Aktivitas Enzimatis Biakan Kapang Aspergillus Section Nigri DUCC (Diponegoro University Culture Collection) Dan Identifikasi Molekuler Isolat Potensial

The enzyme amylase, cellulase and protease are an extracellular enzymes are many produced by microorganisms such as fungi Aspergillus section Nigri. This research aims to find out the potential biakanes of Aspergillus section Nigri DUCC collection (Diponegoro University Culture Collection) which is capable to produce some enzymes that consists of amylase, protease and celullase and also identification in molecular based. Activity enzymatic assay of Aspergillus section Nigri with semi-quantitative method using selective medium, CMC for cellulolytic, Starch agar 1% for amylolytic and Skim Milk agar for proteolytic. Biakane potential is determined by looking at the Enzymatic Index (EI) is highest for all of three enzyme. Molecular identification is using the universal primer ITS4 and ITS5. The results showed that the culture of DUCC K207 has high activity for all of three enzyme. Index enzymatic of isolate DUCC K207 , 1.55 mm for amylolytic, 1.49 mm for cellulolytic and 1, 24 mm for proteolytic. Result of molecular identification DUCC K207 known as Aspergillus niger that has 100% similarity with Aspergillus niger MH 109325.1.

Invasive Factors Recognition in Aspergillus Section Nigri Isolates from Patient and Environmental Samples in the Centre Region, Cameroon

Background: Aspergillus section Nigri species are invasive opportunistic pathogens, seen in individuals with various immune disorders. The invasive capacity involves the production of varieties of enzymes such as lipases, phospholipase, proteases and hyaluronidase. The determination of proteinase, phospholipase, esterase and biofilm production in patient and environmental isolates approve the pathogenic strength of the species. Aims: To evaluate the invasive capacity of Aspergillus section Nigri isolates from patients and environmental samples. Methods: Our study is cross sectional and experimental, performed at the outpatient clinic of the otorhinolaryngology department of Central and University teaching hospital during a period of 12 months from March 2018 to February 2019. To determine the invasive capacity of Aspergillus section Nigri species, 400 samples were evaluated in the study (that is; 250 from patients and 150 samples from environment). Patient samples and hospital environment samples were evaluated by standard phenotypic methods for detecting of Proteinase, phospolipase, esterase and biofilm. The variables were statistically analyzed using Chi-square test of independent and SPSS (Version 16.0). Results: The isolates recovered from the patient sample shows maximum invasive capacity as compared to the environmental isolates, that is for 44 isolates; 42 isolates showed proteinase activity and biofilm production, followed by phospholipase activity 36, and then esterase 32.The isolates recovered from the hospital environment also showed the production of the various invasive factors, that is for 16 isolates; 15 isolates showed biofilm production, followed by proteinase activity 6, phospholipase 5 and esterase 4.The disparities of the invasive capacity in patient and environment isolates virulence were statistically significant for proteinase, phospholipase and esterase ( that is; p-value <0.05). Majority the isolates recovered from patients and the environment were potential producers of biofilm. Conclusion: The isolates recovered from patients sample showed high invasive capacity as compare to the isolates recovered from the environment. This highlights the implications of phospholipase enzyme, proteinase enzyme, esterase enzyme and biofilm used by Aspergillus section Nigri isolates as means of survival in the host system.

Genetic Diversity, Ochratoxin A and Fumonisin Profiles of Strains of Aspergillus Section Nigri Isolated from Dried Vine Fruits

We investigated ochratoxin A (OTA) contamination in raisin samples purchased from Slovak markets and determined the diversity of black-spored aspergilli as potential OTA and fumonisin (FB1 and FB2) producers. The taxonomic identification was performed using sequences of the nuclear ITS1-5.8s-ITS2 region, the calmodulin and beta-tubulin genes. We obtained 239 isolates from eight fungal genera, of which 197 belonged to Aspergillus (82%) and 42 strains (18%) to other fungal genera. OTA contamination was evidenced in 75% of the samples and its level ranged from 0.8 to 10.6 µg/kg. The combination of all three markers used enabled unambiguous identification of A. carbonarius, A. luchuensis, A. niger, A. tubingensis and A. welwitschiae. The dominant coloniser, simultaneously having the highest within-species diversity isolated from our raisin samples, was A. tubingensis. Out of all analysed strains, only A. carbonarius was found to produce OTA, but in relatively high quantity (2477–4382 µg/kg). The production of FB1 and FB2 was evidenced in A. niger strains only.

Polyphasic, Including MALDI-TOF MS, Evaluation of Freeze-Drying Long-Term Preservation on Aspergillus (Section Nigri) Strains

This study aims to evaluate the effect of freeze-drying and long-term storage on the biotechnological potential of Aspergillus section Nigri strains. Twelve selected strains were freeze-dried and aged by accelerated storage, at 37 °C in the dark, for 2 and 4 weeks. To assess possible changes as a consequence of the ageing in the freeze-drying ampoules, morphological characteristics, mycotoxins and enzymes production, matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALTI-TOF MS) spectra, and M13 phage probe fingerprinting were used as part of a polyphasic approach. Phenotypical changes were observed; nevertheless, they did not substantially affect the potential biotechnological use of these strains. The activity of hydrolytic enzymes (protease, carboxymethylcellulase, xylanase, pectinase and mannanase) was maintained or increased after freeze-drying. MALDI-TOF MS data originated spectra that grouped, for the majority of samples, according to strain independently of preservation time point. M13 profiles revealed the presence of some genetic polymorphisms after preservation. However, the three studied times still clustered for more than 50% of strains. Our results show that the studied strains maintain their biotechnological potential after preservation, with minimal phenotypic alterations. These findings provide evidence that freeze-drying preservation is a suitable option to preserve biotechnologically relevant aspergilli strains from section Nigri, and one should consider that the observed effects might be species/strain-dependent.

Molecular characterization of ochratoxin A-producing fungi isolated from poultry feed

Mycotoxins are toxic metabolites mainly produced by Aspergillus spp. fungi. Further fungal characterization and mycotoxin estimation in feed are crucial measures for animal health and safety. The aim of this study was to identify and characterize mycotoxin-producing fungi in poultry feed. Forty poultry ration samples were collected and were subjected to morphological and molecular as well as to phylogenetic analyses of Aspergillus section Nigri. On average, 40% (16 of 40 samples) were contaminated with Aspergillus section Nigri fungi. Moreover, A. niger and A. tubingensis were identified and characterized in the contaminated poultry feed using molecular and phylogenetic analyses. Furthermore, our results showed that ochratoxin A was produced by some fungal isolates indicating their mycotoxigenicity. In conclusion, A. niger and A. tubingensis contaminate poultry feed and can produce mycotoxins under optimal conditions. Furthermore, several preventive and control measures should be undertaken to reduce the occurrence of fungal contamination and toxin production in poultry rations.