Application of nested polymerase chain reaction for the detection of as yet uncultured organisms of the class Actinobacteria in environmental samples

Different groups of as yet uncultured, phylogenetically deeply rooting organisms of the class Actinobacteria have been shown previously by analysis of 16S rDNA clone libraries to be present in different soil and marine environments of worldwide distribution. In order to monitor specifically the occurrence of two of the actinobacterial groups among bacterial DNA extracted from different environments, a nested polymerase chain reaction (PCR) method, aiming at the sensitive detection of 16S rDNA, was used. Analysis of a clone library generated from one of these 16S rDNA-derived, group-specific PCR products of DNA from a forest soil broadened the spectrum of putative taxa within the two phylogenetic groups. Application of this PCR system demonstrated the presence of these organisms in natural environments and in a sample of dried and moistened peat. Fluctuations in the detectability of the target groups in different samples from the same site strongly suggest that the taxon diversity of a 16S rDNA clone library must be regarded as a temporal genomic snapshot taken from this environmental community.     

Bacillus aeolius sp. nov. a novel thermophilic,halophilic marine Bacillus species from Eolian Islands (Italy)

Phylogenetic relationships of a thermophilic, halophilic, aerobic spore-forming strain 4-1(T), isolated from the water of a shallow sea hot spring at Vulcano Island (Italy), revealed its relatedness to members of the genus Bacillus. Chemotaxonomic and phenotypic properties of strain 4-1(T) are sufficiently different from related moderately thermophilic species, e.g., B. smithii, B. fumarioli, B. oleronius, B. sporothermodurans and B. infernus to describe strain 4-1(T) as a new Bacillus species, for which the name Bacillus aeolius sp. nov. is proposed. Strain 4-1(T) is characterised by the potential biotechnological important properties such as exopolysaccharide production, surfactant activity, and utilisation of hydrocarbons.     

Radioisotopic, culture-based, and oligonucleotide microchip analyses of thermophilic microbial communities in a continental high-temperature petroleum reservoir

Activity measurements by radioisotopic methods and cultural and molecular approaches were used in parallel to investigate the microbial biodiversity and its physiological potential in formation waters of the Samotlor high-temperature oil reservoir (Western Siberia, Russia). Sulfate reduction with rates not exceeding 20 nmol of H(2)S liter(-1) day(-1) occurred at 60 and 80 degrees C. In upper horizons (AB, A, and B), methanogenesis (lithotrophic and/or acetoclastic) was detected only in wells in which sulfate reduction did not occur. In some of the wells from deeper (J) horizons, high-temperature sulfate reduction and methanogenesis occurred simultaneously, the rate of lithotrophic methanogenesis exceeding 80 nmol of CH(4) liter(-1) day(-1). Enrichment cultures indicated the presence of diverse physiological groups representing aerobic and anaerobic thermophiles and hyperthermophiles; fermentative organotrophs were predominant. Phylogenetic analyses of 15 isolates identified representatives of the genera Thermotoga, Thermoanaerobacter, Geobacillus, Petrotoga, Thermosipho, and Thermococcus, the latter four being represented by new species. Except for Thermosipho, the isolates were members of genera recovered earlier from similar habitats. DNA obtained from three samples was hybridized with a set of oligonucleotide probes targeting selected microbial groups encompassing key genera of thermophilic bacteria and archaea. Oligonucleotide microchip analyses confirmed the cultural data but also revealed the presence of several groups of microorganisms that escaped cultivation, among them representatives of the Aquificales/Desulfurobacterium-Thermovibrio cluster and of the genera Desulfurococcus and Thermus, up to now unknown in this habitat. The unexpected presence of these organisms suggests that their distribution may be much wider than suspected.     

