Microclimate determines lichens and cyanobacteria distribution in the Negev, with lichens and cyanobacteria inhabit dewy and dewless habitats, respectively. Lichens experiences more frequent and extensive environmental fluctuations than cyanobacteria. The spatial partitioning of chlorolichens (eukaryotes) and cyanobacteria (prokaryotes) are intriguing, especially following recent intense search for extraterrestrial life. This is especially relevant for deserts, where both lithobionts are thought to use rain and dew but may differ in their resilience to environmental extremes and fluctuations. Following the different spatial distribution of lithobionts in a south-facing slope of the Negev Highlands (with cyanobacteria-inhabiting rocks and chlorolichen-inhabiting cobbles), measurements of temperature, non-rainfall water (NRW) and biomass were carried out within the drainage basin aiming to test the hypotheses that (i) cobble-inhabiting lichens may access more water (through NRW) and may be subjected to more extensive environmental fluctuations of temperature and water than bedrock-inhabiting cyanobacteria, and (ii) will therefore have a greater contribution to the ecosystem productivity. In contrast to cyanobacteria, cobble-inhabiting chlorolichens were found to access NRW (up to 0.20 mm of daily amounts in comparison to < 0.04 mm of the cyanobacteria) and to experience higher fluctuations of temperatures (up to 4.1 °C higher and 5.3 °C lower). With lichens and cyanobacteria inhabiting dewy and dewless habitats, respectively, NRW was found responsible for contributing 6.8-fold higher organic carbon to the lithobiontic community. At this site, chlorolichens experience more extensive environmental fluctuations than cyanobacteria, possibly indicating a higher tolerance for environmental fluctuations. These observations may assist in the interpretation of the abiotic conditions responsible for past or present lithobiontic life on Mars.