Microbiological diversity of generalized aggressive periodontitis by 16S rRNA clonal analysis

Background/aim: The purpose of this study was to determine the bacterial diversity in the subgingival plaque of subjects with generalized aggressive periodontitis by using culture-independent molecular methods based on 16S ribosomal DNA cloning. Methods: Samples from 10 subjects with generalized aggressive periodontitis were selected. DNA was extracted and the 16S rRNA gene was amplified with the universal primer pairs 9F and 1525R. Amplified genes were cloned, sequenced, and identified by comparison with known 16S rRNA sequences. Results: One hundred and ten species were identified from 10 subjects and 1007 clones were sequenced. Of these, 70 species were most prevalent. Fifty-seven percent of the clone (40 taxa) sequences represented phylotypes for which no cultivated isolates have been reported. Several species of Selenomonas and Streptococcus were found at high prevalence and proportion in all subjects. Overall, 50% of the clone libraries were formed by these two genera. Selenomonas sputigena, the species most commonly detected, was found in nine of 10 subjects. Other species of Selenomonas were often present at high levels, including S. noxia, Selenomonas sp. EW084, Selenomonas sp. EW076, Selenomonas FT050, Selenomonas sp. P2PA_80, and Selenomonas sp. strain GAA14. The classical putative periodontal pathogens, such as, Aggregatibacter actinomycetemcomitans, was below the limit of detection and was not detected. Conclusion: These data suggest that other species, notably species of Selenomonas, may be associated with disease in generalized aggressive periodontitis subjects.

Characterization of the Phenol Monooxygenase Gene from Chromobacterium violaceum: Potential Use for Phenol Biodegradation

In this work, the biodegradation mechanism of phenol and sub products (such as catechol and hydroquinone) in Chromobacterium violaceum was investigated by cloning and molecular characterization of a phenol monooxygenase gene in Escherichia coli. This gene (Cvmp) is very similar (74 and 59% of similarity and identity, respectively) to the ortholog from Ralstonia eutropha, bacteria capable of utilizing phenol as the sole carbon source. The phenol biodegradation ability of E. coli recombinant strains was tested by cell-growth in a minimal medium containing phenol as the sole source of carbon and release of intermediary metabolites (catechol and hydroquinone). Interestingly, during the growth of these strains on phenol, catechol, and hydroquinone accumulated transiently in the medium. These metabolites were further analyzed by HPLC. These results indicated that phenol can be initially orto or para hydroxylated to produce cathecol or hydroquinone, respectively, followed by meta-cleavage of aromatic rings. To verify this information, the metabolites obtained from HPLC were submitted to LC/MS to confirm their chemical structure, thereby indicating that the recombinant strains utilize two different routes simultaneously, leading to different ring-fission substrates for the metabolism of phenol. (C) KSBB

Screening for endophytic nitrogen-fixing bacteria in Brazilian sugar cane varieties used in organic farming and description of Stenotrophomonas pavanii sp. nov.

A Gram-negative, rod-shaped, non-spore-forming and nitrogen-fixing bacterium, designated ICB 89(T), was isolated from stems of a Brazilian sugar cane variety widely used in organic farming. 16S rRNA gene sequence analysis revealed that strain ICB 89(T) belonged to the genus Stenotrophomonas and was most closely related to Stenotrophomonas maltophilia LMG 958(T), Stenotrophomonas rhizophila LMG 22075(T), Stenotrophomonas nitritireducens L2(T), [Pseudomonas] geniculata ATCC 19374(T), [Pseudomonas] hibiscicola ATCC 19867(T) and [Pseudomonas] beteli ATCC 19861(T). DNA-DNA hybridization together with chemotaxonomic data and biochemical characteristics allowed the differentiation of strain ICB 89(T) from its nearest phylogenetic neighbours. Therefore, strain ICB 89(T) represents a novel species, for which the name Stenotrophomonas pavanii sp. nov. is proposed. The type strain is ICB 89(T) (=CBMAI 564(T) =LMG 25348(T)).

Role of CcpA in Polyhydroxybutyrate Biosynthesis in a Newly Isolated Bacillus sp MA3.3

The ccpA gene was inactivated in the polyhydroxybutyrate (PHB)-producing strain Bacillus sp. MA3.3 in order to reduce glucose catabolite repression over pentoses and develop improved bacterial strains for the production of PHB from lignocellulosic hydrolysates. Mutant Bacillus sp. MSL7 Delta CcpA are unable to grow on glucose and ammonia as sole carbon and nitrogen sources, respectively. Supplementation of glutamate as the nitrogen source or the substitution of the carbon source by xylose allowed the mutant to partially recover its growth performance. RT-PCR showed that CcpA stimulates the expression of the operon (gltAB), responsible for ammonia assimilation via glutamate in Bacillus sp. MA3.3. Moreover, it was demonstrated that the supplementation of xylose or glutamate was capable of stimulating gltAB operon expression independently of CcpA. In PHB production experiments in mineral media, it has been observed that the glucose catabolite repression over the pentoses was partially released in MSL7. Although the carbohydrate consumption is faster in the ccpA mutant, the biomass and PHB biosynthesis are lower, even with supplementation of glutamate. This is attributed to an increase of acetyl-CoA flux towards the tricarboxylic acid cycle observed in the mutant. Copyright (C) 2011 S. Karger AG, Basel