Cacti as low-cost substrates to produce L-asparaginase by endophytic fungi.

This study aimed to select endophytic fungi to produce L-asparaginase and partially optimising the production of the enzyme using cacti as substrate. Seventeen endophytes were assessed for intracellular enzymatic potential in modified Czapek Dox's medium using L-proline as an inducer. The best producer was evaluated for intracellular and extracellular enzymatic activity in modified Czapek Dox's medium using flours of Opuntia ficus-indica and Nopalea cochenillifera as substrate. The biomass and L-asparaginase production profile was analysed and the best conditions for enzyme production were verified using factorial design. Penicillium decaturense URM 7966, Diaporthe ueckerae URM 8321, and Colletotrichum annellatum URM 8538 produced 0.76 U g- 1, 0.87 U g- 1, and 0.74 U g- 1 L-asparaginase, respectively. Diaporthe ueckerae URM 8321 produced only intracellular L-asparaginase, using flours of N. cochenillifera (0.72 U g- 1) and O. ficus-indica (0.90 U g- 1) and the last was selected for the next steps. The ideal time for biomass and L-asparaginase production was 120 h. The best conditions for enzyme production (1.67 U g- 1) were initial pH 4.0, inoculum concentration 1% and cacti flour concentration 0.2%; where was observed an increase of 46.11% in compared to the initial production. Opuntia ficus-indica flour is indicated as an alternative low-cost substrate for the production of L-asparaginase by the endophytic fungus D. ueckerae URM 8321.

Aspergillus and Penicillium (Eurotiales: Trichocomaceae) in soils of the Brazilian tropical dry forest: diversity in an area of environmental preservation.

Soil is a complex biological system that plays a key role for plants and animals, especially in dry forests such as the Caatinga. Fungi from soils, such as Aspergillus and Penicillium, can be used as bioindica- tors for biodiversity conservation. The aim of this study was to isolate and identify species of Aspergillus and Penicillium in soil, from the municipalities of Tupanatinga and Ibimirim, with dry forests, in the Catimbau National Park. Five collections were performed in each area during the drought season of 2012, totaling 25 soil samples per area. Fungi were isolated by suspending soil samples in sterile distilled water and plating on Sabouraud Agar media plus Chloramphenicol and Rose Bengal, and Glycerol Dicloran Agar. Isolates were identified by morphological taxonomy in the Culture Collection Laboratory and confirmed by sequencing of the Internal Transcribed Spacer of rDNA. A total of 42 species were identified, of which 22 belong to the genus Aspergillus and 20 to Penicillium. Penicillium isolates showed uniform distribution from the collecting area in Tupanatinga, and the evenness indices found were 0.92 and 0.88 in Tupanatinga and Ibimirim, respectively. Among isolates of Aspergillus evenness, the value found in Tupanatinga (0.85) was very close to that found in Ibimirim (0.86). High diversity and low dominance of fungi in soil samples was observed. These results con- tributed to the estimation of fungal diversity in dry environments of the Caatinga, where diversity is decreasing in soils that have undergone disturbance.

Antifungal susceptibility of the endophytic fungus Rhinocladiella similis (URM 7800) isolated from the Caatinga dry forest in Brazil.

The present study reports a new occurrence of Rhinocladiella similis isolated as an endophytic fungus in the Caatinga dry tropical forest in Brazil and describes its antifungal susceptibility. The isolate R. similis URM 7800 was obtained from leaves of the medicinal plant Myracrodruon urundeuva. Its morphological characterization was performed on potato dextrose agar medium and molecular analysis using the ITS rDNA sequence. The antifungal susceptibility profile was defined using the Clinical and Laboratory Standards Institute (CLSI) protocol M38-A2. The colony of isolate URM 7800 showed slow growth, with an olivaceous-gray color and powdery mycelium; in microculture, it showed the typical features of R. similis. In the antifungal susceptibility test, isolate URM 7800 showed high minimal inhibitory concentration (MIC) values for amphotericin B (>16 µg/mL), voriconazole (16 µg/mL), terbinafine (>0.5 µg/mL), and caspofungin (>8 µg/mL), among other antifungal drugs. Pathogenic melanized fungi are frequently isolated in environments where humans may be exposed, and these data show that it is essential to know if these isolates possess antifungal resistance.

Untargeted Metabolomics Approach of Cross-Adaptation in Salmonella Enterica Induced by Major Compounds of Essential Oils.

