Water availability acts as the most stringent constraint for life on Earth. Thus, understanding the water relations of microbial extremophiles is imperative to our ability to increase agricultural productivity (e.g., by enhancing the processing and turnover of dead organic matter in soils of arid regions), reduce human exposure to mycotoxins in buildings and our food-supply chain, prevent the spoilage of foods/animal feeds, books, museum specimens and artworks and better control microbiology of industrial fermentations. Only a small number of microbial systems can retain activity at <0.710 water activity (ISME J 2015 9: 1333–1351). It has long-been considered that the most resilient of these is Xeromyces bisporus, which inhabits sugar-rich substrates (Appl Environ Microbiol 1968 16: 1853–1858). The current study focused on germination of Aspergillus penicillioides, a xerophile which is also able to grow under low humidity and saline conditions. Investigations of germination differed from those reported earlier: firstly, aerially borne conidia were harvested, and then used for inoculations, in their dry condition; secondly, cultures were incubated at 24°C, i.e. below optimum germination temperature, to minimize the possibility of water loss from the substrate; thirdly, cultures remained sealed throughout the 73-day study period (microscopic examination was carried out directly 48 through the Petri plate lid); fourthly, the germination parameters determined were: rates and extent of conidial swelling, production of differentiated germination-structures and septate germlings, and subsequent development of mycelium and/or sporulation; fifthly, assessments were carried out over a range of water-activity values and time points to obtain a complete profile of the germination process. Conidia swelled, formed differentiated germination-structures and then produced septate germlings at a water-activity of just 0.585 (≡58.5% relative humidity), outside the currently understood thermodynamic window for life. Furthermore, analyses of these data suggest a theoretical water-activity minimum of 0.565 for germination of A. penicilliodes. In relation to astrobiology, these findings have an application in understanding the limits to life in extraterrestrial environments. In light of current plans for exploration missions to Mars and other places, and the need to safeguard martian scientific sites and potential resources (including water) for future human habitation, a knowledge-based and effective policy for planetary protection is essential. As it is, Mars-bound spacecraft may frequently be contaminated with aspergilli (including A. penicillioides) and other organisms which, when transported to other planetary bodies, pose a contamination risk. In crafting countermeasures to offset this, it is important to know as precisely as possible the capabilities of these potential interplanetary visitors.