bactofection

Successful gene delivery into mammalian cells using bactofection requires entry of the bacterial vector via cell surface integrin receptors followed by release of plasmid DNA into the cellular environment. We show, for the first time, that addition of the DNA transfection reagent Lipofectamine improves entry of invasive Escherichia coli into HeLa cells and enhances up to 2.8-fold green fluorescent protein (GFP) expression from a reporter plasmid. The addition of Lipofectamine may be applicable to other bacterial vectors to increase their DNA delivery efficiency into mammalian cells...

The use of attenuated Salmonella typhimurium as a bactofection vehicle for the oral delivery of a DNA vaccine against rabbit haemorrhagic disease virus (RHDV) was investigated. The DNA vaccine plasmid pcDNA3.1-VP60, which encodes the viral capsid protein VP60, was transformed into the attenuated S. typhimurium strain SL7207. The resulting recombinant bacteria, named as SL/pcDNA3.1-VP60, were orally used to immunise rabbits. The successful delivery of the DNA plasmid was confirmed by the detected VP60 transcription in the rabbit intestines through the reverse transcription polymerase chain reaction...

Bacterial production of visible light is a natural phenomenon occurring in marine (Vibrio and Photobacterium) and terrestrial (Photorhabdus) species. The mechanism underpinning light production in these organisms is similar and involves the oxidation of an aldehyde substrate in a reaction catalysed by the bacterial luciferase enzyme. The genes encoding the luciferase and a fatty acid reductase complex which synthesizes the substrate are contained in a single operon (the lux operon). This provides a useful reporter system as cloning the operon into a recipient host bacterium will generate visible light without the requirement to add exogenous substrate...

A number of genetically modified bacteria able to deliver a therapeutic gene into target cells has already been tested. Apart from the expected effects of bacterial therapy, the therapeutic bacterial strain also mediates a non-specific effect independent of the gene to be delivered. In this regard, we have recently shown that oral administration of the bacterial strain Escherichia coli XL1-Blue via gastric gavage to rats leads to a non-specific decrease in expression of vascular endothelial growth factor (VEGF) in intestinal wall without corresponding changes in other parameters...

The role of innate immunity in the prevention of urinary tract infection is well-documented. Toll-like receptor 4 (TLR4) is a major determinant of innate immune response. In an animal model of urinary tract infection, bactofection-mediated gene transfer of TLR4 was tested in a preventive approach. Bactofection with TLR4 reduced the colonization with uropathogenic Escherichia coli by 91% in the kidney and by 41% in the bladder. Reduced colonization was associated with lower oxidative stress and expression of monocyte chemoattractant protein-1 and myeloperoxidase in the kidney...

Bacterial gene therapy - bactofection is a simple and effective method to deliver plasmid DNA into target tissue. We hypothesize that oral in vivo bactofection can be an interesting approach to influence the course of inflammatory bowel diseases. The aim of this study was to prove the effects of antioxidative and anti-inflammatory bactofection in dextran sulfate sodium (DSS)-treated mice. Attenuated bacteria Salmonella Typhimurium SL7207 carrying plasmids with genes encoding Cu-Zn superoxide dismutase and an N-terminal deletion mutant of monocyte chemoattractant protein-1 were prepared...

Several bacterial species have inherent ability to colonize solid tumors in vivo. However, their natural anti-tumor activity can be enhanced by genetic engineering that enables these bacteria express or transfer therapeutic molecules into target cells. In this review, we summarize latest research on cancer therapy using genetically modified bacteria with particular emphasis on blocking tumor angiogenesis. Despite recent progress, only a few recent studies on bacterial tumor therapy have focused on anti-angiogenesis...

Salmonella typhimurium SL7207 carrying Cu-Zn superoxide dismutase and an N-terminal deletion mutant of monocyte chemoattractant protein-1 genes was applied to dextran sodium sulfate treated female Wistar rats. Stool quality, food and water intake were monitored. Markers of oxidative stress, interleukin 1, interleukin 6 and tumor necrosis factor alpha were quantified. No differences were found in body weights, markers of oxidative stress in plasma and inflammatory markers in colon homogenates. Plasma concentrations of I11, I16 were lower in the treatment groups than in the dextran sodium sulfate group...

Live bacterial vectors may be useful tools for the development of novel cancer therapies that can be added to the repertoire of existing drugs. Several bacterial strains effectively colonize solid tumors and act as antitumor therapeutics. The naturally occurring tumor-colonizing characteristics of bacterial species such as Salmonella sp, Clostridium sp and Escherichia coli can be further modified by genetic manipulations, making these bacterial systems excellent vehicles for the production and targeted delivery of therapeutic molecules into cancer cells...

BACKGROUND AND AIMS: Bactofection is delivery of therapeutic genes into target cells using bacteria penetrating the target cell membrane and releasing the gene into the cell. Hypoxia-inducible factor 1α (HIF-1α) represents a potential therapeutic gene to be used for gene delivery in ischemic diseases. The aim of this study was to prove the effects of bacteria-mediated transfer of hypoxia-inducible factor 1α (HIF-1α) in an experimental model of intestinal ischemia in rats. METHODS: Male Wistar rats with a surgically induced ischemia of colon (cecum) or sham-operated rats were treated by per os application of E...

