Rhizobium cellulosilyticum sp. nov., isolated from sawdust of Populus alba

Abstract

During a study of polysaccharide-hydrolysing bacteria present in different plant sources, two strains were isolated from pulverized decaying wood of Populus alba and classified in the genus Rhizobium on basis of their almost complete 16S rRNA gene sequences. Their closest phylogenetic relatives were Rhizobium galegae USDA 4128T and Rhizobium huautlense S02 T , with 98.2 and 98.1 % 16S rRNA gene sequence similarity, respectively. recA and atpD sequence analysis showed that these species have less than 88 and 92 % similarity, respectively, to the novel strains. In contrast to their closest phylogenetic relatives, the two strains showed strong cellulase activity on plates containing CM-cellulose as a carbon source. They were also distinguishable from these species on the basis of other phenotypic characteristics. The strains were able to induce ineffective nodules on Medicago sativa and the sequence of their nodD gene was phylogenetically close to that of Ensifer meliloti 1021 (99.6 % similarity). DNA-DNA hybridization values ranged from 10 to 22 % with respect to R. galegae USDA 4128T and 14 to 25 % with respect to R. huautlense S02 T , showing that the strains from this study belong to a novel species, for which the name Rhizobium cellulosilyticum sp. nov. is proposed. The type strain is ALA10B2 T (=LMG 23642The species of the genus Rhizobium are traditionally considered as legume endosymbionts and, although Rhizobium daejeonense was isolated from a cyanide-treatment bioreactor (Quan et al., 2005), most of them have been isolated from nodules. Decaying wood is a good source of polysaccharide-hydrolysing micro-organisms, and we have previously isolated several species, mainly actinomycetes, able to hydrolyse xylan and cellulose from similar sources (Rivas et al., 2003(Rivas et al., , 2004a. In this work, from pulverized decaying wood of Populus alba, we isolated two Gram-negative strains, ALA10B2 T and ALA38.2, that actively hydrolyse CM-cellulose in vitro. The 16S rRNA gene sequences of these strains allowed their classification in the genus Rhizobium, near to Rhizobium galegae and Rhizobium huautlense, two species that do not produce cellulases under the conditions used in this study. A polyphasic study of these strains, including phenotypic and molecular taxonomic approaches, showed that they belong to a novel species of the genus Rhizobium.For isolation of strains ALA10B2 T and ALA38.2, a sample of pulverized decaying wood was collected aseptically from a cavity in the trunk of a healthy Populus alba tree in Salamanca, Spain. From this sample, 1 g was ground, placed in 9 ml sterile water and stirred for 60 min. Aqueous portions (100 ml of the mixture) were spread on CEA medium containing 0.7 % CM-cellulose as the only carbon source, 0.3 % yeast extract and 2.5 % agar. Colonies were picked from these plates and inoculated on the same medium. Cellulase activity was detected after 5 days incubation at 28 uC after staining with a 1 % aqueous Congo red solution. Two bacterial strains, ALA10B2T and ALA38.2, showing ...

Rhizobium miluonense sp. nov., a symbiotic bacterium isolated from Lespedeza root nodules

“…3). Taking the recently reported Rhizobium species as references (García-Fraile et al, 2007;Quan et al, 2005;Valverde et al, 2006), the strains in group 7 could be defined as a novel species of the genus Rhizobium based on the results presented in this study and in our previous work (Gu et al, 2007); the name proposed for this species is Rhizobium miluonense sp. nov. Optimum growth temperature is 25-30 u C; can grow at 37 u C, but not 4 u C. Optimum pH is 6-8.…”

“…Among these genera, Rhizobium was the first described species, named 100 years ago. After a series of taxonomic changes since the 1980s, the genus Rhizobium now contains about 20 species, including three recently described species: Rhizobium daejeonense (Quan et al, 2005), Rhizobium lusitanum (Valverde et al, 2006) and Rhizobium cellulosilyticum (García-Fraile et al, 2007).…”

Rhizobium pseudoryzae sp. nov., isolated from the rhizosphere of rice

“…+, Positive; 2, negative; w, weakly positive. Partial recA (530 bp), atpD (533 bp), gyrB (756 bp) and glnII (552 bp) gene sequences were also determined for the new isolate by using the methods and primers reported by Martens et al (2007). Analysis of these housekeeping gene sequences supported the separate phylogenetic position of the novel strain within the genus Rhizobium.…”

Rhizobium tibeticum sp. nov., a symbiotic bacterium isolated from Trigonella archiducis-nicolai (Sirj.) Vassilcz.

“…The current genus Rhizobium includes about 30 species of fast-growing, acid-producing rhizobia, which can be differentiated from Sinorhizobium species on the basis of 16S rRNA gene sequence-based phylogeny. With the exception of Rhizobium cellulosilyticus (Garcia-Fraile et al, 2007), most of them can induce root nodules on a certain range of legume species.In a survey of rhizobial resources in Tibet, 12 strains of root-nodule bacteria were isolated with yeast mannitol agar (YMA) medium according to the method of Vincent (1970) from root nodules of Trigonella archiducis-nicolai (Širj.) Vassilcz., an ephemeral legume growing in grasslands at altitudes of 3700-4000 m. Six of them were identified as Sinorhizobium meliloti, and six as Rhizobium strains (Hou et al, 2009).…”

“…The current genus Rhizobium includes about 30 species of fast-growing, acid-producing rhizobia, which can be differentiated from Sinorhizobium species on the basis of 16S rRNA gene sequence-based phylogeny. With the exception of Rhizobium cellulosilyticus (Garcia-Fraile et al, 2007), most of them can induce root nodules on a certain range of legume species.…”