Flavobacterium Strains Research Articles

Metabolic activity of Flavobacterium strain P25 during starvation and after introduction into bulk soil and the rhizosphere of wheat

The physiological state of introduced Flavobacterium strain P25 cells was determined in starvation cultures, in bulk soil, and in the rhizosphere of wheat using direct viable counts (DVC; based on cell elongation after use of nalidixic acid and substrate addition, resulting in a potential activity measurement) and the redox dye 5-cyano-2, 3-ditolyl tetrazolium chloride (CTC; based on respiration without substrate additions, resulting in an in situ activity measurement). Both methods clearly demonstrated that the metabolic activity of Flavobacterium P25 cells decreased during starvation, followed by increased activity after amendment with substrate. This confirmed the applicability of DVC and CTC methods to Flavobacterium P25. Both DVC and CTC methods showed that the percentage of active cells in an introduced Flavobacterium P25 population in rhizosphere soil was lower than that in bulk soil in the first 1–2 weeks after planting wheat seedlings. After two weeks, the percentage of metabolically active cells in the P25 population in rhizosphere soil was higher than in bulk soil. Since different aspects of cellular physiology are measured when applying DVC and CTC, the impact of variations in environmental factors on the metabolic state of introduced strains may be monitored closely by these methods.

Production of 10-ketostearic acid from oleic acid by a strain of Flavobacterium sp. 12-4A

A new Flavobacterium sp., strain 12-4A, produces solely 10-ketostearic acid (KSA) from oleic acid (OA) in growing cultures. Under optimized conditions 14 mg/ml of KSA was produced from 25 mg/ml of OA after 3 days of cultivation at 30°C.

The nylon oligomer biodegradation system of Flavobacterium and Pseudomonas.

This review article is a compendium of the available information on the degradation of a man-made compound, 6-aminohexanoate-oligomer, in Flavobacterium and Pseudomonas strains, and discusses the molecular basis for adaptation of microorganisms toward these xenobiotic compounds. Three plasmid-encoded enzymes, 6-aminohexanoate-cyclic-dimer hydrolase (EI), 6-aminohexanoate-dimer hydrolase (EII), and endo-type 6-aminohexanoate-oligomer hydrolase (EIII) are responsible for the degradation of the oligomers. Two repeated sequences, designated RS-I and RS-II, are found on plasmid pOAD2, which is involved in 6-aminohexanoate degradation in Flavobacterium. RS-I appears 5 times on the pOAD2, and all copies have the same sequences as insertion sequence IS6100. RS-II appears twice on the plasmid. RS-IIA contains the gene encoding EII, while RS-IIB contains the gene for the analogous EII' protein. Both EII and EII' are polypeptides of 392 amino acids, which differ by 46 amino acid residues. The specific activity of the EII enzyme is 200-fold higher than that of EII'. Construction of various hybrid genes demonstrated that only the combination of two amino acid residues in the EII' enzyme can enhance the activity of the EII' to the same level as that of EII enzyme.

Nylon oligomer degradation gene, nylC, on plasmid pOAD2 from a Flavobacterium strain encodes endo-type 6-aminohexanoate oligomer hydrolase: purification and characterization of the nylC gene product.

A new type of nylon oligomer degradation enzyme (EIII) was purified from an Escherichia coli clone harboring the EIII gene (nylC). This enzyme hydrolyzed the linear trimer, tetramer, and pentamer of 6-aminohexanoate by an endo-type reaction, and this specificity is different from that of the EI (nylA gene product) and EII (nylB gene product). Amino acid sequencing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified EIII demonstrated that the enzyme is made of two polypeptide chains arising from an internal cleavage between amino acid residues 266 and 267.

