Tolerance to pro-oxidant agents of actinomycetes isolated from Andean Wetland

BEQUER URBANO S; CORTEZ N; DI CAPUA C; FARÍAS ME; ALVAREZ HM

Tucumán

Congreso; VII Congreso Argentino de Microbiologia General (SAMIGE); 2011

Sociedad Argentina de Microbiología General (SAMIGE)

High-Altitud Andean Wetland (HAAW) is located at more than 4,400 m above sea level and is characterized by extreme conditions such as high UV radiation, elevated heavy metal content, and high salinity. Desiccation is one of the main factors occurring in these extreme environments and the oxidative stress is one of its consequences. For these reasons, bacteria need to develop efficient mechanisms and physiological strategies to survive in these extreme environments. We investigated the responses of four actinomycetes bacterial strains to the pro-oxidants hydrogen peroxide (H2O2) or methyl viologen (MV). We used in this study two isolates from HAAW; Rhodococcus sp. CH13 and Microbacterium arborescens CH5, and two model bacteria; Rhodococcus jostii RHA1 and Rhodococcus opacus PD630. Tolerance of cells to pro-oxidants was studied in agar plates. Aliquots of 10 uL (OD600 of 0.8) were inoculated onto NB agar or mineral salts media plates and supplemented with hydrogen peroxide (H2O2) or methyl viologen (MV). Lipid accumulation was investigated by thin layer chromatography after extraction with organic solvents. Superoxide dismutase (SOD) activity was analyzed after polyacrylamide gel electrophoresis. The four studied strains posses Mn-SOD type as suggested the inhibition profile of the SOD activity after treatment with H2O2 and KCN. In general, the extremophile strains CH5 and CH13 were more tolerant to oxidant agents than the model strains PD630 and RHA1. Strains CH5 and CH13 were able to grow in the presence of 0.5 mM MV at the 10-4 and 10-2 dilutions, respectively; whereas strains PD630 and RHA1 showed limited growth at the 10-1 dilution. CH13 was the most tolerant strain when cells were challenged with H2O2 (visible growth at 10-1 dilution at 0.5 mM); whereas strain PD630 was the most sensible to this challenge (no growth at 0.25 mM). Since the biosynthesis of triacylglycerols (TAG) by strains PD630 and RHA1 is a NADPH-dependent process as well as the defense mechanisms against the oxidant agents, we investigated the influence of TAG biosynthesis on the antioxidant response of these strains. Cells of both strains were more tolerant to 0.05 mM H2O2 in nitrogen-limiting mineral salts medium (MSM); which promotes TAG accumulation, than those cells cultivated on nitrogen-rich MSM. The results indicated that the extremophile actinomycetes strains CH5 and CH13 isolated from HAAW were more resistant to oxidant agents than the model strains used in this study. Moreover, our results suggested that the TAG-accumulating conditions, such as the cultivation of cells in N-limiting media, enhance the antioxidant response of cells probably due to a high NADPH pool availability.