Characterisation of yeast and filamentous fungi from Brøggerbreen glaciers, Svalbard

ABSTRACTCryoconite holes have ecological and biotechnological importance. This article presents results on culturable cryophilic yeasts and filamentous fungi isolated from cryoconite holes at Austre and Vestre Brøggerbreen glaciers, Svalbard. Based on DNA sequence data, these were identified asRhodotorulasp.,Thelebolussp., andArticulospora tetracladia. Amongst these,Articulospora tetracladia(88.7–89.4% gene similarity with 5.8S rDNA) is a novel species, yet to be described. Filamentous fungusArticulosporasp. Cry-FB1 and Cry-FB2, expressed high amylase, cellulase, lipase and protease activities while yeastRhodotorulasp. Cry-FB3 showed high amylase and cellulase activity.Thelebolussp. Cry-YB 240 and Cry-YB 241 showed protease and urease activities. The effects of temperature, and salt on the growth of the cultures were studied. Optimum temperature of growth was on 10ºC at pH 7.0. Filamentous fungi and yeast in the cryoconite holes possibly drive the process of organic macromolecule degradation through cold-adapted enzyme secretion, thereby assisting in nutrient cycling in these supraglacial environments. Further, these cryophilic fungi, due to their enzyme producing ability, may provide an opportunity for biotechnological research in the Arctic.

A conceptual model describing macromolecule degradation by suspended cultures and biofilms

Macromolecular (> 1,000 daltons) compounds such as proteins and polysaccharides can constitute a significant portion of dissolved organic carbon (DOC) in wastewater, but limited information is available on how these compounds are degraded in suspended and fixed-film biological wastewater treatment systems. Bacteria cannot assimilate intact macromolecules but must first hydrolyze them to monomers or small oligomers. Here, we summarize experiments performed in our laboratory which indicate that the enzymes responsible for hydrolysis are primarily those that remain attached to the cell. In biofilm cultures fed macromolecular substrates, for example, no more than 8% of total hydrolytic activity was found to be located in the cell-free bulk solution. These and other experiments support a generalized mechanism for macromolecule degradation by biofilms that features cell-associated hydrolysis, followed by the release of hydrolytic fragments back into bulk solution. The extent of fragment release is larger for proteins (bovine serum albumin) than for carbohydrates (dextrans).

Lysosomal degradation of glycoproteins and glycosaminoglycans. Efflux and recycling of sulphate and N-acetylhexosamines

Lysosomal degradation of the carbohydrate portion of glycoproteins and glycosaminoglycans produces monosaccharides and sulphate, which must efflux from the lysosomes before re-entering biosynthetic pathways. We examined the degradation of glycoproteins and glycosaminoglycans by lysosomes isolated from cultured human diploid fibroblasts. Cells were grown for 24 h in medium containing [3H]glucosamine and [35S]sulphate. When lysosomes are isolated from these cells, they contain label primarily in macromolecules (glycoproteins and glycosaminoglycans). Glycoprotein degradation by isolated lysosomes was followed by measuring the release of tritiated sugars from macromolecules and efflux of these sugars from the organelles. Glycosaminoglycan degradation was monitored by the release of both tritiated sugars and [35S]sulphate. During macromolecule degradation, the total amounts of free [35S]sulphate, N-acetyl[3H]glucosamine and N-acetyl[3H]galactosamine found outside the lysosome parallels the amounts of these products released by degradation. The total degradation of glycoproteins and glycosaminoglycans by intact cultured cells was also examined. The lysosomal contribution to degradation was assessed by measuring inhibition by the lysosomotropic amine NH4Cl. After 48 h incubation, inhibition by NH4Cl exceeded 55% of glycoprotein and 72% of glycosaminoglycan degradation. Recycling of [3H]hexosamines and [35S]sulphate by intact cells was estimated by measuring the appearance of ‘newly synthesized’ radioactively labelled macromolecules in the medium. Sulphate does not appear to be appreciably recycled. N-Acetylglucosamine and N-acetylgalactosamine, on the other hand, are reutilized to a significant extent.