Paul D. Bridge

Fungal biotechnology in agricultural, food, and environmental applications

Agricultural biotechnology: a biotechnological approach to plant protection with fungi chemical identification of fungi - metabolite profiling and metabolomics isozyme analysis in fungal taxonomy, genetics and population biology molecular methods for identification of plant pathogenic fungi the application of molecular markers in the epidemiology of plant pathogenic fungi molecular biology in the control of mycotoxigenic fungi biotechnological potential of entomopathogenic fungi biotechnological potential of ergot alkaloids fungi as plant growth promoter and disease suppressor challenges and strategies for development of mycoherbicides biofungicides from microbial metabolites molecular biology of biocontrol trichoderma the biological control agent trichoderma - fundamentals to applications biotechnological strategies to control fungal diseases of vegetables control of postharvest fruits using microbes arbuscular mycorrhizal fungi in plant disease control commercialization of arbuscular mycorrhizal biofertilizer control of nematodes by fungi. Food and feeds: fungi in food technology - an overview role of fungi in fermented foods production of edible fungi mycoprotein and related high-value products from yeasts and fungi diversity and genetic variability of yeasts involved in wine fermentation yeast in the dairy industy flavours and aroma antifungal food additives molecular detection of fungi in foods and feeds the role of spoilage fungi in seed deterioration mycotoxin chapter genetics and biochemistry of toxin synthesis. Environmental biotechnology: cellulose degradation by fungi the importance of wood decay fungi in forest ecosystems the biodegradation of lignocellulose by white rot fungi biomineralization of heavy metals decolouration of industrial waste and degradation of dye water bioconversion of distillery waste by fungi degradation of hydrocarbons by yeasts and filamentous fungi biodegradation of azo dyes by fungi fungal degradation of explosives restoration of mycorrhizae in disturbed arid ecosystems.

Microorganisms in the atmosphere over Antarctica

Antarctic microbial biodiversity is the result of a balance between evolution, extinction and colonization, and so it is not possible to gain a full understanding of the microbial biodiversity of a location, its biogeography, stability or evolutionary relationships without some understanding of the input of new biodiversity from the aerial environment. In addition, it is important to know whether the microorganisms already present are transient or resident - this is particularly true for the Antarctic environment, as selective pressures for survival in the air are similar to those that make microorganisms suitable for Antarctic colonization. The source of potential airborne colonists is widespread, as they may originate from plant surfaces, animals, water surfaces or soils and even from bacteria replicating within the clouds. On a global scale, transport of air masses from the well-mixed boundary layer to high-altitude sites has frequently been observed, particularly in the warm season, and these air masses contain microorganisms. Indeed, it has become evident that much of the microbial life within remote environments is transported by air currents. In this review, we examine the behaviour of microorganisms in the Antarctic aerial environment and the extent to which these microorganisms might influence Antarctic microbial biodiversity.

Candida nivariensis, an Emerging Pathogenic Fungus with Multidrug Resistance to Antifungal Agents

ABSTRACT In 2005, Candida nivariensis, a yeast species genetically related to Candida glabrata, was described following its isolation from three patients in a single Spanish hospital. Between 2005 and 2006, 16 fungal isolates with phenotypic similarities to C. nivariensis were submitted to the United Kingdom Mycology Reference Laboratory for identification. The strains originated from various clinical specimens, including deep, usually sterile sites, from patients at 12 different hospitals in the United Kingdom. PCR amplification and sequencing of the D1D2 and internal transcribed spacer 1 (ITS1) regions of the nuclear ribosomal gene cassette confirmed that these isolates from the United Kingdom are genetically identical to C. nivariensis. Biochemically, C. glabrata and C. nivariensis are distinguished by their differential abilities to assimilate trehalose. However, in contrast to the original published findings, we found that C. glabrata isolates, but not C. nivariensis isolates, are capable of assimilating this substrate. Antifungal susceptibility tests revealed that C. nivariensis isolates are less susceptible than C. glabrata isolates to itraconazole, fluconazole, and voriconazole and to have significantly higher flucytosine MICs than C. glabrata strains. Finally, C. nivariensis could be rapidly distinguished from the other common pathogenic fungus species by pyrosequencing of the ITS2 region. In the light of these data, we believe that C. nivariensis should be regarded as a clinically important emerging pathogenic fungus.

Molecular Identification of Unusual Pathogenic Yeast Isolates by Large Ribosomal Subunit Gene Sequencing: 2 Years of Experience at the United Kingdom Mycology Reference Laboratory

ABSTRACT Rapid identification of yeast isolates from clinical samples is particularly important given their innately variable antifungal susceptibility profiles. We present here an analysis of the utility of PCR amplification and sequence analysis of the hypervariable D1/D2 region of the 26S rRNA gene for the identification of yeast species submitted to the United Kingdom Mycology Reference Laboratory over a 2-year period. A total of 3,033 clinical isolates were received from 2004 to 2006 encompassing 50 different yeast species. While more than 90% of the isolates, corresponding to the most common Candida species, could be identified by using the AUXACOLOR2 yeast identification kit, 153 isolates (5%), comprised of 47 species, could not be identified by using this system and were subjected to molecular identification via 26S rRNA gene sequencing. These isolates included some common species that exhibited atypical biochemical and phenotypic profiles and also many rarer yeast species that are infrequently encountered in the clinical setting. All 47 species requiring molecular identification were unambiguously identified on the basis of D1/D2 sequences, and the molecular identities correlated well with the observed biochemical profiles of the various organisms. Together, our data underscore the utility of molecular techniques as a reference adjunct to conventional methods of yeast identification. Further, we show that PCR amplification and sequencing of the D1/D2 region reliably identifies more than 45 species of clinically significant yeasts and can also potentially identify new pathogenic yeast species.

