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Coaggregation of Fusobacterium nucleatum, Selenomonas flueggei, Selenomonas infelix, Selenomonas noxia, and Selenomonas sputigena with strains from 11 genera of oral bacteria.

Twenty-eight strains of Fusobacterium nucleatum and 41 Selenomonas strains, including S. sputigena (24 strains), S. flueggei (10 strains), S. infelix (5 strains), and S. noxia (2 strains), were tested for their ability to coaggregate with each other and with 49 other strains of oral bacteria representing Actinobacillus, Actinomyces, Bacteroides, Capnocytophaga, Gemella, Peptostreptococcus, Porphyromonas, Propionibacterium, Rothia, Streptococcus, and Veillonella species. Selenomonads coaggregated with fusobacteria and with Actinomyces naeslundii PK984 but not with any of the other bacteria, including other selenomonads. In contrast, fusobacteria coaggregated with members of all genera, although not with all strains of each species tested. Each fusobacterium strain appeared to have its own set of partners and coaggregation properties, unlike their partners, whose coaggregation properties in earlier surveys delineated distinct coaggregation groups. Coaggregations of fusobacteria with the 63 gram-negative strains were usually inhibited by EDTA, whereas those with the 27 gram-positive strains were usually not inhibited. Likewise, lactose-inhibitable coaggregations were common among some strains of fusobacteria and some strains from each of the genera containing gram-negative partners but were rarely observed with gram-positive partners. Heating the fusobacteria at 85 degrees C for 30 min completely prevented coaggregation with most partners, suggesting the involvement of a protein on the fusobacteria. Heat treatment of many of the gram-negative partners not only enhanced their coaggregation with the fusobacteria but also changed lactose-sensitive coaggregations to lactose-insensitive coaggregations. Although fusobacteria coaggregated with a broader variety of oral partner strains than any other group of oral bacteria tested to date, each fusobacterium exhibited coaggregation with only a certain set of partner strains, and none of the fusobacteria adhered to other strains of fusobacteria, indicating that recognition of partner cell surfaces is selective. The strains of F. nucleatum are heterogeneous and cannot be clustered into distinct coaggregation groups. Collectively, these results indicate that coaggregation between fusobacteria and many gram-negative partners is significantly different from their coaggregation with gram-positive partners. The contrasting variety of partners for fusobacteria and selenomonads supports the concept of coaggregation partner specificity that has been observed with every genus of oral bacteria so far examined.

Actinomyces↗

Phylogenetic characterization of Centipeda periodontii, Selenomonas sputigena and Selenomonas species by 16S rRNA gene sequence analysis.

The nearly complete 16S rRNA gene sequences for oral Gram-negative anaerobic motile bacteria, Centipeda periodontii, Selenomonas sputigena and Selenomonas species (formerly S. sputigena type strain), were determined in order to unveil their relationship to other oral motile bacteria. To determine the phylogenetic characterization of these bacteria, their 16S rRNA gene sequences were obtained and compared with those from the ribosomal sequence databases previously reported. The 16S rRNA gene sequences of these bacteria were similar to those of Selenomonas ruminantium and Schwartzia succinivorans isolated from rumens, and to Pectinatus cerevisiiphilus isolated from spoiled beer. Among oral bacteria, the nucleotide sequence analysis of these bacteria revealed high nucleotide similarity to Veillonella species, whereas low similarity to oral motile bacteria such as Campylobacter species. Phylogenetic analysis clearly confirmed that C. periodontii and two Selenomonas species were classified as relatives of a group besides Selenomonas, Schwartzia, and Pectinatus species, and not as close relatives to oral motile bacteria, such as Campylobacter species. These results suggest that such oral Gram-negative anaerobic motile bacteria are close relatives of oral bacteria.

Phylogeny↗

Bacteremias caused by Selenomonas artemidis and Selenomonas infelix.

