Maryam I. Daneshvar

Listeria marthii sp. nov., isolated from the natural environment, Finger Lakes National Forest.

Four isolates (FSL S4-120(T), FSL S4-696, FSL S4-710, and FSL S4-965) of Gram-positive, motile, facultatively anaerobic, non-spore-forming bacilli that were phenotypically similar to species of the genus Listeria were isolated from soil, standing water and flowing water samples obtained from the natural environment in the Finger Lakes National Forest, New York, USA. The four isolates were closely related to one another and were determined to be the same species by whole genome DNA-DNA hybridization studies (>82 % relatedness at 55 degrees C and >76 % relatedness at 70 degrees C with 0.0-0.5 % divergence). 16S rRNA gene sequence analysis confirmed their close phylogenetic relatedness to Listeria monocytogenes and Listeria innocua and more distant relatedness to Listeria welshimeri, L. seeligeri, L. ivanovii and L. grayi. Phylogenetic analysis of partial sequences for sigB, gap, and prs showed that these isolates form a well-supported sistergroup to L. monocytogenes. The four isolates were sufficiently different from L. monocytogenes and L. innocua by DNA-DNA hybridization to warrant their designation as a new species of the genus Listeria. The four isolates yielded positive reactions in the AccuProbe test that is purported to be specific for L. monocytogenes, did not ferment L-rhamnose, were non-haemolytic on blood agar media, and did not contain a homologue of the L. monocytogenes virulence gene island. On the basis of their phenotypic characteristics and their genotypic distinctiveness from L. monocytogenes and L. innocua, the four isolates should be classified as a new species within the genus Listeria, for which the name Listeria marthii sp. nov. is proposed. The type strain of L. marthii is FSL S4-120(T) (=ATCC BAA-1595(T) =BEIR NR 9579(T) =CCUG 56148(T)). L. marthii has not been associated with human or animal disease at this time.

Bordetella holmesii Bacteremia: A Newly Recognized Clinical Entity among Asplenic Patients

Bordetella holmesii is a recently identified gram-negative bacterial species associated with bacteremia, endocarditis, and respiratory illness, mainly in immunocompromised patients. From isolates submitted to the Centers for Disease Control and Prevention from 1983 through 2000 for further identification, we identified 30 patients with B. holmesii bacteremia. Of the 26 patients for whom data were available, 22 (85%) were anatomically or functionally asplenic. In 25 (96%) of the 26 patients, B. holmesii was the only organism isolated from blood samples, and 14 patients (54%) had B. holmesii recovered from > or =2 blood cultures. The clinical course of the infection was generally characterized by a nonspecific febrile illness. Twenty-one patients (81%) were treated with various antimicrobial agents, and 20 (77%) were admitted to the hospital. There were no deaths. Our findings support evidence that B. holmesii may be a true pathogen associated with bacteremia among asplenic patients.

Legionella drozanskii sp. nov., Legionella rowbothamii sp. nov. and Legionella fallonii sp. nov. : three unusual new Legionella species

Seven strains of Legionella-like amoebal pathogens (LLAPs) were characterized on the basis of their cultural and staining characteristics, biochemical reactions, serology, cellular fatty acids (CFAs), isoprenoid quinone composition, total DNA relatedness, analysis of 16S rRNA and macrophage infectivity potentiator (mip) gene sequence analyses. All seven strains exhibited limited growth on buffered charcoal yeast extract alpha (BCYE) agar, required cysteine for growth and contained branched-chain CFAs and quinones typical of Legionella species. The bacilli were Gram-negative and catalase-positive. There were varying degrees of serological cross-reactions between these LLAP strains and other previously described Legionella species. Results from the various tests revealed that four LLAP strains represent three unusual new species of Legionella: Legionella drozanskii sp. nov., type strain LLAP-1T; Legionella rowbothamii sp. nov., type strain LLAP-6T; and Legionella fallonii sp. nov., type strain LLAP-10T. Three other LLAP strains, designated LLAP-7FL, LLAP-7NF and LLAP-9, were shown to be members of the species Legionella lytica. The deductions made from the phenetic characteristics of these bacteria were consistent with the phylogenetic relationships inferred from 16S rRNA and mip gene sequence analyses. This study is the first to speciate LLAP strains on the basis of data including quantitative DNA hybridization.

Novel Brucella Strain (BO1) Associated with a Prosthetic Breast Implant Infection

ABSTRACT We report the microbiological, biochemical, and molecular characterization of an unusual Brucella strain (BO1) isolated from a breast implant wound in a 71-year-old woman with clinical symptoms consistent with brucellosis. Initial phenotypic analysis, including biochemical and antimicrobial susceptibility testing, cellular fatty acid analysis, and molecular analysis based on DNA-DNA reassociation and the presence of multiple copies of IS711 element suggested that the isolate was a Brucella-like organism, but species determination using microbiological algorithms was unsuccessful. Furthermore, molecular data based on 16S rRNA gene sequencing and multilocus sequence analysis demonstrated that BO1 was an unusual Brucella strain and not closely related to any currently described Brucella species. However, comparison with equivalent sequences in Ochrobactrum spp. confirms that the isolate is much more closely related to Brucella than to Ochrobactrum spp., and thus the isolate likely represents an atypical and novel strain within the genus Brucella.

