African Journal of Microbiology Research Vol. 7(7), pp. 586-594, 12 February, 2013 Available online at http://www.academicjournals.org/AJMR DOI: 10.5897/AJMR12.2036 ISSN 1996-0808 ©2013 Academic Journals Full Length Research Paper Diversity of endophytic bacteria in the fruits of Coffea canephora Paulo Sérgio Balbino Miguel, Julio Cesar Delvaux, Marcelo Nagem Valério de Oliveira, Larissa Cassemiro Pacheco Monteiro, Fernanda de Souza Freitas, Maurício Dutra Costa, Marcos Rogério Tótola, Célia Alencar de Moraes and Arnaldo Chaer Borges* Departamento de Microbiologia /Instituto de Biotecnologia Aplicada à Agropecuária, Universidade Federal de Viçosa, Viçosa, Minas Gerais, 36570-000, Brazil. Accepted 8 February, 2013 Endophytic bacteria colonize the internal tissues of plants without causing infection or negative effects on their hosts. This study investigates the occurrence and diversity of culturable endophytic bacteria in the fruits of Coffea canephora at three developmental stages. Isolation and quantification were performed in R2A culture medium, and the diversity was established using molecular methods and analysis of fatty acid methyl esters (FAME). α- and γ-Proteobacteria, Actinobacteria, Firmicutes, and Bacteroidetes were identified in the investigated community. Kocuria turfanensis and Pantoea vagans were identified as endophytes for the first time. Of the 18 species that were found, the following seven had not been previously described as endophytic in coffee fruits: Bacillus thuringiensis, Bacillus licheniformis, Agrobacterium tumefaciens, Escherichia coli, Enterobacter hormaechei, Chryseobacterium sp., and Ochrobactrum sp. The diversity of endophytic bacteria varied during the three developmental stages that were investigated, and the diversity was greatest in fruits during the green stage, during which Bacillus subtilis predominated. The number of Gram-positive bacteria was larger than the number of Gram-negative bacteria during the two earliest developmental stages, whereas their numbers were similar during the ripe stage. The diversity was lowest during the ripe stage, and Klebsiella oxytoca was the predominant species at this stage, probably due to the higher caffeine and sugar contents in the fruits. Key words: Coffee, bacterial community, sequencing, 16S rDNA, FAME. INTRODUCTION The economic and social importance of coffee can be assessed using data provided by the International Coffee Organization, which estimates that 75 to 125 million individuals worldwide are dependent on coffee-related activities. In Brazil, the coffee production chain generates more than eight million jobs (MAPA, 2012). Coffee plants belong to the family Rubiaceae, which comprises approximately 650 genera and more than 13,000 species (Delprete and Jardim, 2012). The most widely cultivated are Coffea arabica, which produces superior-quality coffee beverages due to its organoleptic *Corresponding author. E-mail: chaer@ufv.br. Tel: +55(31) 3899 2976. Fax: +55(31) 3899 2573. properties (Judd et al., 2008), and Coffea canephora, which produces neutral, weak-flavored, and very bitter coffee beverages (Ky et al., 2001). The raw material for the instant coffee industry is derived from C. canephora, and this species was used in 50 to 55% of the blends manufactured in Brazil in 2011, which represents a 20% increase compared to 2000 (ABIC, 2012). The Brazilian coffee research program is the largest in the world, and investments in this research have produced significant advances in breeding, plant nutrition, biotechnology, and other areas (MAPA, 2012). Recently, the map of the C. arabica genome was made available for functional genomics studies (Vieira et al., 2006). Although the International Coffee Genome Network - ICGN is currently mapping the genome of C. canephora, no studies investigating the colonization of its fruits by endophytes were