Growth of Bacillus subtilis cells, normally adapted at Earth-normal atmospheric pressure (ϳ101.3 kPa), was progressively inhibited by lowering of pressure in liquid LB medium until growth essentially ceased at 2.5 kPa. Growth inhibition was immediately reversible upon return to 101.3 kPa, albeit at a slower rate. A population of B. subtilis cells was cultivated at the near-inhibitory pressure of 5 kPa for 1,000 generations, where a stepwise increase in growth was observed, as measured by the turbidity of 24-h cultures. An isolate from the 1,000-generation population was obtained that showed an increase in fitness at 5 kPa when compared to the ancestral strain or a strain obtained from a parallel population that evolved for 1,000 generations at 101.3 kPa. The results from this preliminary study have implications for understanding the ability of terrestrial microbes to grow in low-pressure environments such as Mars.Microorganisms grow optimally within a characteristic range of fundamental physical parameters such as temperature, osmolarity, pH, and pressure. Cells that can grow at the extreme limits of these parameters are known collectively as extremophiles (reviewed in reference 8). Thus, in extreme environments on Earth there exist halophiles in hypersaline niches, psychrophiles and thermophiles in extreme cold and hot environments, and acidophiles or alkaliphiles in environments of extreme acidic or basic pH. Regarding extremes of pressure, piezophiles (barophiles) have been isolated from high-pressure (hyperbaric) submarine environments and have been studied rather extensively; in addition, high pressure has been shown to exert lethal and inhibitory effects on various microbial systems not normally adapted to high pressure (reviewed in reference 19). On the opposite extreme, there has been very little investigation concerning the survival and growth of microbes under conditions of extremely low atmospheric pressure (hypobaria), primarily because such environments do not exist in nature on the Earth's surface. However, some investigators in the emerging field of astrobiology have become concerned with microbial survival and/or growth at low pressures because (i) the closest potential life-bearing planet, Mars, contains a low-pressure atmosphere, and (ii) the possibility exists that terrestrial microbes could be transferred from Earth to Mars as a result of natural impacts (23) or human spaceflight activities (24). Most of the experiments conducted have concentrated on testing the ability of terrestrial microbes merely to survive in the Mars surface environment (reviewed in references 6, 23, and 28); relatively few experiments have tested the ability of specific Earth microbes to grow or metabolize at reduced pressure (12,14). We previously reported that growth of at least 37 different microorganisms on semisolid agar medium was inhibited at pressures approaching ϳ2.5 to 3.5 kPa (3,4,27,30). (Note that atmospheric pressures at the surfaces of Earth and Mars average ϳ101.3 kPa and ϳ0.7 kPa, respectively [26].) T...

The demand for packaging materials with low gas permeabilities is increasing, but commonly used petroleum-derived single-use plastics are not renewable, biodegradable, or easy to recycle. Nanomaterials composed of chitin and cellulose, which are abundant in nature, have high crystallinities and low oxygen permeabilities (OPs), providing a viable alternative. In this work, we explore how deacetylation conditions of crab shell chitin can be used to tune the charge and size of the resulting chitin nanowhiskers (ChNWs) and the resulting OP of layered film structures. Three deacetylation factors, the concentration of sodium hydroxide (% NaOH), temperature (T), and reaction time, were explored using a three-factor three-level Taguchi design with an orthogonal array. The resulting ChNW suspensions were sequentially spray-coated with suspensions of cellulose nanocrystals (CNCs) onto cellulose acetate (CA) films to form a multilayer structure. We show that ChNWs prepared under more aggressive deacetylation conditions inside the original orthogonal array (higher % NaOH, T, and time) had shorter lengths but the surface charge was significantly influenced only by more aggressive deacetylation outside of the original design conditions. With only ∼10% decrease in the ultimate tensile strength and no significant loss in failure strain, the ChNW–CNC coating resulted in ∼20% decrease in the water vapor transmission rate in comparison to uncoated CA films. The optimization of process conditions resulted in CA-ChNW-CNC films with a 91–99% decrease in OP (132–16.7 cm3·μm/m2/day/kPa vs 1553 cm3·μm/m2/day/kPa for uncoated CA).

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