Assignment of CDC Weak Oxidizer Group 2 (WO-2) to the Genus Pandoraea and Characterization of Three New Pandoraea Genomospecies

ABSTRACT CDC weak oxidizer group 2 (WO-2) consists of nine phenotypically similar human clinical isolates received by the Centers for Disease Control and Prevention between 1989 and 1998. Four of the isolates were from blood, three were from sputum, and one each was from bronchial fluid and maxillary sinus. All are aerobic nonfermentative, motile gram-negative rods with one to eight polar flagella per cell. All grew at 25 and 35°C and were positive for catalase, urease (usually delayed 3 to 7 days), citrate, alkalinization of litmus milk, oxidization of glycerol (weakly), and growth on MacConkey agar and in nutrient broth without NaCl. All except one strain were oxidase positive with the Kovács method, and all except one isolate weakly oxidized d-glucose. All were negative for oxidation of d-xylose, d-mannitol, lactose, sucrose, maltose, and 20 other carbohydrates, esculin hydrolysis, indole production, arginine dihydrolase, and lysine and ornithine decarboxylase. Only two of nine isolates reduced nitrate. Broth microdilution susceptibilities were determined for all strains against 13 antimicrobial agents. Most of the strains were resistant to ampicillin, extended-spectrum cephalosporins, and aminoglycosides, including gentamicin, tobramycin, and amikacin, but they varied in their susceptibility to fluoroquinolones. High-performance liquid chromatographic and mass spectrometric analyses of the WO-2 group identified ubiquinone-8 as the major quinone component. The percent G+C of the WO-2 strains ranged from 65.2 to 70.7% (thermal denaturation method). All shared a common cellular fatty acid (CFA) profile, which was characterized by relatively large amounts (7 to 22%) of 16:1ω7c, 16:0, 17:0cyc, 18:1ω7c, and 19:0cyc11-12; small amounts (1 to 3%) of 12:0 and 14:0; and eight hydroxy acids, 2-OH-12:0 (4%), 2-OH-14:0 (trace), 3-OH-14:0 (12%), 2-OH-16:1 (1%), 2-OH-16:0 (3%), 3-OH-16:0 (4%), 2-OH-18:1 (2%), and 2-OH-19:0cyc (3%). This profile is similar to the CFA profile of Pandoraea, a recently described genus associated with respiratory infections in cystic fibrosis patients (T. Coenye et al., Int. J. Syst. Evol. Microbiol., 50:887–899, 2000). Sequencing of the 16S rRNA gene (1,300 bp) for all nine strains indicated a high level (≥98.8%) of homogeneity withPandoraea spp. type strains. DNA-DNA hybridization analysis (hydroxyapatite method; 70°C) confirmed the identity of WO-2 with the genus Pandoraea and assigned three strains toPandoraea apista and three to Pandoraea pnomenusa, and identified three additional new genomospecies containing one strain each (ATCC BAA-108, ATCC BAA-109, ATCC BAA-110). This study also shows that Pandoraea isolates may be encountered in blood cultures from patients without cystic fibrosis.

Paracoccus yeeii sp. nov. (Formerly CDC Group EO-2), a Novel Bacterial Species Associated with Human Infection

ABSTRACT CDC eugonic oxidizer group 2 (EO-2) is a group of unclassified gram-negative bacterial strains isolated from various human sources. As determined by biochemical tests and analyses of fatty acid compositions, these organisms form a homogeneous group that appears to be distinct from but related to other Paracoccus species. Molecular studies were performed on a set of 13 EO-2 strains from various clinical sources and geographic locations in the United States and Canada to determine their relationship to the Paracoccus genus. Control strains were Paracoccus denitrificans ATCC 17741T, P. versutus ATCC 25364T, P. aminophilus ATCC 49673T, P. solventivorans ATCC 700252T, and Psychrobacter immobilis ATCC 43116T, which are phenotypically similar to EO-2. Nearly complete (1,500-base) 16S rRNA gene sequencing of eight EO-2 strains showed a high level of sequence similarity (>99.3%) within the group, and a BLAST search of GenBank placed the EO-2 cluster in close proximity to Paracoccus species (95 to 97% similarity). DNA-DNA hybridization studies of 13 of the EO-2 strains showed all to be related at the species level, with >70% relatedness under stringent conditions and a divergence within the group of less than 2%. None of the Paracoccus control strains hybridized at >54% with any of the EO-2 strains. These results indicate that EO-2 represents a new Paracoccus species, the first isolated from human clinical specimens. A new species, Paracoccus yeeii, is proposed for the EO-2 strains. The type strain of P. yeeii is CDCG1212 (ATCC BAA-599 and CCUG 46822), isolated in Pennsylvania from dialysate of a 77-year-old male with peritonitis.

Identification of “Haematobacter,” a New Genus of Aerobic Gram-Negative Rods Isolated from Clinical Specimens, and Reclassification of Rhodobacter massiliensis as “Haematobacter massiliensis comb. nov.”

