Biomedical subjects

J T Trevors

Publications and source records attributed to J T Trevors.

At least 19 recordsLinked to original sources

PubMed2006

Analysis of denitrification genes and comparison of nosZ, cnorB and 16S rDNA from culturable denitrifying bacteria in potato cropping systems.

Bacterial denitrification in agricultural soils is a major source of nitrous oxide, a potent greenhouse gas. This study examined the culturable bacterial population of denitrifiers in arable field soils in potato (Solanum tuberosum L.) production and denitrification genes (nir, nor and nos) and 16S rDNA in those isolates. Enrichments for culturable denitrifiers yielded 31 diverse isolates that were then analysed for denitrification genes. The nitrous oxide reductase (nosZ) gene was found in all isolates. The majority of isolates ( approximately 90%) contained the cnorB nitric oxide reductase gene, with the remainder containing the qnorB gene. Nitrite reductase genes (nirS and nirK) were amplifiable from most of the isolates, and were segregated between species similar to previously isolated denitrifiers. Isolated strains were preliminarily identified using fatty acid methyl ester analysis and further identified using 16S rDNA sequencing. The majority of isolates (21) were classified as Pseudomonas sp., with smaller groups of isolates being most similar to Bosea spp. (4), Achromobacter spp. (4) and two isolates closely related to Sinorhizobium/Ensifer spp. Phylogenetic trees were compared among nosZ, cnorB and 16S rDNA genes for a subset of Pseudomonas strains. The trees were mostly congruent, but some Pseudomonas sp. isolates grouped differently depending on the gene analysed, indicating potential horizontal gene transfer of denitrification genes. Although Bosea spp. are known denitrifiers, to the best of our knowledge this is the first report of isolation and sequencing of denitrification genes from this bacterial genus.

Bacteria↗

PubMed2005

The Big Bang, Superstring Theory and the origin of life on the Earth.

This article examines the origin of life on Earth and its connection to the Superstring Theory, that attempts to explain all phenomena in the universe (Theory of Everything) and unify the four known forces and relativity and quantum theory. The four forces of gravity, electro-magnetism, strong and weak nuclear were all present and necessary for the origin of life on the Earth. It was the separation of the unified force into four singular forces that allowed the origin of life.

Animals↗

PubMed2005

Real-time polymerase chain reaction quantification of the transgenes for roundup ready corn and roundup ready soybean in soil samples.

A method for quantification of recombinant DNA for Roundup Ready (RR) corn and RR soybean in soil samples is described. Soil DNA from experimental field samples was extracted using a soil DNA extraction kit with a modified protocol. For the detection and quantification of recombinant DNA of RR corn and RR soybean, a molecular beacon and two pairs of specific primers were designed to differentially target recombinant DNA in these two genetically modified crops. Soil DNA extracts were spiked with RR corn or RR soybean DNA, and recombinant DNA was quantified using real-time PCR with a molecular beacon. As few as one copy of RR corn genome or one copy of RR soybean genome was detected in the soil DNA extract.

DNA, Plant↗

PubMed2005

Fluorescent methods to study DNA, RNA, proteins and cytoplasmic membrane polarization in the pentachlorophenol-mineralizing bacterium Sphingomonas sp. UG30 during nutrient starvation in water.

