INVERTEBRATE MICROBIOLOGY Bacterial Community and Nitrogen Fixation in the Red Turpentine Beetle, Dendroctonus valens LeConte (Coleoptera: Curculionidae: Scolytinae) Jesús Morales-Jiménez & Gerardo Zúñiga & Lourdes Villa-Tanaca & César Hernández-Rodríguez Received: 23 April 2009 / Accepted: 27 May 2009 / Published online: 20 June 2009 # Springer Science + Business Media, LLC 2009 Abstract The red turpentine beetle, Dendroctonus valens LeConte (Coleoptera: Curculionidae: Scolytinae), colonizes all pines species within its native range throughout North and Central America. Recently, this species was acciden- tally introduced to China, where it has caused severe damage in pine forests. It belongs to a group of beetles that spend most of their lives between the tree bark and sapwood, where it feeds on phloem: a poor substrate with very low nutritional value of nitrogen and toxic properties due to its high content of secondary defensive compounds. The aim of this study was to characterize the bacterial community of the D. valens gut by culture-dependent and - independent methods. Polymerase chain reaction denatur- ing gradient gel electrophoresis and ribosomal gene library analyses revealed that species diversity in the D. valens gut was relatively low, containing between six and 17 bacterial species. The bacterial community associated with larvae and adults was dominated by members of the following genera: Lactococcus, Acinetobacter, Pantoea, Rahnella, Stenothrophomonas, Erwinia, Enterobacter, Serratia, Jan- ibacter, Leifsonia, Cellulomonas, and Cellulosimicrobium. The members of the last four genera showed cellulolytic activity in vitro and could be involved in cellulose breakdown in the insect gut. Finally, nitrogen fixation was demonstrated in live larvae and adults; however, capacity of nitrogen fixing in vitro was not found among enterobacte- rial species isolated in nitrogen-free media; neither were nifD nor nifH genes detected. In contrast, nifD gen was detected in metagenomic DNA from insect guts. The identification of bacterial species and their potential physiological capacities will allow exploring the role of gut symbiotic bacteria in the adaptation and survival of D. valens in a harsh chemical habitat poor in nitrogen sources. Introduction Insects are one of the most successful and diverse groups on earth; they are adapted to a wide variety of diets and live in practically any habitat. Bark beetles have different life styles ranging from highly aggressive (tree killing), facultative (colonizing weak or recently killed trees), parasitic (using living trees), to saprophagous (using dead hosts) [50]. In general, they tunnel through the bark to construct galleries in the phloem and cambial layers upon which they feed [74]. The phloem is a substrate rich in cellulose (22.1% of dry and ethanol-benzene-insoluble material), hemicellulose (15% of dry and ethanol-benzene-insoluble material), and soluble carbohydrates (120–200 mg g −1 ) but poor in assimilable nitrogen (11–13 mg g −1 of dry weight) [66, 70, 72]. This situation requires bark beetle species to have a high phloem consumption rate and to establish symbiotic relationships with microorganisms that supply assimilable carbon and nitrogen sources and growth factors [3, 5]. The microorganisms associated with the insect gut could be Microb Ecol (2009) 58:879–891 DOI 10.1007/s00248-009-9548-2 J. Morales-Jiménez : L. Villa-Tanaca : C. Hernández-Rodríguez (*) Departamento de Microbiología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Prol. De Carpio y Plan de Ayala. Col. Sto. Tomas, Mexico, Distrito Federal CP 11340, Mexico e-mail: chdez38@hotmail.com G. Zúñiga Departamento de Zoología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, Prol. De Carpio y Plan de Ayala. Col. Sto. Tomas, Mexico, Distrito Federal CP 11340, Mexico