Biomedical subjects

W Ludwig

Publications and source records attributed to W Ludwig.

At least 55 records · Page 3Linked to original sources

PubMed1999

Chlorobium ferrooxidans sp. nov., a phototrophic green sulfur bacterium that oxidizes ferrous iron in coculture with a "Geospirillum" sp. strain.

A green phototrophic bacterium was enriched with ferrous iron as sole electron donor and was isolated in defined coculture with a spirilloid chemoheterotrophic bacterium. The coculture oxidized ferrous iron to ferric iron with stoichiometric formation of cell mass from carbon dioxide. Sulfide, thiosulfate, or elemental sulfur was not used as electron donor in the light. Hydrogen or acetate in the presence of ferrous iron increased the cell yield of the phototrophic partner, and hydrogen could also be used as sole electron source. Complexed ferric iron was slowly reduced to ferrous iron in the dark, with hydrogen as electron source. Similar to Chlorobium limicola, the phototrophic bacterium contained bacteriochlorophyll c and chlorobactene as photosynthetic pigments, and also resembled representatives of this species morphologically. On the basis of 16S rRNA sequence comparisons, this organism clusters with Chlorobium, Prosthecochloris, and Pelodictyon species within the green sulfur bacteria phylum. Since the phototrophic partner in the coculture KoFox is only moderately related to the other members of the cluster, it is proposed as a new species, Chlorobium ferrooxidans. The chemoheterotrophic partner bacterium, strain KoFum, was isolated in pure culture with fumarate as sole substrate. The strain was identified as a member of the epsilon-subclass of the Proteobacteria closely related to "Geospirillum arsenophilum" on the basis of physiological properties and 16S rRNA sequence comparison. The "Geospirillum" strain was present in the coculture only in low numbers. It fermented fumarate, aspartate, malate, or pyruvate to acetate, succinate, and carbon dioxide, and could reduce nitrate to dinitrogen gas. It was not involved in ferrous iron oxidation but possibly provided a thus far unidentified growth factor to the phototrophic partner.

Bacteria, Anaerobic↗

PubMed1999

Novel 16S rRNA gene sequences retrieved from highly saline brine sediments of kebrit deep, red Sea

In this study, we report on first 16S rRNA gene sequences from highly saline brine sediments taken at a depth of 1,515 m in the Kebrit Deep, northern Red Sea. Microbial DNA extracted directly from the sediments was subjected to PCR amplification with primers specific for bacterial and archaeal 16S rRNA gene sequences. The PCR products were cloned, and a total of 11 (6 bacterial and 5 archaeal) clone types were determined by restriction endonuclease digestion. Phylogenetic analysis revealed that most of the cloned sequences were unique, showing no close association with sequences of cultivated organisms or sequences derived from environmental samples. The bacterial clone sequences form a novel phylogenetic lineage (KB1 group) that branches between the Aquificales and the Thermotogales. The archaeal clone sequences group within the Euryarchaeota. Some of the sequences cluster with the group II and group III uncultivated archaea sequence clones, while two clone groups form separate branches. Our results suggest that hitherto unknown archaea and bacteria may thrive in highly saline brines of the Red Sea under extreme environmental conditions.

Journal Article↗

PubMed1999

Intracellular expression of CD61 precedes surface expression.

We report a case of acute myeloid leukemia which we have classified as acute megakaryoblastic leukemia because of intracytoplasmic expression of CD61. Light microscopy demonstrated agranular blasts which, by cytochemical staining, were negative for myeloperoxidase. Using flow cytometry, the blast cells were seen to be positive for HLA-DR, CD7, CD13, CD33, and CD34, thus revealing their myeloid origin. Immunophenotyping on fixed blood smears additionally showed positive reaction with the CD61 antibody, demonstrating the megakaryoblastic differentiation of the blasts. After permeabilization of the cell membrane, the intracytoplasmic CD61 expression was confirmed by flow cytometry. Cytogenetic analysis disclosed a del(7)(q21-22). Our findings suggest that cytoplasmic expression of CD61 may precede the cell-surface expression of this antigen and should therefore be investigated in all cases of acute leukemias with undifferentiated morphology and negative cytochemistry to set apart early FAB-M7 from FAB-M0.

Aged↗

PubMed1999

Lactobacillus perolens sp. nov., a soft drink spoilage bacterium.