The effect of sodium pentachlorophenolate (NaPCP) exposure on the nutrient-starved pentachlorophenol (PCP)-mineralizing bacterium Sphingomonas sp. UG30 was assessed using fluorescent methods to measure DNA, RNA, total cellular protein, and cytoplasmic membrane proteins. UG30 cells were inoculated into sterilized Speed River (Guelph, ON, Canada) water samples in the presence of 50, 100, and 250 ppm NaPCP. No marked changes were observed in the total cellular DNA, RNA or protein levels over 90 d, indicating the macromolecular composition of UG30 was not affected by both nutrient limitation and NaPCP. Total cell counts as determined by DAPI staining also did not change over 90 d. Over the same period, viable counts decreased with increasing concentrations of NaPCP. At 250 ppm NaPCP, viable cell counts decreased over 6 orders of magnitude after 1 hr exposure. Cell numbers partially recovered once NaPCP was degraded. The UG30 cytoplasmic membrane polarization ratio also decreased after NaPCP was depleted. The decreased polarization value at the end of the study period suggested the UG30 membrane was more fluid and that this increase in fluidity was due to nutrient starvation effects rather than exposure to NaPCP. These results indicated that UG30 is a robust organism that is able to degrade NaPCP even under adverse conditions and fluorescent methods are useful for determining macromolecular concentrations and cytoplasmic membrane polarization values.

Bacterial Proteins↗

PubMed2004

Persistence of extracellular baculoviral DNA in aquatic microcosms: extraction, purification, and amplification by the polymerase chain reaction (PCR).

Genetically-modified baculoviruses have potential uses as bio-pesticides in forestry. However, the baculoviral occlusion bodies (OBs) may release genetically-modified DNA into the forest environment. In this research, outdoor aquatic microcosms, spiked with 673 microg of genomic DNA (4.4 x 10(12) target copies) from the genetically modified baculovirus Choristoneura fumiferana MNPVegt-/lacZ+, were exposed to natural summer conditions. A 530 bp DNA fragment from the genome of CfMNPVegt-/lacZ+ was detected in field microcosm water samples for about 24 h. The introduced DNA may have persisted for a longer time, but was below the detection limit of the PCR analysis (13.5 pg DNA or 8.9 x 10(4) target copies ml(-1) water). The detection limit of PCR was determined by spiking water samples with a dilution series of CfMNPVegt-/lacZ+ genomic DNA, extracting and purifying the DNA, and then PCR analysis. This study provides some of the first information on the persistence and detection limits of this viral DNA under aquatic ecological conditions, and the methods that can be used to conduct such a molecular-based field study.

Baculoviridae↗

PubMed2004

Cytoplasmic membrane polarization in Gram-positive and Gram-negative bacteria grown in the absence and presence of tetracycline.

The ability of numerous diverse compounds and ions to cross the bacterial cytoplasmic membrane by diffusion and active transport is highly dependent on cytoplasmic membrane fluidity, which can be measured using fluorescent probes to estimate membrane polarization values. However, membrane polarization data are lacking for most bacterial species. The cytoplasmic membrane polarization values for Arthrobacter sp. ATCC 21908, Bacillus cereus NRC 3045, Pseudomonas fluorescens R2F, Pseudomonas putida NRC 2986 and Escherichia coli C600 bacterial cells were spectrofluorometrically measured over a temperature range from 10 to 50 degrees C, and in the absence and presence of 1 microg/ml tetracycline, using the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene (DPH) to obtain new information on their membrane fluidity. At an assay temperature of 10 degrees C, E. coli cells grown in the absence of tetracycline exhibited the highest cytoplasmic membrane polarization value (least fluid membrane) of 0.446, followed by values of 0.392, 0.371, 0.344 and 0.293, respectively, for B. cereus, Arthrobacter sp., P. fluorescens and P. putida. At an assay temperature of 30 degrees C, the polarization values ranged from 0.357 to 0.288 for cells grown in the absence of tetracycline, regardless of the species. B. cereus grown in the presence of 1 microg/ml tetracycline had lower polarization values than when grown in the absence of this antibiotic at all assay temperatures. Regardless of the absence or presence of 1 microg/ml tetracycline in the growth medium, all bacterial species generally exhibited a more fluid membrane as the assay temperature increased from 10 to 50 degrees C. To our knowledge, these are some of the first cytoplasmic membrane polarization values reported for these Gram-negative and Gram-positive bacteria over a broad temperature range and also for cells grown in the presence of tetracycline.

