Michèle Janssens

Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for identification of Nocardia species.

ABSTRACT The identification of Nocardia species, usually based on biochemical tests together with phenotypic in vitro susceptibility and resistance patterns, is a difficult and lengthy process owing to the slow growth and limited reactivity of these bacteria. In this study, a panel of 153 clinical and reference strains of Nocardia spp., altogether representing 19 different species, were characterized by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). As reference methods for species identification, full-length 16S rRNA gene sequencing and phenotypical biochemical and enzymatic tests were used. In a first step, a complementary homemade reference database was established by the analysis of 110 Nocardia isolates (pretreated with 30 min of boiling and extraction) in the MALDI BioTyper software according to the manufacturers recommendations for microflex measurement (Bruker Daltonik GmbH, Leipzig, Germany), generating a dendrogram with species-specific cluster patterns. In a second step, the MALDI BioTyper database and the generated database were challenged with 43 blind-coded clinical isolates of Nocardia spp. Following addition of the homemade database in the BioTyper software, MALDI-TOF MS provided reliable identification to the species level for five species of which more than a single isolate was analyzed. Correct identification was achieved for 38 of the 43 isolates (88%), including 34 strains identified to the species level and 4 strains identified to the genus level according to the manufacturers log score specifications. These data suggest that MALDI-TOF MS has potential for use as a rapid (<1 h) and reliable method for the identification of Nocardia species without any substantial costs for consumables.

Distribution of nocardia species in clinical samples and their routine rapid identification in the laboratory.

ABSTRACT Eighty-six Nocardia strains isolated from clinical samples in Belgium were identified by 16S rRNA gene sequencing. Eighty-three (96%) strains belonged to only six Nocardia species: N. farcinica (38 [44%]), N. nova (19 [22%]), N. cyriacigeorgica (13 [15%]), N. brasiliensis (6 [6.9%]), N. abscessus (5 [5.8%]), and N. paucivorans (2 [2.3%]). A gallery of nine conventional and enzymatic tests was developed for the rapid identification of the most common species isolated during this survey. Pyrrolidonyl aminopeptidase, γ-glutamyl aminopeptidase, α-mannosidase, and α-glucosidase were found to be highly discriminating and could be used to develop an identification scheme.

Yersinia-mollaretii Sp-nov and Yersinia-bercovieri Sp-nov, Formerly Called Yersinia-enterocolitica Biogroups 3a and 3b

Yersinia enterocolitica biogroups 3A and 3B are biochemically, serologically, and ecologically different from biogroup 3 and other Y. enterocolitica biogroups. Both biogroup 3A and biogroup 3B can be characterized by their negative Voges-Proskauer reactions and positive reactions in tests for pyrazinamidase, acid production from mucate, proline peptidase, and acid production from D-xylose. Biogroup 3A ferments L-sorbose but not L-fucose; biogroup 3B has the opposite fermentation pattern. Deoxyribonucleic acid relatedness studies (hydroxyapatite method) indicated that biogroups 3A and 3B are two new species that are about 55% interrelated and 25 to 46% related to other Yersinia species (except Yersinia ruckeri [20 to 22%]). The names Yersinia mollaretii sp. nov. and Yersinia bercovieri sp. nov. are proposed for biogroups 3A and 3B, respectively.

Bacteremia Due to a Novel Microbacterium Species in a Patient with Leukemia and Description of Microbacterium paraoxydans sp. nov.

ABSTRACT A yellow-pigmented coryneform rod was isolated from the blood of a child with acute lymphoblastic leukemia who was perfused with a central venous catheter. The culture bottles were positive twice, at a 2-month interval. The isolate was identified as a Microbacterium sp. and studied along with five other similar strains. Phenotypic, chemotaxonomic, and genetic characteristics indicated that they are closely related to Microbacteriumoxydans but that they belong to a distinct species, for which the name Microbacteriumparaoxydans sp. nov. is proposed. The type strain of M. paraoxydans is CF36T = DSM 15019T. The G+C content of its DNA is 69.9 mol%.

Description of Chryseobacterium anthropi sp. nov. to accommodate clinical isolates biochemically similar to Kaistella koreensis and Chryseobacterium haifense, proposal to reclassify Kaistella koreensis as Chryseobacterium koreense comb. nov. and emended description of the genus Chryseobacterium.

