Reclassification of Micrococcus aloeverae and Micrococcus yunnanensis as later heterotypic synonyms of Micrococcus luteus.

Micrococcus aloeverae,Micrococcus endophyticus, Micrococcus luteus and Micrococcus yunnanensis are phenotypically and genotypically closely related, and together comprise the M. luteus group. In this study, the taxonomic relationships among Micrococcus aloeverae, M. luteus and M. yunnanensis were re-evaluated by using polyphasic approaches. The similarity values of the concatenated housekeeping gene (gyrB, recA and rpoB) sequences shared by the type strains of M. aloeverae, M. luteus and M. yunnanensis ranged from 98.3 to 99.4 %. The average nucleotide identity, average amino acid identity and digital DNA‒DNA hybridization values among these three taxa were greater (97.1‒98.1 %, 96.8‒98.1 % and 75.0‒83.5 %, respectively) than the thresholds for bacterial species delineation, indicating that they belong to the same species, whereas those for M. endophyticus were clearly lower than the thresholds. In addition, phenotypic and chemotaxonomic characterization results also support the synonymy of these three taxa. Therefore, we propose that M. aloeverae and M. yunnanensis should be reclassified as later heterotypic synonyms of M. luteus.

Structural characterization and antioxidant potential of a novel anionic exopolysaccharide produced by marine Microbacterium aurantiacum FSW-25.

An exopolysaccharide (EPS) producing strain FSW-25 was isolated from the Rasthakaadu beach Kanyakumari, Tamil Nadu India. Based on polyphasic taxonomy, the strain FSW-25 was assigned to the genus Microbacterium and found to be the closest relative of the species aurantiacum. Large quantity of EPS (7.81 g/l) was secreted by the strain upon fermentation using Reasoner's 2A medium enriched with 2.5% glucose and was designated as Mi-25. FT-IR spectrum revealed presence of hydroxyl, carbonyl, carboxyl, methyl and sulfate functional groups in purified EPS. The EPS Mi-25 has a molecular weight of 7.0 × 106 Da and mainly comprises of glucuronic acid followed by glucose, mannose and fucose. Rheological study revealed that Mi-25 possesses significant viscosity with pseudoplastic nature. Interestingly, it was observed that the EPS Mi-25 has higher antioxidant activity as compared to xanthan. The characteristics of EPS Mi-25 suggested that, it can be used as a potential antioxidant with viscosifier properties in diverse industrial sectors.

Evaluation of autochthonous micrococcus strains as starter cultures for the production of Kedong sufu.

AIMS: The technological properties of 22 micrococcus strains from traditional fermented Kedong sufu were evaluated in order to develop autochthonous starter cultures. METHODS AND RESULTS: The proteolytic, autolytic and lipolytic activity, salt tolerance, production and degradation of the biogenic amines of six Micrococcus luteus, nine Kocuria kristinae and seven Kocuria rosea were evaluated. The results indicated that these micrococcus strains exhibited a certain technological diversity, and the results also indicated the best properties to be used in mixed starter cultures. Based on the above findings, two sets of autochthonous starters were formulated. Considering the physicochemical properties and sensory characteristics of sufu, the maturation period of sufu was shortened by 30 days. The profiles of free amino acids and peptides partly revealed the mechanism of sensory quality and shorter ripening time of sufu manufactured using autochthonous mixed starters. Compared to back-slopping fermentation, sufu manufactured with selected autochthonous starter cultures exhibited lower levels of total biogenic amines. CONCLUSIONS: The selected strains could be used as starter to avoid the accumulation of high concentrations of biogenic amines while also maintaining typical sensory characteristics and preserving the autochthonous strains of the traditional Kedong sufu. The maturation times of Kedong sufu were shortened by 30 days with application of the autochthonous starter. SIGNIFICANCE AND IMPACT OF THE STUDY: Autochthonous mixed starters can reduce the generation of biogenic amines, speed up the sufu maturation process and preserve typical sensory quality. Furthermore, the rotation of two sets of mixed starter cultures can effectively resist phage attack during the production of sufu.