Cell Division↗

PubMed2004

Chance and necessity do not explain the origin of life.

Where and how did the complex genetic instruction set programmed into DNA come into existence? The genetic set may have arisen elsewhere and was transported to the Earth. If not, it arose on the Earth, and became the genetic code in a previous lifeless, physical-chemical world. Even if RNA or DNA were inserted into a lifeless world, they would not contain any genetic instructions unless each nucleotide selection in the sequence was programmed for function. Even then, a predetermined communication system would have had to be in place for any message to be understood at the destination. Transcription and translation would not necessarily have been needed in an RNA world. Ribozymes could have accomplished some of the simpler functions of current protein enzymes. Templating of single RNA strands followed by retemplating back to a sense strand could have occurred. But this process does not explain the derivation of "sense" in any strand. "Sense" means algorithmic function achieved through sequences of certain decision-node switch-settings. These particular primary structures determine secondary and tertiary structures. Each sequence determines minimum-free-energy folding propensities, binding site specificity, and function. Minimal metabolism would be needed for cells to be capable of growth and division. All known metabolism is cybernetic--that is, it is programmatically and algorithmically organized and controlled.

Animals↗

PubMed2004

Extraction, detection and persistence of extracellular DNA in forest litter microcosms.

A DNA extraction method was developed that preferentially extracted extracellular DNA rather than intracellular DNA from forest litter. The method purposely avoided the use of harsh chemicals and physical disruption steps used in total DNA extraction to release DNA from cells. The detection limit of PCR, determined by spiking forest litter samples with a dilution series of Choristoneura fumiferana MNPVegt(-)/lacZ(+) genomic DNA, was about 1 ng DNA or 6.85 x 10(6) target copies 0.5 g(-1) moist forest litter or 0.14 g(-1) dry forest litter. In this study, outdoor terrestrial microcosms, each spiked with 49.2 microg of genomic DNA (from the baculovirus CfMNPVegt(-)/lacZ(+)), were exposed to summer conditions. A 530 bp DNA fragment from the genome of the baculovirus CfMNPVegt(-)/lacZ(+) was detected in these microcosms for about 3 months. The DNA may have persisted for a longer period but was below the detection limit of the PCR analysis.

Animals↗

PubMed2004

The development of a DNA microarray-based assay for the characterization of commercially formulated microbial products.

Commercially formulated bioproducts containing a complex consortia of bacteria as an active ingredient pose a significant challenge for regulatory agencies and companies seeking to assess the safety and efficacy of these bioproducts. The main challenge stems from how to characterize the bacterial composition of these products, for which there is presently a lack of suitable methods. A prototype DNA microarray composed of oligonucleotide probes for functional genes, virulence factors, and taxonomic genes for a number of bacterial species was developed to examine the utility of microarray technology as a molecular tool for characterizing consortia bioproducts. The genomic DNA from four different products was extracted by two methods and examined with the microarray prototype and by denaturing gradient gel electrophoresis (DGGE). Although the identity of the consortial species remains unknown, the microarray assay provided unique and reproducible hybridization patterns for all four products, and agreed with the fingerprints generated by DGGE. The ability to differentiate between a variety of consortia products demonstrates that DNA microarrays have the potential to be a powerful tool in monitoring complex microbial communities.

Bacteriological Techniques↗

PubMed2004

Biology, persistence and detection of Cryptosporidium parvum and Cryptosporidium hominis oocyst.

Cryptosporidium parvum and Cryptosporidium hominis are obligate enteric protozoan parasites which infect the gastrointestinal tract of animals and humans. The mechanism(s) by which these parasites cause gastrointestinal distress in their hosts is not well understood. The risk of waterborne transmission of Cryptosporidium is a serious global issue in drinking water safety. Oocysts from these organisms are extremely robust, prevalent in source water supplies and capable of surviving in the environment for extended periods of time. Resistance to conventional water treatment by chlorination, lack of correlation with biological indicator microorganisms and the absence of adequate methods to detect the presence of infectious oocysts necessitates the development of consistent and effective means of parasite removal from the water supply. Additional research into improving water treatment and sewage treatment practices is needed, particularly in testing the efficiency of ozone in oocyst inactivation. Timely and efficient detection of infectious C. parvum and C. hominis oocysts in environmental samples requires the development of rapid and sensitive techniques for the concentration, purification and detection of these parasites. A major factor confounding proper detection remains the inability to adequately and efficiently concentrate oocysts from environmental samples, while limiting the presence of extraneous materials. Molecular-based techniques are the most promising methods for the sensitive and accurate detection of C. parvum and C. hominis. With the availability of numerous target sequences, RT-PCR will likely emerge as an important method to assess oocyst viability. In addition, a multiplex PCR for the simultaneous detection of C. parvum, C. hominis and other waterborne pathogens such as Giardia lamblia would greatly benefit the water industry and protect human health.

Animals↗

PubMed2003

Growth and membrane polarization in Pseudomonas aeruginosa UG2 grown in randomized microgravity in a high aspect ratio vessel.

Growth and membrane polarization of Pseudomonas aeruginosa UG2 cells grown under randomized microgravity (RMG) and 1xg were measured in a high aspect ratio vessel (HARV) and also in batch cultures mixed at 12 and 150 rpm in Erlenmeyer shake flasks. Membrane polarization was measured using the fluorescent probe, 1,6-diphenyl-1,3,5-hexatriene (DPH). No differences were observed in the growth curves or membrane polarization values (about 0.300) under all three culture conditions. However, the net effect of RMG at the single cell level may be still unknown. It may be possible that RMG effects are species-dependent or bacterial cells with a small mass and volume may be near the threshold where RMG exerts a minimal effect.

Bioreactors↗

PubMed2003

Fluorescent probes for bacterial cytoplasmic membrane research.

Fluorescent methods in biological and medical research are extremely useful at the cellular and molecular levels. This is due to sensitive and affordable detection equipment and a variety of specific and more general fluorescent probes, and analytical procedures. In this article, I examine the use of fluorescence membrane probes to study the fluidity (membrane polarization) of the bacterial cytoplasmic membrane, central to energy transduction, ion and nutrient transport and diffusion of water and gases.

Bacteria↗

PubMed2003

Polychlorinated biphenyl (PCB) degradation and persistence of a gfp-marked Ralstonia eutropha H850 in PCB-contaminated soil.

Ralstonia eutropha H850 was labelled chromosomally with a gfp marker gene encoding for the green fluorescent protein, and designated R. eutropha H850g13. Visual observation of green fluorescent cells under an epifluorescence microscope, and PCR amplification products, confirmed that the bacterium was labelled with gfp. Southern blot hybridization products further confirmed the gfp was chromosomally labelled. Using resting cell assays, it was determined that insertion of the gfp gene decreased the microorganisms' ability to degrade biphenyl compared to the parent strain. However, this marker facilitated the identification and monitoring of R. eutropha H850g13 survival in soil microcosm experiments. Survival and polychlorinated biphenyl degradation by R. eutropha H850g13 was analysed in soil microcosms spiked with 2,2',5,5'-tetrachlorobiphenyl (TeCB). R. eutropha H850g13 was detected by viable plate counts and most-probable-number/PCR after 102 days in TeCB-contaminated soil microcosms, and was likely outcompeted by indigenous soil microorganisms in microcosms amended with oil and Daramend (an organic amendment, http://www.adventusremediation.com). R. eutropha H850g13 did not degrade TeCB in any of the soil microcosms. This research confirmed that gfp was useful as a marker to distinguish R. eutropha H850g13 from indigenous soil microorganisms over a 102 day period and that, under the experimental conditions used, R. eutropha H850g13 did not degrade TeCB.

Benzidines↗

PubMed2003

Early assembly of cellular life.