The intracellular pathogen Listeria monocytogenes represents a promising therapeutic vector for the delivery of DNA, RNA or protein to cancer cells or to prime immune responses against tumour-specific antigens. A number of biological properties make L. monocytogenes a promising platform for development as a vector for either gene therapy or as an anti-cancer vaccine vector. L. monocytogenes is particularly efficient in mediating internalization into host cells. Once inside cells, the bacterium produces specific virulence factors which lyse the vaculolar membrane and allow escape into the cytoplasm...

Bacteria-mediated transfer of plasmid DNA to mammalian cells (bactofection) has been shown to have significant potential as an approach to express heterologous proteins in various cell types. This is achieved through entry of the entire bacterium into cells, followed by release of plasmid DNA. In a murine model, we show that Listeria monocytogenes can invade and spread in tumors, and establish the use of Listeria to deliver genes to tumors in vivo. A novel approach to vector lysis and release of plasmid DNA through antibiotic administration was developed...

We investigated the use of two previously described attenuated strains of Salmonella enterica subspecies enterica serovar Choleraesuis (S. Choleraesuis), DeltaphoP and DeltarpoS, compared with the commercial attenuated SC-54 strain, as bactofection vehicles, to deliver an epitope model (3xFLAG) to the intestinal immune system. The gene encoding the epitope 3xFLAG was subcloned into the pCMVbetam2A mammalian expression vector (creating pCMV3xFLAGm2A) and introduced into S. Choleraesuis strains. The 3xFLAG epitope was expressed efficiently in murine macrophage J774A...

Protective immune responses in shrimp induced by DNA vaccines against white spot syndrome virus (WSSV) with intramuscular injection have been reported in recent reports. In this study, we investigated the utilities of attenuated Salmonella enterica serovar Typhimurium (Salmonella typhimurium) as a bactofection vehicle for the oral delivery of a DNA vaccine plasmid to crayfish (Cambarus clarkii). The DNA vaccine plasmid pcDNA3.1-VP28, encoding viral envelope protein VP28, was transformed to an attenuated S. typhimurium strain SV4089 and the resulting recombinant bacteria named SV/pcDNA3...

BACKGROUND: Listeria monocytogenes is a highly versatile bacterial carrier system for introducing protein, DNA and RNA into mammalian cells. The delivery of tumor antigens with the help of this carrier into tumor-bearing animals has been successfully carried out previously and it was recently reported that L. monocytogenes is able to colonize and replicate within solid tumors after local or even systemic injection. METHODS: Here we report on the delivery of two prodrug converting enzymes, purine-deoxynucleoside phosphorylase (PNP) and a fusion protein consisting of yeast cytosine deaminase and uracil phosphoribosyl transferase (FCU1) into cancer cells in culture by L...

Bacteria-mediated gene transfer ('bactofection') has emerged as an alternative approach for genetic vaccination and gene therapy. Here, we assessed bactofection of airway epithelial cells in vitro and in vivo using an attenuated Escherichia coli genetically engineered to invade non-phagocytic cells. Invasive E. coli expressing green fluorescent protein (GFP) under the control of a prokaryotic promoter was efficiently taken up into the cytoplasm of cystic fibrosis tracheal epithelial (CFTE29o-) cells and led to dose-related reporter gene expression...

In this study, we investigated the use of Salmonella typhimurium (STM1 strain) as a bactofection vehicle to deliver a transmission-blocking DNA vaccine (TBDV) plasmid to the intestinal immune system. The gene encoding the full length cyst wall protein-2 (CWP2) from Giardia lamblia was subcloned into the pCDNA3 mammalian expression vector and stably introduced into S. typhimurium STM1. Eight-week-old female BALB/c mice were orally immunized every 2 weeks, for a total of three immunizations. Vaccinated and control mice were sacrificed 1 week following the last injection...

The development of an effective therapeutic vaccine to induce cancer-specific immunity remains an unsolved yet pressing priority requiring novel vaccine strategies. Here we have generated a series of vaccines in which bacteria deliver a plasmid encoding a tumor antigen under the control of a mammalian promoter in an attempt to induce an antitumor immune response. Utilizing a plasmid release mechanism involving the suicide of the carrier bacteria, we were able to engineer Listeria monocytogenes to induce antitumor immunity to a physiologically relevant tumor antigen, the cervical cancer oncoprotein E7...

Recent advances in gene therapy can be attributed to improvements of gene delivery vectors. New viral and nonviral transport vehicles that considerably increase the efficiency of transfection have been prepared. However, these vectors still have many disadvantages that are difficult to overcome, thus, a new approach is needed. The approach of bacterial delivery could in the future be important for gene therapy applications. In this article we try to summarize the most important modifications that are used for the preparation of applied strains, difficulties that are related with bacterial gene delivery and the current use of bactofection in animal experiments and clinical trials...

Bacteria-mediated transfer of plasmid DNA into mammalian cells (bactofection) is a potent approach to express plasmid-encoded heterologous proteins (protein antigens, toxins or enzymes) in a large set of different cell types including phagocytic and nonphagocytic mammalian cells. Previously, we have described a Listeria monocytogenes-mediated DNA delivery system, which releases plasmid DNA directly into the cytosol of mammalian cells by partial self-destruction of the carrier bacteria. Here we report on a second generation of this phage lysin supported bactofection system, which is greatly improved with respect to plasmid stability, transfer efficacy and biosafety...