Biodegradation of cypermethrin by immobilized cells of Micrococcus sp. strain CPN 1

Pyrethroid pesticide cypermethrin is a environmental pollutant because of its widespread use, toxicity and persistence. Biodegradation of such chemicals by microorganisms may provide an cost-effective method for their detoxification. We have investigated the degradation of cypermethrin by immobilized cells of Micrococcus sp. strain CPN 1 in various matrices such as, polyurethane foam (PUF), polyacrylamide, sodium alginate and agar. The optimum temperature and pH for the degradation of cypermethrin by immobilized cells of Micrococcus sp. were found to be 30 °C and 7.0, respectively. The rate of degradation of 10 and 20 mM of cypermethrin by freely suspended cells were compared with that of immobilized cells in batches and semi-continuous with shaken cultures. PUF-immobilized cells showed higher degradation of cypermethrin (10 mM and 20 mM) than freely suspended cells and cells immobilized in other matrices. The PUF-immobilized cells of Micrococcus sp. strain CPN 1 were retain their degradation capacity. Thus, they can be reused for more than 32 cycles, without losing their degradation capacity. Hence, the PUF-immobilized cells of Micrococcus sp. could potentially be used in the bioremediation of cypermethrin contaminated water.

Degradation of benzo[a]pyrene by bacterial isolates from human skin.

Polycyclic aromatic hydrocarbons (PAHs) are some of the most widespread xenobiotic pollutants, with the potentially carcinogenic high-molecular-weight representatives being of particular interest. However, while in eukaryotes, the cytochrome P450 (CYP)-mediated activation of benzo[a]pyrene (B[a]P) has become a model for metabolism-mediated carcinogenesis, the oxidative degradation of B[a]P by microorganisms is less well studied. This should be reason for concern as the human organ most exposed to environmental PAHs is the skin, which at the same time is habitat to a most diverse population of microbial commensals. Yet, nothing is known about the skin's microbiome potential to metabolise B[a]P. This study now reports on the isolation of 21 B[a]P-degrading microorganisms from human skin, 10 of which were characterised further. All isolates were able to degrade B[a]P as sole source of carbon and energy, and degradation was found to be complete in at least four isolates. Substrate metabolism involved two transcripts that encode a putative DszA/NtaA-like monooxygenase and a NifH-like reductase, respectively. Analysis of the 16S-rRNA genes showed that the B[a]P-degrading isolates comprise Gram(+) as well as Gram(-) skin commensals, with Micrococci being predominant. Moreover, microbial B[a]P-degradation was detected on all volunteers probed, indicating it to be a universal feature of the skin's microbiome.

Micrococcus cohnii sp. nov., isolated from the air in a medical practice.

Three Gram-reaction-positive bacteria, isolated from the air in a medical practice (strains WS4601(T), WS4602) or a pharmaceutical clean room (strain WS4599), were characterized using a polyphasic approach. Phylogenetic analyses based on 16S rRNA and recA gene sequences of the three novel strains showed that they formed a distinct lineage within the genus Micrococcus, sharing 16S rRNA gene sequence similarities of 96.1-98.0 % with other species of this genus. Chemotaxonomic features also supported the classification of the three novel strains within the genus Micrococcus. The major cellular fatty acids of strain WS4601(T) were anteiso-C(15 : 0) and iso-C(15 : 0), the cell-wall peptidoglycan was of type A3α (L-Lys-L-Ala), and the predominant respiratory quinones were MK-7(H(2)) and MK-8(H(2)). The polar lipid profile contained diphosphatidylglycerol and phosphatidylglycerol, but no phosphatidylinositol. The G+C content of the genomic DNA was 70.4 mol%. Numerous physiological properties were found that clearly distinguished strains WS4599, WS4601(T) and WS4602 from established members of the genus Micrococcus. Based on the phenotypic and phylogenetic data, strains WS4599, WS4601(T) and WS4602 are considered to represent three different strains of a novel species of the genus Micrococcus, for which the name Micrococcus cohnii sp. nov. is proposed. The type strain is WS4601(T) (=DSM 23974(T)=LMG 26183(T)).

Agrococcus terreus sp. nov. and Micrococcus terreus sp. nov., isolated from forest soil.