Selecting inocula for the biodegradation of organic compounds at low concentrations

The inability of many organisms to degrade pollutants at low concentrations is a problem when selecting inocula for bioremediation of sites with these low concentrations. Thus, a study was conducted to determine the effect of low concentrations of p-nitrophenol (PNP) on growth of four PNP-degrading bacteria and their abilities to metabolize low concentrations of the compound in culture and samples from an oligotrophic lake. PNP did not increase the growth rates of Flavobacterium sp. M4, Pseudomonas sp. K, Flavobacterium sp. M1, and Pseudomonas sp. SP3 at concentrations of less than 2, 4, 10, and 100 ng/ml, respectively, when it was the sole added carbon source in culture, but it stimulated multiplication at higher concentrations. In liquid culture with the nitro compound as sole added carbon source, the four bacteria extensively mineralized PNP at 50 and 100 ng/ml, and three of the four degraded much of the substrate at 25 ng/ml. Pseudomonas sp. SP3 mineralized more than 20% but the two Flavobacterium strains converted less than 10% of the substrate to C02 at 10 ng/ml, and none of the three mineralized more than 5% at 1 and 5 ng PNP/ml. Under conditions where more than 99% of the radioactivity from (14)C-PNP added at 1 ng/ml remained in solution, two of the isolates formed organic products. Pseudomonas sp. K had no activity at 1, 5, and 10 ng/ml. In contrast, when each of the bacteria was separately inoculated into samples of water from an oligotrophic lake and from a well in which PNP was not biodegraded, the bacteria were able to mineralize as little as 1 ng PNP/ml. The addition to a salts solution of 10 ng of glucose per ml resulted in mineralization of PNP at concentrations too low to be mineralized when the nitro compound was the sole source of added carbon. Bacteria may thus be able to mineralize substrates in natural waters at concentrations below those suggested by tests conducted in culture media, possibly because of the availability of other carbon sources for the bacteria.

Isolation of a benzene-tolerant bacterium and its hydrocarbon degradation

After treatment of deep-sea sediment samples (collected from a depth of 1,945-m in Suruga Bay) with 20 v/v% benzene followed by 50 v/v% kerosene, fifty-six halotolerant bacteria were isolated as candidate hydrocarbon degraders that also possessed the characteristic of benzene-tolerance. One of the isolates showed halotolerant growth and tolerance to various kinds of organic solvents (OSs) such as 5 v/v% benzene, 10 v/v% toluene and 10 v/v% p-xylene. From these characteristics, in addition to its morphological and biochemical characteristics, it seemed to be a variant strain of Flavobacterium sp. and was tentatively named strain no. DS-711. The correlation between log p, used as a parameter for the degree of toxicity of the OSs, and growth on the OSs was further examined using the benzene-tolerant strain DS-711 and the toluene-tolerant Pseudomonas putida strain IH-2000. The results indicated that the mechanism of OS tolerance of DS-711 was different from that of P. putida strain IH-2000. Furthermore, a hydrocarbon degradation experiment indicated that the benzene-tolerant strain DS-711 had a greater n-alkane degrading ability compared to that of our selected control strains, the OS-sensitive Alteromonas sp. strain no. DS-201 and the toluene-tolerant P. putida strain IH-2000. With stepwise addition of concentrated cell suspension, strain DS-711 could degrade more than 90% of n-alkanes in kerosene.

Removal of Musty Odor Compound in Drinking Water by Biological Filter

In order to remove the musty odor compound 2-methyl isoborneol (MIB) from drinking water resources at Lake Biwa, a pilot plant of a biological filtration was installed and operated. In this study, the effects of water temperature, pH and initial concentration of MIB on the MIB degradation potential of the ceramic media packed column in the biological filter were studied. Microorganisms on the surface of the granular media were observed under the scanning electron microscope (SEM). Bacteria which were responsible for the degradation were isolated, studied and identified. From this experiment, the extent of the effect of water temperature on the degradation potential was found to be similar to that on the biochemical oxygen demand (BOD) removal activity by an activated sludge. The optimum pH for the degradation potential was seen to be between 7 and 9. The MIB degradation reaction was seen to be almost a first order reaction. On the surface of the washed granular media, coccoid and rod-shaped bacteria were observed. Eleven out of 34 bacteria isolated from the washed media were judged as biodegraders. Some characteristics of 7 Gram negative biodegraders were studied and two strains of Pseudomonasaeruginosa,one strain of Flavobacteriummultivorum, one strain of Pseudomonas sp. and one strain of Flavobacterium sp. were identified.

