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PubMed1997

Site-specific endonucleases RspLKI and RspLKII from Rhodococcus species LK2 are isoschizomers of SphI and BamHI.

Two site-specific endonucleases, RspLKI and RspLKII, have been isolated and purified to functional homogeneity from the soil bacterium Rhodococcus species LK2. RspLKI recognizes the 5'-GCATG decreases C-3' DNA sequence and RspLKII recognizes the 5'-G decreases GATCC-3' sequence (arrows indicate DNA cleavage sites). The isolated enzymes are class II site specific endonucleases and are isoschizomers of endonucleases SphI and BamHI, respectively.

Chromatography, Ion Exchange↗

PubMed1997

Nucleic acid amplification for rapid detection of Rhodococcus equi in equine blood and tracheal wash fluids.

OBJECTIVE: To evaluate the ability of nucleic acid amplification techniques to detect Rhodococcus equi in equine buffy coat, blood, and tracheal wash fluid and to differentiate between virulent and avirulent strains of the bacteria. SAMPLE POPULATION: Blood anticoagulated with EDTA and tracheal wash fluid from healthy horses. PROCEDURE: Logarithmic dilutions of virulent and avirulent strains of R equi were added to equine buffy coat and tracheal wash fluid samples. The DNA was extracted and amplified by polymerase chain reaction (PCR), using primers specific for the 16S ribosomal subunit gene and the virulence plasmid of R equi. RESULTS: PCR with 16S ribosomal subunit primers amplified a 441-bp segment of DNA from virulent and avirulent strains of R equi, but not from samples containing other species of bacteria. The virulence plasmid primers amplified an 875-bp segment of DNA from virulent strains of R equi, but not from avirulent R equi, or from other species of bacteria. Virulent strains of R equi could be identified by PCR and differentiated from avirulent strains within 12 to 24 hours after sample collection, with as few as 10 to 100 organisms present. CONCLUSIONS: PCR can be used to rapidly and accurately identify R equi in equine blood and tracheal wash fluid samples and can differentiate between virulent and avirulent strains of the organism. CLINICAL RELEVANCE: Because PCR can confirm a diagnosis of R equi infection in horses more rapidly and specifically than use of standard culture techniques, extrapolation of this assay to soil and fecal samples could be useful in epidemiologic studies and studies of environmental disinfection or decontamination.

Actinomycetales Infections↗

PubMedDate not supplied

[Isolation of nitrile hydratase from Rhodococcus rhodochrous M8 cells and determination of the N-terminal amino acid sequence of its subunits].

Nitrile hydratase was isolated and purified to homogeneity from cells of Rhodococcus rhodochrous M8. This enzyme catalyzes the hydrolysis of acrylic acid nitrile to acrylamide. Nitrile hydratase content in the cell was shown to be 17% of total soluble protein. The molecular weight of the native enzyme was 510 kDa. The enzyme consisted of two subunits with molecular weights of 23.5 kDa and 28.0 kDa. The N-terminal amino acid sequences of these subunits were estimated.

Amino Acid Sequence↗

PubMed1997

Nutritional factors that affect the production of cholesterol oxidase by Rhodococcus equi no. 23.

Some nutritional factors affecting the production of cholesterol oxidase (COX) by Rhodococcus equi no. 23 were investigated. Cholesterol and yeast extract respectively were the best carbon source and nitrogen source for the COX production. The optimum concentration of cholesterol and yeast extract was found to be about 0.1% and 0.4-0.5% (w/v) respectively. In addition, NH4Cl, NaCl and Tween 80 also exhibited enhancing effects on COX production, being maximal at 0.1% (w/v), 0.2% (w/v) and 1.0% (v/v) respectively. Moreover, optimal enzyme production occurred in medium that had an initial pH of 7.0.

Biotechnology↗

PubMed1998

New shuttle vectors for Rhodococcus sp. R312 (formerly Brevibacterium sp. R312), a nitrile hydratase producing strain.

Two shuttle vectors named pRC52 (10.7 Kb) and pRK52 (12.2 Kb) carrying chloramphenicol (Cm) and chloramphenicol plus kanamycin (Km) resistance genes, respectively, were constructed by fusion of a cryptic plasmid pBL13869 (replicon pBL1, 5.8 Kb) from Brevibacterium lactofermentum ATCC13869 with pBR328 E. coli plasmid. Transformation of Rhodococcus sp. R312 (formerly Brevibacterium sp. R312) protoplasts was realised with an efficiency of 28 transformants per micrograms of DNA.

Brevibacterium↗

PubMed1998

Associations between physical examination, laboratory, and radiographic findings and outcome and subsequent racing performance of foals with Rhodococcus equi infection: 115 cases (1984-1992).

OBJECTIVE: To determine whether physical examination, laboratory, or radiographic abnormalities in foals with Rhodococcus equi infection were associated with survival, ability to race at least once after recovery, or, for foals that survived and went on to race, subsequent racing performance. DESIGN: Retrospective study. ANIMALS: 49 Thoroughbreds and 66 Standardbreds admitted to 1 of 6 veterinary teaching hospitals between 1984 and 1992 in which R equi infection was positively diagnosed. PROCEDURE: Results of physical examination, laboratory testing, and thoracic radiography were reviewed. Indices of racing performance were obtained for foals that recovered and eventually raced and compared with values for the US racing population. RESULTS: 83 (72%) foals survived. Foals that did not survive were more likely to have extreme tachycardia (heart rate > 100 beats/min), be in respiratory distress, and have severe radiographic abnormalities on thoracic radiographs at the time of initial examination than were foals that survived. Clinicopathologic abnormalities were not associated with whether foals did or did not survive. Forty-five of the 83 surviving foals (54%) eventually raced at least once, but none of the factors examined was associated with whether foals went on to race. Racing performance of foals that raced as adults was not significantly different from that of the US racing population. CLINICAL IMPLICATIONS: R equi infection in foals is associated with a decreased chance of racing as an adult; however, foals that eventually go on to race perform comparably to the US racing population.

Actinomycetales Infections↗

PubMed1998

Maximization of cholesterol oxidase production by Rhodococcus equi no. 23 By using response surface methodology.

Medium optimization for the production of cholesterol oxidase (EC 1. 1.3.6) by Rhodococcus equi no. 23 was investigated by using response surface methodology and a central composite design. Results revealed that cholesterol and yeast extract had positive effects and the interaction between any two of three factors had no significant effect on cholesterol oxidase production. The optimized medium was the basal medium with the addition of 2.30 g/l cholesterol, 8.18 g/l yeast extract and 4.10 ml/l Tween 80. The peak cholesterol oxidase production (0.242 unit/ml) after 60-72 h cultivation was approx. 4-fold that in control medium.

Carbon↗

PubMed1998

[Other infections (Streptococcus pneumoniae, Haemophilus influenzae, Pseudomonas aeruginosa, Salmonella spp., Campylobacter spp., Nocardia asteroides, Rhodococcus equi and Bartonella spp.)].

People infected with human immunodeficiency virus (HIV) are at increased risk for bacterial infections due to HIV-associated immunologic defects. Bacterial infections were found to be, both a predictor of progression to AIDS and a substantial cause of mortality in pre-AIDS stages. Most bacterial infections are caused by Streptococcus pneumoniae, Haemophilus influenzae, Salmonella spp. and Pseudomonas aeruginosa. Rhodococcus equi, Nocardia spp., Campylobacter spp. and Bartonella spp. are less common. Data derived from two AIDS Clinical Trials Group studies showed that the most common bacterial infections were sinusitis (8.5 per 100 episodes per person years [py]), bacterial pneumonia (5.0 per 100 py), bronchitis (4.1 per 100 py) and soft tissue infections (3.5 per 100 py). In this review clinical characteristics and treatment recommendations according to data available in the literature for these infections are summarized.

AIDS-Related Opportunistic Infections↗

PubMed1998

Etherolytic cleavage of 4-(2,4-dichlorophenoxy)butyric acid and 4-(4-chloro-2-methylphenoxy)butyric acid by species of Rhodococcus and Aureobacterium isolated from an alkaline environment.

Bacterial strains were isolated from the concrete rubble of a demolished herbicide production plant. The predominant feature of these strains was the etherolytic cleavage of 4-(2,4-dichlorophenoxy)butyric acid (DCPB)1) and 4-(4-chloro-2-methylphenoxy)butyric acid (MCPB) while liberating 2,4-dichlorophenol (DCP) and 4-chloro-2-methylphenol (MCP) respectively. Some of the isolates were identified by 16S rDNA sequence analysis and shown to belong to the genera Aureobacterium sp. (strain K2-17) and Rhodococcus (Rh. erythropolis K2-12). The other strains isolated clustered into these two groups according to fatty acid analysis. Etherolytic cleavage proceeded under neutral to alkaline conditions with an optimum at around pH 8.5. With Aureobacterium sp. No. K2-17, the degradation rate was zero at a pH of 6 but as much as 60% of the maximum activity was observed at pH 10.5. With Rh. erythropolis K2-12, by contrast, pronounced activity was detected at pH 6.5 while degradation was no longer observed at pH 10.5. The maximum rates of cleavage were about 1 mmol DCPB/h.g dry mass with Aureobacterium sp. No. K2-17 and about 0.6 mmol DCPB/h.g dry mass with Rh. erythropolis K2-12. DCPB and MCPB were utilized to the same extent. Substrate cleavage and product formation (DCP) proceeded at almost equal rates with Aureobacterium sp. No. K2-17 and Rh. erythropolis K2-12, which indicates that this compound was not further metabolized. Only phenoxybutyric acid compounds served as substrates; phenoxyacetic acid and phenoxypropionic acid derivatives were not utilized by these strains.

2-Methyl-4-chlorophenoxyacetic Acid↗

PubMed1983

Degradation of aniline and monochloroanilines by Rhodococcus sp. An 117 and a pseudomonad: a comparative study.

Two newly isolated aniline-degrading bacterial strains were characterized with regard to their enzyme systems responsible for aniline catabolism. One of them identified as a Rhodococcus sp. metabolized aniline exclusively via the beta-ketoadipate pathway by means of inducible enzymes. The aniline-degrading enzyme system of the second isolate, presumably a pseudomonad, was shown to consist of an inducible aniline-converting enzyme and constitutive meta-pathway enzymes. Both isolates failed to metabolize monochlorinated anilines in the absence of additional carbon sources. To explain this the ring-cleaving enzymes of both isolates were examined for their substrate specificities. Furthermore, the effect of 4-chlorocatechol on the enzymes catalyzing aniline conversion and catechol oxygenation was investigated.

Actinomycetales↗

PubMed1996

2-D Crystallization of the Rhodococcus 20S Proteasome

The 2D crystallization method using a liquid-liquid interface has been applied to the Rhodococcus 20S proteasome. Two types of ordered arrays were obtained, both large enough for high-resolution analysis. The first one is a hexagonal close-packed array, whereas the second one has fourfold symmetry. By image analysis based on a real space correlation averaging technique, the close-packed array was found to be hexagonally packed but the molecules had complete rotational freedom. The fourfold array is, however, a true crystal with p4 symmetry. Lattice constants are a = b = 20.0 nm and the unit cell of this crystal contains two proteasomes. The diffraction pattern computed from the original picture shows the spots up to (4.5) that correspond to 3.1 nm resolution. After applying an unbending procedure, the diffraction pattern shows spots extending to 1.8 nm resolution.

Journal Article↗

PubMed1999

Biosynthesis of polyhydroxyalkanoates from low-rank coal liquefaction products by Pseudomonas oleovorans and Rhodococcus ruber

A screening identified several bacteria that were able to use chemically heterogeneous low-rank coal liquefaction products as complex carbon sources for growth. Pseudomonas oleovorans and Rhodococcus ruber accumulated polyhydroxyalkanoic acids (PHA) amounting to 2%-8% of the cell dry weight when the cells were cultivated on these liquefaction products in the absence of any other carbon source. R. ruber accumulated, in addition to PHA, small amounts of triacylglycerols. The accumulated PHA consisted of 3-hydroxyhexanoate, 3-hydroxydecanoate, and 3-hydroxydodecanoate (P. oleovorans) or 3-hydroxybutyric acid and 3-hydroxyvaleric acid (R. ruber). Low-rank coal liquefaction products obtained from Trichoderma atroviride were better substrates for P. oleovorans than chemically produced fulvic acids.

Journal Article↗

PubMed1989

Composition and toxicity of lipids from Rhodococcus rhodochrous grown on medium containing galactose, glucose or mannose.

Rhodococcus rhodochrous, a producer of mycolic acid of approx. C40, exhibited a higher cellular mass yield when grown on glucose than when grown on galactose or mannose. The cellular content of the diethyl ether-soluble lipids in microorganisms cultivated on glucose or mannose varied with the incubation time, while that of microorganisms grown on galactose remained constant. The lipids extracts from cells cultivated on different hexoses and collected at the exponential phase of growth were more toxigenic; this property was related in general to the content of glycolipid. On the other hand, cells cultivated on galactose or mannose had a higher quantity of glycolipid in the exponential phase, while the glycolipid content of those grown on glucose remained approximately constant. Amongst the components of the lipid extract, the glycolipid fraction was the sole fraction bearing toxic property. Neutral plus fatty acids and phospholipids displayed no similar characteristic.

Animals↗

PubMed1991

Time course dependent changes in contents and physical properties of glycolipid species in Rhodococcus rhodochrous.

The yield of trehalose dimycolate (TDM), the major glycolipid species elaborated by Rhodococcus rhodochrous, a producer of approx. C40-mycolic acid, was not constant in cells cultured for different periods of time. From cells collected at 24, 36, 72, 144 and 172 h of cultivation the following percentages of TDM in diethyl ether soluble lipids (DESL) were found: 10.8%, 23.4%, 10.0%, 9.0% and 5.0%, respectively. In turn, the cellular content accounted for approx. 0.6%, 1.2%, 0.9%, 0.6% and 0.2%, respectively. On the other hand, the yield of galactose monomycolate (GalMM), a minor glycolipid species maintained at approx. 3.4% in DESL during the different periods of time examined; this value represented about 0.3% of the cellular content. The melting temperatures of TDMs fell between 37 degrees C to approximately 97 degrees C with the lowest value from cells grown for 36 h, whereas the melting temperatures of the GalMMs were in a narrow range between 56 degrees C and 64 degrees C. The methyl ester derivatives of the constituent fatty acid moieties of DTMs and GalMMs migrated on thin layer chromatography like methyl esters of C40-C46 mycolic acids, therefore faster than methyl esters of C28-C34 mycolic acids but slower than methyl esters of C50-C56 mycolic acids. Further analysis of the products of pyrolysis of the methyl ester derivatives of the fatty acid moiety released from TDM after alkaline hydrolysis was carried out using gas chromatography combined mass spectrometry.(ABSTRACT TRUNCATED AT 250 WORDS)

Cord Factors↗

PubMed1989

Nitrile hydratase of Rhodococcus sp. N-774. Purification and amino acid sequences.

The nitrile hydratase of Rhodococcus sp. N-774 was purified and crystallized. The enzyme is composed of two different subunits (molecular masses: subunit alpha, 28,500 Da; subunit beta, 29,000 Da). The amino-terminal amino acid sequence of each subunit was determined. There is no sequence homology between the two subunits, suggesting that the peptides originate from different cistrons. The activity of the purified enzyme did not decrease during incubation in the dark, whereas it gradually decreased in intact cells.

Amino Acid Sequence↗

PubMed1986

Ecology of Rhodococcus (Corynebacterium) equi in soil on a horse-breeding farm.

The ecology of Rhodococcus (Corynebacterium) equi in soil was studied on a horse-breeding farm. R. equi was cultured from soil at a depth of 0, 10, and 20 cm on the six sites of the farm at monthly intervals for 10 months from March to December of 1983. The highest numbers of R. equi were found in the surface soil. The mean number of bacteria in soil samples at every depth increased remarkably from 0 or 10(2) to 10(4) colony-forming units (CFU) g-1 of soil in the middle of April, and later decreased gradually. R. equi inoculated into six soil exudate broths prepared from surface soils at separate sites yielded suspensions with different optical densities, indicating differences in growth. The distribution of serotypes in the soil was similar to that in the horses on the farm. These findings indicated that R. equi could multiply in the soil and flourish in the cycle existing between horses and their soil environment.

Actinomycetales↗

PubMed2001

Regioselective biotransformation of the dinitrile compounds 2-, 3- and 4-(cyanomethyl) benzonitrile by the soil bacterium Rhodococcus rhodochrous LL100-21.

The cyanomethyl benzonitrile compounds used for this study contain two cyano groups: a -CH(2)CN side chain, plus a cyano group attached to the benzene ring. The ortho, meta and para -CH(2)CN substituted compounds were biotransformed using whole cell suspensions of the bacterium Rhodococcus rhodochrous LL100-21. The bacterium had previously been grown on the mono-nitrile compounds propionitrile, benzonitrile or acetonitrile, inducing the formation of nitrile hydrolyzing enzymes.Suspensions of R. rhodochrous LL100-21 that had been grown on propionitrile or benzonitrile converted the aliphatic group of 2-(cyanomethyl) benzonitrile (a) to the corresponding carboxylic acid, 2-(cyanophenyl) acetic acid (d) with excellent recovery of the product and no evidence for any other products. Conversely, when grown on acetonitrile the bacterium converted 2-(cyanomethyl) benzonitrile (a) to the amide derivatives 2-(cyanophenyl) acetamide (k) and 2-(cyanomethyl) benzamide (l) but only in low yields.Biotransformations of 3-(cyanomethyl) benzonitrile (b) and 4-(cyanomethyl) benzonitrile (c), by suspensions of bacteria that had been grown on benzonitrile or propionitrile, resulted in hydrolysis of the aromatic nitrile to produce 3- and 4-(cyanomethyl) benzoic acid (j) and (m), respectively, both with a high yield. Low concentrations of other products were also detected, for example the diacids 3- and 4-(carboxyphenyl) acetic acid (h) and (i).When the bacterium was grown on acetonitrile it could biotransform 3- and 4-(cyanomethyl) benzonitrile (b) and (c) to different products indicating less regiospecificity by the nitrile hydratase enzyme.Comparison of the initial rates of conversion of the aliphatic cyano side chain of 2-(cyanomethyl) benzonitrile (a) and other substituted benzonitriles indicated that electronic effects did not affect the initial rate of the reaction as they would require transmission through an SP(3) methylene carbon atom.

Journal Article↗

PubMed1983

Recurrent skin infection with Rhodococcus in an immunosuppressed patient.

A renal transplant patient taking prednisone and azathioprine has had repeated episodes of skin infection with the soil saphrophyte Rhodococcus. Human disease with this organism has not been proved before. Although the lesions have always responded to antibiotics, frequent recurrence makes the long-term outlook uncertain.

Actinomycetales↗