We report two different cases of bacteremia caused by two recently described Selenomonas species, Selenomonas artemidis and Selenomonas infelix. Both species are normally found in human buccal flora. S. artemidis bacteremia appeared in a patient (number 1) who presented with an air-fluid pulmonary cavity and clinical conditions consistent with an anaerobic lung abscess. While the patient improved with antibiotic therapy, cultures of respiratory secretions yielded Mycobacterium tuberculosis. This case demonstrated a strong possibility of a coexisting lung abscess due to S. artemidis. S. infelix bacteremia appeared in a cancer patient (number 2) with heart disease during preterminal acute respiratory distress. It was more difficult in this case to assess the clinical impact of the Selenomonas organisms on the patient.

Adenocarcinoma↗

Phylogenetic relationships of three amino-acid-utilizing anaerobes, Selenomonas acidaminovorans, 'Selenomonas acidaminophila' and Eubacterium acidaminophilum, as inferred from partial 16S rDNA nucleotide sequences and proposal of Thermanaerovibrio acidaminovorans gen. nov., comb. nov. and Anaeromusa acidaminophila gen. nov., comb. nov.

16S rRNA gene sequences of three previously described amino-acid-fermenting anaerobes, Selenomonas acidaminovorans, 'Selenomonas acidaminophila' and Eubacterium acidaminophilum, were determined. All three were found to cluster within the Clostridium and related genera of the subphylum of the Gram-positive bacteria. The thermophile, S. acidaminovorans, formed an individual line of descent and was equidistantly placed between Dethiosulfovibrio peptidovorans and Anaerobaculum thermoterrenum (similarity of 85%), both of which also form single lines of descent. 'S. acidaminophila' was related to Clostridium quercicolum, a member of cluster IX, with a similarity of 90%, whereas E. acidaminophilum was closely related to Clostridium litorale (similarity of 96%) as a member of cluster XI. Based on the phylogenetic data presented in this report and the phenotypic descriptions of these bacteria published previously, it is recommended that S. acidaminovorans be transferred to a new genus, Thermanaerovibrio gen. nov., as Thermanaerovibrio acidaminovorans comb. nov. and 'Selenomonas acidaminophila' be transferred to a new genus, Anaeromusa gen. nov., as Anaeromusa acidaminophila comb. nov. Though the transfer of E. acidaminophilum to a new taxon is justified, this is not recommended until the taxonomic status of all the members of cluster XI has been reviewed.

Amino Acids↗

Identification of Selenomonas species by whole-genomic DNA probes, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, biochemical tests and cellular fatty acid analysis.

Nonisotopic, whole-genomic DNA probes, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), biochemical tests in microtiter trays and cellular fatty acid (CFA) analysis were compared for the identification of 5 oral Selenomonas species. DNA probes were prepared by biotin-labeling DNA extracted from the type strains of Selenomonas noxia, Selenomonas flueggei, Selenomonas artemidis, Selenomonas infelix and Selenomonas sputigena. The probes were hybridized with DNA from 21 reference strains, 18 fresh isolates of Selenomonas species, and 21 strains of other oral gram-negative species. Target DNAs were obtained by in situ extraction of colonies blotted onto filter paper. Streptavidin-linked alkaline phosphatase was used to detect homologous reactions of probe and target DNA. Each Selenomonas species DNA probe reacted with reference strains of only that species. All Selenomonas strains that reacted with the DNA probe for a particular species gave similar biochemical test results, SDS-PAGE protein profiles, and CFA profiles to those of the type strain of the corresponding species. All the methods tested were useful for identifying the species, and all yielded similar identifications of the fresh isolates. The DNA probes, however, had the potential for identifying Selenomonas species directly from primary isolation plates or plaque samples.

Bacteroidaceae↗

Selenomonas lipolytica sp. nov., an obligately anaerobic bacterium possessing lipolytic activity.

