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At least 685 records · Page 38Linked to original sources
PubMed2005

Apical extrusion of intracanal bacteria following use of two engine-driven instrumentation techniques.

AIM: To evaluate the number of bacteria extruded apically from extracted teeth ex vivo after canal instrumentation using the two engine-driven techniques utilizing nickel-titanium instruments (ProTaper and System GT). METHODOLOGY: Forty extracted single-rooted human mandibular premolar teeth were used. Access cavities were prepared and root canals were then contaminated with a suspension of Enterococcus faecalis and dried. The contaminated roots were divided into two experimental groups of 15 teeth each and one control group of 10 teeth. Group 1, ProTaper group: the root canals were instrumented using ProTaper instruments. Group 2, System GT group: the root canals were instrumented using System GT instruments. Group 3, control group: no instrumentation was attempted. Bacteria extruded from the apical foramen during instrumentation were collected into vials. The microbiological samples from the vials were incubated in culture media for 24 h. Colonies of bacteria were counted and the results were given as number of colony-forming units. The data obtained were analysed using the Kruskal-Wallis one-way analysis of variance and Mann-Whitney U-tests, with alpha = 0.05 as the level for statistical significance. RESULTS: There was no significant difference as to the number of extruded bacteria between the ProTaper and System GT engine-driven systems (P > 0.05). CONCLUSIONS: Both engine-driven nickel-titanium systems extruded bacteria through the apical foramen.

Colony Count, Microbial↗
PubMed1989

Production of extracellular nucleic acids by genetically altered bacteria in aquatic-environment microcosms.

The factors which affect the production of extracellular DNA by genetically altered strains of Escherichia coli, Pseudomonas aeruginosa, Pseudomonas cepacia, and Bradyrhizobium japonicum in aquatic environments were investigated. Cellular nucleic acids were labeled in vivo by incubation with [3H]thymidine or [3H]adenine, and production of extracellular DNA in marine waters, artificial seawater, or minimal salts media was determined by detecting radiolabeled macromolecules in incubation filtrates. The presence or absence of the ambient microbial community had little effect on the production of extracellular DNA. Three of four organisms produced the greatest amounts of extracellular nucleic acids when incubated in low-salinity media (2% artificial seawater) rather than high-salinity media (10 to 50% artificial seawater). The greatest production of extracellular nucleic acids by P. cepacia occurred at pH 7 and 37 degrees C, suggesting that extracellular-DNA production may be a normal physiologic function of the cell. Incubation of labeled P. cepacia cells in water from Bimini Harbor, Bahamas, resulted in labeling of macromolecules of the ambient microbial population. Collectively these results indicate that (i) extracellular-DNA production by genetically altered bacteria released into aquatic environments is more strongly influenced by physiochemical factors than biotic factors, (ii) extracellular-DNA production rates are usually greater for organisms released in freshwater than marine environments, and (iii) ambient microbial populations can readily utilize materials released by these organisms.

DNA, Bacterial↗
PubMed1988

Use of microcosms to study genetic interactions between microorganisms.

Microcosms are used regularly in the study of the toxicity of environmental chemicals and their biodegradation. The use of these systems to study genetic interactions and adaptation mechanisms in soil and aquatic microorganisms is discussed, and their application to studies on the fate of genetically engineered organisms and their genes in the natural environment is outlined.

Conjugation, Genetic↗
PubMed2005

Antibody processing and engineering in plants, and new strategies for vaccine production.

The use of transgenic plants for the production of recombinant proteins is not a universal solution for all proteins. The choice of this expression system depends very much on the type of protein and its applications. Many proteins will best be made by conventional microbial fermentation, similarly, we are already identifying proteins where plants represent the only practical option for one reason or another. It will be important to understand better the cellular mechanisms of protein folding, assembly and processing in plants, in order to maximise the potential of transgenic plants as a protein production system. One of the main advantages that plants offer is that they are higher eukaryotic organisms with an endomembrane system. Therefore, they fold and assemble recombinant proteins using protein chaperones that are homologous to those in mammalian cells, and they perform post-translational modifications. This allows, for example, the expression of monoclonal antibodies, first described in 1989, as well as a range of other types of immunoglobulin molecules and multimeric complexes.

Animals↗
PubMed1995

Issues and advances in the use of transgenic organisms for the production of thaumatin, the intensely sweet protein from Thaumatococcus danielli.

