Modeling the inactivation of Aspergillus fischeri and Paecilomyces niveus ascospores in apple juice by different ultraviolet light irradiances.

The present work aimed to evaluate and to model the influence of UV-C light treatments with different irradiances (6.5, 13, 21, and 36 W/m2) on Aspergillus fischeri and Paecilomyces niveus ascospores inactivation in clarified apple juice. Approximately 5.0 and 6.0 log CFU/mL spores of P. niveus and A. fischeri, respectively, were suspended in 30 mL of clarified apple juice (pH 3.8, 12 ± 0.1°Brix) and exposed to UV-C light at different irradiances (as above) and exposure times (0 to 30 min). The first-order biphasic model was able to describe the experimental data with good statistical indices (RMSE = 0.296 and 0.308, R2 = 0.96 and 0.98, for P. niveus and A. fischeri respectively). At the highest irradiance level tested (36 W/m2), the UV-C light allowed the reduction of 5.7 and 4.2 log-cycles of A. fischeri and P. niveus ascospores, respectively, in approximately 10 min. P. niveus was the most UV-C resistant mould. The results showed that, to a defined UV-C fluence, a change in the level of either time or UV-C irradiance did not affect the effectiveness of UV-C light for A. fischeri and P. niveus inactivation. Thus, the modeling of the inactivation as a function of the UV-C fluence allowed the estimation of the primary model parameters with all experimental data and, consequently, no secondary models were needed. The model parameters were validated with experiments of variable UV-C fluences. Accordingly, experimental results allowed to conclude that UV-C treatment at the irradiances tested is a promising application for preventing A. fischeri and P. niveus spoilage of juices.

Insights into the substrate specificity and synergy with mannanase of family 27 α-galactosidases from Neosartorya fischeri P1.

Thermophilic Neosartorya fischeri P1 is an excellent carbohydrate-active enzyme (CAZyme) producer. Two α-galactosidases of GH (glycoside hydrolase ) family 27 with a very low sequence identity (28.7%), Gal27A and Gal27B, were identified in strain P1 and functionally expressed in Pichia pastoris. In comparison to other characterized GH27 fungal counterparts, rGal27B has a higher temperature optimum (75 °C) and better thermostability (>50% activity at 70 °C for 15 min), and rGal27A shows stability over the broadest pH range (pH 2.0-12.0). Moreover, great distinctions lie in the two enzymes. When using pNPG as the substrate, rGal27B had a higher turnover number (1621.4 vs. 368.3 s(-1)) but lower affinity (2.84 vs. 0.8 mM) and catalytic efficiency (460.8 vs. 580.3 s(-1) mM(-1)) than rGal27A. rGal27B acted on galacto-oligosaccharides, whereas rGal27A was active on polymeric substrates. Although both enzymes showed synergy in galactomannan degradation when combined with a ß-mannanase of the same strain, enzyme combinations including rGal27A released more reducing sugars (up to 11.67-fold). Homology modeling predicts different loops in N. fischeri α-galactosidases, highlighting the larger tunnel structure in Gal27A to accommodate/bind branched galactomannan with high galactose contents. Phylogenetic analysis reveals the far relationship of Gal27A and Gal27B that they may evolve in different action modes, and their coexistence widens the substrate spectrum for nutrient utilization. This study illustrates the substrate profiles and synergistic mechanism of GH27 α-galactosidases of different structures.

Misidentification of Neosartorya pseudofischeri as Aspergillus fumigatus in a lung transplant patient.

We present a case of disseminated Neosartorya pseudofischeri infection in a bilateral lung transplant patient with cystic fibrosis. The organism was originally misidentified from respiratory specimens as Aspergillus fumigatus using colonial and microscopic morphology. DNA sequencing subsequently identified the organism correctly as N. pseudofischeri.

Diversity and specificity of microsatellites within Aspergillus section Fumigati.

BACKGROUND: Microsatellites (or short tandem repeats, STRs) are the genetic markers of choice for studying Aspergillus fumigatus molecular epidemiology due to its reproducibility and high discrimination power. However, the specificity of these markers must be investigated in a group of isolates from closely related species. The aim of this work was to test a microsatellite-based PCR multiplex previously designed for A. fumigatus in a set of species belonging to section Fumigati, namely Aspergillus fumigatiaffinis, Aspergillus lentulus, Aspergillus novofumigatus, Aspergillus unilateralis, Aspergillus viridinutans, Neosartorya fischeri, Neosartorya hiratsukae, Neosartorya pseudofischeri and Neosartorya udagawae. RESULTS: The reference A. fumigatus strain ATCC 46645 was easily genotyped in standard conditions showing a final electrophoretic profile of 8 expected peaks corresponding to each microsatellite locus. Inversely, no peaks were observed for all other species from section Fumigati, with an exception for marker MC6b in A. unilateralis. By screening the genome sequence of Neosartorya fischeri NRRL 181, the results showed that MC3, MC6a and MC7 might be employed for N. fischeri genotyping since these markers present several repeats of each motif. The accumulation of insertions and deletions was frequently observed in the genomic regions surrounding the microsatellites, including those where the A. fumigatus primers are located. The amplification of microsatellite markers in less stringent amplification conditions resulted in a distinct electrophoretic profile for species within section Fumigati. CONCLUSIONS: Therefore, the microsatellite-based PCR multiplex allow simple identification of A. fumigatus and, with a slight modification of temperature conditions, it also allows discriminating other pathogenic species within section Fumigati, particularly A. fumigatiaffinis, N. fischeri and N. udagawae.

