Pichia anomala: cell physiology and biotechnology relative to other yeasts
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- Walker, G.M. Antonie van Leeuwenhoek (2011) 99: 25. doi:10.1007/s10482-010-9491-8
Pichia anomala is a most interesting yeast species, from a number of environmental, industrial and medical aspects. This yeast has been isolated from very diverse natural habitats (e.g. in foods, insects, wastewaters etc.) and it also exhibits wide metabolic and physiological diversity. Some of the activities of P. anomala, particularly its antimicrobial action, make it a very attractive organism for biological control applications in the agri-food sectors of industry. Being a ‘robust’ organism, it additionally has potential to be exploited in bioremediation of environmental pollutants. This paper provides an overview of cell physiological characteristics (growth, metabolism, stress responses) and biotechnological potential (e.g. as a novel biocontrol agent) of P. anomala and compares such properties with other yeast species, notably Saccharomyces cerevisiae, which remains the most exploited industrial microorganism. We await further basic knowledge of P. anomala cell physiology and genetics prior to its fuller commercial exploitation, but the exciting biotechnological potential of this yeast is highlighted in this paper.
Pichia spp. represent very interesting yeasts from both fundamental and applied perspectives. For example, from a cell biology viewpoint, they have proved most valuable in studies of organelle biogenesis, structure and function; and from an applied viewpoint, they have widespread biotechnological significance ranging from human therapeutic protein production, food fermentations, biocontrol agents and biofuel production. One particular species, Pichia anomala,1 exhibits great diversity with regard to its natural habitat, growth morphology, metabolism, stress-tolerance, and antimicrobial properties. It has been isolated from the following sources: flowering plants, fruit skins, insect intestinal tracts, human tissue and faeces, dairy and baked food products, contaminated oil, wastewaters, tree exudates, salted foods, and from the marine environment. This represents a wider range of habitats in Nature compared with the well-known Saccharomyces cerevisiae (brewer’s or baker’s yeast). P. anomala also differs from S. cerevisiae with regard to its mode of central carbon metabolism, in that it exhibits an insensitivity to glucose (i.e. is Crabtree negative). An interesting shared characteristic of both species relates to their killer yeast activity. However, do both yeasts kill other yeasts (or fungi) by the same mechanisms? The antifungal activity of P. anomala appears to be linked to cell wall hydrolysis (β glucanase-induced lysis) and/or to production of volatile metabolites (e.g. ethyl acetate), whereas S. cerevisae produces a killer toxin (e.g. K1 toxin peptide) that disrupts plasma membrane integrity. The antimycotic properties of P. anomala have led to this yeast being considered a valuable biocontrol agent against fungi of agronomical importance. Other potential biotechnological applications of P. anomala include environmental bioremediation, biopharmaceuticals and biofuels.
This paper reviews some of the unusual characteristics of P. anomala and will highlight these with reference to its potential biotechnological exploitation. Cell physiology of P. anomala will also be compared with better known yeast species, notably S. cerevisiae.
P. anomala in foods and beverages
Important roles of Picha anomala in foods, feeds and beverages
Examples of beneficial roles
Volatile (e.g. esters) and savoury (e.g. nucleotides) flavours
Biological control of fungi in fruits and cereals
Mo et al. (2004)
Sourdough fermentations (not necessarily beneficial)
Daniel et al. (2010)
Volatile aromas, low alcohol wines, malic acid reduction
Enzymatic food/feed processing
Phytase, amylase, peptidase
Anti-gushing potential in malting barley
There are some detrimental roles of P. anomala in relation to food production and storage (e.g. Deak 2008). As a food spoilage yeast, its contamination of yoghurts, bread, sugary cakes (Lanciotti et al. 1998), and wine (e.g. Rojas et al. 2001) can lead to taints commonly referred to as ‘chemical adulteration’. This may be due to the propensity of P. anomala to produce ethyl acetate (see below). In stored animal feeds such as silage, it can consume lactic acid (Jonsson and Pahlow 1984) and this may elevate pH thus reducing periods of safe feed storage.
Although there are some reports of nosocomial infections caused by P. anomala (e.g. Charkrabarti et al. 2001), with regard to food safety aspects P. anomala is classed at biosafety level 1 (De Hoog, 1996) and is considered safe for healthy individuals. In fact, P. anomala currently has QPS (qualified presumption of safety) status from EFSA (European Food Safety Authority) and this has benefits in terms of public perspectives of food biotechnology and acceptability of novel microorganisms in food (Sundh and Melin 2010).
