The biodegradation of polychlorinated biphenyls (PCBs) has been studied extensively as these compounds are toxic to living organisms. The recent development of a genome-based analysis demonstrated that PCB degraders have various metabolic genes for degrading xenobiotics. The presence of many putative genes coding for oxygenases suggested that these degraders have increased their catabolic versatility. The analysis of crystal structures has also progressed in the past few years, and the analysis of reaction intermediates has revealed reaction mechanisms. These results can be applied to the creation of new catalytic functions by extending the substrate specificity of enzymes. A thermophilic PCB degrader was isolated and characterized. There are still difficulties with using PCB degraders for bioremediation. Physiological as well as ecological studies which clarify suitable conditions for degradation should be done for the development of PCB degradation in contaminated soil and sediments.
Populations of phototrophic purple nonsulfur bacteria (PNSB) of up to 104-107 CFU ml-1 were found in photosynthetic sludge and activated sludge plants treating wastewater having different biological oxygen demand (BOD) levels and containing acetate as a main BOD source. There was a positive correlation between the population density of PNSB and the strength of BOD in the aeration tanks. All of the PNSB strains isolated were identified as species of the genera Rhodobacter and Rhodopseudomonas by 16S rRNA gene sequencing. The Rhodobacter species were abundant in high BOD-loaded wastewater environments, while the Rhodopseudomonas species increased at lower BOD levels. Therefore, the PNSB population structure appeared to be greatly affected by the concentration of lower fatty acids as major BOD sources, varying over time and space. The utilization of lower fatty acids with different carbon numbers (C2 to C6) by aerobically or semi-aerobically grown cells of authentic and isolated PNSB strains was evaluated by monitoring substrate-dependent oxygen uptake. The oxidation of the fatty acids by the Rhodobacter strains depended upon the number of carbons in the substrate, while the Rhodopseudomonas strains utilized all substrates equally. A low carbon number was much preferred by the Rhodobacter strains. The affinity for acetate of the Rhodobacter and Rhodopseudomonas strains ranged from 0.14 to 3.0 mM and 0.032 to 0.096 mM, respectively. These results suggest that the concentration and kind of lower fatty acids as major BOD sources are important factors affecting not only the population level but also the species composition of PNSB in wastwater environments.
A DNA fragment that carries the gene encoding 4-hydroxybenzoate 3-hydroxylase (pobA) together with genes encoding a potential regulator (pobR) and a potential transporter (pobK) was cloned from "Corynebacterium cyclohexanicum" strain ATCC 51369, which is herein reclassified as belonging to the genus Arthrobacter. Nucleotide sequencing revealed that the deduced amino acid sequence encoded by the Arthrobacter pobA gene exhibits 42.0-46.7% identity with that of gram-negative bacteria. The gene organization of the pob cluster differs from that of gram-negative bacteria. The pobA gene product (PobA), expressed in Escherichia coli, preferred NADH over NADPH similar to 4-hydroxybenzoate 3-hydroxylase of ATCC 51369. The Arthrobacter pobA gene was inactivated by insertion of pK19mob. The resultant mutant strain, POBA1, grew on neither cyclohexanecarboxylate nor 4-hydroxybenzoate. These results strongly suggest that the cloned pobA gene plays an essential role in the catabolism not only of 4-hydroxybenzoate but also of cyclohexanecarboxylate in strain ATCC 51369.
We analyzed the changes that occurred in a bacterial community from forest soil after adding 3-chlorobenzoate (3CB) by two different PCR-denaturing gradient gel electrophoreses (DGGEs), targeting the 16S rRNA gene and benA encoding the benzoate 1,2-dioxygenase large subunit. The concentration of 3CB in the soil had decreased to about 40% of that added (500 ppm) after seven days at 28°C. Total DNA was directly extracted from the soil before and after incubation, and subjected to PCR-DGGE. Four new bands clearly appeared after the incubation with 3CB in the DGGE profiles of both the 16S rRNA gene and benA. The major bands were sequenced and compared with sequences obtained from DNA databases by phylogenetic analysis. All four of the new bands in the profiles of the 16S rRNA gene were most closely related to Burkholderia spp. Moreover, three of the four bands that intensified after the addition of 3CB in the profiles of benA were also most closely related to known benA sequences derived from Burkholderia strains. These results suggest that several Burkholderia-related bacteria played significant roles in the degradation of 3CB in the soil.