Cross-adaptation phenomena in bacterial populations, induced by sublethal doses of antibacterial solutions, are a major problem in the field of food safety. In this regard, essential oils and their major compounds appear as an effective alternative to common sanitizers in food industry environments. The present study aimed to evaluate the untargeted metabolomics perturbations of Salmonella enterica serovar Enteritidis that has been previously exposed to the sublethal doses of the major components of essential oils: cinnamaldehyde, citral, and linalool (CIN, CIT, and LIN, respectively). Cinnamaldehyde appeared to be the most efficient compound in the assays evaluating the inhibitory effects [0.06% (v/v) as MBC]. Also, preliminary tests exhibited a phenotype of adaptation in planktonic and sessile cells of S. Enteritidis when exposed to sublethal doses of linalool, resulting in tolerance to previously lethal concentrations of citral. A metabolomics approach on S. Enteritidis provided an important insight into the phenomenon of cross-adaptation induced by sublethal doses of major compounds of some essential oils. In addition, according to the results obtained, when single molecules were used, many pathways may be involved in bacterial tolerance, which could be different from the findings revealed in previous studies regarding the use of phytocomplex of essential oils. Orthogonal projection to latent structures (OPLS) proved to be an interesting predictive model to demonstrate the adaptation events in pathogenic bacteria because of the global engagement to prevent and control foodborne outbreaks.

Genetic Diversity and Pathogenicity of Botryosphaeriaceae Species Associated with Symptomatic Citrus Plants in Europe.

This study represents the first survey studying the occurrence, genetic diversity, and pathogenicity of Botryosphaeriaceae species associated with symptomatic citrus species in citrus-production areas in five European countries. Based on morphological features and phylogenetic analyses of internal transcribed spacer (ITS) of nuclear ribosomal DNA (nrDNA), translation elongation factor 1-alpha (TEF1) and ß-tubulin (TUB2) genes, nine species were identified as belonging to the genera Diplodia, Dothiorella, Lasiodiplodia, and Neofusicoccum. Isolates of Neofusicoccum parvum and Diplodia pseudoseriata were the most frequently detected, while Dothiorella viticola had the widest distribution, occurring in four of the five countries sampled. Representative isolates of the nine Botryosphaeriaceae species used in the pathogenicity tests caused similar symptoms to those observed in nature. Isolates assayed were all re-isolated, thereby fulfilling Koch's postulates. Isolates of Diplodia pseudoseriata and Diplodia olivarum are recorded for the first time on citrus and all species found in our study, except N. parvum, are reported for the first time on citrus in Europe.

Insights into the Bioprospecting of the Endophytic Fungi of the Medicinal Plant Palicourea rigida Kunth (Rubiaceae): Detailed Biological Activities.

A multitude of plants from the Brazilian savanna are known for their medicinal properties. Many plants contain endophytic fungi, which lead to the production of bioactive compounds by both the fungi and their hosts. This study investigated the bioprospecting of endophytic fungi recovered from the leaves of Palicourea rigida, a native medicinal plant of the Brazilian savanna. Four fungal taxa (Colletotrichum sp. SXS649, Pestalotiopsis sp. SXS650, the order Botryosphaeriales SXS651, and Diaporthe sp. SXS652) were recovered. The phenolic, flavonoid, extracellular degrading enzymes (amylase, cellulase, protease, and tannase) and antioxidant activity of these taxa were determined. Evaluation of the antimicrobial activity showed that the Botryosphaeriales SXS651 extract displays a minimum inhibitory concentration (MIC) of 23.20 mg mL-1 against Staphylococcus epidermidis and Pseudomonas aeruginosa, and the Diaporthe sp. SXS652 extract exhibited an MIC of 27.00 mg mL-1 against Escherichia coli. The Colletotrichum sp. SXS649 isolate inhibited tumors in potato discs by 69% at a concentration of 9.70 mg mL-1. All isolates had potential bioremediation criteria against soil contaminated with soybean oil, as proved by a high percentage of germination of Lactuca sativa and a reduction in phytotoxicity. Furthermore, the taxa under investigation demonstrated antagonistic action to phytopathogenic fungi, namely, Aspergillus niger, Inonotus rickii, Pestalotiopsis mangiferae, and Coniophora puteana, with an inhibition range between 34.2% and 76.9%. The preliminary toxicity assessment showed that all isolates possessed an LC50 of less than 100 mg mL-1 to the microcrustacean Artemia salina. These results indicate that the endophytic fungi of the Brazilian savanna are promising candidates for biotechnological and industrial applications and, in agricultural applications, for the biological control of phytopathogenic fungi.