P. anomala in the environment
Some diverse Picha anomala habitats
Homo sapiens (human skin, faeces etc.)
Nagatsuka et al. (2005)
Insects (e.g. Drosophila and malaria mosquito Anopheles)
Kajikazawa et al. (2007)
Recek et al. (2002)
Cereal silos and silage
El-Latif et al. (2006)
Wang et al. (2007)
P. anomala plays certain beneficial roles in the environment. For example, it exhibits (as do many other yeasts) saprophytic roles in the carbon cycle; it can help to alleviate pollution by bioremediation of recalcitrant chemicals/heavy metals in wastewaters; and it can act in the biological control of harmful microbes by combating biodeteriogenic fungi. Walker et al. (1995) showed that P. anomala was able to inhibit certain wood decay basidiomycetous fungi and it also displayed fungistatic acitivity against plant pathogenic fungi, including the causative agent of Dutch Elm disease, Ophiostoma novo ulmi. El-Latif Hesham et al. (2006) have shown that P. anomala can effectively degrade toxic chemicals such as the aromatic hydrocarbons naphthalene and benzopyrene, thus highlighting its potential role in environmental bioremediation processes (e.g. oil-contaminated industrial, terrestrial and marine environments).
P. anomala in industry
P. anomala products of biotechnological potential
Thaniyavarn et al. (2008)
γ-aminobutyric acid, GABA
Pharmaceuticals (GABA acts as a neurotransmitter, improves cerebral blood flow)
Kaku and Hagiwara (2008)
Volatile organic compounds
Buzzini et al. (2003)
US Patent (2009)
Beverage starter culture
Low-alcohol wines; aromas
Ertin and Campbell (2001)
Novel zymocidial agents
Izgü et al.(2006)
Influenza virus therapy
Conti et al. (2008)
Therapy of Acanthamoeba infections
Fiori et al. (2006)
Therapy of Pneumocystis carnii
Seguy et al. (1996)
Therapy of Streptococcal infections
Conti et al. (2002)
Stored grain, vines, fruit
Phytase, esterase, peptidase, β-glucosidase, amylase
Ray and Nanda (1996); Satyanarayana, T (2010)
Maintenance of airtight stored grain (biofuels)
Passoth et al. (2009)
Antimicrobial activity of P. anomala
Summary of antimicrobial properties of P. anomala
Examples of microbes suppressed
Aspergillus, Botrytis, Penicillium, Fusarium
Various yeasts, incl. C. albicans
Sawant et al. (1988)
Erwinia spp.; Enterobacteriaceae; Streptococci
Conti et al. (2008)
P. anomala is a killer yeast and a variety of killer toxins are known in P. anomala strains. With regard to the genetic basis of the killer phenomenon in P. anomala, the killer factor proteins are thought to be chromosomally inherited, unlike S. cerevisiae killer toxins (such as K1) which are encoded on double-stranded RNA virus-like extra-chromosomal elements, or Kluyveromyces lactis toxins which are encoded on linear DNA plasmids. P. anomala killer toxins also differ from those of other killer yeasts in that they exhibit diversity in terms of broad spectrum antimicrobial activity, variable molecular mass (e.g. from 3 to 300 kDa), and different pH and temperature optima (Passoth et al. 2006). Recently, De Ingeniis et al. (2009) have shown that a P. anomala killer toxin (peptide of ~8 kDa) possesses novel ubiquitin-like characteristics.
Antimycotic activity of P. anomala: candidate antifungal agents
Antifungal agents or modes of antifungal action
Likely relative contribution (ranging from ***** predominant to * lesser importance)
Hydrolytic enzymes (e.g. β-glucanase)
Volatile chemicals (e.g. ethyl acetate)
Other antifungal agents
In addition to its action against biodeteriogenic fungi in the agri-food areas, P. anomala also has potential applications in medical mycology. For example, P. anomala has long been recognised as possessing anti-Candida albicans activity (e.g. Hodgson et al. 1995; Polonelli et al. 1983; Sawant et al. (1988); Buzzini and Martini, 2001). Polonelli et al. (1990) were the first to show that P. anomala killer toxin was active in vivo in experimental mice. More recently, Izgü et al. (2006) have shown that the K5 killer toxin of P. anomala displays activity against selected dermatophytes (Microsporum spp. and Trichophyton spp). The K5 killer protein (named ‘panomycin’) was previously shown by Izgü and Altinbay (2004) to exhibit exo-β-1,3-glucanase activity. Magliani et al. (1997) and Polonelli et al. (2010) have discussed medical applications of P. anomala killer toxins, in particular the immunomodulatory activities of ‘antibiobodies’.
