ABSTRACT. Tropical and subtropical plants are rich in endophytic community diversity. Endophytes, mainly fungi and bacteria, inhabit the healthy plant tissues without causing any damage to the hosts. These fungi can be useful for biological control of pathogens and plant growth promotion. Some plants of the genus Piper are hosts of endophytic microorganisms; however, there is little information about endophytes on Piper hispidum, a medicinal shrub used as an insecticide, astringent, diuretic, stimulant, liver treatment, and for stopping hemorrhages. We isolated the fungal endophyte community associated with P. hispidum leaves from plants in a Brazilian forest remnant. The endophytic diversity was examined based on sequencing of the ITS1-5.8S-ITS2 region of rDNA. A high colonization frequency was obtained, as expected for tropical angiosperms. Isolated endophytes were divided into 66 morphogroups, demonstrating considerable diversity. We identified 21 isolates, belonging to 11 genera (Alternaria, Bipolaris, Colletotrichum, Glomerella, Guignardia, Lasiodiplodia, Marasmius, Phlebia, Phoma, Phomopsis, and Schizophyllum); one isolate was identified only to the order level (Diaporthales). Bipolaris was the most frequent genus among the identified endophytes. Phylogenetic analysis confirmed the molecular identification of some isolates to genus level while for others it was confirmed at the species level.

Endophytes are being considered for use in biological control, and the enzymes they secrete might facilitate their initial colonization of internal plant tissues and direct interactions with microbial pathogens. Microbial proteases are also biotechnologically important products employed in bioremediation processes, cosmetics, and the pharmaceutical, photographic and food industries. In the present study, we evaluated antagonism and competitive interactions between 98 fungal endophytes and Alternaria alternata, Colletotrichum sp., Phyllosticta citricarpa and Moniliophthora perniciosa. We also examined the proteolytic activities of endophytes grown in liquid medium and conducted cup plate assays. The results showed that certain strains in the assemblage of P. hispidum endophytes are important sources of antifungal properties, primarily Lasiodiplodia theobromae JF766989, which reduced phytopathogen growth by approximately 54 to 65%. We detected 28 endophytes producing enzymatic halos of up to 16.40 mm in diameter. The results obtained in the present study highlight the proteolytic activity of the endophytes Phoma herbarum JF766995 and Schizophyllum commune JF766994, which presented the highest enzymatic halo diameters under at least one culture condition tested. The increased activities of certain isolates in the presence of rice or soy flour as a substrate (with halos up to 17.67 mm in diameter) suggests that these endophytes have the potential to produce enzymes using agricultural wastes.

Endophytic fungi have been described as producers of important bioactive compounds; however, they remain under-exploited as exopolysaccharides (EPS) sources. Therefore, this work reports on EPS production by submerged cultures of eight endophytes isolated from Piper hispidum Sw., belonging to genera Diaporthe, Marasmius, Phlebia, Phoma, Phyllosticta and Schizophyllum. After fermentation for 96 h, four endophytes secreted EPS: Diaporthe sp. JF767000, Diaporthe sp. JF766998, Diaporthe sp. JF767007 and Phoma herbarumJF766995. The EPS from Diaporthe sp. JF766998 differed statistically from the others, with a higher percentage of carbohydrate (91%) and lower amount of protein (8%). Subsequently, this fungus was grown under submerged culture for 72, 96 and 168 h (these EPS were designated EPSD1-72, EPSD1-96 and EPSD1-168) and the differences in production, monosaccharide composition and apparent molecular were compared. The EPS yields in mg/100 mL of culture medium were: 3.0 ± 0.4 (EPSD1-72), 15.4 ± 2.2 (EPSD1-96) and 14.8 ± 1.8 (EPSD1-168). The EPSD1-72 had high protein content (28.5%) and only 71% of carbohydrate; while EPSD1-96 and EPSD1-168 were composed mainly of carbohydrate (≈95 and 100%, respectively), with low protein content (≈5%) detected at 96 h. Galactose was the main monosaccharide component (30%) of EPSD1-168. Differently, EPSD1-96 was rich in glucose (51%), with molecular weight of 46.6 kDa. It is an important feature for future investigations, because glucan-rich EPS are reported as effective antitumor agents.

Endophytic fungi colonize the interior of plant tissues and organs, establishing an intimate mutualistic association with no visible symptoms. The fungi may help protect the plant against herbivores and pathogens, making them potentially useful endophytes in the biological control of diseases and agricultural pests. The biotechnological interest in these organisms has stimulated research related to the bioprospecting of endophytic fungi. Grapevine is among the oldest of plants cultivated by man, with the grape being one of the most highly consumed fruits in the world. Diseases cause significant damage to grape cultures, making their integrated control important to reduce the use of pesticides and, consequently, environmental and human contamination. The rustic species Vitis labrusca L. (Vitaceae), used in the preparation of juices and wines, is highly resistant to fungal diseases. We isolated leaf endophytic fungi of the Bordô and Concord cultivars (V. labrusca L.), which were ordered into 68 and 62 morpho-groups of the Bordô and Concord cultivars, respectively. We used scanning electron microscopy to confirm the presence of endophytes in the leaves. Endophytic diversity was analyzed based on sequencing the ITS1-5.8S-ITS2 region of rDNA, allowing the identification of fungi belonging to genera including Cochliobolus, Bipolaris, Fusarium, Alternaria, Diaporthe, Phoma and Phomopsis. Phylogenetic analysis confirmed the identity of the endophytes. The biotechnological potential of endophytes was tested in vitro for the control of pathogenic fungi of grapevines including Alternaria sp., Sphaceloma sp. and Glomerella sp. Inhibition percentages above 50 % as demonstrated by some isolates demonstrate their potential for biological control.

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