Lactic acid bacteria that are able to spoil soft drinks with low pH comprise a limited number of acidotolerant or acidophilic species of the genera Lactobacillus, Leuconostoc and Weissella. Various Gram-positive rods causing turbidity and off-flavour were isolated from orange lemonades. Physiological and biochemical studies including SDS-PAGE whole-cell protein analysis showed a homogeneous group of organisms. The 16S rRNA gene sequence analysis of two representatives revealed that they formed a phylogenetically distinct line within the genus Lactobacillus. All strains were facultatively heterofermentative, producing L-lactic acid. Based on the data presented a new species L. perolens is proposed. The name refers to the off-flavour caused by high amounts of diacetyl. The type strain of L. perolens is DSM 12744 (LMG 18936). A rRNA targeted oligonucleotide probe was designed that allows a fast and reliable identification of L. perolens.

Bacterial Proteins↗

PubMed1999

Specific oligonucleotide probes for in situ detection of a major group of gram-positive bacteria with low DNA G + C content.

Almost one thousand 16S rRNA sequences of Gram-positive bacteria with a low DNA G + C content from public databases were analyzed using the ARB software package. A signature region was identified between positions 354 and 371 (E. coli numbering) for the Bacillus sub-branch of the Gram-positive bacteria with a low DNA G + C content, the former orders Bacillales and Lactobacillales. Three oligonucleotide probes, namely LGC354A, LGC354B, and LGC354C, were designed to target this diagnostic site. Their fluorescent derivatives were suitable for whole cell detection by fluorescence in situ hybridization (FISH). Hybridization conditions were adjusted for differentiation of target and related non-target reference species. When applying FISH to whole bacterial cells in a sample of activated sludge from a communal wastewater treatment plant, members of the Bacillus sub-branch were detected at levels from 0.01% of cells in samples fixed with paraformaldehyde to over 8 percent in the same samples fixed with ethanol and treated with lysozyme. The problems of quantitative in situ analysis of Gram-positive bacteria with a low DNA G + C content in biofilm flocs are discussed and recommendations made. Members of the Bacillus sub-branch were detected in different abundances in activated sludge samples from different wastewater plants.

Bacillus↗

PubMed1999

Genotypic diversity of Acidovorax strains isolated from activated sludge and description of Acidovorax defluvii sp. nov.

Fluorescence in situ hybridization of activated sludge samples from a municipal wastewater treatment plant using oligonucleotide probes specific for Acidovorax demonstrated that these bacteria are highly abundant in this environment. For the targeted cultivation of representatives belonging to this genus, isolates grown on agar plates after serial dilution were screened by whole-cell hybridization with specific probes. The obtained strains clustered in two phylogenetic groups as determined by 16S rRNA gene sequence analyses. The isolates of one cluster were phylogenetically and genotypically closely related to A. delafieldii. In contrast, the strains of the other cluster were genotypically and phenotypically distinct from the hitherto known Acidovorax species. Therefore, a new species, Acidovorax defluvii sp. nov., was proposed for these strains. The main characteristics of the newly defined species are as follows: Gram-negative, motile or non-motile rods with rounded ends, often with large polyhydroxybutyrate granules. In broth cultures flocs are formed. Test for cytochrome oxidase is positive with all strains. The majority of strains is catalase positive and reduces nitrate. All strains are metabolically inactive against most carbohydrates and organic acids. Fatty acid patterns are typical for the genus Acidovorax. The guanine-plus-cytosine content of DNAs varies between 62 and 64 mol%. The type strain of A. defluvii is BSB411T (DSM 12644). A new 16S rRNA-targeted oligonucleotide probe reacting by in situ hybridization with all known Acidovorax species, including A. defluvii sp. nov., was designed.

Base Sequence↗

PubMed1999

rRNA probe-based cell fishing of bacteria.

We have developed a new, cultivation-independent, fast and flexible method for the rRNA-targeted probe-based enrichment of bacteria. The target cells were labelled by in situ hybridization with biotinylated polyribonucleotide probes. These probes were generated by in vitro transcription of amplified rDNA of a variable region in domain III of the 23S rRNA molecules. The probes were about 300 nucleotides in length and were labelled by incorporation of biotin-UTP during the transcription. Probes were hybridized with bacterial cells and incubated with paramagnetic streptavidin-coated particles. The labelled target cells can be separated in a column filled with steel wool inserted into the field of a permanent magnet. Unlabelled, non-target cells pass through the column, whereas labelled cells are retained. They were eluted from the column after removal of the magnetic field. Up to now, the method has been tested with mixtures of different pure cultures. For the first time, transcript probes have been used for the labelling of the target cells and for their specific separation. The enrichment of the target cells can be monitored by a streptavidin-fluorescein staining of the biotinylated target cells and/or by a subsequent in situ hybridization with fluorescently labelled oligonucleotide probes. Enrichment rates of up to 90-fold, depending on the original abundance of the cells of interest, could be determined. To demonstrate that the sorted cells were amenable to molecular analysis, we amplified and sequenced a part of the tuf gene of enriched Acinetobacter calcoaceticus cells.