Two bacterial strains, DNG5T and V3M1T, isolated from forest soil of the Changbai mountains in China, were characterized using a polyphasic approach. Analysis of their 16S rRNA gene sequences indicated that strains DNG5T and V3M1T were phylogenetically related to members of the genus Agrococcus (96.0-98.4% similarity) and Micrococcus (96.7-98.0% similarity), respectively, within the order Actinomycetales. Strains DNG5T and V3M1T were Gram-stain-positive and strictly aerobic and formed yellow colonies on LB agar. Cells of strain DNG5T were short, non-motile rods, 0.4-0.5x0.8-1.0 microm. Strain DNG5T contained MK-10 and MK-11 as the major respiratory quinones and anteiso-C15:0 (49.2%) and iso-C16:0 (22.4%) as the major fatty acids. The diamino acid in the peptidoglycan of strain DNG5T was 2,4-diaminobutyric acid and the murein was of the acetyl type. Cells of strain V3M1T were cocci, 0.6-0.7 microm in diameter. The cell-wall peptidoglycan of strain V3M1T contained the amino acids lysine, glutamic acid, alanine and glycine. Strain V3M1T contained MK-7, MK-7(H2), MK-8 and MK-8(H2) as respiratory quinones and anteiso-C15:0 (78.2%) and iso-C15:0 (13.1%) as the major cellular fatty acids. The DNA G+C contents of strains DNG5T and V3M1T were 75.9 and 67.2 mol%, respectively. The DNA-DNA relatedness of strain DNG5T to Agrococcus jejuensis DSM 22002T, A. jenensis JCM 9950T, A. baldri JCM 12132T and A. citreus JCM 12398T was 58.3, 43.9, 36.1 and 54.1%, respectively. The DNA-DNA relatedness of strain V3M1T to Micrococcus luteus CGMCC 1.2299T, M. antarcticus CGMCC 1.2373T and M. lylae CGMCC 1.2300T was 57.5, 45.4 and 39.0%, respectively. Combining phenotypic and genotypic traits, strain DNG5T represents a novel species of the genus Agrococcus, for which the name Agrococcus terreus sp. nov. is proposed, with DNG5T (=CGMCC 1.6960T =NBRC 104260T) as the type strain. Strain V3M1T represents a novel species of the genus Micrococcus, for which the name Micrococcus terreus sp. nov. is proposed, with V3M1T (=CGMCC 1.7054T =NBRC 104258T) as the type strain.

Micrococcus yunnanensis sp. nov., a novel actinobacterium isolated from surface-sterilized Polyspora axillaris roots.

In this study, strain YIM 65004(T), isolated from roots of Polyspora axillaris, was shown to represent a novel species of the genus Micrococcus by means of a polyphasic approach. Chemotaxonomic data gathered for peptidoglycan type, menaquinones, phospholipids and fatty acids strongly supported the classification of this strain within the genus Micrococcus: the cell-wall peptidoglycan contained lysine, glutamic acid, alanine, glycine and aspartic acid, the predominant menaquinones were MK-8(H(2)) (66.97 %) and MK-7(H(2)) (23.26 %), the phospholipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and an unknown ninhydrin-negative phospholipid, and the major cellular fatty acids were anteiso-C(15 : 0) (61.98 %), iso-C(16 : 0) (14.25 %) and iso-C(15 : 0) (13.04 %). The G+C content of the genomic DNA was 71.7 mol%. A number of physiological features were found that clearly distinguished strain YIM 65004(T) from recognized Micrococcus species. DNA-DNA hybridization studies suggested that the novel strain represents a separate genomic species. Based on the above data, a novel species of the genus Micrococcus, Micrococcus yunnanensis sp. nov., is proposed, with the type strain YIM 65004(T) (=CCTCC AA 208060(T)=DSM 21948(T)).

Micrococcus endophyticus sp. nov., isolated from surface-sterilized Aquilaria sinensis roots.

A Gram-positive bacterial strain, designated YIM 56238(T), was isolated from plant roots (Aquilaria sinensis), and characterized by using a polyphasic approach. Strain YIM 56238(T) grew optimally at pH 7.0-8.0 and at 28 degrees C. Analysis of the 16S rRNA gene sequence of strain YIM 56238(T) indicated that it belongs to the genus Micrococcus. Chemotaxonomic data strongly supported the classification of this strain within the genus Micrococcus: the cell-wall peptidoglycan contained lysine, glutamic acid, alanine and glycine; the predominant menaquinones were MK-8(H(2)) (63.6 %) and MK-7(H(2)) (21.1 %); the phospholipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and an unknown ninhydrin-negative phospholipid; and the major cellular fatty acids were iso-C(15 : 0) (30.95 %) and anteiso-C(15 : 0) (53.75 %). The G+C content of the genomic DNA was 72.9 mol%. A number of physiological features were found that clearly distinguished strain YIM 56238(T) from recognized species of the genus Micrococcus. DNA-DNA hybridization studies suggested that the novel strain represents a separate genomic species. On the basis of the data, therefore, strain YIM 56238(T) represents a novel species of the genus Micrococcus, for which the name Micrococcus endophyticus sp. nov. is proposed. The type strain is YIM 56238(T) (=DSM 17945(T)=KCTC 19156(T)).