The thaumatins are a class of intensely sweet proteins isolated from the fruit of the tropical plant Thaumatococcus danielli. Thaumatin is approved for use in many countries and has application as both a flavor enhancer and a high-intensity sweetener. The supply of naturally occurring thaumatin is limited, which has prompted extensive research into its synthesis via transgenic organisms. The gene encoding thaumatin has been introduced into various microorganisms under transcriptional control of heterologous promoters. Yields to date have been low, but the factors governing more efficient microbial production have been identified. Continued research should allow microbial yields to be improved to commercially viable levels. The unique properties of thaumatin as a food additive could well be exploited by the food industry. Alternatively, the thaumatin gene could be engineered directly into selected fruit and vegetable crops to improve their flavor and sweetness.

Africa↗
PubMed1998

Function of root border cells in plant health: pioneers in the rhizosphere.

Plants dedicate a large amount of energy to the regulated production of living cells programmed to separate from roots into the external environment. This unusual process may be worth the cost because it enables the plant to dictate which species will share its ecological niche. For example, border cells can rapidly attract and stimulate growth in some microorganisms and repel and inhibit the growth of others. Such specificity may provide a way to control the dynamics of adjacent microbial populations in the soil to foster beneficial associations and inhibit pathogenic invasion. Plant genes controlling the delivery of border cells and the expression of their unique properties provide tools to genetically engineer plants with altered border cell quality and quantity. Such variants are being used to test the hypothesis that the function of border cells is to protect plant health by controlling the ecology of the root system.

Journal Article↗
PubMed1992

Engineering dehydrated amino acid residues in the antimicrobial peptide nisin.

The small antimicrobial peptide nisin, produced by Lactococcus lactis, contains the uncommon amino acid residues dehydroalanine and dehydrobutyrine and five thio ether bridges. Since these structures are posttranslationally formed from Ser, Thr, and Cys residues, it is feasible to study their role in nisin function and biosynthesis by protein engineering. Here we report the development of an expression system for mutated nisin Z (nisZ) genes, using nisin A producing L. lactis as a host. Replacement by site-directed mutagenesis of the Ser-5 codon in nisZ by a Thr codon, led to a mutant with a dehydrobutyrine instead of a dehydroalanine residue at position 5, as shown by NMR. Its antimicrobial activity was 2-10-fold lower relative to wild-type nisin Z, depending on the indicator strain used. In another mutagenesis study a double mutation was introduced in the nisZ gene by replacing the codons for Met-17 and Gly-18 by codons for Gln and Thr, respectively, as in the third lanthionine ring of the related antimicrobial peptide subtilin from Bacillus subtilis. This resulted in the simultaneous production of two mutant species, one containing a Thr residue and the other containing a dehydrobutyrine residue at position 18, both having different bacteriocidal properties.

Amino Acid Sequence↗
PubMed2004

Rapid preparation of isotopolog libraries by in vivo transformation of 1)C-glucose. Studies on 6,7-dimethyl-8-ribityllumazine, a biosynthetic precursor of vitamin B2.

An Escherichia coli strain engineered for expression of the ribABGH genes of Bacillus subtilis was shown to produce 100 mg of the riboflavin precursor 6,7-dimethyl-8-ribityllumazine per liter of minimal medium. Growth of the recombinant strain in medium supplemented with [U-13C6]glucose and/or 15NH4Cl as single sources of carbon and/or nitrogen afforded 6,7-dimethyl-8-ribityllumazine universally labeled with 13C and/or 15N. The yield of [U-13C13]-6,7-dimethyl-8-ribityllumazine based on [U-13C6]glucose was 25 mg/g. Fermentation with [1-13C1]-, [2-13C1]-, or [3-13C1]glucose afforded mixtures of 6,7-dimethyl-8-ribityllumazine isotopologs, predominantly with 13C enrichment of single carbon atoms. The isotope-labeled samples enabled a comprehensive NMR analysis of 6,7-dimethyl-8-ribityllumazine. Isotopolog libraries of a wide variety of microbial metabolites can be produced by the same experimental approach.

Bacillus subtilis↗
PubMed2000

Evidence of in vivo differential bioavailability of the active forms of matrix metalloproteinases 9 and 2 in parturition, spontaneous rupture of membranes, and intra-amniotic infection.

