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Numerical approach to reference identification of Staphylococcus, Stomatococcus, and Micrococcus spp.

A numerical-code system for the reference identification of Staphylococcus species, Stomatococcus mucilaginosus, and Micrococcus species was established by using a selected panel of conventional biochemicals. Results from 824 cultures (289 eye isolate cultures, 147 reference strains, and 388 known control strains) were used to generate a list of 354 identification code numbers. Each six-digit code number was based on results from 18 conventional biochemical reactions. Seven milliliters of purple agar base with 1% sterile carbohydrate solution added was poured into 60-mm-diameter agar plates. All biochemical tests were inoculated with 1 drop of a heavy broth suspension, incubated at 35 degrees C, and read daily for 3 days. All reactions were read and interpreted by the method of Kloos et al. (G. A. Hebert, C. G. Crowder, G. A. Hancock, W. R. Jarvis, and C. Thornsberry, J. Clin. Microbiol. 26:1939-1949, 1988; W. E. Kloos and D. W. Lambe, Jr., P. 222-237, in A. Balows, W. J. Hansler, Jr., K. L. Herrmann, H. D. Isenberg, and H. J. Shadomy, ed., Manual of Clinical Microbiology, 5th ed., 1991). This modified reference identification method was 96 to 98% accurate and could have value in reference and public health laboratory settings.

Bacterial Typing Techniques↗

Identification of some charcoal-black-pigmented CDC fermentative coryneform group 4 isolates as Rothia dentocariosa and some as Corynebacterium aurimucosum: proposal of Rothia dentocariosa emend. Georg and Brown 1967, Corynebacterium aurimucosum emend. Yassin et al. 2002, and Corynebacterium nigricans Shukla et al. 2003 pro synon. Corynebacterium aurimucosum.

Sixty-three clinical isolates of charcoal-black-pigmented, gram-positive coryneform rods were received for identification by the Centers for Disease Control and Prevention (CDC) and were provisionally designated CDC fermentative coryneform group 4 (FCG4). Forty-five of these were characterized by morphological, physiologic, antimicrobial susceptibility, cellular fatty acids, 16S rRNA gene sequencing, and DNA-DNA hybridization analyses. Nitrate reduction, cellular fatty acid analysis, 16S rRNA gene sequencing, and DNA-DNA hybridization studies segregated these strains into two groups: FCG4a (8 strains) and FCG4b (37 strains). The FCG4a strains, only one of which was from a female genitourinary source, produced cellular fatty acid and biochemical profiles similar to those observed with reference strains of Rothia dentocariosa and Rothia mucilaginosa, while the FCG4b strains were similar to Corynebacterium species. DNA-DNA hybridization analysis demonstrated species-level relatedness among six FCG4a tested strains and showed that they were a charcoal-black-pigmented variant of R. dentocariosa. Sixteen isolates of the FCG4b group, mainly from female genitourinary tract specimens, as well as the type strains of two recently named species, Corynebacterium aurimucosum and Corynebacterium nigricans, were shown by DNA-DNA hybridization analysis and the sequencing of the 16S rRNA gene to be related at the species level and unrelated to the type strain of R. dentocariosa; therefore, the Corynebacterium-like strains were classified as a charcoal-black-pigmented variant of C. aurimucosum, because this name has nomenclatural priority over C. nigricans. These findings indicate that FCG4 represents a heterogeneous group that contains pigmented variants of both R. dentocariosa and C. aurimucosum; hence, the descriptions of both R. dentocariosa and C. aurimucosum have been amended to include charcoal-black-pigmented variants, and C. nigricans is a pro synonym of C. aurimucosum.

Centers for Disease Control and Prevention, U.S.↗

Erroneous reporting of coagulase-negative Staphylococci as Kocuria spp. by the Vitek 2 system.