Identification of aerobic mesophilic bacilli isolated from board and paper products containing recycled fibres

AIMS: To identify aerobic mesophilic bacteria isolated from coreboard, kitchen roll paper and food packaging boards containing recycled fibres and to create a rapid fingerprint-based database for their identification. METHODS AND RESULTS: A total of 197 isolates and 20 relevant type strains were characterized by automated ribotyping and as far as possible identified by the similarities of their riboprints to the relevant type strains. One strain from each unidentified ribotype, a total of 87 strains, was subjected to partial 16S rDNA sequencing and in most cases also to fatty acid analysis and physiological tests. From the isolates 113 and seven different ribotypes were generated belonging to the genera Bacillus and Paenibacillus, respectively. The dominating species, or closest related to them, were B. simplex (22.8% of isolates), B. licheniformis (18.3%) and B. amyloliquefaciens (12.7%); 5.1% of the isolates were identified as B. cereus, a potential food-borne pathogen. In particular, this species was present in one food packaging board (26.3% of isolates). Based on these results, 40.1% of the isolates and 45.0% of ribotypes were so different from the relevant type strains that they may represent novel species. CONCLUSIONS: All isolates were aerobic spore-formers, indicating that all non-spore-formers were eliminated during the drying stage of the processes. Although many isolates could be affiliated to described species of Bacillus or Paenibacillus, a significant proportion of the isolates could not be identified unambiguously as members of a described species. SIGNIFICANCE AND IMPACT OF THE STUDY: A RiboPrint identification database, composed of 120 composite patters, was established for bacteria originating from the pulp and paper industry. Considering the discrimination power of ribotyping, this database will be extremely useful in future for the reliable and rapid identification of bacteria isolated from pulp and paper industrial sources.     

Psychromonas antarcticus gen. nov., sp. nov., a new aerotolerant anaerobic, halophilic psychrophile isolated from pond sediment of the McMurdo Ice Shelf, Antarctica

A gram-negative, rod- to oval-shaped, aerotolerant anaerobic bacterium was isolated from an anaerobic enrichment inoculated with sediment taken from below the cyanobacterial mat of a high-salinity pond near Bratina Island on the McMurdo Ice Shelf, Antarctica. The organism was positive for terminal oxidase and catalase and was motile by means of a polar flagellum. Optimal growth of anaerobic cultures occurred at 12° C, at pH 6.5, and at an NaCl concentration of 3% (w/v). Of a variety of polysaccharides tested, only starch and glycogen supported growth. No growth was observed on cellulosic substrates and xylan, and the organism was unable to attack esculin. Monosaccharides and disaccharides, including the cyanobacterial cell-wall constituent N-acetyl glucosamine, were fermented. Per 100 mol of hexose, the following products (in mol) were formed: acetate, 60; formate, 130; ethanol, 56; lactate, 73; CO₂, 15; and butyrate, 2. Propionate, ethanol, n-propanol, n-butanol and succinate were not detectable in the culture medium (< 1 mol per 100 mol of monomer). Hydrogen was not detected in the head space (detection limit < 10^–5 atm). Growth yields in aerobic static liquid cultures were slightly higher than those in anaerobic culture, and fermentation favoured acetate at the expense of electron sink products. Growth was inhibited in aerobic shaking cultures, and the organism did not utilize nitrate or sulfate as electron acceptors. The G+C content of the DNA from the bacterium was 42.8 mol%. A phylogenetic analysis indicated that the organism is a member of the γ-subgroup of Proteobacteria, but that it is distinct from other members of this group based on the sequence of its 16S rRNA gene, mol% G+C, morphology, and physiological and biochemical characteristics. It is designated as a new genus and species; the type strain is star-1 (DSM 10704). Received: 17 June 1996 / Accepted: 13 October 1997     

The ecology of Biomphalaria sudanica in Lake Ziway, Ethiopia

A longitudinal study was made over a period of 24 months from January 2001 to December 2002 in Lake Ziway, Ethiopia, to elucidate factors influencing changes in the population density of Biomphalaria sudanica and to identify seasons of peak transmission of Schistosoma mansoni. Between January 2001 and December 2002, bimonthly visits were made to the lake to collect data on snail population density and schistosome infection in snails, vegetation and lake water level. Water samples were collected during the dry and rainy seasons and chemical analyses were made for major cations, anions, electrical conductivity, total hardness and pH using standard methods for the examination of water and wastewater. The seasonal fluctuation in the population density of B. sudanica was found to be highly associated with rainfall, water level of the lake and vegetation availability and abundance. Water chemical analysis showed that major cations and anions were all within the range reported for other African freshwaters. Both malacological and meteorological data show that snail population density in Lake Ziway peaks at the end of the long rainy season (June–August), coinciding with a seasonal transmission peak of schistosomiasis.     

