Biomedical subjects

J T Trevors

Publications and source records attributed to J T Trevors.

At least 19 recordsLinked to original sources

PubMed1992

Germanium and silver resistance, accumulation, and toxicity in microorganisms.

Germanium is an inert metal with no known biological function in prokaryotic or eukaryotic organisms. Its toxicity is low compared to that of silver. Germanium is accumulated in certain bacterial strains by either energy-independent passive binding or an energy-dependent mechanism. Little is known about the molecular aspects of silver resistance, toxicity, and accumulation in bacterial strains. This is surprising because silver has been used as an antimicrobial agent in the medical field for centuries. It is likely that silver ions are excluded (resulting in decreased silver accumulation) from certain bacterial strains or immobilized intracellularly to prevent toxic effects from being exerted. These mechanisms of silver resistance have not been fully elucidated. This review examines the toxicity and accumulation of germanium and silver in selected microbial species. In addition, resistance mechanisms to these biologically nonessential metals is discussed, with more emphasis placed on silver-resistant bacteria due to the knowledge available.

Bacteria↗

PubMed1991

Respiratory activity of alginate-encapsulated Pseudomonas fluorescens cells introduced into soil.

Alginate-entrapped cells of Pseudomonas fluorescens were introduced into soil microcosms to evaluate their respiratory activity (O2 consumption and CO2 evolution) and survival during a 14-day incubation period at 20 degrees C. Alginate-entrapped cells and cells resuspended in sterile distilled water and introduced into sterile soil exhibited relatively similar O2 consumption/CO2 evolution and survival over the 14-day period. The same treatments in non-sterile soil exhibited lower respiratory activity and a population density decrease of about 2.0 Log. cfu/g after 14 days. Alginate-entrapped bacterial cells may be a useful method for introducing genetically-engineered and non-engineered bacterial strains into the soil environment.

Alginates↗

PubMed1991

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.

Biodegradation, Environmental↗

PubMed1991

Occurrence of antibiotic and metal resistance and plasmids in Bacillus strains isolated from marine sediment.

Eleven hundred Bacillus strains isolated from marine sediment from the Minas Basin, Nova Scotia, Canada, were purified on LB agar supplemented with ampicillin, chloramphenicol, erythromycin, streptomycin, tetracycline, or mercuric chloride. Seventy-seven isolates were examined for plasmid DNA, and for resistance to 11 antibiotics, HgCl2, and phenylmercuric acetate. Minimum inhibitory concentrations of Ag, Cd, Co, Cu, and Zn were also determined. Forty-three percent of antibiotic- and mercury-resistant strains contained one or more plasmids ranging from 1.9 to 210 MDa. Fifty-four percent carried plasmids greater than 20 MDa, and 97% were resistant to two or more metals. There was no correlation between plasmid content and resistance either to antibiotics or to mercurial compounds in these strains. Mercury-resistant isolates were unable to transform Hg2+ to volatile Hg0 by virtue of a mercuric reductase enzyme system (mer). Strains resistant to Hg2+ were investigated for their ability to produce H2S and intracellular acid-labile sulfide when grown in the absence and presence of HgCl2. Lower levels of H2S and intracellular sulfide were detected only in metal-resistant strains grown in the presence of HgCl2, suggesting that cellular sulfides complexed with Hg2+ in these strains.

Bacillus↗

PubMed1990

Electroporation and expression of plasmid pBR322 in Klebsiella aerogenes NCTC 418 and plasmid pRK2501 in Pseudomonas putida CYM 318.

Klebsiella aerogenes NCTC 418 and Pseudomonas putida CYM 318 were transformed via high-voltage electroporation with plasmids pBR322 and pRK2501, respectively. The number of transformants obtained was dependent on the applied voltage, capacitance, and cell recovery procedure. For example, 7.87 x 10(4) transformants/micrograms DNA were obtained at 2500 V, 25 muF when K. aerogenes cells were electroporated with pBR322 DNA. A lower voltage (1500) and capacitance (3 muF) yielded 2.4 x 10(3) transformants/micrograms DNA. P. putida CYM 318 required a 24 h outgrowth period to assist in the recovery of transformants containing pRK2501. Electroporation may be a useful protocol to transform bacterial strains that are not easily transformed by traditional methods.

Electricity↗

PubMed1990

Occurrence of aminoglycoside phosphotransferase subclass I and II structural genes among Enterobacteriaceae spp. isolated from meat samples.