The catabolism of aromatic compounds in Corynebacterium glutamicum was investigated by genome data mining and by experimental analysis. Results indicated that C. glutamicum assimilated different aromatic compounds such as phenol, p-cresol, benzoate, 4-hydroxybenzoate, vanillate, vanillin, resorcinol, 3,5-dihydroxytoluene and 2,4-dihydroxybenzoate. Genome data indicated, and enzyme assays confirmed; the existence of multiple ring-cleavage pathways for the catabolism of central aromatic intermediates; the protocatechuate and catechol branches of the β-ketoadipate pathway, two similar hydroxyquinol pathways and the gentisate pathway. Two putative hydroxyquinol 1,2-dioxygenase genes (ncg11113 and ncg12951) were cloned and functionally identified in Escherichia coli. The genes encoding enzymes for the conversion of phenol, benzoate, 3-hydroxybenzoate, 4-hydroxybenzoate and vanillate in the central β-ketoadipate pathways were mapped on the chromosome of C. glutamicum. A unique 30-kb (approximately 1% of the entire genome) catabolic island that channels the degradation of various aromatic compounds was mapped to position 2525-2555 kb of the genome. The global analysis and characterization of aromatic degradation pathways provided new insights into the metabolic ability of C. glutamicum in addition to its well-known ability to produce various amino acids and vitamins.
Endophytic bacteria in the seeds of rice plants (Oryza sativa, cultivar Nipponbare) were studied during the rice maturation process by comparing them with the bacteria at the surface of rice seeds. The endophytic and surface bacteria were isolated from cultivated rice seeds by using a nutrient broth (NB) and a diluted nutrient broth (DNB) agar medium. The endophytes at the early stages of maturation were relatively diverse, consisting of strains closely related to the genera Bacillus, Sphingomonas and Pantoea. In contrast, the endophytic and surface bacteria at the middle and late stages of the maturation process (39 strains altogether) were all closely related only to the genus Bacillus with the exception of one isolate, and almost all of these strains were motile and spore-forming. It was deduced that these bacteria might have migrated into the rice seeds before they hardened that is, during the early stage (the endophytes at the early stage were also all motile and about half of them were spore-forming), subsisting as endospores during the middle and late stages of the maturation process. Furthermore, most of the isolates (19 of 23) from the surface bacteria and the endophytic bacteria grew better when sucrose (1.2 M) was added to the NB liquid medium. These bacteria may be able to adapt to a high osmotic pressure.
The effects of four strains of Gram positive bacteria (PS2, PS9, PS11, and PS79) isolated from the sponge Pseudoceratina pupurea on the growth of a set of fouling bacteria and standard strains of genera Vibrio-Photobacterium were examined. Using a paper disc diffusion method, 4 sponge isolates were found to have inhibitory activity against 4 of 12 isolates of the fouling bacteria. All of the four bacterial supernatants also showed inhibitory activity against Vibrio alginolyticus and V. fischeri, while sponge extracts inhibited every strain tested. Phylogenetic analysis using 16S rRNA gene sequences revealed that three of the sponge isolates (PS2, PS9 and PS79) belonged to the genus Bacillus and one (PS11) belonged to the genus Virgibacillus. The isolation of strains possessing inhibitory activity against fouling bacteria suggests that sponge-associated bacteria could be potential sources of novel natural products.
The cellular slime mold Dictyostelium discoideum, a lower eukaryote feeding on bacteria in soil, is highly resistant to DNA damaging agents. D. discoideum expresses at least two RecA homologs, Rad51 and mitochondria localizing RecA. Rad51 consisting of 351 amino acid residues is 69% identical to human Rad51. UV light-irradiation and H2O2-treatment increased the expression level of Rad51, suggesting that Rad51 is required for DNA repair in D. discoideum. A gene disruption mutant was not viable, suggesting that Rad51 is essential for D. discoideum.