Isolation of Uncultured Bacteria from Antarctica Using Long Incubation Periods and Low Nutritional Media

Uncultured microorganisms comprise most of the microbial diversity existing on our planet. Despite advances in environmental sequencing and single-cell genomics, in-depth studies about bacterial metabolism and screening of novel bioproducts can only be assessed by culturing microbes in the laboratory. Here we report uncultured, or recalcitrant, microorganisms from an Antarctic soil sample, using relatively simple methods: oligotrophic media, extended incubation periods, observation under stereo microscopy, and selection of slow-growing bacteria. We managed to isolate several rare microorganisms belonging to infrequently isolated or recently described genera, for example Lapillicoccus, Flavitalea, Quadrisphaera, Motilibacter, and Polymorphobacter. Additionally, we obtained isolates presenting 16S rRNA sequence similarity ranging from 92.08 to 94.46% with any other known cultured species, including two distinct isolates from the class Thermoleophilia, that although common in Antarctic soils (as identified by metagenomics), was never reported to be isolated from such samples. Our data indicates that simple methods are still useful for cultivating recalcitrant microorganisms, even when dealing with samples from extreme environments

Surviving in hot and cold: psychrophiles and thermophiles from Deception Island volcano, Antarctica

Polar volcanoes harbor unique conditions of extreme temperature gradients capable of selecting different types of extremophiles. Deception Island is a marine stratovolcano located at Maritime Antarctica that is notable for its pronounced temperature gradients over very short distances, reaching values up to 100 °C in the fumaroles, and subzero temperatures next to the glaciers. Due to these characteristics, Deception can be considered an interesting analogue of extraterrestrial environments. Our main goal in this study was to isolate thermophilic and psychrophilic bacteria from sediments associated with fumaroles and glaciers from two geothermal sites in Deception Island, comprising temperatures between 0 and 98 °C, and to evaluate their survivability to desiccation and UV-C radiation. Our results revealed that culturable thermophiles and psychrophiles were recovered among the extreme temperature gradient in Deception volcano, which indicates that these extremophiles remain alive even when the conditions do not comprise their growth range. The viability of culturable psychrophiles in hyperthermophilic environments is still poorly understood and our work showed the importance of future studies about their survival strategies in high temperatures. Finally, the spore-forming thermophilic isolates which we found have displayed good survival to desiccation and UV-C irradiation, which suggests their potential to be further explored in astrobiological studies

Survival of extremophilic yeasts to the stratospheric environment on balloon flights and laboratory simulations

The high-altitude atmosphere is a harsh environment with extremely low temperatures, low pressure, and high UV irradiation. For this reason, it has been proposed as an analogue for Mars, presenting deleterious factors similar to those on the surface of that planet. We evaluated the survival of extremophilic UV-resistant yeasts isolated from a high-elevation area in the Atacama Desert under stratospheric conditions. As biological controls, intrinsically resistant Bacillus subtilis spores were used. Experiments were performed in two independent stratospheric balloon flights and with an environmental simulation chamber. The three following different conditions were evaluated: (i) desiccation, (ii) desiccation plus exposure to stratospheric low pressure and temperature, and (3) desiccation plus exposure to the full stratospheric environment (UV, low pressure, and temperature). Two strains, Naganishia (Cryptococcus) friedmannii 16LV2 and Exophiala sp. strain 15LV1, survived full exposures to the stratosphere in larger numbers than did B. subtilis spores. Holtermanniella watticus (also known as Holtermanniella wattica) 16LV1, however, suffered a substantial loss in viability upon desiccation and did not survive the stratospheric UV exposure. The remarkable resilience of N. friedmannii and Exophiala sp. 15LV1 under the extreme Mars-like conditions of the stratosphere confirms its potential as a eukaryotic model for astrobiology. Additionally, our results with N. friedmannii strengthen the recent hypothesis that yeasts belonging to the Naganishia genus are fit for aerial dispersion, which might account for the observed abundance of this species in high-elevation soils