Analysis of the Interaction between Pisum sativum L. and Rhizobium laguerreae Strains Nodulating This Legume in Northwest Spain.

Pisum sativum L. (pea) is one of the most cultivated grain legumes in European countries due to the high protein content of its seeds. Nevertheless, the rhizobial microsymbionts of this legume have been scarcely studied in these countries. In this work, we analyzed the rhizobial strains nodulating the pea in a region from Northwestern Spain, where this legume is widely cultivated. The isolated strains were genetically diverse, and the phylogenetic analysis of core and symbiotic genes showed that these strains belong to different clusters related to R. laguerreae sv. viciae. Representative strains of these clusters were able to produce cellulose and cellulases, which are two key molecules in the legume infection process. They formed biofilms and produced acyl-homoserine lactones (AHLs), which are involved in the quorum sensing regulation process. They also exhibited several plant growth promotion mechanisms, including phosphate solubilization, siderophore, and indole acetic acid production and symbiotic atmospheric nitrogen fixation. All strains showed high symbiotic efficiency on pea plants, indicating that strains of R. laguerreae sv. viciae are promising candidates for the biofertilization of this legume worldwide.

Paenibacillus endophyticus sp. nov., isolated from nodules of Cicer arietinum.

A bacterial strain, designated PECAE04(T), was isolated from root nodules of Cicer arietinum in Spain. Phylogenetic analysis based on 16S rRNA gene sequence placed the isolate into the genus Paenibacillus with its closest relative being Paenibacillus castaneae Ch-32(T) with 98.4 % 16S rRNA gene sequence similarity followed by Paenibacillus glycanilyticus DS-1(T), Paenibacillus prosopidis PW21(T), Paenibacillus xinjiangensis B538(T) and Paenibacillus catalpae D75(T) with similarities ranging from 97.9 to 96.8 %. DNA-DNA hybridization measurements showed values lower than 20 % between the strain PECAE04(T) and any of these species. The isolate was a Gram-stain-positive, motile, sporulating rod. Catalase and oxidase activities were positive. Aesculin was hydrolysed but casein and gelatin were not. Acetoin production, H2S production, nitrate reduction and urease and caseinase production were negative. Growth was supported by many carbohydrates and organic acids as carbon sources. MK-7 was the predominant menaquinone and anteiso-C15 : 0, iso-C16 : 0 and C16 : 0 were the major fatty acids. Major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, a glycolipid, three phospholipids and an unidentified lipid. Meso-diaminopimelic acid was not detected in the peptidoglycan. The DNA G+C content was 52.9 mol%. Phylogenetic, chemotaxonomic and phenotypic analyses showed that strain PECAE04(T) should be considered to be a representative of a novel species of the genus Paenibacillus, for which the name Paenibacillus endophyticus sp. nov. is proposed. The type strain is PECAE04(T) ( = LMG 27297(T) = CECT 8234(T)).

Phyllobacterium endophyticum sp. nov., isolated from nodules of Phaseolus vulgaris.

A strain, PEPV15(T), was isolated from a nodule on Phaseolus vulgaris grown in soil in northern Spain. Phylogenetic analyses of 16S rRNA and atpD genes showed that this strain belongs to the genus Phyllobacterium. The most closely related species were, in both cases, Phyllobacterium brassicacearum, Phyllobacterium bourgognense and Phyllobacterium trifolii, the type strains of which gave sequence similarities of 98.9, 98.6 and 98.4 %, respectively, in the 16S rRNA gene and 88.1, 87.5 and 88.7 %, respectively, in the atpD gene. PEPV15(T) contained Q-10 as the major quinone (88 %) and low amounts of Q-9 (12 %). It differed from its closest relatives in its growth in diverse culture conditions and in the assimilation of several carbon sources. The strain was not able to produce nodules in Phaseolus vulgaris. The results of DNA-DNA hybridization, phenotypic tests and fatty acid analyses confirmed that this strain represents a novel species of the genus Phyllobacterium for which the name Phyllobacterium endophyticum sp. nov. is proposed; the type strain is PEPV15(T) ( = LMG 26470(T) = CECT 7949(T)). An emended description of the genus Phyllobacterium is also provided.