Elsevier

Fungal Biology

Volume 120, Issue 5, May 2016, Pages 752-763
Fungal Biology

Preliminary studies on the evolution of carbon assimilation abilities within Mucorales

https://doi.org/10.1016/j.funbio.2016.02.004Get rights and content

Highlights

  • Carbon assimilation profiles of Mucorales are strongly variable.

  • The observed pattern reflects several gains and losses of particular characters.

  • None of the analyzed substrates showed dependence on the Mucorales evolutionary history.

  • Carbon assimilation capacities are shaped by environmental conditions.

Abstract

Representatives of Mucorales belong to one of the oldest lineages of terrestrial fungi. Although carbon is of fundamental importance for fungal growth and functioning, relatively little is known about enzymatic capacities of Mucorales. The evolutionary history and the variability of the capacity to metabolize different carbon sources among representatives of the order Mucorales was studied using Phenotypic Microarray Plates. The ability of 26 strains belonging to 23 nonpathogenic species of Mucorales to use 95 different carbon sources was tested. Intraspecies variability of carbon assimilation profiles was lower than interspecies variation for some selected strains. Although similarities between the phylogenetic tree and the dendrogram created from carbon source utilization data were observed, the ability of the various strains to use the analyzed substrates did not show a clear correlation with the evolutionary history of the group. Instead, carbon assimilation profiles are probably shaped by environmental conditions.

Introduction

Representatives of Mucorales belong to one of the oldest lineages of terrestrial fungi. They belong to subphylum Mucoromycotina, which is divided into three lineages: Mortierellales, Endogonales, and Mucorales (Hibbett et al. 2007). However, the order Mortierellales is currently placed within a new, separate subphylum: Mortierellomycotina (Hoffmann et al. 2011). All of these fungi probably appeared on Earth in the Precambrian, and they constituted a significant component of the Carboniferous terrestrial biota (Berbee and Taylor, 2010, Krings et al., 2013). Although commonly known as ubiquitous saprotrophs, Mucorales representatives may also be pathogens of animals, including humans. Recently, their role in the aetiology of fungal infections in humans has been considered increasingly important, particularly in warm-climate countries (Voigt et al., 2009, Muszewska et al., 2014).

For more than 150 y, Mucorales were identified using only morphological features. However, recent phylogenetic studies proved that this order is polyphyletic (Kirk et al., 2008, O'Donnell et al., 2001, Voigt and Wöstemeyer, 2001). Hoffmann et al. (2013) proposed a new taxonomic structure for the families within the order Mucorales, which better reflects the phylogenetic relationships within the group. Several traditionally-used morphological characters were revealed to be a result of convergent evolution (homoplasy). Therefore, finding new, synapomorphic diagnostic features for the clades are of particular importance. In certain groups of fungi such as yeasts, carbon assimilation profiles are now a standard feature included in species description (Yurlowa & de Hoog 1997).

Carbon is of fundamental importance for fungal growth and functioning because it is a backbone component of all organic compounds. However, it occurs mainly in complex forms that are not easily accessible for fungi and other microorganisms. The main sources of nutrients for fungi are plant-, animal-, and fungal-derived organic matter, which are accessed by fungal saprotrophs, pathogens or symbionts. However it is the plant material, composed mainly of cellulose and other polysaccharides (such as hemicelluloses and pectin) and complex aromatic compounds such as lignin, that is most commonly used. Fungi produce a wide variety of extracellular enzymes that are responsible for the decomposition of polymers into smaller compounds that can be assimilated by the cells. Fungi vary in their abilities to use different carbon compounds. In general, these differences are due to either the permeability of the cell wall or the presence of specific enzymes. It has been shown that fungi differ strongly in their carbon nutrition capacities (Madan & Thind 1998). The ability of the fungus to use different carbon sources available in the substrate can therefore be perceived as one of the main factors shaping the potential for a given fungal taxon to occupy a particular niche, in addition to abiotic factors such as temperature, humidity, and pH.

Mucorales are commonly called ‘sugar fungi’. They grow well on media rich in simple organic compounds, but they are unable to assimilate more complex organic compounds such as cellulose and lignin, in contrast to Ascomycetes and Basidiomycetes (Hesseltine & Ellis 1973). However, some recent reports suggest that Mucorales are also able to produce enzymes that allow for the degradation of more complex polymers. These include cellobiohydrolases, endoglucanases, and β-glucosidases in Mucor circinelloides (Saha 2004), and tyrosinases and lignin peroxidases in Rhizopus oryzae (León-Santiesteban et al. 2008). Recently, the ability of Umbelopsis isabellina to degrade some phenolic compounds was shown (Janicki et al. 2015). Based on these findings, the traditional opinion that Mucorales are only ‘sugar fungi’ should be revised, as their role during the decomposition of organic matter is much more important than previously thought.

Although some mucoralean fungi are commonly used in biotechnology, fermentation processes, and production of steroids or biotransformation of carotenoids (Feofilova et al., 2009, Voigt et al., 2009), relatively little is known about their enzymatic capacity. The existing knowledge is restricted to only a few well-studied taxa.

To date, the most comprehensive analyses of carbon assimilation profiles within Mucorales were made by Schwartz et al. (2007) and Vastag et al. (1998). They included a total of 62 strains (57 in the first study and five in the second study) representing 15 species from Mucorales. The main objective of both studies was to develop species-specific carbon assimilation profiles, enabling the identification of pathogenic taxa. They tested a total of approximately 80 different carbon sources and determined the carbon assimilation profiles characteristic for most of the analyzed species (including Lichtheimia corymbifera and Rhizomucor miehei). However, both studies were focused mainly on pathogenic species. The development of similar profiles for more species, especially non-pathogenic species, is needed.

The main objective of this study is to explore diversity in carbon utilization capacities and possible correlation of the metabolic pattern with phylogeny.

Section snippets

Isolates and culture techniques

This study included 26 strains of Mucorales belonging to 23 species, including 16 type strains provided by the Centraalbureau voor Schimmelcultures and the Warsaw University Herbarium. The phylogenetic position of selected strains is shown in Fig 1. The positions of samples used in earlier studies of carbon assimilation profiles in Mucorales are also indicated. Most of the selected strains were representatives of Lichtheimiaceae (containing genera: Circinella, Thamnostylum, Fennelomyces, Zychaea

Carbon assimilation profiles

The analyzed strains differed significantly in their ability to use different carbon sources. All three repetitions for each strain were relatively consistent. None of the analyzed strains was able to use all 95 carbon sources. Two strains, Thamnostylum repens and Helicostylum cordense, had the highest capacities to assimilate different carbon sources (approx. 80 % of all substrates). On average, approximately 40 substrates were absorbed per strain, i.e., less than 50 % (Fig 2).

The metabolic

Discussion

We observed a similarity between our results and those of Schwartz et al. (2007). The carbon assimilation profiles of Lichtheimia corymbifera were 89 % similar, and after removing substrates that varied at the intraspecific level in our observations the similarity was raised to 100 %. Identification of mucoralean representatives based on standard microbiological procedures is time consuming and often uncertain (Schwartz et al. 2007). Recently, molecular techniques have been used for species

Acknowledgements

The study was supported by the National Science Centre, Poland under grant No. 2015/17/D/NZ8/00778 and by the Ministry of Science and Higher Education through the Faculty of Biology, University of Warsaw intramural grant DSM 501/86-110131. The sending of many mucoralean strains from the Centraalbureau voor Schimmelcultures, Utrecht to the authors is gratefully acknowledged. Finally, we wish to thank the reviewers for critically reviewing and valuable comments on the manuscript.

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