3'-Aminoglycoside phosphotransferase [APH(3')] enzymes are a group responsible for resistance to the antibiotics kanamycin (Km) and neomycin (Nm) in bacteria. Escherichia coli ECT24, originally isolated from a meat sample, harboured an 83-kb conjugative R-plasmid (pRPJ24) that carries transferable resistance to Km and Nm. Plasmid pRPJ24 was transferred by conjugation to Enterobacter cloacae 94R, which was used as the source of plasmid DNA in development of a probe for the Km-resistance determinant. Random cloning of BamHI and HindIII double-digest restriction fragments of pRPJ24 in the pUC18 vector plasmid produced clones resistant to both Nm and Km carrying a 1.9-kb DNA insert. Southern hybridization of pRPJ24 cloned chimeric plasmid DNA (pKPJ94) showed homology with the APH(3')II gene from transposon Tn5. A PstI digest of pKPJ94 produced a 920-bp fragment which hybridized with the APH(3')II structural gene, and was used as a DNA probe for the APH(3')II subclass gene. A 980-bp BamHI fragment from plasmid pGH54 carrying the APH(3')I gene from transposon Tn903 was used as a subclass I probe. Total DNA from 206 randomly screened Km-resistant Enterobacteriaceae isolates from raw ground beef and chicken meat samples were examined for the occurrence of APH(3') subclass I and II using non-radioactively-labelled DNA probes. Thirty-six percent and 60% of the isolates examined carried subclass I and II resistances, respectively, in the isolates from chicken meat samples. The corresponding values for bacterial strains from raw ground beef samples were 51% and 72%, respectively. Four percent of the resistant bacterial isolates from chicken samples did not display homology to either probe.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1990

Bacterial interactions with silver.

This review examines interactions between bacteria and the biologically non-essential metal, silver. Aspects of silver toxicity, tolerance and accumulation (possible binding and uptake as opposed to energy-dependent transport) in bacteria are discussed. In addition, plasmid biology is examined briefly since little information is available on the exact mechanism(s) of plasmid-endoced silver resistance in bacteria.

Bacteria↗

PubMed1990

Mercury resistance determined by a self-transmissible plasmid in Bacillus cereus 5.

Inducible mercuric reductase activity in Bacillus cereus 5 was plasmid-encoded. Plasmid analysis revealed three plasmids with molecular masses of 2.6, 5.2 and 130 MDa. A mating system permitted transfer of the resistance determinant among strains of B. cereus and B. thuringiensis. Transfer of mercury resistance from B. cereus 5 to B. cereus 569 and B. thuringiensis occurred during mixed culture incubation on agar surfaces. The 130-MDa plasmid (pGB130) was responsible for transfer; frequencies ranged from 10(-5) to 10(-4). B. cereus 569 transconjugants inheriting pGB130 were also effective donors. High transfer frequencies and the finding that cell-free filtrates of donor cultures were ineffective in mediating transfer suggested mercury-resistance transfer was not phage-mediated. Transfer was also insensitive to DNase activity. Further evidence that pGB130 DNA carried the mercury-resistance determinant was transformation of B. cereus 569 by electroporation with pGB130 DNA isolated from B. cereus 5 and a mercury-resistant B. cereus 569 transconjugant. Mercury-resistant transconjugants and transformants exhibited mercuric reductase activity. Plasmid pGB130 also conferred resistance to phenylmercuric acetate.

Bacillus cereus↗

PubMed1990

Mobilization of Escherichia coli R1 silver-resistance plasmid pJT1 by Tn5-Mob into Escherichia coli C600.

Escherichia coli R1 is an Ag(+)-resistant strain that, as we have shown recently, harbours at least two large plasmids, pJT1 (83 kb) and pJT2 (77 kb). Tn5-Mob was introduced into the E. coli R1 host replicon via conjugation on membrane filters. The transfer functions of plasmid RP4-4 were provided in this process and Tn5-Mob clones mated with E. coli C600 yielded Ag(+)-resistant transconjugants. This mobilization procedure allowed transfer and expression of pJT1 Ag+ resistance in E. coli C600. Prior to use of Tn5-Mob mobilization, it was not possible to transfer Ag(+)-resistant determinant(s) into E. coli by conjugation or transformation including high-voltage electroporation. E. coli C600 containing PJT1 and PJT2 displayed decreased accumulation of Ag+ similar to E. coli R1.E. coli C600 could not tolerate 0.1 and 0.5 mM Ag+, rapidly accumulated Ag+ and became non-viable. Tn5-Mob mobilization may be useful in the study of metal resistance in bacteria, especially in strains not studied for resistance mechanisms.

Conjugation, Genetic↗

PubMed1990

Production of extracellular emulsifying agent by Pseudomonas aeruginosa UG1.

