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At least 217 records · Page 12Linked to original sources
PubMed1997

Genetic engineering of baker's and wine yeasts using formaldehyde hyperresistance-mediating plasmids.

Yeast multi-copy vectors carrying the formaldehyde-resistance marker gene SFA have proved to be a valuable tool for research on industrially used strains of Saccharomyces cerevisiae. The genetics of these strains is often poorly understood, and for various reasons it is not possible to simply subject these strains to protocols of genetic engineering that have been established for laboratory strains of S. cerevisiae. We tested our vectors and protocols using 10 randomly picked baker's and wine yeasts all of which could be transformed by a simple protocol with vectors conferring hyperresistance to formaldehyde. The application of formaldehyde as a selecting agent also offers the advantage of its biodegradation to CO2 during fermentation, i.e., the selecting agent will be consumed and therefore its removal during down-stream processing is not necessary. Thus, this vector provides an expression system which is simple to apply and inexpensive to use.

Drug Resistance, Microbial↗
PubMed2007

The future prospects of microbial cellulose in biomedical applications.

Microbial cellulose has proven to be a remarkably versatile biomaterial and can be used in wide variety of applied scientific endeavors, such as paper products, electronics, acoustics, and biomedical devices. In fact, biomedical devices recently have gained a significant amount of attention because of an increased interest in tissue-engineered products for both wound care and the regeneration of damaged or diseased organs. Due to its unique nanostructure and properties, microbial cellulose is a natural candidate for numerous medical and tissue-engineered applications. For example, a microbial cellulose membrane has been successfully used as a wound-healing device for severely damaged skin and as a small-diameter blood vessel replacement. The nonwoven ribbons of microbial cellulose microfibrils closely resemble the structure of native extracellular matrices, suggesting that it could function as a scaffold for the production of many tissue-engineered constructs. In addition, microbial cellulose membranes, having a unique nanostructure, could have many other uses in wound healing and regenerative medicine, such as guided tissue regeneration (GTR), periodontal treatments, or as a replacement for dura mater (a membrane that surrounds brain tissue). In effect, microbial cellulose could function as a scaffold material for the regeneration of a wide variety of tissues, showing that it could eventually become an excellent platform technology for medicine. If microbial cellulose can be successfully mass produced, it will eventually become a vital biomaterial and will be used in the creation of a wide variety of medical devices and consumer products.

Acetobacter↗
PubMed1995

Survival and activity of Pseudomonas sp. strain B13(FR1) in a marine microcosm determined by quantitative PCR and an rRNA-targeting probe and its effect on the indigenous bacterioplankton.

Genetically engineered Pseudomonas sp. strain B13(FR1) was released into laboratory-scale marine ecosystem models (microcosms). Survival of the introduced population in the water column and the sediment was determined by plating on a selective medium and by quantitative competitive PCR. The activity of the released bacteria was determined by in situ hybridization of single cells with a specific rRNA-targeting oligonucleotide probe. Two microcosms were inoculated with 10(6) cells ml-1, while an uninoculated microcosm served as a control. The number of Pseudomonas sp. strain B13(FR1) cells decreased rapidly to ca. 10(2) cells ml-1 within 2 days after the release, which is indicative of grazing by protozoa. Three days after the introduction into seawater, cells were unculturable, but PCR continued to detect cells in low numbers. Immediately after the release, the ribosomal content of Pseudomonas sp. strain B13(FR1) corresponded to a generation time of 2 h. The growth rate decreased to less than 0.04 h-1 in 5 days and remained low, probably because of carbon limitation of the cells. Specific amendment of the microcosms with 10 mM 4-chlorobenzoate resulted in a rapid increase of the growth rate and an exponentially increasing number of cells detected by PCR, but not in resuscitation of the cells to a culturable state. The release of Pseudomonas sp. strain B13(FR1) into the microcosms seemed to affect only the indigenous bacterioplankton community transiently. Effects on the community were also apparent from the handling of water during filling of the microcosms and the amendment with 4-chlorobenzoate.

Animals↗
PubMedDate not supplied

Characterization of oxygen transfer in miniature and lab-scale bubble column bioreactors and comparison of microbial growth performance based on constant k(L)a.