Misidentification of coagulase-negative staphylococci (CoNS) may delay appropriate treatment. We investigated 20 clinical isolates identified as Kocuria spp. by the Vitek 2 system. All were identified as CoNS by 16S rRNA gene sequencing (18 Staphylococcus epidermidis, 1 Staphylococcus haemolyticus). Four Kocuria isolates were shown to be identical to CoNS from the same patient by pulsed-field gel electrophoresis. Isolates identified by Vitek 2 as Kocuria most likely represent misidentified CoNS, and if clinically indicated, should be investigated further by genomic methods.

Bacteriological Techniques↗

Comparative ultrastructure of methanogenic bacteria.

Electron-microscopic studies using thin sections revealed that methane-producing bacteria were an ultrastructurally diverse group. Fine structure and morphological characteristics separated these bacteria into four discrete cell types. Methanogenic bacteria displayed a gram-positive cell wall that varied considerably among different cell types. Differences in granular inclusions, reserve materials, and intracytoplasmic membranes were observed. Unique ultrastructural features were not shared by all methanogenic species studies.

Bacteria↗

Microbial ecology of the skin.

Humans exist in an environment replete with microorganisms, yet only a few of these microorganisms become residents on the skin surface. These resident flora and the skin constitute a complex ecosystem in which organisms adapt to changes in the microenvironment and to coactions among microorganisms. The skin possesses an assortment of protective mechanisms to limit colonization, and the survival of organisms on the surface lies in part in the ability of the organisms to resist these mechanisms. Microbial colonization on the skin adds to the skin's defense against potentially pathogenic organisms. Although microbes normally live in synergy with their hosts, at times colonization can lead to clinical infection. Common infections consist of superficial infections of the stratum corneum or appendages, which can respond dramatically to therapy but commonly relapse. In rare circumstances these infections can be severe, particularly in immunocompromised patients or hospitalized patients with indwelling foreign devices.

Actinomycetales↗

New species definitions in phages of gram-positive cocci.

About 290 phages of the bacterial genera Leuconostoc, Micrococcus, Staphylococcus, and Streptococcus were surveyed. The phages belong to the Myoviridae, Siphoviridae, and Podoviridae families and comprise seven basic morphotypes. Fourteen new species are proposed, bringing the number of phage species for gram-positive cocci to 27. The distribution of particular phage types may indicate phylogenetic relationships between host bacteria.

Bacteriophages↗

Assessment of acid production by various human oral micro-organisms when palatinose or leucrose is utilized.

One promising way of reducing caries is by using sucrose substitutes in food, e.g., palatinose or leucrose. Previous experiments addressing cariogenic potential of sucrose substitutes have focused mainly on Streptococcus mutans. However, given the many other micro-organisms in the oral cavity, this study compared the acid production of 100 bacterial strains representing 44 different species, by batch fermentation in a test tube containing, as a sole carbohydrate source, glucose, sucrose, palatinose, or leucrose. Selected strains were further analyzed in a fermenter. Additionally, 30 yeast strains were tested by an auxanographic sugar assimilation test. Only Lactobacillus spp., Stomatococcus mucilaginosus, Leuconostoc mesenteroides, and Weissella paramesenteroides, and some of the yeasts studied-i.e., Candida albicans, C. tropicalis, C. parapsilosis, and Saccharomyces cerevisiae-utilized leucrose and/or palatinose well. Strikingly, Stomatococcus mucilaginosus produced water-insoluble polysaccharides by fermentation of leucrose and palatinose. In the fermenter, the respective sucrose substitutes were not only cleaved but also utilized. Thus, extracellular cleavage by autochthonous micro-organisms may produce cariogenic cleavage products (glucose, fructose) that can be used by other well-characterized cariogenic bacteria found in the oral flora. Therefore, the anticariogenic potential of sucrose substitutes in food might be limited.

Bacteria↗

Kocuria kristinae infection associated with acute cholecystitis.