OBJECTIVE: Matrix metalloproteinases (MMP-9 and MMP-2) have been implicated in the digestion of fetal membranes. The purpose of this study was to determine the amniotic fluid concentrations of active forms of MMP-2 and MMP-9 and to explore the participation of these enzymes in labor (term and preterm), rupture of membranes (term and preterm), and microbial invasion of the amniotic cavity. STUDY DESIGN: A cross-sectional study was conducted with 291 women in the following categories: (1) term not in labor, (2) term in labor, (3) preterm labor and intact membranes who delivered at term, (4) preterm labor who delivered preterm, (5) preterm labor with microbial invasion of the amniotic cavity, (6) preterm premature rupture of membranes without microbial invasion of the amniotic cavity, (7) preterm premature rupture of membranes with microbial invasion of the amniotic cavity, (8) term premature rupture of membranes not in labor, and (9) mid trimester. Active forms of MMP-2 and MMP-9 were measured by a novel assay that uses a substrate developed by protein engineering. RESULTS: (1) MMP-2 and MMP-9 were detected in 88% and 96% of amniotic fluid samples, respectively (255/291 and 279/291). (2) The concentrations of active forms of MMP-2 and MMP-9 changed with advancing gestational age. (3) Spontaneous term parturition was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) and a significant decrease in the median concentration of the active forms of MMP-2 (P <.003). (4) Preterm labor with intact membranes leading to preterm delivery in the absence of infection was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.2). (5) Rupture of membranes (either term or preterm) was associated with a significant increase in the concentration of the active forms of MMP-9 and with a significant decrease in the concentration of the active forms of MMP-2 (P <.005 for term and P <.03 and P <.003 for preterm, respectively). (6) Microbial invasion of the amniotic cavity in women with preterm premature rupture of membranes was also associated with a significant increase in the concentration of the active forms of MMP-9 (P <.03) and a decrease in the concentration of the active forms of MMP-2 (P <.05). (7) Microbial invasion of the amniotic cavity in patients with preterm labor was associated with a significant increase in the median concentration of the active forms of MMP-9 (P <.005) but not of the active forms of MMP-2 (P =.6). CONCLUSION: Spontaneous rupture of membranes (either term or preterm), parturition (either term or preterm), and microbial invasion of the amniotic cavity were associated with significant increases in the amniotic fluid concentration of the active forms of MMP-9. In contrast, the concentration of the active forms of MMP-2 either decreased or remained the same in these conditions. Our observations provide evidence for a novel regulation of gelatinolytic activity in vivo.

Amnion↗
PubMed2005

Functional expression of a P450 flavonoid hydroxylase for the biosynthesis of plant-specific hydroxylated flavonols in Escherichia coli.

Flavonols are plant polyphenolic compounds that belong to the class of molecules collectively known as flavonoids. Because of their demonstrated health benefits towards a wide array of human pathological conditions, a great interest has emerged for their biosynthesis from well-characterized microbial hosts. We present the functional expression in Escherichia coli of a plant P450 flavonoid 3', 5'-hydroxylase (F3'5'H) as a fusion protein with a P450 reductase. This expression allowed metabolic engineering of E. coli to produce the flavonol kaempferol and the 3', 4' B-ring hydroxylated flavonol quercetin from the p-coumaric acid precursor by simultaneously co-expressing the fusion protein with 4-coumaroyl:CoA-ligase (4CL), chalcone synthase (CHS), chalcone isomerase (CHI), flavanone 3beta-hydroxylase (FHT) and flavonol synthase (FLS). Biosynthesis of the B-ring tri-hydroxylated flavonol myricetin from the engineered strains was accomplished when flavanones rather than phenylpropanoid acids were used as precursor molecules. Cultivation of the recombinant strains in rich medium increased the synthesis of all flavonoids with the exception of myricetin. The present work opens the possibility of the future production of several other hydroxylated flavonoid molecules in E. coli.

Cytochrome P-450 Enzyme System↗
PubMed1999

Molecular characterization and analysis of the biosynthetic gene cluster for the antitumor antibiotic mitomycin C from Streptomyces lavendulae NRRL 2564.

BACKGROUND: The mitomycins are natural products that contain a variety of functional groups, including aminobenzoquinone- and aziridine-ring systems. Mitomycin C (MC) was the first recognized bioreductive alkylating agent, and has been widely used clinically for antitumor therapy. Precursor-feeding studies showed that MC is derived from 3-amino-5-hydroxybenzoic acid (AHBA), D-glucosamine, L-methionine and carbamoyl phosphate. A genetically linked AHBA biosynthetic gene and MC resistance genes were identified previously in the MC producer Streptomyces lavendulae NRRL 2564. We set out to identify other genes involved in MC biosynthesis. RESULTS: A cluster of 47 genes spanning 55 kilobases of S. lavendulae DNA governs MC biosynthesis. Fourteen of 22 disruption mutants did not express or overexpressed MC. Seven gene products probably assemble the AHBA intermediate through a variant of the shikimate pathway. The gene encoding the first presumed enzyme in AHBA biosynthesis is not, however, linked within the MC cluster. Candidate genes for mitosane nucleus formation and functionalization were identified. A putative MC translocase was identified that comprises a novel drug-binding and export system, which confers cellular self-protection on S. lavendulae. Two regulatory genes were also identified. CONCLUSIONS: The overall architecture of the MC biosynthetic gene cluster in S. lavendulae has been determined. Targeted manipulation of a putative MC pathway regulator led to a substantial increase in drug production. The cloned genes should help elucidate the molecular basis for creation of the mitosane ring system, as well efforts to engineer the biosynthesis of novel natural products.

