Selection and molecular identification of fungal isolates that produce xylanolytic enzymes.

Xylan is a heteropolysaccharide and its complete hydrolysis involves a complex set of xylanolytic enzymes. Fungal xylanases have been widely used in the holocellulose industry to obtain by-products or for its elimination. The aim of this study was to select and identify filamentous fungi from different ecosystems that produce extracellular xylanases showing biotechnological potential. One hundred three fungal isolates were obtained from orchard, horticultural, and forest ecosystems. The ability of fungi to degrade xylan was measured by quantifying their xylanolytic indices after growth on solid culture media and their extracellular xylanolytic and cellulolytic activities after submerged fermentation. All fungal isolates grew on solid medium supplemented with xylan as the sole carbon source, but only 44% of isolates showed xylanolytic indices greater than 1.0. In submerged fermentation, 39% of the fungi tested showed no cellulolytic activity. Filamentous fungi were chosen from correspondence analysis and were identified by molecular tools using internal transcribed spacers. One of the 9 isolates selected belonged to the Phoma genus and the remaining were from the Fusarium genus. Fusarium solani (isolate 59) showed the highest xylanolytic index (0.964 ± 0.042), rapid growth on solid medium (1.233 ± 0.050 cm/day), significant xylanolytic activity (3.823 ± 0.210 U/mg), and a total deficiency of cellulolytic activity compared to other fungal isolates. In the zymogram, a clear zone was observed, indicating that F. solani possesses at least 1 xylanase. Fusarium solani was selected for its ability to produce extracellular xylanases with biotechnological potential.

Mechanisms of interaction of chromium with Aspergillus niger var tubingensis strain Ed8.

Experiments were conducted to determine the mechanisms of interaction with chromium of Aspergillus niger var tubingensis strain Ed8 in batch culture and in bioreactor experiments. Results obtained in this work showed that the interaction of A. niger var tubingensis Ed8 with Cr(VI) is based mainly in a reduction process and also, secondly, in a sorption process. Using electron microscopy techniques the ultrathin sections obtained from the mycelium biomass produced by the fungus in batch cultures showed the ability to incorporate Cr intracellulary, into low electron-dense inclusions, but not extracellularly. On the other hand, cultures without Cr(VI) of A. niger var tubingensis Ed8, grown in a bubble column bioreactor, reduced Cr(VI) immediately after repeated addition of this oxyanion; after six loads, 460 mg Cr(VI) was reduced to Cr(III) in 60 h, corresponding to a reduction rate of 2.62 mg Cr(VI)g(-1) dry biomass h(-1).

Hexavalent chromium reduction by bacterial consortia and pure strains from an alkaline industrial effluent.

AIMS: To characterize the bacterial consortia and isolates selected for their role in hexavalent chromium removal by adsorption and reduction. METHODS AND RESULTS: Bacterial consortia from industrial wastes revealed significant Cr(VI) removal after 15 days when incubated in medium M9 at pH 6·5 and 8·0. The results suggested chromium reduction. The bacterial consortia diversity (T-RFLP based on 16S rRNA gene) indicated a highest number of operational taxonomic units in an alkaline carbonate medium mimicking in situ conditions. However, incubations under such conditions revealed low Cr(VI) removal. Genomic libraries were obtained for the consortia exhibiting optimal Cr(VI) removal (M9 medium at pH 6·5 and 8·0). They revealed the dominance of 16S rRNA gene sequences related to the genera Pseudomonas/Stenotrophomonas or Enterobacter/Halomonas, respectively. Isolates related to Pseudomonas fluorescens and Enterobacter aerogenes were efficient in Cr(VI) reduction and adsorption to the biomass. CONCLUSIONS: Cr(VI) reduction was better at neutral pH rather than under in situ conditions (alkaline pH with carbonate). Isolated strains exhibited significant capacity for Cr(VI) reduction and adsorption. SIGNIFICANCE AND IMPACT OF STUDY: Bacterial communities from chromium-contaminated industrial wastes as well as isolates were able to remove Cr(VI). The results suggest a good potential for bioremediation of industrial wastes when optimal conditions are applied.

Differences in DNA methylation patterns are detectable during the dimorphic transition of fungi by amplification of restriction polymorphisms.

A modification of the amplified fragment length polymorphism technique was developed for the determination of DNA methylation in dimorphic fungi representative of three of the major fungal taxa: Mucor rouxii, a zygomycete; Yarrowia lipolytica, an ascomycete; and Ustilago maydis, a basidiomycete. DNA obtained from the yeast or mycelial stages of the fungi was digested with a mixture of EcoRI, and one of the isoschizomers MspI and HpaII, whose ability to cleave at the sequence CpCpGpG is affected by the methylation state. The resulting fragments were ligated to primers and subjected to a double round of amplification by the polymerase chain reaction, radiolabeled in the second round, and separated by polyacrylamide gel electrophoresis. Comparison of patterns revealed differences indicative of fragments whose methylation state did or did not change during the dimorphic transition. These results indicate the usefulness of the method for the study of DNA methylation, demonstrate the universality of DNA methylation in fungi, and confirm that differential DNA methylation occurs during fungal morphogenesis.