Stress tolerance of P. anomala
Cell physiological aspects of P. anomala
The morphology of P. anomala exhibits diversity in terms of various cellular shapes with budding cells and branched pseudohyphae being evident in both liquid and solid culture media (Kurtzman 1998). As with other yeasts, it is possible that Quorum-sensing mechanisms may be involved in governing morphological changes and cell density related growth inhibition in this yeast (Walker 1998). Sexual reproduction in P. anomala is characterised by formation of hat-shaped spores (Kurtzman 1998).
Carbon and nitrogen growth source diversity in P. anomala
Saccharides: hexoses (glucose, galactose, fructose); pentoses (arabinose, xylose); disaccharides (sucrose, lactose), polysaccharides (starch; β-glucans)
Alcohols: ethanol, glycerol
Organic acids: acetate, citrate, lactate, malate, succinate
Fatty acids: oleate, palmitate
Aromatics: naphthalene, benzopyrene
The practical manifestation of these metabolic differences means that P. anomala can grow aerobically with high sugar concentrations at a relatively high growth rate and to higher cell densities than S. cerevisiae. The lack of a Crabtree Effect in P. anomala means that (unlike S. cerevisiae), there is no real necessity to keep sugar levels low and consequently no need to conduct fed-batch yeast propagation systems to control sugar feeding rates when attempting to maximise yeast biomass production.
Cell physiological and other characteristic differences between P. anomala and S. cerevisiae
Glucose uptake by H+ symport
Facilitated glucose diffusion
Malic acid utilisation
Malate only utilised with glucose
Several enzymes secreted
Few enzymes secreted
Rarely antifungal (some strains)
High ethyl acetate
Low ethyl acetate
Widespread in Nature
Not widespread in Nature
Not very halotolerant
Moderate ethanol tolerance
Opportunistic pathogen (some strains)
Doubtful opportunistic pathogenicity
Conclusions and future perspectives
Pichia anomala exhibits interesting and potentially exploitable physiological and metabolic characteristics. These include: morphological diversity (budding, pseudomycelial); stress tolerance (to low pH, high osmotic pressure, low O2, low aw); enzyme secretion (invertase, lipase, peptidase, amylase, phytase); nutritional diversity (range of C, N, and P sources); biodegradation (of polyaromatic hydrocarbons, naphthalene, benzopyrene,); Crabtree negativity (glucose insensitivity); antimicrobial activity (yeasts, fungi, bacteria, viruses); and production of potential commercial metabolites.
Although S. cerevisiae remains the world’s most exploited organism in industrial bioprocesses, other non-Saccharomyces yeasts like Pichia spp. have fantastic potential in biotechnology. Nevertheless, we still have much to learn about physiology and metabolism in non-Saccharomyces yeasts, including P. anomala, and enhancement of cell physiological knowledge in this yeast is a prerequisite for its fuller industrial exploitation. There are still several unresolved questions regarding carbon metabolism and its regulation in P. anomala. For example, it is conceivable that there is variability of the expression of the Crabtree effect in P. anomala strains, as previously demonstrated in Kluyveromyces lactis by Liti et al. (2001), and the underlying mechanisms of such metabolic phenomena and their practical significance require further research. Other areas of P. anomala research worthy of future investigation include: determination of modes of antimicrobial/antiviral action; and molecular understanding of the underlying reasons for stress tolerance. Stress tolerance and antimicrobial action are especially important P. anomala characteristics that can be exploited for future biotechnological applications.
Throughout this paper the species will be referred to as Pichia anomala, rather than Wickerhamomyces anomalus (see Kurtzman et al. 2008 and Kurtzman, 2010), mainly because it was the nomenclature used in the symposium from which this manuscript emanated (1st International Pichia anomala mini-Symposium).