Emended descriptions of the genus Micrococcus, Micrococcus luteus (Cohn 1872) and Micrococcus lylae (Kloos et al. 1974).

Nine yellow-pigmented, spherical bacterial strains isolated from a medieval wall painting (strain D7), from indoor air (strains 3, 6, 7, 13C2, 38, 83 and 118) and from an activated-sludge plant (strain Ballarat) were classified by a polyphasic approach. Analyses of the 16S rRNA gene sequences of three representatives (strains D7, 118 and Ballarat) indicated that they all belong to the genus Micrococcus. The three isolates shared the highest sequence similarities with Micrococcus luteus DSM 20030T (97.9-98%), Micrococcus antarcticus AS 1.2372T (97.9-98.3%) and Micrococcus lylae DSM 20315T (97.5-97.9%). DNA-DNA reassociation studies clearly demonstrated that all nine isolates belong to the species M. luteus. However, neither their chemotaxonomic features nor their physiological and biochemical properties were consistent with those of M. luteus DSM 20030T. In contrast to M. luteus DSM 20030T, all isolates investigated possessed MK-8(H2) as the major respiratory quinone, and strain Ballarat had an A4alpha peptidoglycan type. On the basis of analyses of their Fourier transform-infrared spectroscopy spectra, isolates D7, 3, 6, 7, 13C2, 38, 83 and 118 could be grouped into a single cluster separate from M. luteus DSM 20030T, strain Ballarat and M. lylae DSM 20315T. In addition, all these isolates could be distinguished from M. luteus DSM 20030T by their ability to assimilate D-maltose, D-trehalose, DL-3-hydroxybutyrate, DL-lactate, pyruvate and L-histidine and to hydrolyse casein. Strains D7, 3, 6, 7, 13C2, 38, 83 and 118 differed from both M. luteus DSM 20030T and strain Ballarat by their ability to assimilate acetate, L-phenylalanine, L-serine and phenylacetate. Furthermore, REP-PCR fingerprinting yielded one common band for these strains, whereas this band was not observed for M. luteus DSM 20030T, strain Ballarat or M. lylae DSM 20315T. On the basis of these data, the species M. luteus can be divided into three biovars that are distinguished by several chemotaxonomic and biochemical traits: biovar I, represented by M. luteus DSM 20030T; biovar II, represented by strains D7 (= DSM 14234 = CCM 4959), 3, 6, 7, 13C2, 38, 83 and 118; and biovar III, represented by strain Ballarat (= DSM 14235 = CCM 4960). On the basis of the results generated in this study, emended descriptions of the genus Micrococcus and the species M. luteus and M. lylae are given.

Characters of coagulase-negative staphylococci and micrococci from cases of endocarditis.

Strains of coagulase-negative staphylococci and micrococci submitted to a reference laboratory from 137 cases of endocarditis over a 5 year period were reviewed. Staphylococcus epidermidis biotype 1 (SII) was the commonest biotype in all groups of patients but exceeded 80 per cent in the 61 patients who had undergone prosthetic valve surgery and the 16 patients who had undergone other forms of surgery. Biotype 4 (SVI) was recovered from 10 of the 34 patients without surgery. Strains from prosthetic valve endocarditis were frequently resistant to many antibiotics while strains from natural valve endocarditis were frequently sensitive to all, or resistant only to penicillin. The value of bacteriophage typing was confirmed.

Modified oxidase and benzidine tests for separation of staphylococci from micrococci.

