Gene expression profiling of microbial activities and interactions in sediments under haloclines of E. Mediterranean deep hypersaline anoxic basins

Abstract

Deep-sea hypersaline anoxic basins (DHABs) in the Eastern Mediterranean Sea are considered some of the most polyextreme habitats on Earth. In comparison to microbial activities occurring within the haloclines and brines of these unusual water column habitats near the Mediterranean seafloor, relatively little is known about microbial metabolic activities in the underlying sediments. In addition, it is not known whether activities are shaped by the unique chemistries of the different DHAB brines and whether evidence exists for active microbial eukaryotes in those sediments. Metatranscriptome analysis was applied to sediment samples collected using ROV Jason from underneath the haloclines of Urania, Discovery and L'Atalante DHABs and a control site. We report on expression of genes associated with sulfur and nitrogen cycling, putative osmolyte biosynthetic pathways and ion transporters, trace metal detoxification, selected eukaryotic activities (particularly of fungi), microbe-microbe interactions, and motility in sediments underlying the haloclines of three DHABs. Relative to our control sediment sample collected outside of Urania Basin, microbial communities (including eukaryotes) in the Urania and Discovery DHAB sediments showed upregulation of expressed genes associated with nitrogen transformations, osmolyte biosynthesis, heavy metals resistance and metabolism, eukaryotic organelle functions, and cell-cell interactions. Sediments underlying DHAB haloclines that have cumulative physico-chemical stressors within the limits of tolerance for microoorganisms can therefore be hotspots of activity in the deep Mediterranean Sea.

Figure 1

Underwater images taken by ROV Jason of (a) Discovery DHAB and (b) Urania DHAB haloclines. Lightest color sediments delineate the upper boundary of the halocline as it impinges on the seafloor. Note that all sampling for this study was from areas that were much flatter than that shown in panel b.

Figure 2

Relative expression (presented as RPKM values) of genes associated with nitrogen cycling in the Urania Control (UC), Urania Middle Halocline (UMH), Discovery Upper Halocline (DUH), Discovery Lower Halocline (DLH) and L'Atalante Lower Halocline (LLH) sediment samples. Numbers following legend label refer to total salinity (PSU), oxygen (μM), and Mg²⁺ (mM) and Na⁺ (mM) concentrations in sediments as explained in Table 1. Unk=not measured.

Figure 3

Relative expression (presented as RPKM values) of genes associated with osmotic stress in the Urania Control (UC), Urania Middle Halocline (UMH), Discovery Upper Halocline (DUH), Discovery Lower Halocline (DLH) and L'Atalante Lower Halocline (LLH) sediment samples. Numbers following legend label refer to total salinity (PSU), oxygen (μM), and Mg²⁺ (mM) and Na⁺ (mM) concentrations in sediments as explained in Table 1. Unk=not measured.

Figure 4

Relative expression (presented as RPKM values) of genes associated with processing (a) mercury and (b) arsenic in the Urania Control (UC), Urania Middle Halocline (UMH), Discovery Upper Halocline (DUH), Discovery Lower Halocline (DLH) and L'Atalante Lower Halocline (LLH) sediment samples. Numbers following legend label refer to total salinity (PSU), oxygen (μM), and Mg²⁺ (mM) and Na⁺ (mM) concentrations in sediments as explained in Table 1. Unk=not measured.

Figure 5

Light micrographs of protists from DHAB haloclines. (a, b) Paired DIC (a) and epifluorescence (b; DAPI labeling) micrographs of unknown protist from Urania halocline (core 607-608 cE). (c, d) Paired phase contrast (c) and epifluorescence (d; DAPI labeling) micrographs of ciliate from Urania upper halocline (core 607 c10). (e) DIC image of ciliate from L'Atalante mid-halocline (core 611 c10). (f) Phase contrast image of ciliate from L'Atalante mid-halocline (core 611 c10). e and f are specimens also illustrated in Bernhard et al., 2014. a=20 μm, c=10 μm, e=50 μm; f=25 μm.

Figure 6

Relative expression (presented as RPKM values) of genes associated with eukaryotic intracellular transport in the Urania Control (UC), Urania Middle Halocline (UMH), Discovery Upper Halocline (DUH), Discovery Lower Halocline (DLH) and L'Atalante Lower Halocline (LLH) sediment samples. Numbers following legend label refer to total salinity (PSU), oxygen (μM), and Mg²⁺ (mM) and Na⁺ (mM) concentrations in sediments as explained in Table 1. Unk=not measured.

Figure 7

Relative expression (presented as RPKM values) of genes associated with antibiotic/antimicrobial production in the Urania Control (UC), Urania Middle Halocline (UMH), Discovery Upper Halocline (DUH), Discovery Lower Halocline (DLH) and L'Atalante Lower Halocline (LLH) sediment samples. Numbers following legend label refer to total salinity (PSU), oxygen (μM), and Mg²⁺ (mM) and Na⁺ (mM) concentrations in sediments as explained in Table 1. Unk=not measured.