Herbaspirillum hiltneri sp. nov., isolated from surface-sterilized wheat roots

The genus Herbaspirillum of the Betaproteobacteria mainly comprises diazotrophic bacteria with a potential for endophytic and systemic colonization of a variety of plants. The plant-associated bacterial isolates N3(T), N5 and N9 were derived from surface-sterilized wheat roots. After phylogenetic analysis of 16S rRNA gene sequence data the isolates could be allocated to the genus Herbaspirillum, and 99.9 % similarity to the sequence of Herbaspirillum lusitanum P6-12(T) was found. A set of 16S rRNA gene-targeted oligonucleotide probes was developed for the identification of the three novel isolates and H. lusitanum (Hhilu446), and for the specific detection of several other Herbaspirillum species described recently. For higher phylogenetic resolution, the 23S rRNA gene sequences of all members of the genus was sequenced and used to construct a phylogenetic tree. Isolates N3(T), N5 and N9 formed a group that was distinct from all other Herbaspirillum species. In addition, isolate N3(T) and H. lusitanum P6-12(T) exhibited a DNA-DNA hybridization value of only 25 %. The value for DNA-DNA hybridization between N3(T) and other members of the genus Herbaspirillum was between 14 and 32 %; DNA-DNA hybridization between strain N3(T) and isolates N5 and N9 produced values above 95 %. This places the three isolates as representatives of a novel species within the genus Herbaspirillum. A Biolog GN2 assay supported this conclusion. The major fatty acids were C(16 : 1)omega7c, C(16 : 0) and C(18 : 1)omega7c, and the DNA G+C content ranged from 60.9 to 61.5 mol%. Therefore these three isolates should be classified within a novel species, for which the name Herbaspirillum hiltneri sp. nov. is proposed. The type strain is N3(T) (=DSM 17495(T)=LMG 23131(T)).