Quadruple sulfur and triple oxygen isotope constraints on biological processes in Mars-analog gypsum

A primary objective of Mars sample return missions is the detection and characterization of potential biosignatures. Sulfates, among the most abundant hydrated minerals on Mars, represent promising targets for such investigation. Terrestrial studies have shown that gypsum can preserve life-associated organic matter and that its δ 34 S signatures provide key evidence for microbial sulfate reduction as early as ∼ 3.5 billion years ago. This study analyzes quadruple sulfur (δ 34 S, Δ 33 S, Δ 36 S) and triple oxygen (δ 18 O, Δ́ 17 O) isotopes in gypsum from three strong evaporation basins on the Qinghai-Tibet Plateau, where sulfate deposition is extensive but surface environments are only marginally habitable due to severe limitations in water, soil development, and nutrient availability. We focus on the Qaidam Basin as a Mars analog environment and place it in context through comparison with a higher-elevation region (Shuanghu) and a magmatic rock-dominated region (Yushu). Together with isotopic mixing models, an open-system steady-state sulfur cycle model, and triple-oxygen isotope oxidation pathway calculations, our quadruple sulfur and triple oxygen isotope measurements reveal that the studied gypsum deposits preserve distinct isotopic biosignatures. In particular, the isotopic compositions of the samples reflect microbial sulfur disproportionation and cryptic sulfur cycling, both of which extend beyond widespread microbial sulfate reduction, demonstrating how sulfur can be internally recycled and energetically utilized to sustain life in Mars-analog environments. This integrated isotopic approach significantly advances our capability to decipher sulfur cycling in extreme terrestrial environments, including Mars analog systems, and provides critical methodology for interpreting potential biosignatures in returned Martian samples.