A novel, oligately anaerobic bacterium capable of hydrolysing lipids was isolated from a tropical anaerobic lagoon receiving waste water from an edible oil mill. The isolate had many characteristics similar to those of members of the genus Selenomonas. The isolate showed lipolytic activity on tributyrin, triolein and groundnut oil in qualitative plate clearance assays, which has not been reported for the type strain of the genus Selenomonas. It did not require n-valerate supplementation for growth on glucose. Acetate and propionate were the only volatile fatty acids produced from glucose fermentation with propionate as the major end product. The isolate could grow optimally at pH 6.8 and at a temperature of 40 degrees C. It could tolerate NaCl concentrations of up to 40 g l-1. The G&C content of the DNA was 40 mol% as determined by thermal denaturation analysis. Comparison of partial 165 rRNA gene sequences revealed that the isolate was most closely related to genus Selenomonas with 91% sequence similarity (250 bp compared) to Selenomonas ruminantium strain GA 192. On the basis of the results obtained in the present investigation, it is suggested that a new species of Selenomonas should be created for this novel isolate and the name Selenomonas lipolytica is proposed for this new species. The type strain is strain CF1BT (= MCMB 505T).

Anti-Bacterial Agents↗

Isolation of Selenomonas spp. from lesions and non-digestive organs of cows, pigs and man.

Selenomonas spp. were isolated for the first time from lesions and non-digestive organs which were apparently normal in 2 cows, 6 pigs, and 1 human being. Identification as Selenomonas was based firstly on electron microscopical observation and secondly on fermentation products. They were divided into 3 major groups by biological properties, as well as by the patterns of these products. It has been confirmed that the habitats of organisms of the genus Selenomonas are generally digestive organs, including the rumen of the ruminant, the cecum of the guinea pig, and the oral cavity of man. The existence of these organisms in lesions and non-digestive organs in such animals and man, however, has been unknown as yet. Moreover, it has been completely unknown about the habitat of these organisms in swine. The findings obtained suggested the possibility of invasion of Selenomonas into other parts than the digestive organ in some animals and the presumable existence of the organism in the swine digestive organs. The role of Selenomonas as a secondary invader into some animals was proposed.

Animals↗

Selenomonas bacteraemia--case report and review of the literature.

Selenomonas species are crescent shaped Gram-negative bacilli with a characteristic tuft of flagella located on the concave surface. They are normally found in human gingiva or the rumen of herbivores. The first case of Selenomonas bacteraemia to be reported in a patient immunocompromised by malignant disease is described and the two previously reported cases of Selenomonas bacteraemia as reviewed. The importance of careful anaerobic culturing to recover the organism and special diagnostic techniques to classify the bacteria as Selenomonas species are emphasised. These organisms may cause serious human disease including bacteraemia.

Aged↗

Characterization of a major envelope protein from the rumen anaerobe Selenomonas ruminantium OB268.

Cell envelopes from the Gram-negative staining but phylogenetically Gram-positive rumen anaerobe Selenomonas ruminantium OB268 contained a major 42 kDa heat modifiable protein. A similarly sized protein was present in the envelopes of Selenomonas ruminantium D1 and Selenomonas infelix. Sodium dodecyl sulfate polyacrylamide gel electrophoresis of Triton X-100 extracted cell envelopes from S. ruminantium OB268 showed that they consisted primarily of the 42 kDa protein. Polyclonal antisera produced against these envelopes cross-reacted only with the 42 kDa major envelope proteins in both S. ruminantium D1 and S. infelix, indicating a conservation of antigenic structure among each of the major envelope proteins. The N-terminus of the 42 kDa S. ruminantium OB268 envelope protein shared significant homology with the S-layer (surface) protein from Thermus thermophilus, as well as additional envelope proteins containing the cell surface binding region known as a surface layer-like homologous (SLH) domain. Thin section analysis of Triton X-100 extracted envelopes demonstrated the presence of an outer bilayer over-laying the cell wall, and a regularly ordered array was visible following freeze-fracture etching through this bilayer. These findings suggest that the regularly ordered array may be composed of the 42 kDa major envelope protein. The 42 kDa protein has similarities with regularly ordered outer membrane proteins (rOMP) reported in certain Gram-negative and ancient eubacteria.