Peptidoglycantyp und Zusammensetzung der Zellwandpolysaccharide vonCellulomonas cartalyticum und einigen coryneformen Organismen

Cellulomonas cartalyticum was found to contain a peptidoglycan type different from that of the other species ofCellulomonas. The diamino acid is lysine instead of ornithine and the interpeptide bridge consists ofd-Asp-d-Ser. The same peptidoglycan type occurs inCorynebacterium manihot, Brevibacterium liticum andArthrobacter luteus. These non cellulolytic organisms are most likely not closely related withCellulomonas cartalyticum, as indicated by the very different G+C content of their DNA, although they formed a narrow cluster includingC. cartalyticum when numeric taxonomical methods were applied.

     

DNS/DNS-Homologiestudien innerhalb der Gattung Pediococcus

Zusammenfassung An 17 Stämmen der Gattung Pediococcus, die auf Grund charakteristischer physiologisch-biochemischer Merkmale aus einer Anzahl von etwa 500 Kulturen unterschiedlicher Herkunft ausgewählt wurden, werden mit Hilfe der DNS/DNS-Hybridisierung die verwandtschaftlichen Beziehungen untersucht. Dabei wird die Eigenständigeit der Arten P. pentosaceus, P. acidilactici, P. parvulus, P. damnosus und P. halophilus bestätigt. Der für P. acidilactici vorgeschlagene Neotyp-Stamm NCDO 1859 ist jedoch nach den vorliegenden Ergebnissen ein echter P. pentosaceus. Ein Stamm von P. dextrinicus, der früher bereits als P. cerevisiae var. dextrinicus beschrieben wurde, zeigt zu keiner untersuchten Art eine genetische Verwandtschaft auf der Basis der DNS/DNS-Homologie. Zwei aus Bier und Bierhefe isolierte Stämme, die hinsichtlich des Phänotyps P. parvulus nahestehen, zeigen weder zu dieser Art noch zu allen anderen bisher beschriebenen Arten hinsichtlich des Genotyps einen hohen Verwandtschaftsgrad.

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DNA-DNA homology studies on the genus Pediococcus

17 strains of Pediococcus, including new isolates and strains from various collections, were investigated by nucleic acid hybridisation. Most of the newly isolated strains, selected from about 500 strains according to physiological and biochemical characters, could be attributed to known species. The reassociation data demonstrate the separate status of P. pentosaceus, P. acidilactici, P. damnosus, P. parvulus and P. halophilus. The proposed neotype strain of P. acidilactici NCDO 1859, however, has been found to be in reality a genuine P. pentosaceus. P. dextrinicus, formerly described as P. cerevisiae var. dextrinicus showed a remarkable lack of relationship to any of the strains tested. Two isolates exhibiting a high phenotypic similarity to P. parvulus show little genetic relationship to this and especially to all other species.

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Abkürzungen CTAB N-cetyl-N,N,N-trimethyammoniumbromid - EDTA Äthylendiamintetraessigsäure - PPO 2,5-Diphenyloxazol - POPOP 1,4-Bis-(5-phenyl-2-oxazolyl)-benzol - SSC Saline-Trinatriumcitrat (0,15M NaCl, 0,015M Trinatriumcitrat, pH 7,0) - Tris Tris(hydroxymethyl)-aminomethan - T _m mittlere Schmelztemperatur der DNS Adresse für Sonderdruck-Anforderungen     

The relatedness of Prochloron sp. Isolated from different didemnid ascidian hosts

The relationship of Prochloron sp. isolated from four different didemnid ascidian hosts, namely Lissoclinum patella, Lissoclinum voeltzkowi, Diplosoma virensand Trididemnum cyclops was elucidated by comparative analysis of their 16S ribosomal ribonucleic acid (RNA). The oligonucleotide catalogues of the 16S rRNA obtained are almost identical, indicating a very close relationship among the prochlorophytes investigated. Phylogenetically Prochloron is a member of Cyanobacteriales.