Twenty-three bacterial strains were isolated from oil-contaminated soil samples. Of these, 20 displayed some ability to effect oil dispersion and they were screened quantitatively for the ability to emulsify 0.5% (v/v) reference oil. One strain, identified as Pseudomonas aeruginosa UG1, produced extracellular material that emulsified reference oil, hexadecane and 2-methylnaphthalene at concentrations as high as 6% (v/v) in nutrient broth. Emulsification activity increased during a 10 day incubation period at 30 degrees C. The activity was not influenced by pH over the range 5 to 9. The emulsifying agent was precipitated by cold ethanol. The highest emulsifying activity was detected in the extracellular fraction precipitated between 30 and 50% (v/v) ethanol. A linear relationship was observed between emulsifier concentration (mg/ml) and emulsifying activity. Genetic analysis showed that the Pseudomonas aeruginosa UG1 strain did not carry extrachromosomal plasmids, suggesting that the gene(s) coding for emulsifying activity was carried on the chromosome.

Alkanes↗

PubMed1990

Electroporation and expression of the broad host-range plasmid pRK2501 in Azotobacter vinelandii.

Azotobacter vinelandii cells were transformed via high-voltage electroporation, with the broad host-range plasmid pRK2501. The number of transformants was dependent on the applied voltage, capacitance, and recovery procedure after electroporation. For example, Log, 4.44 transformants microgram-1 DNA were recovered in the A. vinelandii cell suspension electroporated at 1500 V and 25 microF capacitance (time constant 29.0 ms) and recovered on LB agar amended with 0.5 microgram/ml-1 kanamycin (pRK2501 encodes for both kanamycin and tetracycline resistance). Electroporation at 2500 V and capacitance settings of 25 and 3 microF did not produce any transformants. Cell survival was also poor at high voltages. A. vinelandii transformants were not recovered on N-free agar medium. In addition, no viable cells were recovered on N-free agar after electroporation at 2500 V, 25 microF; 2500 V, 3 microF; and 1500 V, 25 microF. Electroporation may be a useful method to genetically transform Azotobacter species for use in physiological and/or genetic studies.

Azotobacter↗

PubMed1990

Silver accumulation and resistance in Escherichia coli R1.

E. coli R1 contains at least 2 large plasmids (83 and 77 kb) while E. coli S1 was previously cured of the 83 kb plasmid. Transformation using artificial competence, high-voltage electroporation, and plasmid mobilization experiments with the mobilizing plasmid RP4, failed to ascertain the 83 kb plasmid was responsible for Ag(+)-resistance. Silver accumulation by an Ag(+)-sensitive E. coli S1 strain was 5-fold higher than an Ag(+)-resistant E. coli R1 strain. The Ag(+)-resistant E. coli R1 strain produced 33% more H2S and 32% more intracellular acid labile SH than the Ag(+)-sensitive E. coli R1 strain when grown in the absence of AgNO3. Hydrophobic interaction chromatography revealed E. coli R1 displayed higher cell surface hydrophobicity than E. coli S1. HPLC protein analysis of cell-free extracts also revealed differences between protein fractions in E. coli R1 and S1 strains.

Chromatography, Gel↗

PubMed1990

Influence of freezing-thawing and refrigeration on R-plasmid (pRPJ24) stability in Enterobacter cloacae 94R.

The effect of twenty freezing (-20 degrees C) and thawing cycles of Enterobacter cloacae 94R cells containing the R-plasmid pRPJ24 inoculated into broth and ground beef meat samples revealed no loss of resistance due to plasmid instability. In addition, low temperature storage at 4 degrees C did not produce any significant loss of the tetracycline and kanamycin resistances encoded on the pRPJ24 plasmid. The results of this study indicated that indigenous R-plasmids like pRPJ24 are stable in resident recipients like E. cloacae 94R in ground beef. However, the proportion of viable cells containing the pRPJ24 plasmid decreased significantly after 20 freezing-thawing cycles over 14 days incubation at 4 degrees C.

Animals↗

PubMed1990

Transport of a genetically engineered Pseudomonas fluorescens strain through a soil microcosm.

Vertical soil microcosms flushed with groundwater were used to study the influence of water movement on survival and transport of a genetically engineered Pseudomonas fluorescens C5t strain through a loamy sand and a loam soil. Transport of cells introduced into the top 1 cm of the vertical soil microcosms was dependent on the flow rate of water and the number of times microcosms were flushed with groundwater. The presence of wheat roots growing downward in the microcosms contributed only slightly to the movement of P. fluorescens C5t cells to lower soil regions of the loamy sand microcosms, but enhanced downward transport in the loam microcosms. Furthermore, the introduced P. fluorescens C5t cells were detected in the effluent water samples even after three flushes of groundwater and 10 days of incubation. As evidenced by a comparison of counts from immunofluorescence and selective plating, nonculturable C5t cells occurred in day 10 soil and percolated water samples, primarily of the loamy sand microcosms. Vertical soil microcosms that use water movement may be useful in studying the survival and transport of genetically engineered bacteria in soil under a variety of conditions prior to field testing.

Colony Count, Microbial↗

PubMed1989

Silver accumulation in Pseudomonas stutzeri AG259.

Silver toxicity to Pseudomonas stutzeri AG259 was strongly dependent on the NaCl concentration in the medium, which reduced the availability of Ag+ by precipitation as AgCl. Accumulation of Ag by growing cultures was low being less than or equal to 7.5 nmol (mg dry mass)-1 over all treatments examined. The presence of NaCl in the growth medium did not markedly affect the amounts of Ag accumulated by the cells but influenced toxicity as manifest by a lag period which was greatest at low NaCl concentrations (less than or equal to 0.1% mass/vol.). In NaCl-free medium, P. stutzeri did not grow in the presence of 0.5 mM AgNO3 in contrast to Ag-free controls. The majority of Ag accumulation by resting cells of P. stutzeri occurred within 1 min of incubation and there was little difference in uptake capacities between cells previously grown in the absence or presence of AgNO3. Lowest amounts of Ag uptake by resting cells occurred when suspended in 1 mM Mes pH 6.5, containing 1% (mass/vol.) NaCl. Prior exposure of P. stutzeri to Cu(NO3)2 resulted in a marked reduction in Ag uptake when suspended in 1 mM Mes pH 6.5, containing 0.5 mM AgNO3.

Binding Sites↗

PubMed1989

Silver resistance in Escherichia coli R1.

Escherichia coli strain R1, originally isolated from a patient whose burns were treated with silver sulphadiazine, contained two large plasmids of 83 kb (pJT1) and 77 kb (pJT2), and was resistant to 1 mM AgNO3. A silver-sensitive derivative, E. coli S1, cured of the 83-kb plasmid pJT1, was obtained by growth at 46 degrees C. Studies with an Ag+-specific ion electrode showed no significant differences in Ag+ binding by washed resting cell suspensions of strains R1 and S1, with and without glucose. However, transmission electronmicroscopy and energy dispersive X-ray analysis of whole cell mounts from actively growing cultures showed that the Ag+-resistant strain did not accumulate Ag+, whereas the sensitive strain contained dense silver particles. Both strains produced H2S, detected by blackening of lead acetate paper above inoculated broth, and reducing substances (possibly H2S) were detected only around E. coli R1 colonies when methylene blue was used as a indicator in LB agar, which may be a less sensitive assay. The mechanism of silver resistance is not known, but actively growing cells of E. coli R1 did not accumulate silver.

Bacterial Outer Membrane Proteins↗

PubMed1989

Transformation of Bacillus cereus vegetative cells by electroporation.

Transformation of untreated vegetative cells of Bacillus cereus 569 with plasmid pC194 (1.8 megadaltons) by high-voltage electroporation resulted in a maximum of 2 x 10(-5) transformants per viable cell. Transformation of a 130-megadalton plasmid occurred at a comparable frequency. The method was simple, rapid, and yielded transformant colonies in 14 to 24 h. Transformation was obtained with unpurified total plasmid DNA.

Bacillus cereus↗

PubMed1989

Survival of and plasmid stability in Pseudomonas and Klebsiella spp. introduced into agricultural drainage water.

Cell survival and plasmid stability in Pseudomonas fluorescens R2f and Pseudomonas putida CYM 318 containing respectively, plasmid RP4 and pRK2501, and Klebsiella aerogenes NCTC 418 harboring plasmid pBR322 were studied in sterile and nonsterile agricultural drainage water under both aerobic and anaerobic conditions and in the absence and presence of added nutrients. Both Pseudomonas strains survived well in sterile drainage water incubated aerobically, with or without added nutrients. However, Klebsiella aerogenes NCTC 418 (pBR322) only survived in the presence of added nutrients. Pseudomonas fluorescens R2f (RP4) and K. aerogenes NCTC 418 (pBR322) did not survive under anerobic conditions without added nutrients, but showed good survival in the presence of nutrients. Survival of all three strains was negatively affected in nonsterile agricultural drainage water when compared with survival in sterile water. Maintenance of the three plasmids was host, plasmid, and environment dependent. Plasmid pBR322 was not stably maintained in K. aerogenes NCTC 418 under all conditions used in the study, and pRK2501 was readily lost from P. putida CYM 318. Maintenance of RP4 by P. fluorescens R2f was markedly influenced by added nutrients, which caused a loss of the plasmid from cells. The results of the present study demonstrate the influence of nutrients, O2, and native microorganisms on the survival of introduced bacterial strains and plasmid stability in agricultural drainage water.

Klebsiella↗