ORIGINAL PAPER Characterization of root-nodule bacteria isolated from Vicia faba and selection of plant growth promoting isolates Sabrine Saı ¨di • Samir Chebil • Maher Gtari • Ridha Mhamdi Received: 28 November 2012 / Accepted: 30 January 2013 / Published online: 9 February 2013 Ó Springer Science+Business Media Dordrecht 2013 Abstract A collection of 104 isolates from root-nodules of Vicia faba was submitted to 16S rRNA PCR–RFLP typing. A representative sample was further submitted to sequence analysis of 16S rRNA. Isolates were assigned to 12 genera. All the nodulating isolates (45 %) were closely related to Rhizobium leguminosarum USDA2370 T (99.34 %). The remaining isolates, including potential human pathogens, failed to nodulate their original host. They were checked for presence of symbiotic genes, P-solubilization, phytohormone and siderophore produc- tion, and then tested for their growth promoting abilities. Results indicated that 9 strains could induce significant increase (41–71 %) in shoot dry yield of faba bean. A Pseudomonas strain was further assessed in on-farm trial in combination with a selected rhizobial strain. This work indicated that nodule-associated bacteria could be a valu- able pool for selection of effective plant growth promoting isolates. Nevertheless, the possible involvement of nodules in increasing risks related to pathogenic bacteria should not be neglected and needs to be investigated further. Keywords Nodule endophytes Á Inoculation Á PGPR Á Rhizobia Á Vicia faba Introduction Although root nodules represent a minor fraction of the rhizosphere, they are a wealthy interface for microbial resources and molecular connections. It was admitted for longtime that root-nodule occupation was restricted to rhizobia. However, an increasing number of a- b- and c- Proteobacteria have been isolated from root nodules of a wide range of legumes and reported as nodulating bacteria (Moulin et al. 2001; Sy et al. 2001; Rivas et al. 2002; van Berkum and Eardly 2002; Vandamme et al. 2002; Valverde et al. 2005; Lin et al. 2008) or nodule-associated bacteria (Sturz et al. 1997; Gao et al. 2001; Zakhia et al. 2006; Kan et al. 2007). The non-symbiotic nodule endophytes have been poorly studied compared to symbiotic bacteria. Rhi- zobia were also frequently isolated from roots of non- legume plants and consequently could also be included as non-symbiotic endophytes (Antoun and Pre ´vost 2005). Generally, endophytes are defined as microorganisms able to successfully colonize tissues of higher plants and cause unapparent and asymptomatic infections. They may establish neutral, detrimental or beneficial interactions with plants. Rhizobacteria that exert beneficial effects on plant development are termed plant growth-promoting rhizo- bacteria (PGPR). Although the mechanisms by which they promote plant growth are not yet fully understood, multiple modes of action were revealed to be responsible for growth promotion activities. They include direct and indirect mechanisms. Direct effects result from their ability to fix nitrogen, to increase nutrient uptake through solubilization of minerals, or to produce siderophores and phytohormones Electronic supplementary material The online version of this article (doi:10.1007/s11274-013-1278-4) contains supplementary material, which is available to authorized users. S. Saı ¨di Á R. Mhamdi (&) Laboratory of Legumes, Centre of Biotechnology of Borj-Ce ´dria, BP 901, 2050 Hammam-Lif, Tunisia e-mail: ridhamhamdi@yahoo.fr; ridha.mhamdi@cbbc.rnrt.tn S. Chebil Laboratory of Molecular Physiology of Plants, Centre of Biotechnology of Borj-Ce ´dria, Hammam-Lif, Tunisia M. Gtari Laboratory of Microorganisms and Active Biomolecules, Faculty of Sciences of Tunis, University of Tunis El-Manar, Tunis, Tunisia 123 World J Microbiol Biotechnol (2013) 29:1099–1106 DOI 10.1007/s11274-013-1278-4