Acinetobacter calcoaceticus↗

PubMed1999

Coenonia anatina gen. nov., sp. nov., a novel bacterium associated with respiratory disease in ducks and geese.

Taxon 1502 was originally described as a Riemerella anatipestifer-like bacterium causing exudative septicaemia in ducks and geese. In the present study, an integrated genotypic and phenotypic approach was used to elucidate the phylogenetic affiliation and taxonomic relationships of 12 strains of taxon 1502. Whole-cell protein and fatty acid analyses and an extensive biochemical examination by using conventional tests and several API microtest systems indicated that all isolates formed a homogeneous taxon, which was confirmed by DNA-DNA hybridizations. 16S rDNA sequence analysis of a representative strain (LMG 14382T) indicated that this taxon belongs to the Cytophaga-Flavobacterium-Bacteroides phylum and revealed a moderate but distinct relationship to species of the genus Capnocytophaga (overall 16S rDNA sequence identities were 88.8-90.2%). Taxon 1502 is concluded to represent a single species that should be allocated to a novel genus, and the name Coenonia anatina gen. nov., sp. nov. is proposed. The DNA G + C content of representative strains was 35-36 mol% and the type strain is LMG 14382T.

Animals↗

PubMed1999

In situ identification of cyanobacteria with horseradish peroxidase-labeled, rRNA-targeted oligonucleotide probes.

Individual cyanobacterial cells are normally identified in environmental samples only on the basis of their pigmentation and morphology. However, these criteria are often insufficient for the differentiation of species. Here, a whole-cell hybridization technique is presented that uses horseradish peroxidase (HRP)-labeled, rRNA-targeted oligonucleotides for in situ identification of cyanobacteria. This indirect method, in which the probe-conferred enzyme has to be visualized in an additional step, was necessary since fluorescently monolabeled oligonucleotides were insufficient to overstain the autofluorescence of the target cells. Initially, a nonfluorescent detection assay was developed and successfully applied to cyanobacterial mats. Later, it was demonstrated that tyramide signal amplification (TSA) resulted in fluorescent signals far above the level of autofluorescence. Furthermore, TSA-based detection of HRP was more sensitive than that based on nonfluorescent substrates. Critical points of the assay, such as cell fixation and permeabilization, specificity, and sensitivity, were systematically investigated by using four oligonucleotides newly designed to target groups of cyanobacteria.

Bacterial Typing Techniques↗

PubMed1998

MLL and CALM are fused to AF10 in morphologically distinct subsets of acute leukemia with translocation t(10;11): both rearrangements are associated with a poor prognosis.

The translocation t(10;11)(p13;q14) has been observed in acute lymphoblastic leukemia (ALL) as well as acute myeloid leukemia (AML). A recent study showed a MLL/AF10 fusion in all cases of AML with t(10;11) and various breakpoints on chromosome 11 ranging from q13 to q23. We recently cloned CALM (Clathrin Assembly Lymphoid Myeloid leukemia gene), the fusion partner of AF10 at 11q14 in the monocytic cell line U937. To further define the role of these genes in acute leukemias, 10 cases (9 AML and 1 ALL) with cytogenetically proven t(10;11)(p12-14;q13-21) and well-characterized morphology, immunophenotype, and clinical course were analyzed. Interphase fluorescence in situ hybridization (FISH) was performed with 2 YACs flanking the CALM region, a YAC contig of the MLL region, and a YAC spanning the AF10 breakpoint. Rearrangement of at least one of these genes was detected in all cases with balanced t(10;11). In 4 cases, including 3 AML with immature morphology (1 AML-M0 and 2 AML-M1) and 1 ALL, the signals of the CALM YACS were separated in interphase cells, indicating a translocation breakpoint within the CALM region. MLL was rearranged in 3 AML with myelomonocytic differentiation (2 AML-M2 and 1 AML-M5), including 1 secondary AML. In all 3 cases, a characteristic immunophenotype was identified (CD4+, CD13-, CD33+, CD65s+). AF-10 was involved in 5 of 6 evaluable cases, including 1 case without detectable CALM or MLL rearrangement. In 2 complex translocations, none of the three genes was rearranged. All cases had a remarkably poor prognosis, with a mean survival of 9.6 +/- 6.6 months. For the 7 AML cases that were uniformly treated according to the AMLCG86/92 protocols, disease-free and overall survival was significantly worse than for the overall study group (P = .03 and P = .01, respectively). We conclude that the t(10;11)(p13;q14) indicates CALM and MLL rearrangements in morphologically distinct subsets of acute leukemia and may be associated with a poor prognosis.