Proposals of Sphingobacterium faecium sp. nov., Sphingobacterium piscium sp. nov., Sphingobacterium heparinum comb. nov., sphingobacterium thalpophilum comb. nov. and two genospecies of the genus Sphingobacterium, and synonymy of Flavobacterium yabuuchiae and Sphingobacterium spiritivorum.

Seventeen strains of Cytophaga, Flavobacterium and Actinobacillus species were taxonomically characterized. DNA base composition of these strains ranged from 39.0 to 42.3mol%, and the menaquinone system was MK-7. All the strains contained sphingolipids with long-chain bases, which are characteristic of the genus Sphingobacterium. Based on their chemotaxonomic and physiological characteristics together with DNA/DNA hybridization studies, we propose two new species and two new combinations, Sphingobacterium faecium sp. nov., Sphingobacterium piscium sp. nov., Sphingobacterium heparinum comb. nov., Sphingobacterium thalpophilum comb. nov. and two genospecies of the genus Sphingobacterium. Flavobacterium yabuuchiae was found to be a synonym of Sphingobacterium spiritivorum.

Biodegradation of pentachlorophenol in soil: the response to physical, chemical, and biological treatments

The effects of physical, chemical, and biological treatments on biodegradation of pentachlorophenol (PCP) were studied in a silt-loam soil contaminated with 175 mg PCP/kg and uniformly 14C-labelled PCP. Biodegradation of 14C-labelled PCP and technical-grade PCP were monitored over 210 days incubation. Mineralization of labelled PCP was significantly (p=0.05) influenced by soil treatments. Negligible biodegradation occurred in either the sterile control soil or the uninoculated control soil, with less than 1% of added 14C recovered as 14 CO2. Inoculation of unamended soil with a strain of Flavobacterium (ATCC 39723) known to degrade PCP increased biodegradation of PCP; approximately 60% of the [14C]PCP was recovered as 14CO2. Increased soil water content (60% versus 30% w/w) enhanced biodegradation (67% recovery of 14C as CO2), while increased chloride ion concentration and anoxic conditions were inhibitory (20 and 1% recoveries, respectively). Residual soil PCP concentrations were also influenced by various treatments. In the sterile control soil and noninoculated control, after 210 days incubation, concentrations of PCP were 143 and 1223 mg/kg, respectively, while the PCP concentration in the inoculated soil was 21 mg/kg. When soil organic matter was increased by adding finely ground red clover leaf and stem material, the residual PCP concentration was reduced to 6 mg/kg after 210 days. Increased soil water content resulted in a residual PCP concentration of 5 mg/kg. High-pressure liquid chromatography of soil extracts revealed no accumulation of partial PCP degradation products. The results indicated that biodegradation of PCP in soil was significantly influenced by various soil amendments.

Cloning, sequencing, and expression of the N-acyl-D-mannosamine dehydrogenase gene from Flavobacterium sp. strain 141-8 in Escherichia coli

The gene coding for N-acyl-D-mannosamine dehydrogenase (NAM-DH) from Flavobacterium sp. strain 141-8 was cloned and expressed under the control of a lac promoter in Escherichia coli JM109. The DNA sequence of the gene was determined, and an open reading frame encoding a polypeptide composed of 272 amino acid residues (Mr, 27,473) was identified. The E. coli transformants which showed over 200-fold higher NAM-DH activity than did the Flavobacterium strain produced the enzyme as a protein fused with beta-galactosidase. Despite being a fusion, NAM-DH produced by E. coli transformants appeared unchanged in pH optimum, Km, and substrate specificity from Flavobacterium sp. strain 141-8. This newly recombinant enzyme may be applicable to the quantitative determination of sialic acid in serum.

Enzymatic alteration in the shikimate pathway during derivation of menaquinone-4-producing mutants of Flavobacterium sp. 238-7.

