Diversity of anaerobic gut fungal populations analysed using ribosomal ITS1 sequences in faeces of wild and domesticated herbivores.

Gut fungal-specific PCR primers have been used to selectively amplify the ITS1 region of gut fungal rDNA recovered from faeces of domestic and wild animals to investigate population diversity. Two different gel-based methods are described for separating populations of gut fungal rDNA amplicons, namely (1) denaturing gradient gel electrophoresis (DGGE) and (2) separation according to small size differences using Spreadex, a proprietary matrix for electrophoresis. Gut fungal populations were characterised by analysis of rDNA in faeces of seventeen domesticated and ten wild herbivores.

The rapid assignment of ruminal fungi to presumptive genera using ITS1 and ITS2 RNA secondary structures to produce group-specific fingerprints.

Identification of microbial community members in complex environmental samples is time consuming and repetitive. Here, ribosomal sequences and hidden Markov models are used in a novel approach to rapidly assign fungi to their presumptive genera. The ITS1 and ITS2 fragments from a range of axenic, anaerobic gut fungal cultures, including several type strains, were isolated and the RNA secondary structures predicted for these sequences were used to generate a fingerprinting program.

Molecular analysis of the anaerobic rumen fungus Orpinomyces - insights into an AT-rich genome.

The anaerobic gut fungi occupy a unique niche in the intestinal tract of large herbivorous animals and are thought to act as primary colonizers of plant material during digestion. They are the only known obligately anaerobic fungi but molecular analysis of this group has been hampered by difficulties in their culture and manipulation, and by their extremely high A+T nucleotide content. This study begins to answer some of the fundamental questions about the structure and organization of the anaerobic gut fungal genome.

Glucoamylase::green fluorescent protein fusions to monitor protein secretion in Aspergillus niger.

A glucoamylase::green fluorescent protein fusion (GLA::sGFP) was constructed which allows the green fluorescent protein to be used as an in vivo reporter of protein secretion in Aspergillus niger. Two secretory fusions were designed for secretion of GLA::sGFP which employed slightly different lengths of the glucoamylase protein (GLA499 and GLA514). Expression of GLA::sGFP revealed that fluorescence was localized in the hyphal cell walls and septa, and that fluorescence was most intense at hyphal apices.