Amino Acid Sequence↗

[The occurrence and properties of Selenomonas ruminantium bacteria in calves during the time of milk feedings].

The occurrence, morphological properties, urease and alpha-amylase activities of Selenomonas ruminantium were investigated in calves in the period of milk nutrition. The average number of the organisms was found to range between 10(7) and 10(8) per 1 millilitre of rumen contents. The alpha-amylase activity of the Selenomonas strains ranged from 0.1 to 8.8 ncat.ml-1 whereas their urease activity ranged from 6.3 to 261 ncat.ml-1 of nutrient medium. The presence of morphologically typical Selenomonas ruminantium and spontaneous formation of spheroplasts were found by electron microscopic examination.

Age Factors↗

Cloning of the L-lactate dehydrogenase gene from the ruminal bacterium Selenomonas ruminantium HD4.

A clone from a Selenomonas ruminantium HD4 Lambda ZAP II genomic library was isolated by its ability to complement the anaerobic growth deficiency of an Escherichia coli (pfl, ldh) double mutant. The 1.0-kb insert from the clone was sequenced and revealed a single open reading frame (ORF, 957-bp) which was preceded by a putative Shine-Dalgarno (SD) sequence (AGGGGG). The potential SD sequence corresponded to 3' 16S rRNA sequences of various Selenomonas strains. The ORF was predicted to encode a protein of 318 amino acids with a calculated molecular mass of 34,975 Da and an isoelectric point of 5.54. In addition, the ORF contained 51 mol % G + C and this is consistent with the average G + C content (54%) of the S. ruminantium chromosome. The cloned S. ruminantium gene exhibited 59% nucleotide identity and 61% deduced amino acid similarity with L-lactate dehydrogenases (L-LDH) of Pediococcus acidilactici and Bacillus megaterium, respectively. Incorporation of the cloned S. ruminantium gene into E. coli DC1368 (pfl, ldh) restored anaerobic growth on glucose and L-LDH activity was detected in cell extracts. Because lactate accumulation within the rumen can be detrimental to animal performance, characterizing the gene(s) involved in lactate production by predominant ruminal bacteria will lead to a better understanding of lactate metabolism within the rumen.

Amino Acid Sequence↗

Cloning of the O-acetylserine lyase gene from the ruminal bacterium Selenomonas ruminantium HD4.

The gene coding for O-acetylserine lyase (OASL) was cloned from a Selenomonas ruminantium HD4 Lambda ZAP II genomic library by degenerative probe hybridization and complementation. Sequence analysis revealed a 933 bp ORF with a G + C content of 53%. The ORF had significant homology with enzymes involved in cysteine biosynthesis. A CuraBLASTN homology search showed that the ORF shared 59% nucleotide identity with the cysK of Bacillus subtilis. The deduced amino acid sequence exhibited high (>70%) similarity with the CysK of B. subtilis and other cysteine synthesis proteins from Mycobacterium tuberculosis, Mycobacterium leprae, and Spinacia oleracea. Further analysis predicted that the gene product was a member of the pyridoxal phosphate enzyme family and of cytoplasmic origin. Phylogenetic analysis clustered the S. ruminantium gene product with the OASLa isoform of B. subtilis and the OASLb isoforms of Streptococcus suis, Escherichia coli, and Campylobacter jejuni. The OASL of S. ruminantium HD4 was also able to complement the cysM cysK double mutations in Escherichia coli NK3 and allow for growth on minimal media that contained either sulfate or thiosulfate as the sole source of sulfur. These results suggest that the gene functions as a cysM in S. ruminantium HD4. In conclusion, this research describes the cloning and expression of an O-acetylserine lyase gene from the predominant ruminal anaerobe S. ruminantium HD4. To our knowledge, this is the first report characterizing genes involved in sulfur metabolism from the genus Selenomonas.