First evidence of mineralization of petroleum asphaltenes by a strain of Neosartorya fischeri.

A fungal strain isolated from a microbial consortium growing in a natural asphalt lake is able to grow in purified asphaltenes as the only source of carbon and energy. The asphaltenes were rigorously purified in order to avoid contamination from other petroleum fractions. In addition, most of petroporphyrins were removed. The 18S rRNA and ß-tubulin genomic sequences, as well as some morphologic characteristics, indicate that the isolate is Neosartorya fischeri. After 11 weeks of growth, the fungus is able to metabolize 15.5% of the asphaltenic carbon, including 13.2% transformed to CO(2) . In a medium containing asphaltenes as the sole source of carbon and energy, the fungal isolate produces extracellular laccase activity, which is not detected when the fungus grow in a rich medium. The results obtained in this work clearly demonstrate that there are microorganisms able to metabolize and mineralize asphaltenes, which is considered the most recalcitrant petroleum fraction.

Neosartorya hiratsukae peritonitis through continuous ambulatory peritoneal dialysis.

Fungal peritonitis is a rare, potentially lethal, complication of continuous ambulatory peritoneal dialysis (CAPD). We report what we believe to be the first confirmed Neosartorya hiratsukae CAPD-related peritonitis case in Europe. The patient died, despite early removal of the peritoneal catheter and antifungal therapy. This report highlights the impact of emerging fungal pathogens and the importance of early diagnosis on the outcome in CAPD-related fungal peritonitis.

Identification of two marine fungi and evaluation of their antivirus and antitumor activities.

OBJECTIVE: To identify two marine fungi and evaluate the inhibitory effects of their crude extracts on Tobacco mosaic virus and two tumor cell lines. METHODS: Crude extracts was obtained by extracting with MeOH and evaporated in vacuo. The extracts was water-soluble fraction which was dissolved in water, and the other fraction was water insoluble. The fungi were identified by morphology and Internal Transcribed Spcer (ITS) rDNA molecular methods. The inhibitory effect on Tobacco mosaic virus was evaluated by indirect enzyme linked immunosorbent assay, and the anti-tumor activity was tested by methyl thiazolyl tetrazolium method. RESULTS: The fungi were identified as Penicillium oxalicum and Neosartorya fischeri. There crude extracts inhibited Tobacco Mosaic Virus and two tumor cell lines. The active fraction named 0312F1 inhibited Tobacco Mosaic Virus and tumor cell lines and was water-soluble. The fraction named 1008F1 inhibited Tobacco Mosaic Virus and was insoluble in water, whereas the fraction inhibited tumor cell lines was water-soluble. CONCLUSION: The active fraction named 0312F1 inhibited Tobacco Mosaic Virus was different from that named 1008F1 inhibited Tobacco Mosaic Virus. The active fraction named 0312F1 inhibited tumor cell lines was the same as that named 1008F1. Furthermore, the inhibitory activity of water-soluble fraction named 0312F1 against BEL-7404 cell line was much higher than that against SGC-7901 cell lines, whereas the inhibitory activity of active fraction named 1008F1 against SGC-7901 cell line was much higher.

Invasive aspergillosis due to Neosartorya udagawae.

BACKGROUND: Invasive aspergillosis (IA) is most commonly caused by the morphospecies Aspergillus fumigatus. However, genetic-based methods indicate that organisms phenotypically identified as A. fumigatus actually constitute a mold complex, designated Aspergillus section fumigati subgenus fumigati. METHODS: Multilocus sequencing and analysis was performed on fungi identified as A. fumigatus from the clinical culture collection maintained at the National Institutes of Health from 2000 through 2008, with a focus on the internal transcribed spacer 1 and 2 regions of ribosomal DNA (rDNA), beta-tubulin, and rodlet A genes. We reviewed the medical records, radiology, and histopathology of corresponding patients. To confirm identification of Neosartorya udagawae isolates, mating studies were performed with reference strains. Antifungal susceptibility testing was performed by broth microdilution and read at 48 hours. RESULTS: Thirty-six cases of infection attributed to A. fumigatus were identified; 4 were caused by N. udagawae (3 in patients with chronic granulomatous disease and 1 in a patient with myelodysplastic syndrome). Disease due to N. udagawae was chronic, with a median duration of 35 weeks, compared with a median duration of 5.5 weeks for patients with chronic granulomatous disease who had infection due to A. fumigatus sensu stricto (P < .05 , Mann-Whitney U test). Infection spread across anatomical planes in a contiguous manner and was refractory to standard therapy. Two of the 4 patients died. N. udagawae demonstrated relatively higher minimum inhibitory concentrations to various agents, compared with those demonstrated by contemporary A. fumigatus sensu stricto isolates. CONCLUSIONS: To our knowledge, this is the first report documenting infection due to N. udagawae. Clinical manifestations were distinct from those of typical IA. Fumigati-mimetics with inherent potential for antifungal resistance are agents of IA. Genetic identification of molds should be considered for unusual or refractory IA.