Suborder Corynebacterineae Research Articles

Amycolicicoccus subflavus gen. nov., sp. nov., an actinomycete isolated from a saline soil contaminated by crude oil

Two novel actinomycetes, designated DQS3-9A1(T) and DQS3-9A2, were isolated from a saline soil contaminated with crude oil in the Shengli Oilfield in China. On the basis of 16S rRNA gene sequence analysis, the two strains were most closely related to Mycobacterium species (92.7-94.9 % similarities), and formed a distinct lineage in the suborder Corynebacterineae . In addition, the major sugars in the cell wall, arabinose and galactose, supported the affiliation of strain DQS3-9A1(T) with members of the family Mycobacteriaceae. However, strain DQS3-9A1(T) did not contain mycolic acids and MK-8 (85.5 %) was the major menaquinone for both isolates. The major cellular fatty acids for strain DQS3-9A1(T) were C(16 : 0) (20.5 %), 10-methyl C(17 : 0) (19.3 %), 10-methyl C(18 : 0) (16.1 %), summed feature 3 (11.4 %), C(15 : 0) (11.3 %), C(17 : 0) (5.0 %) and C(17 : 1)omega8c (5.0 %). The polar lipids of strain DQS3-9A1(T) consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, phosphatidylinositol and an unknown glucosamine-containing phospholipid. These chemotaxonomic data indicated that strain DQS3-9A1(T) differs from the present members of the suborder Corynebacterineae. Therefore, the creation of Amycolicicoccus subflavus gen. nov., sp. nov. is proposed, with DQS3-9A1(T) (=DSM 45089(T)=CGMCC 4.3532(T)) as the type strain.

Hoyosella altamirensis gen. nov., sp. nov., a new member of the order Actinomycetales isolated from a cave biofilm

A novel actinomycete, strain OFN S31(T), was isolated from a complex biofilm in the Altamira Cave, Spain. A polyphasic study was carried out to clarify the taxonomic position of this strain. Phylogenetic analysis with 16S rRNA gene sequences of representatives of the genera Corynebacterium, Dietzia, Gordonia, Millisia, Mycobacterium, Nocardia, Rhodococcus, Segniliparus, Skermania, Tsukamurella and Williamsia indicated that strain OFN S31(T) formed a distinct taxon in the 16S rRNA gene tree that was more closely associated with the Mycobacterium clade. The type strain of Mycobacterium fallax was the closest relative of strain OFN S31(T) (95.6 % similarity). The cell wall contained meso-diaminopimelic acid, arabinose and galactose, which are characteristic components of cell-wall chemotype IV of actinomycetes. The sugars of the peptidoglycan were acetylated. The polar lipid pattern was composed of phosphatidylinositol, phosphatidylglycerol, phosphatidylethanolamine and diphosphatidylglycerol. Strain OFN S31(T) is characterized by the absence of mycelium and mycolic acids. Strain OFN S31(T) had MK-8 as the major menaquinone. The DNA G+C content was 49.3 mol%, the lowest found among all taxa included in the suborder Corynebacterineae. Based on morphological, chemotaxonomic, phenotypic and genetic characteristics, strain OFN S31(T) is considered to represent a novel species of a new genus, for which the name Hoyosella altamirensis gen. nov., sp. nov. is proposed. The type strain of Hoyosella altamirensis is strain OFN S31(T) (=CIP 109864(T) =DSM 45258(T)).

The Reductase That Catalyzes Mycolic Motif Synthesis Is Required for Efficient Attachment of Mycolic Acids to Arabinogalactan

