The effects of the interaction between medium and yeast dose on NDF-kd (rate of digestion for Neutral Detergent Fibre), when pooling forages (KS: Kansas State medium; GV: Goering and Van Soest medium; MD: McDougall medium). The interaction mediumyeast for NDF-kd was significant with P-value = 0.0239. Different lowercase letters indicate significant differences (P < 0.05).
The South African Proteaceae comprises 14 genera, of which 7 genera are commercially utilised (Vogts 1982, Littlejohn 1999, Paterson-Jones 2000, Rebelo 2001). Three of these, Leucadendron, Leucospermum and Protea are of the greatest commercial value and are grown for their exotic, brightly coloured and textured flowers or bracts, which are in high demand on the world floriculture market (Coetzee & Littlejohn 2001). Fungal pathogens, however, create a serious problem in cultivating flawless blooms. Several groups of fungal pathogens of Proteaceae have in recent years been characterised phylogenetically, e.g. Botryosphaeria stem cankers (Denman et al. 1999, 2000, 2003, Crous et al. 2006b, Marincowitz et al. 2008b), Armillaria and Cylindrocladium root rot (Schoch et al. 1999, Coetzee et al. 2003, Lombard et al. 2010a, b, c), Elsinoë scab disease (Swart et al. 2001) Phomopsis cankers (Mostert et al. 2001a, b), and leaf spots caused by species of Mycosphaerella and Teratosphaeria (Crous et al. 2008, 2009a, b, 2011b). Several other pathogenic fungi on Proteaceae however, have never been studied from a phylogenetic perspective based on DNA analyses, and in light of new knowledge are now suspected to represent species complexes.
Very little is known, however, about the fungi occurring in the CFR, and recent studies have highlighted the unusual diversity that occur in saprobic (Schubert et al. 2007, Visagie et al. 2009, Bensch et al. 2010), and plant pathogenic fungi (Crous et al. 2009a, 2011b). Mycological exploration in South Africa commenced in the late 1700s, and thus later than plant collections, which began in the 1600s (Eicker & Baxter 1999, Rong & Baxter 2006). For more than a century, both collections and studies on fungi were largely conducted by European visiting scientists. It was only in the late 1800s that the first resident mycologists were appointed in South Africa. A third generation mycologist, Dr Ethel M. Doidge, compiled the list of fungi and lichens collected in South Africa to the end of 1945. In this compilation, about 30 fungi are listed associated with members of Proteaceae (Doidge 1950), while based on the known plant biodiversity of South Africa, Crous et al. (2006a) estimated the potential fungal biodiversity as approximately 200 000 species, many of which would occur on Proteaceae.
The first fungus described from Proteaceae in the Western Cape was Pseudocercospora protearum (as Cercospora protearum) (Cooke 1883). After many decades of work, the plant pathogenic fungi associated with the family, especially on Protea, Leucospermum and Leucadendron was published (Crous et al. 2004a), as well as the common saprobic fungi occurring on twig and leaf litter (Marincowitz et al. 2008a).
Fungi are well-known agents of decomposition of organic matter in general and of cellulosic substrates in particular (94, 462). Fungal taxonomy is based largely on the morphology of mycelia and reproductive structures during various stages of the fungal life cycle rather than on substrate utilization capability. Indeed, systematic characterization of growth substrates has not been carried out for many described fungal species. Therefore, it is currently unclear how broadly and deeply cellulolytic capability extends through the fungal world, and a consideration of the taxonomy of cellulolytic fungi may ultimately prove to be only a slightly narrower topic than consideration of fungal taxonomy in its entirety. Nevertheless, some generalizations can be made regarding the distribution of cellulolytic capabilities among these organisms.
A number of species of the most primitive group of fungi, the anaerobic Chytridomycetes, are well known for their ability to degrade cellulose in gastrointestinal tracts of ruminant animals. Although taxonomy of this group remains controversial (94), members of the order Neocallimastigales have been classified based on the morphology of their motile zoospores and vegetative thalli; they include the monocentric genera Neocallimastix, Piromyces, and Caecomyces and the polycentric genera Orpimomyces and Anaeromyces (376). Cellulolytic capability is also well represented among the remaining subdivisions of aerobic fungi. Within the approximately 700 species of Zygomycetes, only certain members of the genus Mucor have been shown to possess significant cellulolytic activity, although members of this genus are better known for their ability to utilize soluble substrates. By contrast, the much more diverse subdivisions Ascomycetes, Basidiomycetes, and Deuteromycetes (each of which number over 15,000 species ), contain large numbers of cellulolytic species. Members of genera that have received considerable study with respect to their cellulolytic enzymes and/or wood-degrading capability include Bulgaria, Chaetomium, and Helotium (Ascomycetes); Coriolus, Phanerochaete, Poria, Schizophyllum and Serpula (Basidiomycetes); and Aspergillus, Cladosporium, Fusarium, Geotrichum, Myrothecium, Paecilomyces, Penicillium, and Trichoderma (Deuteromycetes). For a more detailed consideration of fungal taxonomy and some of its unresolved issues, see reference 94.
