Investigation on the activities of yeasts in the post harvest spoilage of sweet potato(Ipomea batatas L.)

Postharvest losses due to the activities of indigenous microorganisms occur in many crops such as sweet potato (Ipomoea batatas) which result in heavy financial losses for farmers. Correct identification of the pathogen responsible for postharvest infection is central to adopting an appropriate control strategy. Samples of sweet potato tubers were purchased from the local market and stored at three temperatures: 13°C, 21°C and 29° for four weeks. Spoilage yeasts were isolated from the tuber samples. The isolates were characterized using yeast genomic DNA extraction, polymerase chain amplification of rRNA and sequence determination. The yeasts were identified on the basis of the 26S rDNA. Six yeast species were identified as Rhodotorula mucilaginosa, R. minuta, Pichiaguilliermondii, P. anomala, Sporobolomyces marcillae and Saccharomycopsis fibuligera. The results indicated that the D1 and D2 domains of the 5’ end of the 26S rDNA showed a high degree of interspecies sequence variation for the isolates. Two of the yeasts; P. anomala and R. minuta were selected for further investigations namely pathogenicity testing and assay for extracellular enzymes. Results of the pathogenicity tests showed that P. anomala and R. minuta were clearly able to infect the sweet potato tubers. The results of the enzyme assay revealed that P. anomala and R. minuta were able to secrete varying amounts of eight extracellular enzymes: cellulose, amylase, polygalacturonase, glucanase, xylanase, xylosidase, arabinofuranosidase and ferulic acid esterases. These enzymes have the capacity to degrade plant cell walls and possibly enhanced the pathogenicity of the yeasts. Key Words: Sweet-potato, yeast, Postharvest, Pathogenicity, Ipomoea batata

Functional classification of the microbial feruloyl esterases.

Feruloyl esterases have potential uses over a broad range of applications in the agri-food industries. In recent years, the number of microbial feruloyl esterase activities reported has increased and, in parallel, even more related protein sequences may be discerned in the growing genome databases. Based on substrate utilisation data and supported by primary sequence identity, four sub-classes have been characterised and termed type-A, B, C and D. The proposed sub-classification scheme is discussed in terms of the evolutionary relationships existing between carbohydrate esterases

Identification of a type-D feruloyl esterase from Neurospora crassa.

Feruloyl esterases constitute an interesting group of enzymes that have the potential for use over a broad range of applications in the agri-food industries. In order to expand the range of available enzymes, we have examined the presence of feruoyl esterase genes present in the genome sequence of the filamentous fungus Neurospora crassa. We have identified an orphan gene (contig 3.544), the translation of which shows sequence identity with known feruloyl esterases. This gene was cloned and the corresponding recombinant protein expressed in Pichia pastoris to confirm that the enzyme (NcFaeD-3.544) exhibits feruloyl esterase activity. Unusually the enzyme was capable of p-coumaric acid release from untreated crude plant cell wall materials. The substrate utilisation preferences of the recombinant enzyme place it in the recently recognised type-D sub-class of feruloyl esterase

A regulator gene for acetate utilisation from Neurospora crassa.

The Neurospora crassa homologue of the Aspergillus nidulans regulatory gene facB has been cloned. The gene encodes a putative transcriptional activator of 865 amino acids that contains a DNA-binding domain with a Zn(II)(2)Cys(6) binuclear cluster, a linker region and a leucine zipper-like heptad repeat. Two internal amino acid sequences are identical to peptide sequences determined from proteolytic fragments of a DNA-binding protein complex specific for genes involved in acetate utilisation and expressed in acetate-induced mycelia of N. crassa. Recombinant expression of the predicted DNA-binding domain demonstrates that it is capable of independent recognition of a subset of the promoter sequences that bind the protein complex from N. crassa. A duplication-induced mutation in the corresponding gene results in an acetate non-utilising phenotype that is characterised by inefficient induction of the enzymes required for acetate utilisation. The new gene does not fall into any existing complementation group and has been designated acu-15