A Harsh Life to Indigenous Proteobacteria at the Andean Mountains: Microbial Diversity and Resistance Mechanisms Towards Extreme Conditions

Abstract

High-altitude Andean lake (HAAL) ecosystems of the South American Andes are almost unexplored systems of shallow lakes formed during the Tertiary geological period, distributed in the geographical area called the Puna at altitudes from 3,000 to 6,000 m above sea level, and isolated from direct human activity. They present a broad range of extreme conditions which makes the indigenous microbial communities exceptionally interesting to study physiological mechanisms of adaptation to chemical and physical stresses such as hypersalinity and high levels of UV radiation. Previous work have revealed the outstanding diversity of these environments, being Proteobacteria the most extended and best represented microbial taxa within the extremophilic communities. The aim of this work is to review the microbial diversity of Proteobacteria present at the HAAL and to describe their multiple resistance properties towards the extreme factors that these microbial communities thrived in their natural environments. A special reference to the representatives of the genus Acinetobacter found at the HAAL is also presented. Due to the isolation program held at LIMLA (www.limla.com.ar) during the past four years a one-of-a-kind collection of extremophilic strains from the HAAL was assembled. HAAL microbial diversity was investigated by sampling bacterioplankton, benthonic microorganisms, microbial-mat associated microbes as well as gastrointestinal symbiotic organisms from flamingoes living at the lakes. Representatives of Proteobacteria has been profusely isolated from these samples, more exactly from Lakes: Azul, Verde, Negra, Vilama, Aparejos, Chaxas, Salina Grande, Socompa, Dead Man Salar, Tolar Grande, Brava, Diamante, Huaca-Huasi, all of them located above 4000 m, at the Northwest of Argentina. In addition, a more extended coverage of Proteobacteria was detected by non-culture dependent techniques (mainly DGGE), suggesting that much more efforts will be needed to isolate most novel Proteobacteria present at the HAAL. Within Proteobacteria, all the four main groups were represented in our culture collection being the Gammaproteobacteria the class with better coverage. The gammaproteobacteria strains were classified as belonging mainly to Pseudomonas Acinetobacter, Halomonas, Stenotrophomonas, Moraxella, Enterobacter, Serratia, Salinivibrio, Pseudoalteromonas, Aeromonas and Marinobacter. 16S rDNA gene sequence comparison of some isolates with the ones presented at the database indicated an identity lower that 94%, which should point out that these extremophilic communities harbour yet unraveled species. The extreme conditions suffered by these microorganisms at the HAAL made them resistant to factors present as well as not present in their natural environments. Exposure to UV-B radiation during 24 h revealed that most isolates were highly resistant: 33.3% of betaproteobacteria, 44.4% of gammaproteobacteria, 40% of alphaproteobacteria were able to survive through the whole exposition time. In addition, resistance to hipersalinity in most isolates was also observed. Interestingly, antibiotic resistance was also observed in spite of the pristinely and isolation of these lakes. In light of the great adaptability strength of the strains to changing conditions in their original environment, antibiotic resistance may be considered as a consequence of a high frequency of mutational events, which also, may be enhanced by the intense solar irradiation present at the HAAL (UV index in summer: 16- 18). A special reference can be made to the representatives of the genus Acinetobacter isolated from the HAAL. Most of these strains appeared to have multiple resistance profiles to hipersalinity, UV-B irradiation, antibiotics and even arsenic. These ?superbugs? can be subjected to further studies as they can be clues to discover new ways of surviving at extreme conditions, a matter that has applications in astrobiology. On the other hand, it will be very interesting to further research on these strains biotechnological potential because as extremophiles they can be source of novel bioactive compounds.