Development of sequence-characterized amplified region (SCAR) markers for accurate and differential identification of multienzyme-producing and non-enzymatic Aspergillus strains of industrial importance.

Aspergillus strains are known to produce multiple enzymes of industrial importance. To screen Aspergillus isolates and select a strain with the ability to produce multiple enzymes and discriminate it from non-enzymatic strains, a rapid and accurate approach is required. With this background, a DNA fingerprinting-based study was conducted to develop a simple but accurate molecular detection method with the potential to discriminate multienzyme-producing Aspergillus strains from non-enzymatic strains, irrespective of species. To achieve this, Enterobacterial Repetitive Intergenic Consensus (ERIC) PCR was employed to derive group-specific Sequence Characterized Amplified Region (SCAR) markers (i.e., markers corresponding to PCR amplicons of known DNA sequence). To this end, both group-specific (multienzyme-producing and non-enzymatic Aspergillus group) SCAR markers were sought by comparing the ERIC fingerprint profiles and used to develop primers for use in specific and differential identification of multienzyme-producing Aspergillus isolates. As an outcome, the two SCAR-PCR formats were developed. One format is for specific identification of multienzyme-producing Aspergillus strains (SCAR-PCR1), and the other for identifying non-enzymatic Aspergillus strains (SCAR-PCR2). Both SCAR-PCRs were able to discriminate between these two contrasting groups. These formats are simple but accurate and rapid compared to the time-consuming and laborious conventional methods. Therefore, they could be efficient as an alternative strategy for the high-throughput screening of industrially important Aspergillus strains.

Isolation and characterisation of nitrilase producing Aspergillus species recovered from solid waste leachates at two dump sites in Lagos, Nigeria

@#Aims: Nitrile compounds are cyanogenic intermediates, products, byproducts and waste products of agriculture, chemical and pharmaceutical industries and fossil fuels degradation. The enzymatic hydrolysis of nitriles to non-toxic carboxylic acids or amides plays an increasingly important role in environment remediation. This study aimed at exploring the potential of Aspergilli in the detoxification nitrile compounds at two selected dump sites in Lagos, Nigeria. Methodology and results: Decomposing solid waste leachate samples (SWL) were randomly collected at two selected dump sites namely Olusosun (Ojota) and Isolo (Oke-Afa). Samples per site were pooled, processed by selective enrichment and screened for the presence of Aspergilli by culture technique and intergenic spacer sequencing (ITS). Biomass generation and pH changes in the culture fluids were monitored at 4-days interval by dry weight measurements. Nitrilase production was determined spectrophotometrically. Two nitrilase producing Aspergillus strains: Aspergillus fumigatus strain WO2 with accession number MF78882 and Aspergillus niger strain WO7 with accession number MH542673 were identified. Growth investigation revealed biomass generations of 17.8 g and 23.8 g dry weight per one liter media for A. fumigatus strain WO2 and A. niger strain WO7 respectively. Progressive pH monitoring showed decline from 7.2 to 4.5 and 7.2 to 6.2 was obtained for strains WO2 and WO7 respectively, during nitrilase production at different yields of 0.0150 and 0.0161 mg/mL/min respectively. Conclusion, significance and impact of study: This study supports the studied dump sites as important sources of nitrilase-producing A. fumigatus and A niger strains with potentials as cost-effective environmental bioremediation agents in Nigeria.

The potency of luliconazole against clinical and environmental Aspergillus nigri complex.

BACKGROUND AND OBJECTIVES: Black Aspergillus strains including, Aspergillus niger and A. tubingensis, are the most cause of otomycosis with worldwide distribution. Although, amphotericin B was a Gold standard for the treatment of invasive fungal infection for several decades, it gradually replaced by fluconazole and /or voriconazole. Moreover, luliconazole, appears to offer the best potential for in vitro activity against black Aspergillus strains. The aim of the present study was to compare the in vitro activity luliconazole, with commonly used antifungals against clinical and environmental strains of black Aspergillus. MATERIALS AND METHODS: Sixty seven (37 clinical and 30 environmental) strains of black Aspergillus were identified using morphological and molecular technique (ß-Tubulin gene). In addition, antifungal susceptibility test was applied according to CLSI M38 A2. The results were reported as minimum inhibitory concentration (MIC) or minimum effective concentration (MEC) range, MIC50 or MEC50, MIC90 or MEC90 and MIC geometric (GM) or MECGM. RESULTS: Aspergillus niger was the common isolate followed by, A. tubingensis in both clinical and environmental strains. The lowest MIC range, MIC50, MIC90, and MICGM was attributed to luliconazole in clinical strains. The highest resistant rate was found in amphotericin B for both clinical (86.5%) and environmental (96.7%) strains whereas 54.1% of clinical and 30% of environmental isolates were resistant to caspofungin. Clinical strains of Aspergillus were more sensitive to voriconazole (86.7%) than environmental strains (70.3%). On the other hand, 83.8% of clinical and 70% of environmental isolates were resistant to posaconazole. CONCLUSION: Luliconazole versus amphotericin B, voriconazole, posaconazole and caspofungin is a potent antifungal for Aspergillus Nigri complex. The in vitro extremely antifungal efficacy against black Aspergillus strains of luliconazole, is different from those of other used antifungals.

Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels

Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE), in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (β-glucosidase, endoglucanase and filterpaperase) using a novel substrate, Banana Peels (BP) for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes.

Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels.

Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE), in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (ß-glucosidase, endoglucanase and filterpaperase) using a novel substrate, Banana Peels (BP) for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes.