Characteristics of myxobacteria isolated from the surface of freshwater fish

A study was made of 32 nonpathogenic myxobacterial isolates obtained from a variety of fish taken in the Pacific Northwest. Morphological, cultural, biochemical, and serological studies were carried out on these strains. All were found to be members of the genus Cytophaga. Two myxobacterial strains pathogenic to fish were also included in this study for comparative purposes. These pathogenic organisms were found to be culturally and physiologically similar to some of the nonpathogenic strains. Antiserum against the pathogenic species, however, showed no cross agglutination when tested against the other myxobacterial isolates. As a result, serological procedures appear promising as a rapid means for distinguishing pathogenic myxobacteria from one another and from saprophytic myxobacteria commonly found on fish.     

Identification of a slime exopolysaccharide depolymerase in mucoid strains ofPseudomonas aeruginosa

Strains ofPseudomonas aeruginosa recovered from pulmonary infections in cystic fibrosis (CF) patients are often mucoid in appearance owing to the secretion of a viscous slime exopolysaccharide (EPS). Unlike most mucoid isolates, strains WcM#2, P10, and P11 produce mucoid colonies after 24 h of incubation at 37°C, which become nonmucoid upon further incubation; this suggests the presence of a slime-degrading enzyme or depolymerase. Using both qualitative and quantitative assays, the presence of a slime EPS depolymerase was confirmed in each of these three strains as well as in four of four additional mucoid strains. Depolymerase activity was lower but still detectable in four of four nonmucoid strains. Enzyme preparations from strains WcM#2, P10, and P11 were active on most, but not all, slime EPS preparations fromP. aeruginosa strains, as well as sodium alginate; greater activity was observed on substrates after deacetylation. Comparisons are made between the enzyme described in this study and previous reports of slime EPS depolymerase in mucoid strains ofP. aeruginosa.     

Polysaccharide-producing bacteria isolated from paper machine slime deposits

Development of novel enzymatic methods for slime deposit control in paper mills requires knowledge of polysaccharide-producing organisms and the polysaccharide structures present in deposits. In this work, 27 polysaccharide-producing bacteria were isolated from slime samples collected from different parts of a paper machine. Most of the isolates produced polysaccharides in liquid culture and nine of them were selected for production of polysaccharides for characterisation. The selected isolates belonged to seven different genera: Bacillus, Brevundimonas, Cytophaga, Enterobacter, Klebsiella, Paenibacillus and Starkeya. Using ribotyping, partial 16S rDNA sequencing, physiological tests and fatty acid analysis, four of the nine isolates: Bacillus cereus, Brevundimonas vesicularis, K. pneumoniae and P. stellifer were identified to the species level. Production of polysaccharides by the selected isolates varied between 0.07 and 1.20 g L^–1, the highest amount being produced by B. vesicularis. The polysaccharides were heteropolysaccharides with varying proportions of galactose, glucose mannose, rhamnose fucose and uronic acids.     

Biodegradation of dissolved jet fuel in chemostat by a mixed bacterial culture isolated from a heavily polluted site

A mixed bacterial culture capable of biodegrading of jet fuel was isolated from a heavily polluted site in Tapa, Estonia. Residual concentrations of pollutants in the chemostat culture were determined. The total residual concentrations of dissolved jet fuel in culture medium were 0.42 and 2.1 μg l^-1 at the dilution rates 0.1 and 0.17 h^-1respectively. Benzene, toluene, ethylbenzene, and xylenes were completely degraded and thus not detected in culture broth (detection limit 0.1 μg l^-1)at the dilution rates 0.1 and 0.17 h^-1. The values of apparent substrate saturation constant(K_Sapp) in multisubstrate growth conditions were estimated from the experimental data. The residual concentrations satisfy the regulations in the Republic of Estonia for petroleum hydrocarbons (0.00 mg l^-1 – ‘very good’). Results obtained indicate that use of the biodegradation could be sufficient for the treatment of polluted with kerosene-type jet fuel groundwater up to the acceptable quality. This revised version was published online in August 2006 with corrections to the Cover Date.     

Characterization of mucoid Pseudomonas aeruginosa strains isolated from technical water systems

The occurrence of mucoid Pseudomonas aeruginosa strains was investigated in water samples and surface material from non-clinical aquatic environments. Ten of 81 environmental isolates displayed a mucoid colony type after incubation at 36°C for 24 h on Pseudomonas Isolation Agar. The mucoid strains obtained exclusively from surfaces of technical water systems were characterized in terms of medium-dependent expression of mucoid colonial phenotype, exoenzyme profile, pigment production and O-antigen type. The mucoid strains secreted substantially higher quantities of carbohydrate and uronic acid-containing material compared to non-mucoid environmental isolates. Major slime components of the mucoid strains were identified as O-acetylated alginates that contained higher proportions of mannuronate than guluronate monomer residues and were composed of blocks of poly-mannuronate and poly-mannuronate/guluronate, whereas blocks of poly-guluronate were absent. The results suggest that surfaces in aquatic environments may represent a natural habitat for mucoid (i.e. alginate-overproducing) strains of Ps. aeruginosa with properties similar to clinical mucoid strains.     

Some properties of vacuoles isolated from Neurospora crassa slime variant

A method for the isolation of vacuoles based on polybase induced lysis of protoplasts of the cell wall deficient Neurospora crassa slime variant is described. Isolated vacuoles are characterized by 12 to 50 times increased specific activities of several hydrolases as compared with the total homogenate of protoplasts. Total -amino nitrogen, arginine, and polyphosphate are also greatly enriched in these vacuoles. Vacuoles are equipped with a permease for the transport of basic amino acids across the tonoplast.Non-Standard Abbreviation DEAE-dextran diethylaminoethyl-dextran     

Bacteriophages of methanotrophs isolated from fish

Bacteriophages of methanotrophic bacteria were isolated from 67 fish. Only two phages isolated from two fish species specifically lysed Methylocystis sp. and Flavobacterium gasotypicum. The phages lysing these species were designated 63-F and CMF-1-F, respectively. The isolated phages differed greatly in the fine structure of the virion, plaque morphology, spectrum of lytic action, serological properties, and UV sensitivity. At the same time, they had identical one-step growth characteristics: their latent period equalled 5 h, lysis time was 3 to 4 h, and burst size was about 240 virions. The phages had guanine- and cytosine-rich double-stranded DNAs consisting of common nitrogen bases. The molecular masses of the DNAs as determined by the sums of restriction endonuclease cleavage fragments were 28 X 10(6) daltons for phage 63-F and 31 X 10(6) daltons for phage CMF-1-F.     

Two new species ofPichia isolated from slime fluxes of deciduous trees

Two new homothallic species ofPichia, associated with exudates ofPopulus trichocarpa andSalix sp. have been described.Pichia trehalophila sp.n. was isolated from slime exudates in two widely separatedPopulus trichocarpa trees andPichia salictaria sp.n. was isolated from two exudates ofSalix sp. The latter species was previously present as an unidentified strain ofPichia in the culture collection of the Centraalbureau voor Schimmelcultures and had been isolated from a patient in Germany. The natural habitat ofP. salictaria is considered to be in association with willow trees.     

Characteristics of a new reovirus isolated from epizootic ulcerative syndrome infected snakehead fish.

Epizootic ulcerative syndrome (EUS) has been infecting a wide range of fishes in the South and Southeast Asia for the last 2 decades. One reovirus-like agent (snakehead reovirus, SKRV), isolated from an EUS-infected snakehead fish and investigated in the present study, is the only reovirus so far isolated from an EUS-infected fish. SKRV was characterised by the presence of a double-stranded RNA genome with icosahedral symmetry and double capsid. The virus had an average size of 71 nm, a buoyant density of 1.36 g ml(-1) in CsCl and lacked a lipid-containing envelope. Apart from the above, the presence of a segmented genome and structural proteins falling into 3 specific size classes confirmed that the virus belongs to the family Reoviridae. SKRV differed from aquareoviruses by the lack of a cytopathic effect (CPE) with syncitium formation and in the segmentation pattern of RNA genome. The resistance to pH (3.0 to 9.0) and heat treatment and inability to multiply in mammalian cell lines and haemagglutinate human 'O' red blood cells (RBCs) differentiated SKRV from the rest of the similar genera in the family Reoviridae. Serological comparison indicated the antigenic distinctness of the isolate from selected American and European aquareoviruses. SKRV grew well in SSN-1 and SSN-3 cells at 25 to 30 degrees C but not in the most common Aquareovirus susceptible coldwater fish cell line--CHSE-214.     

Psychrophilic yeasts isolated from marine fish

Yeasts were isolated from marine fish using low incubation temperatures. The isolates were screened for their ability to grow at 4 C and 30 C. Isolates growing at 4 C but not at 30 C were considered to be psychrophilic and were selected for further study. Approximately 25% of the isolates were in this category. The isolates comprised the following genera:Candida, Cryptococcus, Debaryomyces, Rhodotorula, Torulopsis andTrichosporon. Rhodotorula infirmo-miniata accounted for 50% of the isolates andTrichosporon pullulans 10%. Other species were present in numbers below the 10% level. The distribution of the species between the two sample areas is discussed. The vitamin requirements and optimum growth temperatures of the isolates were assessed.     

Bacteriophages of methanotrophs isolated from fish.

Bacteriophages of methanotrophic bacteria were isolated from 67 fish. Only two phages isolated from two fish species specifically lysed Methylocystis sp. and Flavobacterium gasotypicum. The phages lysing these species were designated 63-F and CMF-1-F, respectively. The isolated phages differed greatly in the fine structure of the virion, plaque morphology, spectrum of lytic action, serological properties, and UV sensitivity. At the same time, they had identical one-step growth characteristics: their latent period equalled 5 h, lysis time was 3 to 4 h, and burst size was about 240 virions. The phages had guanine- and cytosine-rich double-stranded DNAs consisting of common nitrogen bases. The molecular masses of the DNAs as determined by the sums of restriction endonuclease cleavage fragments were 28 X 10(6) daltons for phage 63-F and 31 X 10(6) daltons for phage CMF-1-F.     

Siderophore synthesis by mucoid Pseudomonas aeruginosa strains isolated from cystic fibrosis patients.

Nonmucoid Pseudomonas aeruginosa responds to iron deprivation by synthesizing the siderophores pyochelin and pyoverdine. When grown in iron-deficient medium, six mucoid P. aeruginosa strains isolated from cystic fibrosis patients synthesized copious amounts of the exopolysaccharide alginate. A procedure that eliminated the interference of alginate was developed so that siderophores could be extracted from the growth medium. All six isolates were then noted to produce both pyoverdine and pyochelin. This report thus confirms that mucoid P. aeruginosa, like its nonmucoid counterparts, elicits the siderophores commonly cited as those of the microbe.     

Chararacteristics of Vagococcus salmoninarum isolated from diseased salmonid fish

Isolates of the salmonid pathogen Vagococcus salmoninarum were recovered from Atlantic salmon, rainbow trout and brown trout with peritonitis. The phenotypes of these isolates and the type strain of Vag. salmoninarum NCFB 2777 were determined by morphological, biochemical and physiological tests and whole cell protein profiles by SDS-PAGE. There was a high level of phenetic similarity between the salmonid isolates and the type strain. The species forms short Gram-positive rods, hydrolyses L-pyrrolidonyl-β-naphthylamide, is α-haemolytic on sheep's blood agar, grows at pH 9·6 and 10°C but not at 40°C or in 6·5% NaCl and is catalase-negative; a Lancefield group N antigen is not present. Vagococcus salmoninarum can be distinguished phenetically from similar fish pathogens including Carnobacterium piscicola, Enterococcus seriolicida and Lactococcus piscium.     

The biological characteristics of micromycetes isolated from fish roe

The fungi Aspergillus fumigatus Fres., and Fusarium sporotrichiella Bilai were studied for their effect on certain biochemical indices of two-year carp and spawn of grass carp under laboratory conditions. The intraperitoneal introduction of A. fumigatus in a dose of 5 and 20 min conidia per individual to two-year carp decreased significantly the protein level in blood serum, the decrease being more considerable with the introduction of the smaller dose of fungal conidia. Joint incubation of the grass carp spawn and F. sporotrichiella induced changes in the protein level, amylolytic and phosphate (acid phosphatase) activity in spawn. In that case the time of the fungus action on grass carp spawn was a decisive factor. Activity of certain hydrolytic enzymes in mycelium and conidia of A. fumigatus and F. sporotrichiella was determined. The amylolytic activity was not revealed in the checked samples. The proteolytic activity was established in all samples of fungi and culture liquid, the highest level being observed in mycelia and conidia of A. fumigatus. The alkaline and acid phosphatase activity was found in F. sporotrichiella: the acid phosphatase activity was higher in mycelium, the acid phosphatase one in the fungus conidia. The problem on the A. fumigatus ability to produce extracellular enzymes is under discussion.     