BACKGROUND: Kocuria, previously classified into the genus of Micrococcus, is commonly found on human skin. Two species, K. rosea and K. kristinae, are etiologically associated with catheter-related bacteremia. CASE PRESENTATION: We describe the first case of K. kristinae infection associated with acute cholecystitis. The microorganism was isolated from the bile of a 56-year old Chinese man who underwent laparoscopic cholecystectomy. He developed post-operative fever that resolved readily after levofloxacin treatment. CONCLUSION: Our report of K. kristinae infection associated with acute cholecystitis expands the clinical spectrum of infections caused by this group of bacteria. With increasing number of recent reports describing the association between Kocuria spp. and infectious diseases, the significance of their isolation from clinical specimens cannot be underestimated. A complete picture of infections related to Kocuria spp. will have to await the documentation of more clinical cases.

Acute Disease↗

Identification and NH2-terminal amino acid sequences of DnaK and groEL homologues in moderate eubacterial halophiles.

We have identified 2 DnaK and 3 GroEL homologues from moderately halophilic Acinetobacter, Pseudomonas, and Planococcus species by partial purification using an ATP-agarose column and by the analysis and similarity search of these NH2-terminal amino acid sequences. Although these bacteria required 1 to 2M NaCl for growth, these DnaK and GroEL homologues did not require high salt to bind to the ATP column, thus suggesting that these chaperones did not require high salts for their biochemically activities.

Acinetobacter↗

Stereochemistry of 2-phenylethylamine oxidation catalyzed by bacterial copper amine oxidase.

The stereochemical course of the reaction catalyzed by a copper amine oxidase from Arthrobacter globiformis has been investigated using 2-phenylethylamine stereospecifically deuterium-labeled at the C1 position. Measurements of deuterium content in the product, phenylacetaldehyde, by gas chromatography-mass spectrometry revealed stereospecific abstraction of the pro-S hydrogen during the enzymatic oxidation, as predicted from the structure modeling for the enzyme-bound substrate.

Acetaldehyde↗

Molecular cloning and expression of the hyu genes from Microbacterium liquefaciens AJ 3912, responsible for the conversion of 5-substituted hydantoins to alpha-amino acids, in Escherichia coli.

A DNA fragment from Microbacterium liquefaciens AJ 3912, containing the genes responsible for the conversion of 5-substituted-hydantoins to alpha-amino acids, was cloned in Escherichia coli and sequenced. Seven open reading frames (hyuP, hyuA, hyuH, hyuC, ORF1, ORF2, and ORF3) were identified on the 7.5 kb fragment. The deduced amino acid sequence encoded by the hyuA gene included the N-terminal amino acid sequence of the hydantoin racemase from M. liquefaciens AJ 3912. The hyuA, hyuH, and hyuC genes were heterologously expressed in E. coli; their presence corresponded with the detection of hydantoin racemase, hydantoinase, and N-carbamoyl alpha-amino acid amido hydrolase enzymatic activities respectively. The deduced amino acid sequences of hyuP were similar to those of the allantoin (5-ureido-hydantoin) permease from Saccharomyces cerevisiae, suggesting that hyuP protein might function as a hydantoin transporter.

Amino Acid Sequence↗

Purification and characterization of hydantoin racemase from Microbacterium liquefaciens AJ 3912.

A hydantoin racemase that catalyzed the racemization of 5-benzyl-hydantoin was detected in a cell-free extract of Microbacterium liquefaciens AJ 3912, a bacterial strain known to produce L-amino acids from their corresponding DL-5-substituted-hydantoins. This hydantoin racemase was purified 658-fold to electrophoretic homogeneity by serial chromatography. The N-terminal amino acid sequence of the enzyme showed homology with known hydantoin racemases from other microorganisms. The apparent molecular mass of the purified enzyme was 27 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and 117 kDa on gel-filtration in the purification conditions, indicating a homotetrameric structure. The purified enzyme exhibited optimal activity at pH 8.2 and 55 degrees C, and showed a chiral preference for L-5-benzyl- rather than D-5-benzyl-hydantoin.

Amino Acid Sequence↗