Méndez-Morales, Emanuel

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    Diversity of Actinobacteria associated with Nasutitermes costalis termite and their ability to inhibit the growth of entomopathogenic fungi
    (2013) Méndez-Morales, Emanuel; Cafaro, Matías J.; College of Arts and Sciences - Sciences; Ríos Hernández, Luis A.; Montalvo Rodríguez, Rafael; Department of Biology; López, Martha L.
    The Actinobacteria are Gram-positive bacteria with high GC content, spore producers, and have the ability to produce antimicrobial agents. They are capable of degrading structural polysaccharides such as cellulose, lignocellulose and chitin. Previous studies have found this group of bacteria associated with different groups of insects including ants, bees, wasps, beetles, and termites. Actinobacteria have two primary roles in these associations: (1) defend the host from pathogenic organisms and (2) decompose organic matter in the gut. However, is known that the genus Streptomyces (Actinobacteria) is naturally selected to perform these symbiotic relationships. In Attine ants and Beewolf wasps, Actinobacteria protect them against pathogenic microbes. At present it is unknown which Actinobacteria could be associated with termites and if they possess the ability of inhibiting the micelial growth of entomopathogenic fungus Metarhizium anisopliae and Beauveria bassiana, which also affect termites. On the other hand, the termite gut has been extensively explored. In termites gut have been identified Actinobacteria of the genus Streptomyces are believed to help degrade organic matter. In this study we characterized the diversity of Actinobacteria associated with Nasutitermes costalis, a common arboreal termite in Puerto Rico, and evaluated their antifungal capacity against Metarhizium anisopliae and Beauveria bassiana. Two castes of termites (workers and soldiers) and nest material were collected in Miradero forest in Mayagüez, Puerto Rico. We used culture-dependent methods (chitin agar, YMEA) for isolation of Actinobacteria from the nest material, the gut, and the termites exoskeleton . Morphological and molecular techniques (sequencing of 16S rDNA gene) were later used for identification. We isolated 1342 Actinobacteria represented by the genera Streptomyces, Amycolatopsis, Lentzea, Saccharothrix, Pseudonocardia, Microbacterium, and Gordonia. Sequence analysis revealed that the most common genus found in this association is Streptomyces from 73% to 97% of frequency among samples and presents great morphological variation. Phylogenetic analysis shows 15 monophyletic clades that separate the Streptomyces sequences isolated from N. constalis with no match to GenBank database. The most frequent and constant strains in the nest material and exoskeleton of termites were closely related to Streptomyces cavourensis (GenBank, HQ610450.1). Strains closely related to Streptomyces cavourensis (HQ610450.1) were not found in nests abandoned by termites. All selected strains were able to inhibit the growth of M. anisopliae and B. bassiana, but 73% of them were effective in inhibiting micelial growth of B. bassiana from 90% to 100%. The strain W1OE185[6] closely related to Streptomyces cavourensis (HQ610450.1) and the second most abundant in worker exoskeletons was the highest ability to inhibit both fungi. The results indicate that the 15 clades of Streptomyces can be unique in termites, but another gene should be used to confirm this hypothesis. Our results support the existence of Streptomyces symbionts in the system of the termite N. costalis and that one of its roles is the production of antifungal agents.