A collection of eight strains, NF 1366(T), NF 450, NF 1101, NF 1107, NF 1123, NF 1413, CCUG 15260 and CCUG 15624, from various clinical origins, were characterized biochemically as similar to Kaistella koreensis and Chryseobacterium haifense. They differed from K. koreensis, which is unable to alkalinize acetate, and from C. haifense, which is ONPG-positive (beta-galactosidase) and acidifies sucrose, fructose and lactose. Based on 16S rRNA gene sequence comparisons, this collection of strains was most closely related to the type strains of K. koreensis (97.3-97.5 %) and C. haifense (99.1 %). Representative strain NF 1366(T) showed only 41.8 % DNA-DNA relatedness with K. koreensis DSM 12107(T) and only 51.9 % with C. haifense DSM 19056(T). DNA-DNA hybridization of strains NF 450 and CCUG 15624 to strain NF 1366(T) was 41.7 and 74.6 %, respectively, and relatedness of these strains with C. haifense DSM 19056(T) was 72.6 and 70.2 %. With the present information, these two strains must be classified as intermediate between C. haifense and strain NF 1366(T). The fatty acid composition and polar lipid profile of strain NF 1366(T) were similar to those reported for other Chryseobacterium species. Like other chryseobacteria, strain NF 1366(T) exhibited a polyamine pattern with the predominant compound sym-homospermidine and a quinone system consisting of menaquinone MK-6 only. For this collection of clinical strains, the name Chryseobacterium anthropi sp. nov. is proposed, with NF 1366(T) (=CCUG 52764(T) =CIP 109762(T)) as the type strain. K. koreensis was shown to be very similar genotypically and phenotypically to Chryseobacterium. Its polar lipid profile exhibited the major characteristics shown for recently described Chryseobacterium species and the fatty acid profile of K. koreensis was also very similar to those of the Chryseobacterium species. Hence, no striking genotypic or phenotypic differences could be found that could justify the classification of this species into a separate genus, and we therefore propose to reclassify Kaistella koreensis in the genus Chryseobacterium as Chryseobacterium koreense comb. nov. (type strain Chj707(T) =IAM 15050(T) =JCM 21512(T) =KCTC 12107(T) =NBRC 103027(T)). An emended description of the genus Chryseobacterium is also proposed.

Short-comings of Irgasan Ticarcillin Chlorate Broth for the Enrichment of Yersinia-enterocolitica Biotype-2, Serotype-9 From Meat

Recovery of Yersinia enterocolitica serotype 0:9 from artificially contaminated minced pork with the enrichment medium irgasan ticarcillin chlorate broth (ITC) was poor. This was due to the lower growth rate of this serotype in comparison with serotype 0:3. Tests on the behaviour of serotype 0:3 and 0:9 towards some selective agents in ITC broth showed that the minimal inhibitory concentration of chlorate was much lower for serotype 0:9 than for serotype 0:3. Omitting chlorate in the ITC enrichment stimulated the growth of pure cultures. Reduction of MgCl2 and malachite green to 80% of the original concentration further increased the growth, so that after 48 h of incubation similar counts (10(7)-10(8) cfu/ml) as for serotype 0:3 were obtained.

Identification of a Novel Brevibacterium Species Isolated from Humans and Description of Brevibacterium sanguinis sp. nov.

ABSTRACT Six coryneforms isolated from blood and dialysate fluid were phenotypically similar to Brevibacterium casei, but 16S rRNA gene sequencing and DNA-DNA hybridization indicate that they belong to a new species for which the name Brevibacterium sanguinis is proposed.

Brevibacterium paucivorans sp. nov., from human clinical specimens.

Seven isolates from various human body sites displayed general chemotaxonomic and phenotypic characteristics of the genus Brevibacterium. This was corroborated by the 16S rRNA gene sequence analysis of strain CF62T, showing a sequence similarity of 99% to Brevibacterium mcbrellneri. However, DNA-DNA hybridization, a peculiar amino acid content of the cell wall and some phenotypic properties clearly suggested that these strains belong to a new species, for which the name Brevibacterium paucivorans sp. nov. is proposed. The type strain of B. paucivorans is CF62T (= DSM 13657T = LMG 19814T). The DNA G+C content of the type strain is 55.8 mol%.

Propionic acid-producing strains previously designated as Corynebacterium xerosis, C. minutissimum, C. striatum, and CDC group I2 and group F2 coryneforms belong to the species Corynebacterium amycolatum

Propionic acid-producing Corynebacterium strains that lacked mycolic acids and were formerly identified as Corynebacterium minutissimum, Corynebacterium xerosis, Corynebacterium striatum, and CDC group I-2 and F-2 strains were studied to determine their relatedness to Corynebacterium amycolatum, A total of 60 strains were used for phenotypic characterization studies, and 26 of these strains were used for genetic studies, DNA-DNA hybridization experiments performed at 65 degrees C revealed that the levels of relatedness between the propionic acid-producing strains and the type strain of C. amycolatum were more than 70% and that the Delta T-m, values ranged from 0 to 5 degrees C (Delta T-m is the difference between the denaturation temperature of a homoduplex and the denaturation temperature of a heteroduplex); these values are consistent with inclusion of these strains in the species C. amycolatum, Currently used conventional tests, such as urease, nitrate reduction, and sugar fermentation tests, were not suitable for accurate identification of C. amycolatum, Phenotypic differentiation of this species from related taxa should be based on the following characteristics in addition to propionic acid production: lipid requirement, Tween esterase activity, tyrosine clearing, alkaline phosphatase activity, alpha-glucosidase activity, and beta-glucuronidase activity.

Biochemical and susceptibility tests useful for identification of nonfermenting gram-negative rods.

ABSTRACT Six hundred nineteen strains of nonfermenting gram-negative rods were tested for alkaline phosphatase, benzyl-arginine arylamidase, pyrrolidonyl arylamidase, ethylene glycol acidification, and susceptibility to desferrioxamine and colistin. The results were highly discriminant. Therefore, the proposed tests may be helpful for the identification of this group of organisms.