Mycolic acids are essential components of the cell walls of bacteria belonging to the suborder Corynebacterineae, including the important human pathogens Mycobacterium tuberculosis and Mycobacterium leprae. Mycolic acid biosynthesis is complex and the target of several frontline antimycobacterial drugs. The condensation of two fatty acids to form a 2-alkyl-3-keto mycolate precursor and the subsequent reduction of this precursor represent two key and highly conserved steps in this pathway. Although the enzyme catalyzing the condensation step has recently been identified, little is known about the putative reductase. Using an extensive bioinformatic comparison of the genomes of M. tuberculosis and Corynebacterium glutamicum, we identified NCgl2385, the orthologue of Rv2509 in M. tuberculosis, as a potential reductase candidate. Deletion of the gene in C. glutamicum resulted in a slow growing strain that was deficient in arabinogalactan-linked mycolates and synthesized abnormal forms of the mycolate-containing glycolipids trehalose dicorynomycolate and trehalose monocorynomycolate. Analysis of the native and acetylated trehalose glycolipids by MALDI-TOF mass spectrometry indicated that these novel glycolipids contained an unreduced beta-keto ester. This was confirmed by analysis of sodium borodeuteride-reduced mycolic acids by gas chromatography mass spectrometry. Reintroduction of the NCgl2385 gene into the mutant restored the transfer of mature mycolic acids to both the trehalose glycolipids and cell wall arabinogalactan. These data indicate that NCgl2385, which we have designated CmrA, is essential for the production of mature trehalose mycolates and subsequent covalent attachment of mycolic acids onto the cell wall, thus representing a focus for future structural and pathogenicity studies.

Smaragdicoccus niigatensis gen. nov., sp. nov., a novel member of the suborder Corynebacterineae

A polyphasic taxonomic approach was applied to determine the taxonomic position of a hydrocarbon-degrading actinomycete, strain Hou_blueT, which was isolated from soil samples collected from an oil spring in Niigata, Japan. The results of 16S rRNA and gyrB gene sequence comparisons indicated that strain Hou_blueT represented a novel lineage in the suborder Corynebacterineae. Colonies were malachite green-like in colour on 1/10 trypticase soy agar and the cell morphology was coccoid in all growth phases. The cell-wall diamino acid and sugar indicated chemotype IV and variation A1gamma. The sugars of the peptidoglycan were glycolated. The polar lipids were composed of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannoside and some unspecified glycolipids. The organism contained two novel cyclic forms of menaquinone, smaragdiquinone A-8(H4, omega-cycl) and smaragdiquinone B-8(H4, dicycl). The major fatty acids were cis-9-18 : 1 (34.46 %) and 16 : 0 (25.1 %). Small amounts of 10-methyl-branched fatty acids were also present (10-methyl-17 : 0, 0.17 %), but not tuberculostearic acid (10-methyl-18 : 0), which has been shown to be present in all nocardiae. Gas-chromatographic analysis of the mycolic acid revealed a carbon-chain length of C43-C49. The DNA G+C content was 63.7 mol%. On the basis of phenotypic and phylogenetic distinctness, the organism is proposed to represent a novel genus and species, Smaragdicoccus niigatensis gen. nov., sp. nov., with the type strain Hou_blueT (=MBIC 06267T=DSM 44881T).

Gordonia defluvii sp. nov., an actinomycete isolated from activated sludge foam

Three strains of non-motile, Gram-positive, filamentous actinomycetes, isolates J4(T), J5 and J59, initially recognized microscopically in activated sludge foam by their distinctive branching patterns, were isolated by micromanipulation. The taxonomic positions of the isolates were determined using a polyphasic approach. Almost-complete 16S rRNA gene sequences of the isolates were aligned with corresponding sequences of representatives of the suborder Corynebacterineae and phylogenetic trees were inferred using three tree-making algorithms. The organisms formed a distinct phyletic line in the Gordonia 16S rRNA gene tree. The three isolates showed 16S rRNA gene sequence similarities within the range 96.9-97.2 % with their nearest phylogenetic neighbours, namely Gordonia bronchialis DSM 43247(T) and Gordonia terrae DSM 43249(T). Strain J4(T) was shown to have a chemotaxonomic profile typical of the genus Gordonia and was readily distinguished from representatives of the genus on the basis of Curie-point pyrolysis mass spectrometric data. The isolates shared nearly identical phenotypic profiles that distinguished them from representatives of the most closely related Gordonia species. It is evident from the genotypic and phenotypic data that the three isolates belong to a novel Gordonia species. The name proposed for this taxon is Gordonia defluvii sp. nov.; the type strain is J4(T) (=DSM 44981(T)=NCIMB 14149(T)).