Amino Acid Sequence↗
PubMed1999

An approach to management of critical indoor air problems in school buildings.

This study was conducted in a school center that had been the focus of intense public concern over 2 years because of suspected mold and health problems. Because several attempts to find solutions to the problem within the community were not satisfactory, outside specialists were needed for support in solving the problem. The study group consisted of experts in civil engineering, indoor mycology, and epidemiology. The studies were conducted in close cooperation with the city administration. Structures at risk were opened, moisture and temperature were measured, and the causes of damage were analyzed. Microbial samples were taken from the air, surfaces, and materials. Health questionnaires were sent to the schoolchildren and personnel. Information on the measurements and their results was released regularly to school employees, students and their parents, and to the media. Repairs were designed on the basis of this information. Moisture damage was caused mainly by difficult moisture conditions at the building site, poor ventilation, and water leaks. Fungal genera (concentrations <200 colony-forming units (cfu)/m(3), <3000 cfu/cm(2)) typical to buildings with mold problems (e.g., Aspergillus versicolor, Eurotium) were collected from the indoor air and surfaces of the school buildings. Where moisture-prone structures were identified and visible signs of damage or elevated moisture content were recorded, the numbers of microbes also were high; thus microbial results from material samples supported the conclusions made in the structural studies. Several irritative and recurrent symptoms were common among the upper secondary and high school students. The prevalence of asthma was high (13%) among the upper secondary school students. During the last 4 years, the incidence of asthma was 3-fold that of the previous 4-year period.

Adolescent↗
PubMed1996

Developments in biotechnological research in Austria.

Austria is a small European country with a small number of universities and biotechnological industries, but with great efforts in the implementation of environmental consciousness and corresponding legal standards. This review attempts to describe the biotechnological landscape of Austria, thereby focusing on the highlights in research by industry, universities, and research laboratories, as published during 1990 to early 1995. These will include microbial metabolite (organic acids, antibiotics) and biopolymer (polyhydroxibutyrate, S-layers) production; enzyme (cellulases, hemicellulases, ligninases) technology and biocatalysis; environmental biotechnology; plant breeding and plant protection; mammalian cell products; fermenter design; and bioprocess engineering.

Academies and Institutes↗
PubMed1999

Antibodies to a microbial peptide sharing sequence homology with betaA3-crystallin damage lens epithelial cells in vitro and in vivo.

Circulating auto-antibodies (Abs) against lens antigens (Ags) are highly prevalent in patients with cataract, but their origin and pathogenic significance are unknown. We hypothesized that Abs raised after exposure to infectious microbes could cross-react with lens Ags. To test this hypothesis, we generated a monoclonal Ab to human betaA3-crystallin. Epitope analysis indicated that the ETQAE sequence in the N-terminus region of betaA3-crystallin was critical for mounting a humoral response. Similar sequences were found in three microbial Ags. Mice injected with a microbial oligopeptide containing ETQAE emulsified with complete Freund's adjuvant (CFA) raised Abs which cross-reacted with betaA3-crystallin and developed lens epithelial cell (LEC) damage in vitro. We also genetically engineered an betaA3-crystallin-expressing E. coli. Mice immunized with the recombinant E. coli developed LEC damage. These results support the hypothesis that exposure to microbes having Ags homologous to self Ags can trigger a humoral immune response that leads to LEC damage in mice.

Amino Acid Sequence↗
PubMed2026

A voyage of reprogrammable metabolic bioengineering reshapes plant defense: from editing tools to synthetic systems.