This work describes the engineering characterization of miniature (2 mL) and laboratory-scale (100 mL) bubble column bioreactors useful for the cultivation of microbial cells. These bioreactors were constructed of glass and used a range of sintered glass gas diffusers with differently sized pores to disperse humidified air within the liquid biomedium. The effect of the pressure of this supplied air on the breakthrough point for gas diffusers with different pore sizes was examined and could be predicted using the Laplace-Young equation. The influence of the superficial gas velocity (u(g)) on the volumetric mass transfer coefficient (k(L)a) was determined, and values of up to 0.09 s(-1) were observed in this work. Two modeling approaches were considered in order to predict and provide comparison criteria. The first related the volumetric power consumption (P/V) to the k(L)a and a good correlation was obtained for differently sized reactors with a given pore size, but this correlation was not satisfactory for bubble columns with different gas diffusers. Values for P/V ranged from about 10 to 400 W.m(-3). Second, a model was developed predicting bubble size (d(b)), bubble rising velocity (u(b)), gas hold-up (phi), liquid side mass transfer coefficient (k(L)), and thus the k(L)a using established theory and empirical correlations. Good agreement was found with our experimental data at different scales and pore sizes. Values for d(b) varied from 0.1 to 0.6 mm, and k(L) values between 1.7 and 9.8 x 10(-4) m.s(-1) were determined. Several E. coli cultivations were performed in the miniature bubble column at low and high k(L)a values, and the results were compared to those from a conventional stirred tank operated under identical k(L)a values. Results from the two systems were similar in terms of biomass growth rate and carbon source utilization.

Bioreactors↗
PubMed2002

Rock phosphate solubilization and colonization of maize rhizosphere by wild and genetically modified strains of Penicillium rugulosum.

Maize root colonization and phosphate solubilizing activity of the fungus Penicillium rugulosum were assessed in a greenhouse trial using soil-plant microcosms. The bacterial gene hph conferring resistance to hygromicin B was introduced by electroporation in the wild-type strain IR-94MF1 of P. rugulosum and one transformant, w-T3, was selected. Maize plants were grown for 5 weeks in a P-poor soil and fertilized with a Florida apatite mineral, with Navay, an apatite rock deposit from Venezuela, or with simple superphosphate. Inoculation treatments included strain IR-94MF1, transformant w-T3 and two IR-94MF1 UV-induced mutants with enhanced (Mps++) or reduced (Mps-) in vitro mineral phosphate solubilizing activity. In the absence of P fertilization, inoculation with any P. rugulosum isolate significantly reduced the size of the total and P-solubilizing bacterial community present in maize rhizosphere. The bacterial community significantly increased in maize inoculated with IR-94MF1 and w-T3 when P was added as apatites Navay or Florida. All P. rugulosum strains were able to stimulate the growth of maize plants as indicated by 3.6 to 28.6% increases in dry matter yields. In the presence of rock phosphate, P uptake by maize plants inoculated with the two mutants Mps++ and Mps- was not always in agreement with their P-solubilizing phenotypes. Strain IR-94MF1 and transformant w-T3 increased P assimilation by the plants fertilized with Navay rock phosphate by 26 and 38%, respectively. In this treatment, w-T3 showed its highest significant maize rhizosphere colonization. With the simple superphosphate treatment, w-T3 increased P uptake in plants by 8% over the uninoculated control and also decreased significantly the community size of total bacteria, total fungi, and P-solubilizing fungi in the rhizosphere.

Biological Transport↗
PubMed2005

Surpassing nature: rational design of sterile-surface materials.

The rise of multidrug-resistant pathogens and recalcitrance of biofilm infections present a formidable challenge to combating infectious diseases. There are numerous disinfectants and antiseptics for treating materials in hospitals and community settings, and devices such as catheters impregnated with anti-infectives have been introduced into practice. However, there are many limitations of materials impregnated with a leaching antibacterial agent. Recently, non-leaching, permanent, sterile-surface materials have been developed in which one end of a long-chained hydrophobic polycation containing antimicrobial monomers is attached covalently to the surface of a material, for example, cotton or plastic. The polymeric chain allows the antimicrobial moieties to permeate into, and kill, the cells of the pathogen. These sterile-surface materials kill both air- and waterborne pathogens and are not susceptible to existing resistance mechanisms.

Biomedical Engineering↗
PubMed2006

Redirection of cytosolic or plastidic isoprenoid precursors elevates terpene production in plants.