Activities of seven enzymes of the shikimate pathway were compared between the wild-type strain and menaquinone-4-producing mutant strains of Flavobacterium sp. 238-7. Activity of 3-deoxy-d-airbino-heptulosonate-7-phosphate (DAHP) synthase, the first regulatory enzyme in the shikimate pathway, was almost the same in these strains in the stationary phase, but the activity of a sulfonamide-resistant strain, SP0736, was 2 times higher than that of other strains in the logarithmic phase. Shikimate dehydrogenase activity was almost the same among these strains during cultivation. Other enzyme activities of mutants were 1.5-4 fold higher than those of the wild-type strain. DAHP synthase and shikimate kinase were found to be inhibited by chorismate and MK-4. Feedback inhibition for shikimate kinase by chorismate was partially removed in strain SP0736.

Specificity of aFlavobacterium in the metabolism of substituted chlorobenzoates

A strain ofFlavobacterium breve capable of utilizing 3,5-dichlorosalicylate as a sole source of carbon and energy was identified. Degradation of 3,5-dichlorosolicylate, was specific as this strain did not metabolize dicamba (3,6-dichloro-2-methoxybenzoic acid), 3,5-dicamba (3,5-dichloros2-methoxybenzoic acid), or 3,6-dichlorosalicylate. The organism was able to remove completely 3,5-dichlorosalicylate in the presence of three times as much 3,6-dichlorosalicylate being degraded. The organism was able to utilize 3,5-dichlorosalicylate at concentrations up to 1000μg/ml. A mixture of 3,5 and 3,6-dichlorosalicylate isomers purified by biological destruction of the unwanted isomer (3,5-dichlorosalicylate) would be useful for producing isomerically pure dicamba, an important herbicide.

Occurrence of homospermidine as a major polyamine in the authentic genus Flavobacterium

Polyamines in various Flavobacterium strains and related eubacteria were analyzed. The major polyamine was homospermidine in all 12 strains of the eight authentic species of the genus Flavobacterium studied (F. aquatile, F. breve, F. balustinum, F. gleum, F. meningosepticum, F. odoratum, F. multivorum, and F. spiritivorum), and in F. ferrugineum, whose taxonomic position is uncertain but related to Flavobacterium. Spermidine was not detected in these flavobacteria. In another seven Flavobacterium species that are taxonomically different from Flavobacterium and belong to other genera, homospermidine was not detected. Key words: Flavobacterium, homospermidine, spermidine, arginine decarboxylase.

A Numerical Taxonomic Study of Pelagic and Benthic Surface-layer Bacteria in Seasonally-cold Coastal Waters

Summary Heterotrophic bacteria from the pelagic zone and the benthic surface-layer of the continental shelf, near Newfoundland, were isolated. Using numerical taxonomy, clusters of related strains were identified and the characteristics of the clusters determined. The majority of the pelagic strains were facultatively anaerobic and were identified as Vibrio, although a few strains were not identified and others were assigned to the family Vibrionaceae. The few strictly aerobic strains from the pelagic zone were Alteromonas. The benthic bacterial population was more diverse and bacteria with strictly respiratory metabolism predominated. Among these were strains of Pseudomonas, Flavobacterium, and Alteromonas. The remaining strains were Vibrio, and some Vibrionaceae. Although some pelagic and benthic strains were fastidious, most utilized a broad range of substrates. This was most evident among, although not restricted to, the marine pseudomonads and the flavobacteria of the benthos. In seasonally-cold oceans the benthic surface-layer is an important zone for detrital breakdown and nutrient recycling. The nutritional versatility of Pseudomonas and Flavobacterium may, in part, account for their abundance in the substrate enriched benthic surface-layer.

Superoxide dismutase in strains of the genus Flavobacterium: isolation and characterization.

Two electrophoretically different forms of superoxide dismutase, one of them containing manganese-protein and the other iron-protein, were detected in eleven different strains of the genus Flavobacterium. The activities of the different strains were similar to those described for other bacteria. The two molecular forms of the enzyme differed clearly with regard to activity, electrophoretic behaviour, sensitivity to cyanide and peroxide, and NaCl requirement. Both molecular forms were isolated from Flavobacterium halmephilum. Molecular mass absorption spectra, metal content, optimum pH, heat-sensitivity and stability were described.

