Hoeflea anabaenae sp nov., an epiphytic symbiont that attaches to the heterocysts of a strain of Anabaena

The heterotrophic, epiphytic, symbiotic bacterial strain WH2K(T) was previously isolated from a two-member culture in which it was attached to the heterocysts of a strain of Anabaena (SSM-00). Analysis of its 16S rRNA gene sequence demonstrated that the symbiont was most closely related to the type strain of Hoeflea marina (96.9% similarity), which belongs to the family Phyllobacteriaceae within the order Rhizobiales of the class Alphaproteobacteria. A polyphasic taxonomic study was performed on strain WH2K(T), which consisted of irregular rods (2-5 mu m long, 0.2 mu m wide) that appeared to be narrower at one pole. Optimal growth was obtained in complex media with 15 g sea salts l(-1), at 18-34 degrees C (30 degrees C optimum) and at pH 6.0-8.0 (optimum pH 6.5). Unknown growth requirements were provided by small amounts of yeast extract but not by standard vitamin and trace metal solutions. Of the substrates tested, WH2K(T) was able to utilize only acetate, pyruvate, malate and fumarate. Growth was observed only under aerobic and microaerobic conditions, and nitrate was not reduced. No photosynthetic pigments were detected under any of the growth conditions tested. The predominant fatty acids were a summed feature that comprises C(18:1)omega 7c, C(18:1)omega 9t, C(18:1)omega 12t or any combination of these (64.0%) and an unidentified fatty acid of equivalent chain length 17.603 (13.5%). The polar lipid profile consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, phosphatidylcholine, phosphoglycolipid, unknown lipids and an unidentified aminolipid. The only respiratory ubiquinone detected was Q-10. The DNA G+C content of the strain was 58.1 mol%. The organism can form a site-specific attached symbiotic relationship with a species of Anabaena. Based on phylogenetic and phenotypic evidence, it is proposed that strain WH2K(T) be classified within a novel species of the genus Hoeflea, for which the name Hoeflea anabaenae sp. nov. is proposed. The type strain is WH2K(T) (=CCUG 56626(T) =NRRL B-59520(T)).