Terpenes constitute a distinct class of natural products that attract insects, defend against phytopathogenic microbes and combat human diseases. However, like most natural products, they are usually made by plants and microbes in small amounts and as complex mixtures. Chemical synthesis is often costly and inefficient, and may not yield enantiomerically pure terpenes, whereas large-scale microbial production requires expensive feedstocks. We engineered high-level terpene production in tobacco plants by diverting carbon flow from cytosolic or plastidic isopentenyl diphosphate through overexpression in either compartment of an avian farnesyl diphosphate synthase and an appropriate terpene synthase. Isotopic labeling studies suggest little, if any, metabolite exchange between these two subcellular compartments. The strategy increased synthesis of the sesquiterpenes patchoulol and amorpha-4,11-diene more than 1,000-fold, as well as the monoterpene limonene 10-30 fold, and seems equally suited to generating higher levels of other terpenes for research, industrial production or therapeutic applications.

Carbon↗
PubMed2004

Acinetobacter sp. ADP1: an ideal model organism for genetic analysis and genome engineering.

Acinetobacter sp. strain ADP1 is a naturally transformable gram-negative bacterium with simple culture requirements, a prototrophic metabolism and a compact genome of 3.7 Mb which has recently been sequenced. Wild-type ADP1 can be genetically manipulated by the direct addition of linear DNA constructs to log-phase cultures. This makes it an ideal organism for the automation of complex strain construction. Here, we demonstrate the flexibility and versatility of ADP1 as a genetic model through the construction of a broad variety of mutants. These include marked and unmarked insertions and deletions, complementary replacements, chromosomal expression tags and complex combinations thereof. In the process of these constructions, we demonstrate that ADP1 can effectively express a wide variety of foreign genes including antibiotic resistance cassettes, essential metabolic genes, negatively selectable catabolic genes and even intact operons from highly divergent bacteria. All of the described mutations were achieved by the same process of splicing PCR, direct transformation of growing cultures and plating on selective media. The simplicity of these tools make genetic analysis and engineering with Acinetobacter ADP1 accessible to laboratories with minimal microbial genetics expertise and very little equipment. They are also compatible with complete automation of genetic analysis and engineering protocols.

Acinetobacter↗
PubMed1993

Variable stability of antibiotic-resistance markers in Bacillus cereus UW85 in the soybean rhizosphere in the field.

We compared the stability of antibiotic-resistance markers in strains derived from Bacillus cereus UW85 in culture media and in the soybean rhizosphere in a growth chamber and in the field. We studied two independent, spontaneous mutants resistant to neomycin, three independent, spontaneous mutants resistant to streptomycin, and strains carrying plasmid pBC16, which encodes tetracycline resistance. Antibiotic-resistance markers were maintained in populations of all UW85 derivatives in culture and in the rhizosphere of soybeans grown in soil in a growth chamber. In two field experiments, antibiotic resistance was substantially lost in rhizosphere populations of B. cereus as early as 14 or as late as 116 days after planting. To distinguish between death of the inoculated strain and loss of its marker, we tested populations of B. cereus for other phenotypes (orange pigmentation, plasmid-borne resistance to tetracycline, and biocontrol activity) that are typical of UW85-derivatives used as inoculum, but atypical of the indigenous populations of B. cereus, and these phenotypes were maintained in populations from which the marker was lost. In general, neomycin-resistance markers were maintained at a higher frequency than streptomycin-resistance markers, and maintenance of antibiotic-resistance markers varied with position on the root and with the year of the experiment. In a semi-defined medium, the UW85 derivatives grew at the same rate as the wild type at 28 degrees C, but most grew more slowly than the wild type at 16 degrees C, demonstrating that antibiotic resistance can affect fitness under some conditions. The results suggest that the stability of antibiotic-resistance markers should be assessed in the ecosystems in which they will be studied.

Bacillus cereus↗
PubMed2006

System for expression of microsporidian methionine amino peptidase type 2 (MetAP2) in the yeast Saccharomyces cerevisiae.