Two simple and rapid methods for the separation of staphylococci from micrococci are described. They are based on modified oxidase and benzidine tests. Micrococci and Staphylococcus sciuri yield a blue color with a 6% solution of tetramethylphenylenediamine in dimethyl sulfoxide, whereas all of the other staphylococci exhibit no coloration. Best's results were obtained with overnight cultures on blood agar. The presence of c-type cytochromes i micrococci and S. sciuri could be detected with benzidine; all noncovalently linked heme groups are removed before the addition of the benzidine reagent. The oxidase test is the simplest and most rapid method for the separation of staphylococci (except S. sciuri) from micrococci, if the nutritional requirements and the time of incubation are strictly followed. This test is especially recommended for the examination of clinical material in which S. sciuri is usually not found.

Sulfur amino acid auxotrophy in Micrococcus species isolated from human skin.

Since methionine and (or) cysteine are required by a large percentage of natural auxotrophic Micrococcus strains isolated from human skin, investigations were directed to determine the specific enzymes affected in sulfur amino acid biosynthesis. Known intermediates in the interrelated cysteine and methionine biosynthetic pathways were tested as growth stimulants. Based on these growth studies, sulfur amino acid auxotrophs were grouped into three cysteine classes and five methionine classes. Selected auxotrophs of M. luteus had deficiencies in ATP sulfurylase (EC 2.7.7.4) and adenosine-5-sulfatophosphate (APS) kinase (EC 2.7.1.25), sulfite reductase (EC 1.8.1.2), serine transacetylase (EC 2.3.1.30), or beta-cystathionase (EC 4.4.1.8) activity; auxotrophs of M. lylae had deficiencies in sulfite reductase and serine transacetylase, beta-cystathionase, or N5, N10-methyltetrahydrofolate reductase (EC 1.1.1.68) activity; all auxotrophs of M. sedentarius tested had deficiencies in N5,N10-methyltetrahydrofolate reductase activity; auxotrophs of M. nishinomiyaensis had deficiencies in adenosine-3-phosphate-5-sulfatophosphate (PAPS) reductase, sulfite reductase, serine transacetylase, or N5,N10-methyltetrahydrofolate reductase activity; auxotrophs of M. varians had deficiencies in APS kinase, PAPS reductase, sulfite reductase, homoserine omicron-transsuccinylase, beta-cystathionase, or N5,N10-methyltetrahydrofolate reductase activity; auxotrophs of M. kristinae had deficiencies in serine transacetylase or cystathionine-gamma-synthase (EC 4.2.99.9) activity; auxotrophs of M. roseus had deficiencies in PAPS reductase, sulfite reductase, or serine transacetylase activity. Results of studies with various mutagens suggested that sulfur amino acid auxotrophy was primarily the result of a single base substitution in usually one or two of the genes controlling biosynthesis. A preliminary study of the amino acid composition of sweat suggested that this important source of nutrients does not contain adequate amounts of cysteine for the growth of cysteine auxotrophs but contains methionine that may be utilized in place of cysteine.

Gram-positive, motile, cluster-forming cocci as a cause of urinary infection.

One hundred and thirteen strains of motile, Gram-positive, catalase-positive, cluster-forming cocci were isolated from patients with urinary infection attending a private surgery. They constituted 1% of the total 11 302 positive cultures. The biochemical characteristics and the drug sensitivities of the strains are described. The significance of motility for organisms which cause urinary infections is pointed out.At the present time the organisms isolated are orphans in the controversial classification of staphylococci and micrococci.

Use of antibody to membrane adenosine triphosphatase in the study of bacterial relatioships.

An antiserum to Ca(2+)-activated adenosine triphosphatase from membranes of Micrococcus lysodeikticus cross-reacted in agar gels with membrane adenosine triphosphatases from other pigmented micrococci and related species. Species of Micrococcus and Sarcina showed different levels of inhibition of adenosine triphosphatase activities in heterologous reactions with antiserum. Inter- and intraspecific relationships based on the inhibition reaction were compared with an independent parameter, namely the quantitative and qualitative composition of the bacterial membrane phospholipids and fatty acids. The guanine plus cytosine contents in the deoxyribonucleic acid of the species studied correlated well with the serological cross-reactivity of adenosine triphosphatases from their membranes. The types of cross-bridges found in the peptidoglycans of these cocci were also compared with the other properties. The results suggest that an antiserum specific for a major membrane protein may be a reliable and most useful adjunct in studying bacterial serotaxonomy.