Acute Disease↗

PubMed1998

Bacterial phylogeny based on comparative sequence analysis.

Comparative sequence analysis of small subunit rRNA is currently one of the most important methods for the elucidation of bacterial phylogeny as well as bacterial identification. Phylogenetic investigations targeting alternative phylogenetic markers such as large subunit rRNA, elongation factors, and ATPases have shown that 16S rRNA-based trees reflect the history of the corresponding organisms globally. However, in comparison with three to four billion years of evolution the phylogenetic information content of these markers is limited. Consequently, the limited resolution power of the marker molecules allows only a spot check of the evolutionary history of microorganisms. This is often indicated by locally different topologies of trees based on different markers, data sets or the application of different treeing approaches. Sequence peculiarities as well as methods and parameters for data analysis were studied with respect to their effects on the results of phylogenetic investigations. It is shown that only careful data analysis starting with a proper alignment, followed by the analysis of positional variability, rates and character of change, testing various data selections, applying alternative treeing methods and, finally, performing confidence tests, allows reasonable utilization of the limited phylogenetic information.

Bacteria↗

PubMed1998

Dehalobacter restrictus gen. nov. and sp. nov., a strictly anaerobic bacterium that reductively dechlorinates tetra- and trichloroethene in an anaerobic respiration.

The highly enriched anaerobic bacterium that couples the reductive dechlorination of tetrachloroethene to growth, previously referred to as PER-K23, was obtained in pure culture and characterized. The bacterium, which does not form spores, is a small, gram-negative rod with one lateral flagellum. It utilized only H2 as an electron donor and tetrachloroethene and trichloroethene as electron acceptors in an anaerobic respiration process; it could not grow fermentatively. Acetate served as a carbon source in a defined medium containing iron as the sole trace element, the two vitamins thiamine and cyanocobalamin, and the three amino acids arginine, histidine, and threonine. The cells contained menaquinones and b-type cytochromes. The G+C content of the DNA was 45.3 +/- 0.3 mol%. The cell wall consisted of type-A3gamma peptidoglycan with ll-diaminopimelic acid and one glycine as an interpeptide bridge. The cells are surrounded by an S-layer; an outer membrane was absent. Comparative sequence analysis of the 16S rRNA sequence showed that PER-K23 is related to gram-positive bacteria with a low G+C content of the DNA. Based on the cytological, physiological, and phylogenetic characterization, it is proposed to affiliate the isolate to a new genus, Dehalobacter, with PER-K23 as the type strain of the new species Dehalobacter restrictus.

Anaerobiosis↗

PubMed1998

The fermenting bacterium Malonomonas rubra is phylogenetically related to sulfur-reducing bacteria and contains a c-type cytochrome similar to those of sulfur and sulfate reducers.

Malonomonas rubra is a microaerotolerant fermenting bacterium which can maintain its energy metabolism for growth by decarboxylation of malonate to acetate. 16S rRNA sequence analysis revealed that M. rubra is closely related to the cluster of mesophilic sulfur-reducing bacteria within the delta subclass of the Proteobacteria, with the fermenting bacterium Pelobacter acidigallici and the sulfur reducers Desulfuromusa kysingii, D. bakii and D. succinoxidans as closest relatives. The cells contain high amounts (up to 12% of the total cell protein content) of a c-type cytochrome which is present mainly (> 60%) in the cytoplasm and to minor parts in the periplasm (> 20%) and associated with the membrane fraction (> 10%), independent of the growth substrate. This cytochrome is a tetraheme cytochrome of 13,700 Da molecular mass with a midpoint redox potential of -0.210 V.M. rubra does not reduce sulfur or ferric iron compounds. Since this cytochrome appears not to be involved in the energy metabolism it is concluded that it is a remnant of sulfur-reducing ancestors of this bacterium, without a conceivable physiological function in its present energy metabolism.

Cytochrome c Group↗

PubMed1998

Lactobacillus amylolyticus sp. nov., isolated from beer malt and beer wort.

Some of the strains used for the biological acidification in breweries belong to L. delbrueckii subsp. delbrueckii, L. delbrueckii subsp. lactis or L. fermentum. However, more recent studies showed that most strains isolated are physiologically different from the above mentioned species and were tentatively allocated to Lactobacillus amylovorus. Genotypic studies of 25 strains exclusively isolated from beer malts and beer worts, showed, that there were differences to the type strain of L. amylovorus concerning DNA-DNA similarities and the sequences of their 16S and 23S rRNA genes. Therefore, we propose to combine these strains in a new species of the genus Lactobacillus, namely L. amylolyticus. Strain DSM 11664 is proposed as the type strain. An rRNA targeted oligonucleotide probe was designed that allows a fast and reliable identification of Lactobacillus amylolyticus.