Amino Acid Sequence↗

16S-23S rDNA spacer of Pectinatus, Selenomonas and Zymophilus reveal new phylogenetic relationships between these genera.

The 16S-23S rDNA spacer regions of two Pectinatus species, two Zymophilus species and one Selenomonas species were cloned after PCR amplification. The results of PCR amplification showed that these species had two types of spacer regions which differ in molecular size (long and short). Only the long spacer regions in these bacteria contained one or two tRNA genes (alanine and/or isoleucine). The spacer regions in these bacteria had a relatively high level of homology. Homology was particularly high for bacteria belonging to the same genus. Interestingly, the order of the two tRNA genes present in the long spacer regions of Pectinatus and Selenomonas was the reverse of that which had been previously reported for other bacteria. The results of spacer homology analysis and the order of the tRNA genes suggest that the taxonomic classification of anaerobic bacteria isolated from the brewing process should be re-examined.

Beer↗

Detection of N6-methyladenine in GATC sequences of Selenomonas ruminantium.

The presence of N6-methyladenine in GATC sequences in DNA of Selenomonas ruminantium was investigated using sensitive methylation discriminating isochizomeric restriction enzymes analysis. Methylated adenine was detected in 8 out of 18 tested strains belonging to the subsp. lactilytica of S. ruminantium. No corresponding restriction activity was detected in three tested strains. No GATC methylation was detected in 3 analysed S. ruminantium subsp. ruminantium strains. Sustainable progress was achieved in the molecular biology of ruminal microorganisms in the last decade. Many different genes acting in the cell wall degradation were cloned and characterized. As practically all cloning experiments were done in Escherichia coli cells, there is a lack of data about regulation of gene(s) expression in the natural hosts. However, much better understanding of molecular genetics of ruminal bacteria is required for improving rumen functions by genetic modifications of rumen bacteria. DNA methylation is main mechanism of the control of gene expression in eukaryotes. In prokaryotes, DNA methylation influences wide variety of important cellular functions as accessibility of DNA to digestion by restriction endonucleases, control of replication initiation, transposition, phage DNA packaging, including positive and negative regulation of gene expression. Most of the DNA methyltransferases; enzymes which facilitate methylation; identified in prokaryotes are part of restriction modification systems (WILSON and MURRAY 1991. Another class of methyltransferase are independent methylases like Dam and Dcm in Escherichia coli. Dam methylase recognizes the sequence GATC and methylates adenine at N6 position (BARRAS and MARINUS 1989). The methylation is the only documented case of prokaryotic methylation involved in the regulation of cellular process (NOYER-WIEDNER and TRAUTNER 1993). Screening of large number of bacteria have detected the presence of Dam methylation in cyanobacteria as well as in the group of related families of Enterobacteriaceae (which includes E. coli), Parvobacteriaceae and Vibrionaceae. Methylated GATC sequences have been found in several other bacterial species (NOYER-WIEDNER and TRAUTNER 1993), however, it could be assumed that some of these methylations are due to the presence of restriction modification systems. There is no data about the presence of Dam methylation neither in Selenomonas ruminantium nor in any other ruminal bacteria. Bacteria of S. ruminantium species are known to contain frequently restriction modification systems. Several restriction endonucleases were characterized from these bacteria (VANAT et al., 1993, PRISTAS et al. 1994, 1995). During characterization of modification activities associated with these endonucleases we have detected the modification of adenine in GATC sequences of DNA of this species.

Adenine↗

SruI restriction endonuclease from Selenomonas ruminantium.

