichment culture of the methanotrophic bacterial community. The contribution of methanotrophic bacteria ranged from 2.614% in the environmental sample to 64.696% in the bacterial culture. The methanotrophic enrichment culture was predominantly composed of methanotrophs from the genera Methylomonas (48.848%) and Methylomicrobium (15.636%) with methane oxidation rates from 3.353 ± 0.105 to 4.200 ± 0.505 µmol CH_{4 mL}^-1 day^-1.

Keywords: Methylomonas; enrichment culture; methane; methanotrophs; salt mine.

Figure 1

Location of Miocene evaporite deposits in the Carpathian Foredeep (modified after [32]).

Figure 2

Stages of enrichment of the methane-oxidizing bacteria from environmental material being rocks associated with salt deposits at the Wieliczka Salt Mine.

Figure 3

Gas dynamics during incubation of environmental samples (W4) (A) and incubation of rock material in NMS medium (preculture) and during the first 3 days of methanotrophic enrichment culture growth (B). Growth curve of the enrichment culture and its methanotrophic activity (MA) (C).

Figure 4

Venn diagram showing the number of genera specific to a given sample and common to all samples.

Figure 5

Qualitative and quantitative changes in the structure of methanotrophs during enrichment culture of the methanotrophic community from rocks accompanying salt beds.

Figure 6

The taxonomic composition of the methanotrophic enrichment culture juxtaposed with the original environmental material (W4) and the preculture. The bacterial genera depicted in the presentation account for over 0.1% of the entire bacterial community within the enrichment culture. The heatmap presents normalized data.