Microcalorimetric study of mitochondria isolated from fish liver tissue

We determined the thermogenesis curves of mitochondria isolated from fish liver tissue by using an LKB 2277 Bioactivity Monitor. After isolation from the fish liver, mitochondria still have activity and can live for a long time by using the stored nutrients. We calculated the recovery rate constants of mitochondria. We found that the thermogenesis curves of mitochondria are similar to those obtained from prokaryotic cells, but not similar to those obtained from eukaryotic cells. We determined the metabolic thermogenesis curves of mitochondria isolated from two kinds of carp liver tissue, scattered-scaled mirror carp and harvest carp. There are some important similarities and some important differences between these thermogenesis curves.     

Identification and characterization of Carnobacteria isolated from fish intestine

Eleven bacterial strains were isolated from the gastrointestinal tract of four fish species, Atlantic salmon (Salmo salar L.), Arctic charr (Salvelinus alpinus L.), Atlantic cod (Gadus morhua L.) and wolffish (Anarhichas lupus L.). All the strains were Gram-positive rods, non-sporing, catalase and oxidase-negative, able to grow at pH 9.0 but not on acetate containing media (pH < or = 5.4), and were fermentative. They had a high content of oleic acid (18:1 n-9) in cellular lipid, and were found to belong to the genus Carnobacterium by phenotypic criteria. The eleven carnobacteria strains were further identified on the basis of 16S rDNA sequence analysis and AFLP(TM) fingerprinting.     

鱼源溶藻弧菌胞外产物的特性研究

溶藻弧菌经胰胨大豆胨琼脂培养基(TSA 2%NaCl)培养48h后,培养物用PBS洗脱、离心,再经孔径0.22μm微孔滤膜过滤,制得胞外产物(Extracellular product,ECP)。经检测,该ECP具有淀粉酶、酪蛋白酶、卵磷脂酶和脂肪酶等4种酶活性,其中淀粉酶的活性最高,酪蛋白酶、脂肪酶活性次之,卵磷脂酶活性较低,未检测出明胶酶及脲酶活性;能溶解断斑石鲈(Pomadasys hasta)、眼斑拟石首鱼(Sciaenops ocellatus)和赤点石斑鱼(Epinephelus akaara)的红细胞,而不能溶解鸡、羊及O型人红细胞;对小白鼠和石斑鱼的LD50分别为0.08mg/g和0.17mg/g,95%可信限分别为0.06—0.12mg/g和0.07—0.28mg/g;50mmol/L EDTA和100mmol/L苯甲基磺酰氟能使胞外蛋白酶(Extracellular pro-teinase,ECPase)的活性分别下降83.40%和51.32%;ECPase对热稳定性较差,碱性条件下活性较高,其最适温度为50℃,最适pH为8.0。     

First bacterial chalcone isomerase isolated from Eubacterium ramulus

The human fecal anaerobe Eubacterium ramulus is capable of degrading various flavonoids, including the flavone naringenin. The first step in the proposed degradation pathway is the isomerization of naringenin to the corresponding chalcone. Cell-free extracts of E. ramulus displayed chalcone isomerase activity. The enzyme from E. ramulus was purified to homogeneity. Its apparent molecular mass was estimated to be 136 and 129 kDa according to gel filtration and native polyacrylamide gel electrophoresis, respectively. Chalcone isomerase is composed of one type of subunit of 30 kDa. The purified enzyme catalyzed the isomerization of naringenin chalcone, isoliquiritigenin, and butein, three chalcones that differ in their hydroxylation pattern. N-bromosuccinimide, but also naringenin and phloretin, inhibited the purified enzyme considerably. This is the first report on a bacterial chalcone isomerase. The physiological function of the purified enzyme is unclear, but an involvement in the conversion of the flavanone naringenin to the chalcone is proposed.