ABSTRACT Twelve strains of gram-negative, nonfermenting rods recovered mainly from septicemic patients were studied using conventional and molecular methods. The phenotypic profiles of these strains most closely resembled Psychrobacter phenylpyruvicus. They produced catalase, oxidase, urease, and H2S (lead acetate paper) but did not produce indole, reduce nitrate or nitrite, or hydrolyze gelatin or esculin. No acid production was observed in a Kings oxidation-fermentation base containing d-glucose, d-xylose, d-mannitol, sucrose, lactose, or maltose. All strains were nonmotile and nonpigmented. Most strains produced green discoloration on blood agar. All strains grew at 25°C and 35°C and most grew on MacConkey agar. They shared a common cellular fatty acid (CFA) profile characterized by large amounts (56% to 90%) of 18:1ω7c and the presence of 3-OH-10:0, 16:1ω7c, 16:0, and 19:0cycω8c that overall was most similar to that of Rhodobacter species but was quite distinct from that of P. phenylpyruvicus. The MICs for most β-lactams, fluoroquinolones, aminoglycosides, and carbapenems were low. MICs for aztreonam and piperacillin were higher, with MICs for some strains of > 64 mg/liter and > 128 mg/liter, respectively. Polyphasic analysis of these strains, including morphological, biochemical, CFA composition, DNA-DNA hybridization, 16S rRNA gene sequencing, and percent guanine-plus-cytosine (G+C) content analysis, demonstrated that these strains and Rhodobacter massiliensis represent a new genus, “Haematobacter” (proposed name), with the species H. missouriensis (type strain H1892T = CCUG 52307T = CIP 109176T) and H. massiliensis comb. nov. (type strain FramboiseT = CCUG 47968T = CIP 107725T) and an unnamed genomospecies.

Phenotypic and Genetic Characterization of Clinical Isolates of CDC Coryneform Group A-3: Proposal of a New Species of Cellulomonas, Cellulomonas denverensis sp. nov.

ABSTRACT CDC coryneform group A-3 bacteria are rare human pathogens. In this study, six group A-3 isolates (two from blood, one from cerebrospinal fluid, and one each from homograft valve, lip wound, and pilonidal cyst) were compared to the type strains of phenotypically related organisms, Cellulomonas fimi, Cellulomonas hominis, Oerskovia turbata, and Sanguibacter suarezii, and characterized by phenotypic, chemotaxonomic, and genotypic studies. DNA-DNA reassociation analysis identified two genomic groups, and phylogenetic analysis of the 16S rRNA gene sequence identified the taxonomic positions of these groups to genus level. Two groups were defined, and both were more closely related to Cellulomonas species: one group of three strains, for which we propose the new species Cellulomonas denverensis sp. nov., with the type strain W6929 (ATCC BAA-788T or DSM 15764T), was related to C. hominis ATCC 51964T (98.5% 16S rRNA gene sequence similarity), and the second group of three strains was related to C. hominis ATCC 51964T (99.8 to 99.9% 16S rRNA gene sequence similarity). The definition of this new Cellulomonas species and the confirmation of three strains as C. hominis serve to further clarify the complex taxonomy of CDC coryneform group A-3 bacteria and will assist in our understanding of the epidemiology and clinical significance of these microorganisms.

Characterization of Burkholderia rhizoxinica and B. endofungorum Isolated from Clinical Specimens

Eight isolates submitted to CDC from 1989 to 2006 from clinical specimens were initially identified as members of the genus Burkholderia based on preliminary cellular fatty acid analysis and/or 16S rRNA gene sequencing. With the recent descriptions of the new species B. rhizoxinica and B. endofungorum, which are considered endosymbiotic bacteria in Rhizopus microsporus fungi, we now identify seven of these clinical isolates as B. rhizoxinica and one as B. endofungorum based on biochemical testing, 16s rRNA, and DNA-DNA hybridization results. We also further characterize these isolates by assessing toxin production and/or by multiple locus sequence typing.

Cellular fatty acid composition of Lautropia mirabilis.

ABSTRACT Ten strains of Lautropia mirabilis (ATCC 51599T and nine phenotypically similar clinical isolates) were examined for cellular fatty acid (CFA) composition to evaluate their chemical relatedness to known bacterial species and groups. The CFAs were liberated from whole cells by base hydrolysis, methylated, and analyzed by gas-liquid chromatography. CFA profiles were generated by using a commerical software package (MIDI, Newark, Del.). All strains tested had an identical CFA profile characterized by major amounts of 16:1ω7c (41%) and 16:0 (44%); smaller amounts (1 to 4%) of 3-OH-10:0, 12:0, 14:0, 15:0, and 18:1 ω7c; trace amounts (<1%) of 10:0, 18:2 and 18:0; and no cyclopropane acids. This profile was similar to the CFA profiles of Acidovorax delafieldii, Comamonas terrigena, and strains of an unclassified Centers for Disease Control group designated weak oxidizer group 1. CFA analysis, when supplemented by phenotypic characterization, is useful for the identification of L. mirabilis isolates.