Characterization of biological ice nuclei from a lichen

Abstract

Biological ice nuclei (active at approximately-40C) were extracted from cells of the lichen Rhizoplaca chrysokuca by sonication. Sensitivity to proteases, guanidine hydrochloride, and urea showed these nuclei to be proteinaceous. The nuclei were relatively heat stable, active from pH 1.5 to 12, and active without lipids, thereby demonstrating significant differences from bacterial ice nuclei.It has recently been discovered that many species of lichens (symbiotic associations of fungi and algae) are capable of nucleating ice at relatively warm temperatures, (7). This activity is similar to the ability of certain strains of several species of epiphytic bacteria (e.g., Pseudomonas syringae, Pseudomonas fluorescens, and Erwinia herbicola) to initiate freezing of supercooled water at temperatures as warm as -2C (11, 12). Field-collected lichens which have been found to have ice nucleation activity (INA) at temperatures above -5°C include the genera Rhizoptaca, Xanthoparmelia, and,Xanthoria. Of these, Rhizoplaca chrysoleuca has the warrhest INA temperature (-2.3°C). Attempts to isolate bacteria with INA from lichens have been unsuccessful, suggesting a nonbacterial source of INA. An axenic culture of the lichen fungus R. chrysoleuca showed warmtemperature INA (-2°C), further indicating a nonbacterial source of lichen INA and implicating the fungal partner as the producer of lichen ice nuclei (7). In a survey of axenic cultures of lichen fungi (mycobionts) and lichen algae (photobionts), many of the mycobionts exhibited INA at temperatures above -5°C, but none of the photobionts was an active nucleator above -5°C, providing further evidence that INA in the symbiotic association is due s'olely to the fungal partner (8).The process of isolating and characterizing biological ice nuclei is made difficult by the low numbers of nucleator molecules per cell (15,16,19); however, this difficulty is partially offset by the ease of detecting and quantifying INA. Bacterial ice nuclei from P. syringae have been shown to be proteins located in the gram-negative outer membrane (13).These proteins are thought to consist of monomers which have molecular weights of approximately 150,000 and which aggregate to form larger nucleation sites, and. the INA temperature appears to be directly related to the overall size of these aggregated protein monomers (4, 22). Similar protein monomers which mediate INA in P. fluorescens have been found to have molecular weights of.180,000 (1). Membrane-bound bacterial nuclei are active. only when associated with membrane lipids, as shown.by the fact that delipidating agents such as phospholipases or nonpolar solvents eliminate warm-temperature INA in .these organisms (3). The ice-nucleating bacterium E. herbicola releases membrane vesicles with associated INA (17). Inactivation of bacterial INA by lectins has been reported, suggesting that carbohydrate groups are also involved in ice nucleation sites (9, 10). Bacterial INA is known to be sensitive to heat (14) is inactivated by pHs below 5 (10). It has a...

Global fungal spore emissions, review and synthesis of literature data

“…In order to assess their activity as CCN or IN it is not relevant whether they are alive or not. The important feature is the shape and presence of the active proteins which actually act as nuclei (Kieft and Ruscetti, 1990). Therefore, it is not the viability of the fungal spore that affects its IN activity, but whether the ice nucleation active proteins on its surface are denatured or not.…”

“…Lichen were found to nucleate ice at temperatures higher than −8 • C and some even at temperatures higher than −5 • C (Kieft, 1988). The lichen fungus Rhizoplaca chrysoleuca was even found to be an active ice nucleus at temperatures as high as −2 • C (Kieft, 1988;Kieft and Ruscetti, 1990). To date, only a few fungus species have been found to be active ice nucleators: besides the above mentioned lichen these are Fusarium avenaceum and Fusarium acuminatum (Pouleur et al, 1992).…”

Sources of organic ice nucleating particles in soils

“…INPs produced by bacteria and fungi are, however, often unaffected by this temperature. This includes leaf-derived ice nuclei (Schnell and Vali, 1973), most strains of IN M. alpina , IN lichen (Kieft and Ruscetti, 1990), and IN Fusarium spp. (Pouleur et al, 1992), although the onset temperature of activity of cell-free INPs of F. avenaceum was lowered when heated > 40 • C (Hasegawa et al, 1994).…”

The ice nucleation ability of one of the most abundant types of fungal spores found in the atmosphere

“…Similarly, several ice nucleation experiments were carried out with species of lichens (Ashworth and Kieft, 1992;Kieft, 1988;Kieft and Ahmadjian, 1989;Kieft and Ruscetti, 1990), which are symbiotic organisms wherein fungi and algae and/or cyanobacteria form a single biological entity (Nash, 2008). In one particular study, Rhizoplaca chrysoleuca, a species of lichen, was found to induce freezing at approximately −4 • C (Kieft and Ruscetti, 1990). However, the ice nucleation properties of spores from lichens were not addressed.…”