Microsporidia are parasitic protists of all classes of vertebrates and most invertebrates. They recently emerged as important infections in various immunosuppressed and immunocompetent patient populations. They are also important veterinary and agricultural pathogens. Current therapies for microsporidiosis include benzimidazoles, which bind tubulin-inhibiting microtubule assembly, and fumagillin and its derivatives, which bind and inhibit methionine amino peptidase type 2 (MetAP2). Benzimidazoles are not active against Enterocytozoon bieneusi, the most common cause of human microsporidiosis. Fumagillin is active against most microsporidia, including E. bieneusi, but thrombocytopenia has been a problem in clinical trials. There is a pressing need for more-specific microsporidian MetAP2 inhibitors. To expedite and facilitate the discovery of safe and effective MetAP2 inhibitors, we have engineered Saccharomyces cerevisiae to be dependent on Encephalitozoon cuniculi MetAP2 (EcMetAP2) for its growth, where EcMetAP2 is harbored on an episomal uracil-selectable tetracycline-regulated plasmid. We have also constructed a leucine-selectable tetracycline-regulated expression plasmid into which any MetAP2 gene can be cloned. By utilizing a 5-fluoroorotic acid-mediated plasmid shuffle in the EcMetAP2 yeast strain, a yeast strain can be generated whose growth is dependent on MetAP2 from any organism. The level of heterologous MetAP2 gene expression can be controlled by the addition of tetracycline to the growth medium. These yeast strains should permit high-throughput screening for the identification of new inhibitors with high specificity and activity toward microsporidian MetAP2.

Aminopeptidases↗
PubMed1990

Effect of aerosolization on subsequent bacterial survival.

To determine whether aerosolization could impair bacterial survival, Pseudomonas syringae and Erwinia herbicola were aerosolized in a greenhouse, the aerosol was sampled at various distances from the site of release by using all-glass impingers, and bacterial survival was followed in the impingers for 6 h. Bacterial survival subsequent to aerosolization of P. syringae and E. herbicola was not impaired 1 m from the site of release. P. syringae aerosolized at 3 to 15 m from the site of release at a temperature of 12 degrees C and a relative humidity of 80% survived 35- to 65-fold better than P. syringae released at 27 degrees C and a relative humidity of 40%. No difference was observed in the survival of P. syringae and E. herbicola following aerosolization at the same temperature and relative humidity. Bacteria sprayed directly onto bean and oat plants established stable populations at comparable numbers on both plants over an 8-day period following inoculation. Bacteria that inoculated adjacent plants by drifting downwind up to 5 m were detectable at an initial population of 10(2) CFU/g on oats and 10(5) CFU/g on beans 2 h after the spray. However, bacterial populations on both plants were undetectable within 48 h.

Aerosols↗
PubMed1992

Segregation of altered parental properties in fusions between Saccharomyces cerevisiae and the D-xylose fermenting yeasts Candida shehatae and Pichia stipitis.

A prototrophic strain of Saccharomyces cerevisiae CSIR Y190 MATa xyl-, resistant to high levels of ethanol, was hybridized with xylose-fermenting, auxotrophic mutants of Candida shehatae and Pichia stipitis through polyethylene glycol-induced protoplast fusion in an attempt to produce ethanol-tolerant, xylose-fermenting hybrids. Mononucleate fusants were obtained, but these dissociated into a mixture of parental-type segregants. Purified Candida- and Pichia-resembling segregants failed to acquire improved ethanol tolerance but expressed other novel properties of S. cerevisiae, suggesting that karyogamy was impaired after internuclear gene transfer.

Candida↗
PubMed2000

Biology in politics.

Increases in biological knowledge lead to increasing potential for practical applications of this knowledge. Politicians need to stand at the boundary between science and society, understanding the biological issues in order to make informed decisions on behalf of the public.

Biological Warfare↗
PubMed2002

RMS Titanic and the emergence of new concepts on consortial nature of microbial events.

The RMS Titanic sank in 1912 and created a historical event that still ripples through time. Stories were told and lessons learned but the science has only just begun. Today the fading remains of the ship resemble the hanging gardens of Babylon except that it is not plants that drape the walls but complex microbial growths called rusticles. These organisms have been found to be not a species, like plants and animals, but to be structures created by complex communities of bacterial species. Like the discovery of tube worms in the mid-oceanic vents, the nature of these rusticles presents another biological discovery of a fundamental nature. Essentially these microbial consortia on the RMS Titanic have generated structures of a mass that would rival whales and elephants while gradually extracting the iron from the steel. Rusticle-like consortia appear to play many roles within the environment, and it is perhaps the RMS Titanic that is showing that there is a new way to understand the form, function, and nature of microorganisms. This understanding would develop by considering the bacteria not as individual species functioning independently but as consortia of species functioning in community structures within a common habitat. This concept, if adopted, would change dramatically the manner in which a microbial ecologist and any scientist or engineer would view the occurrence of a slime, encrustation, biocolloid, rust flake, iron pan, salt deposit, and perhaps even some of the diseases that remain unexplained as a disease of unknown cause.

Animals↗