SruI, specific restriction endonuclease, has been characterized from Selenomonas ruminantium isolated from the rumen of fallow deer. Results from the study demonstrate that S. ruminantium 18D possesses a type II restriction endonuclease, which recognizes the sequence 5'-TTT decreases AAA-3'. The recognition sequence of SruI was identified using digestions on pBR322, pBR328, pUC18, M13mp18RF, pACYC184 and lambda DNA. The cleavage patterns obtained were compared with computer-derived data. SruI recognises the palindromic hexanucleotide sequence and cleaves DNA after the third T in the sequence, producing blunt ends. The purification and characterization of restriction endonuclease SruI presented here is the first described for Selenomonas ruminantium spp. and demonstrates that this microorganism possesses a DNA-cleaving enzyme with the same specificity as DraI or AhaIII.

Animals↗

Taxonomic study of anaerobic, gram-negative, rod-shaped bacteria from breweries: emended description of Pectinatus cerevisiiphilus and description of Pectinatus frisingensis sp. nov., Selenomonas lacticifex sp. nov., Zymophilus raffinosivorans gen. nov., sp. nov., and Zymophilus paucivorans sp. nov.

A collection of 47 strains of obligately anaerobic, gram-negative, rod-shaped bacteria that were isolated mainly from spoiled beer and pitching yeast was studied to learn more about their taxonomic positions. A new species of the genus Pectinatus, Pectinatus frisingensis, a new species of the genus Selenomonas, Selenomonas lacticifex, and a new genus comprising two species, Zymophilus raffinosivorans and Zymophilus paucivorans, are described. All of the strains contained directly cross-linked meso-diaminopimelic acid-containing peptidoglycan and in addition the diamine cadaverine or (rarely) putrescine. The diamine was covalently linked to the alpha-carboxyl group of D-glutamic acid in the peptide subunit of peptidoglycan. Lipid F was also found as a characteristic cellular compound. The phylogenetic relationships of members of these new species were examined by reverse transcriptase sequencing of 16S rRNA or by DNA-DNA hybridization studies or both. All of the organisms belong to the subdivision containing species with gram-negative cell walls within the phylum of gram-positive bacteria. This finding is in good agreement with the presence of a peptidoglycan that contains diamine.

Anaerobiosis↗

Fatty acid composition of oral isolates of Selenomonas.

Fatty acids of 16 strains of Selenomonas isolated from the human oral cavity were examined by gas-liquid chromatography. The strains showed similar patterns, characterized by the presence of straight-chain fatty acids in the range C11 to C18. Fatty acids of odd-numbered carbon atoms dominated and the major acids were n-pentadecanoate and 3-hydroxytridecanoate. The general fatty acid pattern of Selenomonas differed distinctly from those of other previously analysed anaerobic or microaerophilic Gram-negative bacilli.

Bacteroidaceae↗

Characterization of Egg Yolk Antibodies for Detection and Quantification of Selenomonas ruminantium by Using an Enzyme-Linked Immunosorbent Assay.

The specificity of polyclonal antibodies prepared against strains of Selenomonas ruminantium, the effect of assay conditions, and quantification of individual strains in mixed-cell suspensions of selenomonad strains were examined in this study. Whole-cell suspensions were prepared with pure cultures of S. ruminantium PC18, HD(4), GA192, and D. Each cell suspension was injected into a Leghorn laying hen, and polyclonal antibodies were harvested from eggs laid in week 3 or 7 following initial immunization. Antibodies made to the S. ruminantium strains readily discerned the homologous strain from the heterologous strains. Cross-reactivity among antibodies and the heterologous S. ruminantium strains ranged from 5 to 26%. Among non-S. ruminantium species, cross-reactivity of S. ruminantium antibodies was greatest with Selenomonas sputigena (3 to 34%) and Succinivibrio dextrinosolvens (0 to 37%). Antibodies made to strains GA192 and D were used to quantify a mixture of the two strains. Both antibodies responded to graded concentrations of the homologous antigen in the biculture mixtures in accord with the change in the direct cell counts for each strain (strain D, R = 0.92; strain GA192, R = 0.90). This enzyme-linked immunosorbent assay enabled concurrent and accurate quantification of two strains of S. ruminantium subsp. ruminantium in a mixed-cell suspension with a precision of much less than 1 order of magnitude.

Journal Article↗