Popular hypotheses that attempt to explain the origin of prebiotic molecules and cellular life capable of growth and division are not always agreed upon. In this manuscript, information on early bacterial life on Earth is examined using information from several disciplines. For example, knowledge can be integrated from physics, thermodynamics, planetary sciences, geology, biogeochemistry, lipid chemistry, primordial cell structures, cell and molecular biology, microbiology, metabolism and genetics. The origin of life also required a combination of elements, compounds and environmental physical-chemical conditions that allowed cells to assemble in less than a billion years. This may have been widespread in the subsurface of the early Earth located at microscopic physical domains.

Adaptation, Physiological↗

PubMed2003

Effect of selected environmental and physico-chemical factors on bacterial cytoplasmic membranes.

Membranes lipids are one of the most adaptable molecules in response to perturbations. Even subtle changes of the composition of acyl chains or head groups can alter the packing arrangements of lipids within the bilayer. This changes the balance between bilayer and nonbilayer lipids, serving to affect bilayer stability and fluidity, as well as altering lipid-protein interactions. External factors can also change membrane fluidity and lipid composition; including temperature, chemicals, ions, pressure, nutrients and the growth phase of the microbial culture. Various biophysical techniques have been used to monitor fluidity changes within the bacterial membrane. In this review, bacterial cytoplasmic membrane changes and related functional effects will be examined as well as the use of fluorescence polarization methods and examples of data obtained from research with bacteria.

Bacteria↗

PubMed2003

Possible origin of a membrane in the subsurface of the Earth.

A location for the origin of life on Earth could have been an oil/water interface in the warm, subsurface environment of the Earth. The physico-chemical conditions of the subsurface would include elevated, but eventually cooling temperatures, anaerobic conditions, and protection from intense surface radiation. This type of subsurface oil/water environment may have been ideal for the assembly of the first simple membrane(s), where no enzyme catalysis was needed. Once a stable, simple, continuous closed membrane was formed, one central component of the first cell(s) would have been present; a semi-permeable open system that allowed the passage of both matter and energy in and out of the cell. Such an open system could also acquire novel functions, whereas a closed system would be unable to evolve.

Cell Membrane↗

PubMed2002

Characterization of tetrachlorohydroquinone reductive dehalogenase from Sphingomonas sp. UG30.

Tetrachlorohydroquinone reductive dehalogenase (PcpC) is the second of three enzymes that catalyze the initial degradation of pentachlorophenol in Sphingomonas sp. UG30 and several other bacterial strains. The UG30 PcpC shares a high degree (94%) of primary sequence identity with the well-studied PcpC from Sphingobium chlorophenolicum ATCC 39723. Significant differences, however, were observed between the two PcpC enzymes in some of their functional and kinetic properties. The temperature optimum of the UG30 PcpC is 10 degrees C higher and the pH optimum is approximately 2 units higher than the S. chlorophenolicum PcpC. In addition, the S. chlorophenolicum PcpC is subject to inhibition by the substrate tetrachlorohydroquinone (TCHQ), and this has necessitated the use of a mutant enzyme, which was not inhibited by TCHQ, for kinetic studies. In contrast, the UG30 PcpC was not inhibited by TCHQ and this may allow detailed kinetic and mechanistic studies using the wild-type enzyme.

Amino Acid Sequence↗

PubMed2002

Bacterial genomics: the use of DNA microarrays and bacterial artificial chromosomes.

Immense amounts of genetic information are contained within microbial genomes. As the number of completely sequenced microbial genomes is increasing, functional and comparative genomic techniques will be employed for sequence analysis and gene characterization. Sequence comparison and expression profiling by DNA microarrays can determine phylogenetic relationships and identify genes while bacterial artificial chromosomes (BACs) allow the study of entire biochemical pathways and permit the expression of bacterial genes in a foreign host.

Anti-Bacterial Agents↗