Pneumonia and meningitis caused by a new nonfermentative unknown gram-negative bacterium

Seven isolates of an unclassified bacterium resembling Flavobacterium spp. were characterized by growth requirements, microscopic examination, biochemical characteristics, antimicrobial susceptibility tests, protein profile analysis, and serologic data. The unclassified isolates were differentiated from Flavobacterium meningosepticum, Flavobacterium odoratum, Flavobacterium balustinum, Flavobacterium strain IIb, Chromobacterium violaceum, Aquaspirillum serpens, and Pseudomonas spp. The bacterium was a gram-negative rod with a polar flagellum. Protein profile analysis demonstrated two major protein bands present in the unclassified isolates that were absent from the Flavobacterium and Pseudomonas controls but present in the Aquaspirillum and Chromobacterium controls. However, no serologic cross-reactions were observed. Our results showed that the unclassified bacterium was distinct from any previously known genus of bacterium.

Flavobacterium meningosepticum infection in a neonatal ward

An outbreak of infections due to Flavobacterium meningosepticum type C in a neonatal intensive care unit is described. During a period of two weeks, two infants developed meningitis and a third was colonized in the respiratory tract and had transient bacteremia. The two meningitis patients were treated with clindamycin, rifampicin and cefotaxime systemically, plus rifampicin intraventricularly. Bacteriological eradication was achieved within 48 h, and both infants recovered from the meningitis without apparent neurological sequelae; however, one infant died two months later of unrelated causes. Environmental surveillance cultures failed to demonstrate a reservoir for the epidemic strain, but other Flavobacterium strains were recovered. Two clinically healthy infants were found to be colonized in the nasopharynx with strains that were extremely difficult to differentiate phenotypically from the epidemic strain. Extensive characterization of strains is necessary in order to differentiate between strains and subsequently to determine a certain source of infection.

Identification of Flavobacterium strains by gas liquid chromatographic analysis of volatile fatty acids produced in culture

The classification previously established for 74 Flavobacterium strains by gas liquid chromatographic (GLC) analysis of volatile fatty acids (VFA) produced in culture allowed the recovery of 9 groups ( J. gen. Microbiol., 1986, 132, 2723–2732). Since graphic representation of the strains based on the first 3 factors obtained by principal component analysis (PCA) clearly separated these groups, we tried to identify 80 new strains by comparing their positions with those of the 9 groups, on the basis of both hierarchical classification and PCA methods. Of the 153 strains studied, only 12 were not allocated to a group corresponding to their original biochemical identification Thus, on the whole, this characterization method by GLC analysis seemed satisfactory, although it could not be established whether the method was adequate for routine identification, or would serve merely as a complement.

Characterization of Flavobacterium species by analysis of volatile fatty acid production.

Seventy-four Flavobacterium strains were characterized by gas-liquid chromatographic analysis of volatile fatty acids produced in the culture medium. Principal components analysis permitted the graphic representation of the relative positions of the different strains, and aggregation according to the variance enabled a hierarchical classification to be established. The study revealed three subgroups each for F. meningosepticum and F. odoratum. Our F. breve, Flavobacterium sp. group IIb and F. multivorum strains appeared to be homogeneous. These results tallied with those of previous studies on DNA base composition and reassociation, electrophoretic protein profiles and cellular fatty acid composition.

Pentachlorophenol degradation: a pure bacterial culture and an epilithic microbial consortium.

The steady-state growth of a Flavobacterium strain known to utilize pentachlorophenol (PCP) was examined when cellobiose and PCP simultaneously limited its growth rate in continuous culture. A concentration of 600 mg of PCP per liter in influent medium could be continuously degraded without affecting steady-state growth. We measured specific rates of PCP carbon degradation as high as 0.15 +/- 0.01 g (dry weight) of C per h at a growth rate of 0.045 h-1. Comparable specific rates of PCP degradation were obtained and maintained by PCP-adapted, natural consortia of epilithic microorganisms. The consortium results suggest that a fixed-film bioreactor containing a PCP-adapted natural microbial population could be used to treat PCP-contaminated water.