In Mars-analogous hyperarid soils of the Atacama Desert, phospholipid fatty acids (PLFAs) as the main component of biomembrane play a role in reliably determining viable microbes. PLFA analyses illustrated a rise of the microbial abundance (from 5.0 × 106 to 4.2 × 107 cells g−1) and biodiversity (from 7 to 15 different individual PLFAs) from the north hyperarid core of the Atacama Desert to the southern arid region. Abundant cyclopropyl PLFAs (47.2 ± 4.6%) suggested the resistance to oligotrophic and hypersaline environments by Atacama microbial communities. The southernmost arid site had the highest proportion (8.7%) of eukaryotic and fungal lipid biomarkers. Different precipitations (ranging from 0.7 to 2 mm year−1) in the hyperarid core exerted different effects on microbial biomass, PLFA diversity, bacteria and microeukaryotes. By principal component analysis (cumulative 74.6% of variance), the dominance of PLFA hydroxylation was associated with the microbial viability; bacteria rich in C16:0, C18:0 and C16:1ω9 favoured higher soil conductivity and nitrate; and other PLFAs contributed more to the organic content. Additionally by comparing the ratios of PLFAs to well-preserved organics (e.g., mineral-bound organic carbon and microbial phosphorus), I found that the degradation of PLFAs decreased to a minimum when the mean annual precipitation is lower than 2 mm. These findings may further specify identifiable biomarkers on Mars, if potentially extant Martian microbes possess comparable phospholipid membrane structure.