Streptococcus sinensis sp. nov., a Novel Species Isolated from a Patient with Infective Endocarditis

Abstract

A bacterium was isolated from the blood culture of a patient with infective endocarditis. The cells were facultative anaerobic, nonsporulating, gram-positive cocci arranged in chains. The bacterium grows on sheep blood agar as alpha-hemolytic, gray colonies of 0.5 to 1 mm in diameter after 24 h of incubation at 37°C in ambient air. Growth also occurs in 10 or 40% bile and on bile esculin agar but not in 6% NaCl. No enhancement of growth is observed in 5% CO 2 . It is nongroupable with Lancefield groups A, B, C, D, F, or G antisera and is resistant to optochin and bacitracin. The organism is aflagellated and is nonmotile at both 25 and 37°C. It is Voges-Proskauer test positive. It produces leucine arylamidase and ␤-glucosidase but not catalase, urease, lysine decarboxylase, or ornithine decarboxylase. It hydrolyzes esculin and arginine. It utilizes glucose, lactose, salicin, sucrose, pullulan, trehalose, cellobiose, hemicellulase, mannose, maltose, and starch. 16S rRNA gene sequencing showed that there were 3.6, 3.7, 4.3, 4.7, and 5.9% differences between the 16S rRNA gene sequence of the bacterium and those of Streptococcus gordonii, Streptococcus intermedius, Streptococcus constellatus, Streptococcus sanguis, and Streptococcus anginosus, respectively. The G؉C content of it (mean ؎ standard deviation) was 53.0% ؎ 2.9%. Based on phylogenetic affiliation, it belongs to the mitis or anginosus group of Streptococcus. For these reasons a new species, Streptococcus sinensis sp. nov., is proposed, for which HKU4 is the type strain. Further studies should be performed to ascertain the potential of this bacterium to become an emerging cause of infective endocarditis.Since the discovery of PCR and DNA sequencing, comparison of the gene sequences of bacterial species showed that the 16S rRNA gene is highly conserved within a species and among species of the same genus, and hence it can be used as the new gold standard for speciation of bacteria. Using this new standard, phylogenetic trees, based on base differences between species, are constructed, and bacteria are classified and reclassified into new genera (9, 10). Furthermore, noncultivable organisms and organisms with ambiguous biochemical profiles can be classified and identified (11,12). Recently we have reported the use of this technique for the identification of bacterial strains with ambiguous biochemical profiles (14,15,16,19,23), species that are rarely encountered clinically (6,18,20), and a bacterium that is noncultivable (2); discovery of a novel clinical syndrome (22) and a novel genus and species (24); and characterization of beta-hemolytic Lancefield group G streptococcal bacteremia (21).In this study, we report the isolation of a bacterial strain from the blood culture of a patient with infective endocarditis. The strain, named HKU4, exhibited phenotypic characteristics that do not fit into patterns of any known species. 16S rRNA gene sequencing showed that there was only 96.4% base identity between the 16S rRNA gene of HKU4 and that of the most closely r...

Anaerospora hongkongensis Gen. Nov. Sp. Nov., a Novel Genus and Species with Ribosomal DNA Operon Heterogeneity Isolated from an Intravenous Drug Abuser with Pseudobacteremia

“…With the whole genome sequencing data of more than 100 bacteria and archaea available, it has been noted that there are relatively small interoperon heterogeneities for 16S rDNAs. In our experience in 16S rDNA sequencing, both for the known species and novel genera and species, direct sequencing of the PCR products has always been successful (16,(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)30). In this study, when we first amplified the 16S rDNA of HKU15 T and sequenced the PCR product directly, it was repeatedly observed that the 5' end of the PCR product could not be sequenced clearly, with the presence of multiple peaks.…”

“…Bacterial DNA extraction was performed according to our previous publications (24,30). Briefly, 80 µl of NaOH (0.05 M) was added to 20 µl of bacterial cells suspended in distilled water and the mixture was incubated at 60 C for 45 min, followed by addition of 6 µl of Tris-HCl (pH 7.0), achieving a final pH of 8.0.…”

“…Preparation of genomic DNA and determination of the GϩC content were performed according to our previous publications (24,27). Briefly, the temperature of the genomic DNA in SSC (0.15 M NaCl with 0.015 M sodium citrate) buffer (25 µg/ml) was increased slowly (0.5 C/min) from 25 C and the absorbance of the solution at 260 nm was monitored continuously against a blank containing SSC buffer only.…”

“…PCR amplification and DNA sequencing of the 16S rDNA was performed according to our previous publications (19,24). Briefly, DNase I-treated distilled water and PCR master mix [which contains deoxynucleoside triphosphates (dNTPs), PCR buffer, and Taq polymerase] were used in all PCR reactions by adding 1 U of DNase I (Pharmacia, Sweden) to 40 µl of distilled water or PCR master mix, incubating the mixture at 25 C for 15 min, and subsequently at 95 C for 10 min to inactivate the DNase I.…”

Use of Phylogenetic and Phenotypic Analyses To Identify Nonhemolytic Streptococci Isolated from Bacteremic Patients

“…The inherent problem of this approach is the large number of species relative to the limited number of biochemical traits that can be analyzed, the variability of several traits within species (33,35,36,44), the poor reproducibility of some tests (12,17,26,36,44), and the lack of sufficient phenotypic data on more recently described species in the underlying databases. The last problem applies to the species S. cristatus (23), S. peroris, S. infantis (31), S. australis (55), S. sinensis (57), Streptococcus macedonicus (51), Streptococcus infantarius, Streptococcus lutetiensis, Streptococcus gallolyticus (42), and S. pseudopneumoniae (1).…”