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Due to their extreme aridity, high rate of UV irradiation and low soil carbon (C) content, the soils of the Atacama Desert represent one of the world’s most hostile environments for microbial life and its survival. Although infrequent, climatic conditions may, however, prevail which temporarily remove these stresses and allow life to briefly flourish. In this study we investigated the response of soil microbial communities to water and C availability across an aridity gradient (semi-arid, arid, hyper-arid) within the Atacama Desert. We simulated the impact of hyper-dry spells, humid fogs and precipitation events on the activation of the microbial community and the subsequent mineralization of low (glucose) and high (plant residues) molecular weight C substrates. Our results showed that mineralization rate followed the trend: semi-arid > arid > hyper-arid. Some glucose mineralization was apparent under hyper-arid conditions (water activity, a(w) = 0.05), although this was 10-fold slower than under humid conditions and ca. 200-fold slower than under wet conditions. A lag phase in CO2 production after glucose-C addition in the hyper-arid soils suggested that mineralization was limited by the low microbial biomass in these soils. No lag phase was apparent in the corresponding semi-arid or arid soils. In contrast, the breakdown of the plant residues was initially much slower than for glucose and involved a much longer lag phase in all soils, suggesting that mineralization was limited by low exoenzyme activity, particularly in the humid and hyper-dry soils. Our results also showed that microbial C use efficiency followed the trend: hyper-arid > arid > semi-arid. In conclusion, we have shown that even under hyper-arid conditions, very low levels of microbial activity and C turnover do occur. Further, the microbial communities are capable of rapidly responding to available C once water becomes more abundant, however, this response is both biomass and metabolically limited in hyper-arid soils.