Bacterial Typing Techniques↗

PubMed1998

Hymenobacter roseosalivarius gen. nov., sp. nov. from continental Antartica soils and sandstone: bacteria of the Cytophaga/Flavobacterium/Bacteroides line of phylogenetic descent.

Aseptically collected sandstone and soil samples from the antarctic Dry Valleys were inoculated into oligotrophic media and incubated under low light intensities. A total of 41 Gram-negative isolates were obtained with reddish colonies spreading on agar. A sandstone isolate and four soil strains were characterized further. They were nearly identical in morphological, physiological, biochemical and chemotaxonomic properties. They produced large amounts of extracellular polymer and utilized for growth: glucose, saccharose, mannitol, sorbitol, L-aspartate, malate and acetate, but not D-ribose, adonitol, DL-alanine, glutamate, glycolate, lactate or succinate. All strains hydrolyzed gelatin, starch, casein, xylan, Tweens 80 or 60 and dead or living yeast cells, but not cellulose or pectin. Nitrate was not reduced, ethanol was not oxidized and acid was not produced from maltose, mannitol or dulcitol. Ammonia was not produced from peptone. They were strictly aerobic. Major fatty acids were n 16:1 d 9, n 16:1 d 11, n 17:1 d 11, and i 15:0. The strains contained the quinone MK-7 and phosphatidylethanolamine as the main phospholipid. The base ratio ranged from 55 to 61 mol% G+C. A 16S rRNA sequence analysis of strains AA-688 and AA-718 showed these to be identical and to represent a special phylogenetic group within the Cytophaga/Flavobacterium/Bacteroides major line of descent. Three soil strains labeled "Taxeobacter" Txc1, Txg1, and Txo1 (Reichenbach, 1992) belonged to the same group but had lower sequence similarities (<95%). Some of their characteristics were different from those of the antarctic strains: the utilization of C-compounds, hydrolysis of polymers, temperature tolerances, major fatty acids and base ratios. Txc1 and Txg1 may later have to be considered as members of this group, possibly on the species level, while Txo1 could represent a different related genus. It is concluded that the five antarctic strains represent a new genus and species for which the name of Hymenobacter roseosalivarius is proposed. The type strain is AA-718T (DSM 11622T).

Antarctic Regions↗

PubMed1998

Application of 23S rDNA-targeted oligonucleotide probes specific for enterococci to water hygiene control.

Identification of enterococci species by hybridization with recently designed species-specific and group-specific 23S rDNA-targeted oligonucleotide probes was superior to results obtained with a common biochemical test panel. Considering these findings, a molecular biological procedure for the detection of enterococci in water samples was developed. A short enrichment is followed by an amplification step and a hybridization reaction in microtiter plate format. The detection limit is about 1 CFU/ml, and results are available within 26 h.

Bacterial Typing Techniques↗

PubMed1998

Phylogenetic identification of two major nitrogen-fixing bacteria associated with sugarcane.

Acetobacter diazotrophicus and Herbaspirillum seropedicae were identified by genetic methods based on 16S rRNA sequences. A specific PCR method in combination with probing was developed for A. diazotrophicus. The PCR system includes four primers, of which the primers named AC (CTGTTTCCCGCAAGGGAC) and DI (GCGCCCCATTGCTGGGTT) generated an 445 bp amplicon in all of the 11 A. diazotrophicus strains tested. The phylogenetic position of H. seropedicae was determined. H. seropedicae forms with Oxalobacter formigenes a separate lineage in the beta-subclass of Proteobacteria.

Acetobacter↗

PubMed1998

Identification of streptococci from Greek Kasseri cheese and description of Streptococcus macedonicus sp. nov.

Taxonomic studies were performed on some Streptococcus-like organisms isolated from naturally fermented Greek Kasseri cheese. By SDS-PAGE analysis of whole-cell proteins the group was found to be quite different from Streptococcus thermophilus. Comparative 16S and 23S rRNA sequence analyses showed that the isolates represent a new species within the genus Streptococcus, where they are most closely related to the Streptococcus bovis cluster. On the basis of these phylogenetic results and some phenotypic differences, a new species, Streptococcus macedonicus, is proposed. The type strain is ACA-DC 206.

Bacterial Proteins↗