Phylogenetic analysis of members of the metabolically diverse genus Gordonia based on proteins encoding the gyrB gene

Members of the metabolically diverse genus Gordonia were isolated from various biotopes including pristine and polluted sites around Taiwan. Identification, comparison and diversity assessment based on the gyrB gene were carried out using a newly developed primer pair for gyrB. The 16S rRNA gene was also sequenced for comparison. A 1.2-kb fragment of the gyrB gene of 17 Gordonia strains including type strains was determined by direct sequencing of PCR amplified fragments. A total of 25 strains (8 of which were retrieved from a public database) of the genus Gordonia form a distinct phyletic line in the GyrB-based tree and are separated from other closely related species of genera of the suborder Corynebacterineae. Sequence similarity of the gyrB sequence from twelve Gordonia type strains ranged from 79.3 to 97.2%, corresponding to between 270 and 41 nucleotide differences, while there was only a 0.3–3.8% difference in 16S rRNA gene sequence similarity at the interspecies level. Phylogenetic analysis based on the GyrB sequence deduced from the gyrB gene is consistent with that of DNA–DNA hybridization results and provides a better discrimination within the species of Gordonia compared to the 16S rRNA gene. The present study demonstrates that gyrB gene analysis will aid in describing novel species belonging to the genus Gordonia.

Williamsia muralisPulmonary Infection

<i>Williamsia muralis</i>Pulmonary Infection

Screening for exopolysaccharide-producing bacteria from sub-tropical polluted groundwater.

A selection of exopolysaccharide (EPS)--producing bacterial strains was conducted in groundwater adjacent to an old controlled landfill in the City of São Carlos (São Paulo, Brazil). The strains were isolated in P and E media under aerobic and microacrophilic conditions at 25 degrees C. A total of 26 strains were isolated and based on the mucoid mode of the colonies, 6 were selected and their morphological, physiological and biochemical aspects were characterized. All strains presented pigmentation, ranging from yellow to orange and from pink to salmon, with a shiny glistening aspect in all tested media. Strains Lb, Lc and Lg, which excelled the others with regard to the mucoid mode of the colonies, were selected to be cultured in E medium with alternate sucrose and glucose as carbon sources in anaerobiosis at 25 degrees C to analyze the production of EPS. Strains Lc and Lg were classified as being of order Actinomycelates, suborder Corynebacterineae. Lg strain was identified as Gordonia polyisoprenivorans and Lc strain did not correspond to a known description and therefore a more detailed study is under preparation. Considering all ecological aspects and the metabolic potential associated with the microorganisms of the environment studied, as well as the capacity to produce pigment and EPS, and the presence of G. polyisoprenivorans, a rubber degrader bacterium, the potential of the groundwater analyzed is evident as a source of microorganisms to be utilized in studies related to environmental remediation.

Williamsia muralis gen. nov., sp. nov., isolated from the indoor environment of a children's day care centre.

The taxonomic status of an actinomycete (MA 140/96T) isolated from indoor building materials of a children's day care centre was studied using the polyphasic approach. The cell morphology was atypical for an actinomycete, electron microscopy revealed a hairy surface, highly unusual for Gram-positive bacteria. The organisms grew at 10-37 degrees C, no growth was visible at 5 degrees C and 45 degrees C in 5 d. The cell wall contained the diamino acid meso-diaminopimelic acid and the sugars arabinose, galactose, mannose and ribose. The phospholipids phosphatidylethanolamine, phosphatidylinositol, phosphatidylglycerol and diphosphatidylglycerol were detected. The only menaquinone found was MK-9(H2). The fatty acid pattern was composed of palmitic acid (23.6%) palmitoleic acid (16.5%) and another hexadecenoic acid 16:1cis11 (1.4%), oleic acid (29.9%), stearic acid (2.9%) and the 10-methyl-branched tuberculostearic acid (23.3%). A gas-chromatographic analysis of the mycolic acid revealed a carbon-chain length of C50-C56. The G + C was 64.8 mol%. The results of 16S rDNA sequence comparisons revealed that strain MA 140/96T represents a new lineage in the suborder Corynebacterineae of the order Actinomycetales. Therefore, it was concluded that strain MA 140/96T should be assigned to a new genus and species, for which the name Williamsia muralis gen. nov., sp. nov. is proposed. The type strain of the species is MA 140/96T (= DSM 44343T).