Endolithic life in the Atacama desertmicrobial ecology and adaptation strategies through a multidisciplinary approach

Resumen

The hyper-arid Atacama Desert (North Chile) is one of the most challenging polyextreme environments on Earth due to its hyper-aridity, extreme solar irradiance, both ultraviolet radiation (UVR) and photosynthetic active radiation (PAR), high day/night temperature fluctuations and, in some cases, high salinity. Despite the combination of extreme environmental conditions, microbial life has found a refuge in endolithic (inside rocks) microhabitats, as in other arid deserts, in diverse lithic substrates (gypcrete, granite, calcite, gypsum crust, halite and ignimbrite). Endolithic microhabitats are constituted by a network of pores and fissures connected to the surface within semi-translucent rock known to provide protection from lethal UVR and excess of PAR, as well as enhance moisture availability. Despite the fact that the light regime within rocks seems very scarce, the microbial communities in these microhabitats are photosynthetic-based, so that oxygenic phototrophic primary producers (mainly cyanobacteria) support a diversity of heterotrophic microorganisms. Due to the harsh conditions the cyanobacterial community is usually dominated by members of the Chroococcidiopsis genus, characterized by their extremely high resistance to ionizing radiation and desiccation. The scope of this doctoral thesis is to explore the endolithic life, paying special attention to the phototrophic fraction that supports the microbial community and its adaptations strategies and acclimation capacity, by adopting a multidisciplinary approach. This main goal is addressed in four chapters. In Chapter 1 the biogeography of microbial communities in three different endolithic microhabitats of gypcrete was investigated through a pioneering approach, on a micro scale. Microhabitat architecture was characterized using Scanning Electron Microscopy in BackScattered Electron mode (SEM-BSE) and Computerized Tomography (CT-Scan) along with the exploration of the composition and diversity of the endolithic microbial communities by means of high-throughput sequencing. These results demonstrated that microhabitats less exposed to sun radiation showed a lower diversity in the endolithic community, as well as the presence of unique cyanobacteria taxa. Thus, it was suggested that the differences in the habitable architecture of a microhabitat, even within the same piece of lithic substrate, determines microbial community diversity and composition. Chapter 2 was focused on describing the cyanobacterial community inhabiting the hypersaline endolithic microhabitat of halite in one of the driest locations on Earth, Yungay. The morphology and ultrastructure of the obtained cyanobacterial isolates from Chroococcidiopsis genus along with the cyanobacterial aggregates was characterized by means of a combination of microscopy techniques (light, fluorescence and transmission electron microscopy (TEM)). The development of a specific DNA isolation protocol allowed to perform a phylogenetic study and describe the adaptation strategies to osmotic stress through whole genome sequencing of the Chroococcidiopsis isolated strain. The comparison of the Chroococcidiopsis isolate features with those of the major cyanobacteria observed inhabiting the halite endolithic habitat pointed to Chroococcidiopsis as the dominant cyanobacterium in the halite endolithic habitat in Yungay. In Chapter 3, the response to UVR and PAR of Chroococcidiopsis strains isolated from the chasmoendolithic microhabitat of calcite and the cryptoendolithic microhabitat of halite was addressed. Oxidative stress was evaluated by studying Reactive Oxygen Species (ROS) accumulation, through spectrofluorometric measurements and microscopy observations, and the adaptation strategies of the cyanobacterial isolates to oxidative stress were described thanks to whole genome sequencing. In addition, the accumulation of the UV-screening compound scytonemin was evaluated through high performance liquid chromatography (HPLC) along with the metabolic activity and ultrastructural characterization studied by microscopy techniques (fluorescence microscopy and TEM). The results revealed differences in their acclimation to similar microhabitats exposed to slightly different light conditions. This suggested specific adaptation strategies related to their original microhabitat and a strain-specific environmental pressure selection. In Chapter 4, the bioactive compounds present in four cyanobacterial strains from the Chroococcidiopsis and Gloeocapsopsis genera isolated from different endolithic microhabitats and lithic substrates were explored. Bioassays (antibacterial and cytotoxicity), liquid chromatography tandem mass spectrometry (LC-MS/MS) and genomic tools (identification, annotation and analysis of secondary metabolites gene clusters-antiSMASH) were used to determine the actual and potential activity of each strain. The results showed a large number of compounds actually and potentially produced by the studied cyanobacterial strains with weak antibacterial activity and important cytotoxic activity against cancer cells. This suggests that cyanobacterial strains from this polyextreme environment constitute a promising source of natural products of biomedical interest. The set of results presented in this thesis suggests the importance of a “micro” perspective when analyzing the distribution and composition of microbial communities colonizing such a restrictive microhabitat as the endolithic. The microenvironmental conditions in each microhabitat and substrate and the specific biotic interactions determine the whole selection of genotypes and phenotypes able to colonize efficiently each microhabitat and lithic substrate in a polyextreme environment. This thesis, which aims to understand the ecology and behavior of endolithic microbial communities in extreme environments, encourages further studies towards understanding the limits of life, offering a new perspective on environmental selection due to the abiotic and biotic factors that occur in a microhabitat as particular as the endolithic.