The impact of molecular data in fungal systematics

Molecular data has been used in fungal systematics since the 1970s, and its rate of incorporation has increased significantly in recent years. In phylogeny molecular data has already been used to clarify major evolutionary lines, and has aided in the delineation of higher taxonomic groups including the kingdom Fungi, and the main phyla within it. Molecular data has been used at all taxonomic levels and has allowed for a greater phylogenetic signal to be represented within systematic groups. At the higher levels this has led to the re-evaluation of some orders and families, and at lower taxonomic levels it has helped in the identification of species, particular populations and possibly individuals. There are however some limitations to the widespread use of molecular data. Some of these relate to the comparability and utility of methods between different fungal groups, some relate to the wide diversity of life cycles adopted by fungi, and others are due to the paucity of comparable definitive evolutionary markers. A significant limitation to the wider application of molecular data is the restricted range of data currently available, and the relation of this to the as yet unquantified numbers of undescribed species. Despite these limitations molecular data has had a very significant effect on our understanding of fungal systematics, and many further systematic aspects are likely to be elucidated in the future.

VNTR derived oligonucleotides as PCR primers for population studies in filamentous fungi

One M13 phage derived and three simple sequence repeat primers were assessed for the development of a standard set of PCR primers for population studies with filamentous fungi. Fungal isolates from five plant or insect pathogenic genera with ascomycete and basidiomycete affinities were screened. Three of the four primers were found to be suitable for generating ‘genetic fingerprints’ for all of the fungal genera tested. Strain groupings obtained from the individual primers were found to be supportive through constrained two‐dimensional cluster analysis.

Minimal influence of water and nutrient content on the bacterial community composition of a maritime antarctic soil.

Bacterial community composition was determined by culture-independent PCR-based methods in two soils differing markedly in their water, C, N and P contents sampled from Mars Oasis on Alexander Island, western Antarctic Peninsula. 16S rRNA sequences of the phyla Actinobacteria, Cyanobacteria, α-Proteobacteria and Acidobacteria were commonly (> 8% frequency) obtained from soil. Those of β-, γ- and δ-Proteobacteria, Chloroflexi, Bacteroidetes, Verrucomicrobia, Planctomycetes, Gemmatimonadetes and Firmicutes were less frequent. Comparisons of slopes of collectors curves and the Shannon-Weiner diversity index indicated no difference in overall bacterial diversity between the two soils, although sequences of δ-Proteobacteria and the cyanobacterial genus Leptolyngbya were more commonly derived from the soil with the higher water and nutrient content. The data suggest that different levels of soil water, C, N and P have only a minor effect on the bacterial community composition of maritime Antarctic soils.

Polycytella Hominis is a mutated form of Scedosporium Apiospermum

PCR amplification and sequencing of two separate regions of the nuclear ribosomal repeat region revealed that Polycytella hominis, a hyphomycete isolated from a human case of mycetoma, was genetically indistinguishable from Scedosporium apiospermum (the anamorph of Pseudallescheria boydii). These organisms also exhibited remarkably similar susceptibility profiles to common antifungal agents. P. hominis is thus likely to be a mutant of S. apiospermum showing abnormalities of sporulation, for which a possible mechanism is discussed. Polycytella hominis should thus be regarded as a synonym of Scedosporium apiospermum.

Sebacinales are associates of the leafy liverwort Lophozia excisa in the southern maritime Antarctic.

The leafy liverwort Lophozia excisa, which is colonised by basidiomycete fungi in other biomes and which evidence suggests may be colonised by mycorrhizal fungi in Antarctica, was sampled from Léonie Island in the southern maritime Antarctic (67°36′ S, 68°21′ W). Microscopic examination of plants indicated that fungal hyphae colonised 78% of the rhizoids of the liverwort, apparently by entering the tips of rhizoids prior to growing into their bases, where they formed hyphal coils. Extensive colonisation of stem medullary cells by hyphae was also observed. DNA was extracted from surface-sterilised liverwort tissues and sequenced following nested PCR, using the primer set ITS1F/TW14, followed by a second round of amplification using the ITSSeb3/TW13 primer set. Neighbour-joining analyses showed that the sequences obtained nested in Sebacinales clade B as a 100% supported sister group to Sebacinales sequences from the leafy liverworts Lophozia sudetica, L. incisa and Calypogeia muelleriana sampled from Europe. Direct PCR using the fungal specific primer set ITS1F/ITS4 similarly identified fungi belonging to Sebacinales clade B as the principal colonists of L. excisa tissues. These observations indicate the presence of a second mycothallus in Antarctica and support the previous suggestion that the Sebacinales has a wide geographical distribution.

Arthroderma olidum, sp. nov. A new addition to the Trichophyton terrestre complex

In 1981, four fungal isolates from hair of the European badger (Meles meles) were examined by Dr Phyllis Stockdale at the Commonwealth Mycological Institute, Kew, and deposited in the UK National Collection of Pathogenic Fungi as an undescribed member of the Trichophyton terrestre complex. The present paper formalizes the complete description of a new ascomycete taxon, Arthroderma olidum following successful recent attempts to re-isolate the same fungus from the soil of Badger holes in South West England. Furthermore, using ribosomal RNA gene sequencing, we show that the asexual form of A. olidum is conspecific with the recently described Trichophyton eboreum1 isolated from a human skin specimen in Germany.