A Gram-negative, rod-shaped bacterium, designated H63T, was isolated from aortic valve cells of an individual with native valve endocarditis. period of composing, the genus Tbx1 comprised 54 species with validly 1037624-75-1 published brands and one genomospecies (Benson stress (Pearce or a novel species (Pearce was initially isolated from cooling tower drinking water in Czechoslovakia and provides been isolated once from a case of Legionnaires disease in European countries (Ricketts species. Phenotypic profiling revealed a number of differences between H63T and its phylogenetically nearest neighbours. Furthermore, H63T replicated in a human macrophage cell line and in the murine lung, indicating that H63T represents a virulent strain. To establish the placement of the novel strain in the genus on the basis of 16S rRNA gene sequence similarity as recommended for the calculation of pairwise percentage similarity values (Chun (98.80?%), followed by (97.03?%), (96.76?%), (96.20?%), (95.99?%), (95.73?%), (95.72?%), (95.71?%) and (95.71?%) (Table 1). To determine whether H63T represents a novel species, DNACDNA hybridization was performed at 30 C using the microplate technique with photobiotin-labelled DNA as described previously (Ezaki ATCC 43878T, in reciprocal associations, well below the 70?% cut-off for species delineation (Table 1). Furthermore, H63T exhibited less than 20?% relatedness 1037624-75-1 to all eight of the remaining type strains tested (Table 1). The difference between reciprocal hybridizations was within 20?% and the standard deviation among replicates was 7?%, both of which are acceptable deviations for the microplate technique (Ezaki sp. nov. H63TClinicalPearce (2011)(100)(100)(100)41.8ATCC 43878TEnvironmentalWilkinson (1988)98.817.35.820.37.240.5ATCC 49505TEnvironmentalDennis (1993)97.05.72.42.41.143.0ATCC 33623TEnvironmentalCherry (1982)96.815.71.84.53.045.0ATCC 35303TEnvironmentalBrenner (1985)96.24.30.67.56.651.0NCTC 13409TEnvironmentalLck (2010)96.05.00.48.72.642.5ATCC 35304TEnvironmentalBrenner (1985)95.76.22.69.34.752.0ATCC 35072TEnvironmentalHerwaldt (1984)95.73.72.26.42.846.0ATCC 33152TClinicalBrenner (1979)95.75.32.315.15.739.0ATCC 43702TClinicalWilkinson (1987)95.76.32.04.52.642.9 Open in a separate window *Meanssd of at least triplicate hybridization experiments are shown. To define the relationship between H63T and other species further, a 584 bp portion of the gene and a 327 bp portion of the gene of H63T were sequenced as described previously (Kuroki Infections (EWGLI) gene sequence database was used to determine the similarity based on (Altschul gene sequence of strain H63T was most similar to that of ATCC 43878T (85.49?%), followed by ATCC 35250T (85.11?%), ATCC 35298T (85.11?%), ATCC 35072T (83.95?%) and ATCC 49751T (83.56?%). Based on analysis of sequences, strain H63T was again most similar to the type strain of (91.2?%), followed by the type strains of (89.4?%), (89.7?%), (88.3?%) and (88.3?%). For phylogenetic analyses, the 16S rRNA, and sequences of type strains of species and the nearest other relative within the and gene sequences indicated that strain H63T is usually most closely related to and (Fig. 1). The strength 1037624-75-1 of the association was confirmed by bootstrap values 80 based on 100 replicates. For completeness, DNACDNA hybridizations were performed comparing H63T with both ATCC 35250T and ATCC 35298T, because they were closely related according to the consensus tree. According to hybridization analysis, ATCC 35250T was 8.0?% (0.9) related to H63T when H63T DNA served as the probe and 31.7?% (0.4) similar when H63T represented the covalent DNA. ATCC 35298T was 10.1?% (1.2) similar to H63T when H63T DNA was the probe and 7.7?% (3.3) similar when H63T DNA was the covalent DNA. The topologies of the individual gene trees support the consensus assignment of H63T and as sister taxa (Figs S1CS3, available in IJSEM Online). Open in a separate window Fig. 1. Neighbour-joining tree showing relationships between strain H63T and all previously sequenced type strains of species based on the consensus sequence of the 16S rRNA, and loci. Bootstrap values greater.