Aerobic cellulose degraders, both bacterial and fungal, utilize cellulose through the production of substantial amounts of extracellular cellulase enzymes that are freely recoverable from culture supernatants (554, 606), although enzymes are occasionally present in complexes at the cell surface (67, 715). The individual enzymes often display strong synergy in the hydrolysis of cellulose. While many aerobic bacteria adhere to cellulose, physical contact between cells and cellulose does not appear to be necessary for cellulose hydrolysis. Kauri and Kushner (322) have shown that separating Cytophaga cells from cellulose via an agar layer or membrane filters appears to enhance cellulose utilization; they suggest that this separation may dilute hydrolytic products, thus relieving catabolite repression of enzyme synthesis. Aerobic cellulolytic bacteria and fungi produce high cell yields characteristic of aerobic respiratory growth, and this has led to considerable technological interest in producing microbial cell protein from waste cellulosic biomass (175, 567, 594, 623). In addition, many studies of aerobic cellulolytic microbes have focused on improving the yield and characteristics of cellulase enzymes. The physiology of the organisms themselves has received surprisingly little study, apart from studies on the effect of growth conditions on enzyme secretion (see, e.g., reference 236).
The production of at least two β-glucosidases by T. reesei facilitates the hydrolysis of cellobiose and small oligosaccharides to glucose. Both BGLI and BGLII have been isolated from culture supernatants, but a large fraction of these enzymes remains cell wall bound (442, 690). The presence of β-glucosidases in close proximity to the fungal cell wall may limit loss of glucose to the environment following cellulose hydrolysis. T. reesei produces β-glucosidases at low levels compared to other fungi such as Aspergillus species (560). Furthermore, the β-glucosidases of T. reesei are subject to product (glucose) inhibition (102, 217, 417) whereas those of Aspergillus species are more glucose tolerant (138, 231, 724, 768). The levels of T. reesei β-glucosidase are presumably sufficient for growth on cellulose, but not sufficient for extensive in vitro saccharification of cellulose. T. reesei cellulase preparations, supplemented with Aspergillus β-glucosidase, are considered most often for cellulose saccharification on an industrial scale (560, 644).
The best-studied species of cellulolytic aerobic bacteria belong to the genera Cellulomonas and Thermobifida (formerly Thermomonospora). Cellulomonas species are coryneform bacteria that produce at least six endoglucanases and at least one exoglucanase (Cex) (99). The individual cellulases of Cellulomonas resemble the cellulase systems of aerobic fungi and contain CBMs; however, cellulosome-like protuberant structures have been noted on Cellulomonas cells grown with cellulose and cellobiose as carbon sources (370, 714). The thermophilic filamentous bacterium Thermobifida fusca (formerly Thermomonospora fusca) is a major cellulose degrader in soil. Six cellulases, three endoglucanases (E1, E2, and E5), two exoglucanases (E3 and E6), and an unusual cellulase with both endoglucanase and exoglucanase activity (E4) have been isolated. The latter enzyme has high activity on BMCC and also exhibits synergism with both the other T. fusca endoglucanases and exoglucanases (304). The E4 enzyme also contains a family III CBM that assists the enzyme in processivity (303). Factorial optimization of the T. fusca cellulase system was undertaken, and the highest synergistic effect was shown with the addition of CBHI from T. reesei (335).
Anaerobic chytrid fungi are only found in the rumens of herbivorous animals (509) and produce highly active cellulases (68, 103, 745, 759). High-molecular-weight complexes with high affinity for microcrystalline cellulose have been isolated from Piromyces sp. strain E2. Conserved noncatalytic repeat peptide domains have been identified in cellulases and xylanases from Neocallimastix and Piromyces species and are thought to provide a docking function (180, 385). Recently, Steenbakkers et al. (639) used PCR primers based on DNA sequences that encode these 40-amino-acid cysteine-rich docking domains to recover the genes of several cellulosome-like components. Preliminary data indicate the presence of multiple scaffoldins; however they have not yet been isolated from culture fractions (639). Evidence is thus mounting that anaerobic fungi also utilize cellulosomes for hydrolysis of crystalline cellulose. Evolutionary convergence might have occurred between the anaerobic fungi and clostridia. However, the 40-amino-acid dockerin sequence of the anaerobic fungi differs significantly from those of the clostridia, suggesting independent development of the cellulosomes of anaerobic fungi. 153554b96e