Metabolic bioengineering has emerged as a transformative approach for reshaping plant defense by targeting intrinsic biosynthetic pathways to enhance immunity in modern agriculture. Moving beyond proof-of-concept metabolomics to broad-spectrum programmable pathway engineering addresses gaps in plant rational design and optimizes resilience in response to diverse environmental cues. This review aims to comprehensively highlight the transition of innovative approaches to phenolics, alkaloids, flavonoids, terpenoids, and benzoxazinoids, inferring adaptive reprogramming that mediates the growth-defense balance and functions as molecular sentinels in plants. Furthermore, decoding the volatile metabolome reveals a dynamic signaling interface that influences defense responses and stress-induced plant-microbe interactions, with the shikimate, jasmonate, and salicylate pathways functioning as central hubs for microbial deterrence and priming immune memory. Recent developments in multi-scalar genome-editing strategies, including CRISPR-driven combinatorial edits, enzyme orthogonalization, fluxomics, and spatially resolved multi-omics, reconfigure central and specialized metabolic fluxes toward improved defense function and regulation. Additionally, emerging tools, such as WUSCHEL2 and BABY BOOM transcriptional modules, and artificial engineering strategies integrating deep learning model-driven predictions facilitate rapid development of synthetic genetic circuits and support a predictive engineering of plants. Moreover, Mass spectrometry imaging (MSI) in spatial metabolomics enables to obtain structures and locations of unidentified endogenous metabolites within cells and tissues. Overall, this review emphasizes a diverse array of primary and secondary metabolites, spanning molecular concepts to recent advances in plant immune mechanisms. It also illustrates new frontiers in programmable metabolic engineering that accelerate the understanding of plant-microbe-metabolite cross-talks, offering strategies to improve plant resistance and advance sustainable agricultural solutions.

metabolic bioengineering↗
PubMed1988

Conjugal transfer of R68.45 and FP5 between Pseudomonas aeruginosa strains in a freshwater environment.

Recent concern over the release of genetically engineered organisms has resulted in a need for information about the potential for gene transfer in the environment. In this study, the conjugal transfer in Pseudomonas aeruginosa of the plasmids R68.45 and FP5 was demonstrated in the freshwater environment of Fort Loudoun Resevoir, Knoxville, Tenn. When genetically well defined plasmid donor and recipient strains were introduced into test chambers suspended in Fort Loudoun Lake, transfer of both plasmids was observed. Conjugation occurred in both the presence and absence of the natural microbial community. The number of transconjugants recovered was lower when the natural community was present. Transfer of the broad-host-range plasmid R68.45 to organisms other than the introduced recipient was not observed in these chambers but was observed in laboratory simulations when an organism isolated from lakewater was used as the recipient strain. Although the plasmids transferred in laboratory studies were genetically and physically stable, a significant number of transconjugants recovered from the field trials contained deletions and other genetic rearrangements, suggesting that factors which increase gene instability are operating in the environment. The potential for conjugal transfer of genetic material must be considered in evaluating the release of any genetically engineered microorganism into a freshwater environment.

Colony Count, Microbial↗
PubMed2001

Bacterial contamination associated with electronic faucets: a new risk for healthcare facilities.

OBJECTIVE: To investigate the safety of the hospital water supply following a major flood. DESIGN: Surveillance was conducted of the hospital water supply as it entered the hospital and at randomly selected water faucets throughout the facility. SETTING: A newly constructed surgical critical-care unit in a 265-bed community hospital that had to be evacuated and was out of operation for 6 weeks following a major flood of the city. METHODS: Random water samples throughout the facility were analyzed for heterotrophic plate counts (HPCs), chlorine, and coliforms utilizing standard methods. RESULTS: Water samples entering the hospital met appropriate standards, indicating the city water distribution system was not contaminated. Of 169 faucets tested, 13 (22%) of 59 electronic faucets exceeded the HPC threshold, and 12 (11%) of 110 manual faucets exceeded the HPC threshold (P<.14). A comparison of two brands of electronic faucets with manual faucets and with each other revealed that the HPC threshold was exceeded by 11 (32%) of 34 brand A faucets as compared to 12 (11%) of 110 manual faucets (P<.006). The HPC threshold was exceeded by 2 (8%) of 25 brand B faucets compared to 12 (11%) of 110 manual faucets (P<.94). Contamination rates of brand A and brand B faucets differed significantly (P<.003). Similar testing 2 months after hyperchlorination of the water supply indicated that the HPC threshold was exceeded by 16 (52%) of 31 brand A faucets compared to 10 (9.%) of 110 manual faucets (P<.0000003) and by 2 (18%) of 25 brand B faucets compared to 10 (9%) of 110 manual faucets (P=1.0). CONCLUSIONS: A certain brand of electronic water faucet used in the hospital was associated with unacceptable levels of microbial growth in water and was a continuing source of bacteria potentially hazardous to patients.

Chlorine↗