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Shiv Bharadwaj, Robert J Mitchell, Anjum Qureshi, Javed H Niazi
Electronic-cigarettes (e-cigarette) are widely used as an alternative to traditional cigarettes but their safety is not well established. Herein, we demonstrate and validate an analytical method to discriminate the deleterious effects of e-cigarette refills (e-juice) and soluble e-juice aerosol (SEA) by employing stress-specific bioluminescent recombinant bacterial cells (RBCs) as whole-cell biosensors. These RBCs carry luxCDABE-operon tightly controlled by promoters that specifically induced to DNA damage (recA), superoxide radicals (sodA), heavy metals (copA) and membrane damage (oprF)...
April 15, 2017: Biosensors & Bioelectronics
Leon H Oosterik, Huruma N Tuntufye, Jessica Tsonos, Tom Luyten, Sam Noppen, Sandra Liekens, Rob Lavigne, Patrick Butaye, Bruno M Goddeeris
Avian pathogenic Escherichia coli (APEC) are responsible for significant economic losses in the poultry industry. In this study, a model for investigating the pathogenesis of APEC infections was established. APEC strain CH2 (O78) was marked with the luciferase operon (luxCDABE) using a Tn7 transposon and tissues of experimentally infected chickens were analysed for a correlation between the bioluminescent signal and the number of bacteria. Transposition of the lux operon into the chromosome of the APEC isolate did not affect sensitivity to lytic bacteriophages and there was no effect on virulence in an intratracheal infection model in 1-day-old chicks, although results with a subcutaneous infection model were inconclusive...
October 2016: Veterinary Journal
Olga E Melkina, Ignatiy I Goryanin, Gennadii B Zavilgelsky
The antirestriction proteins ArdA ColIb-P9, Arn T4 and Ocr T7 specifically inhibit type I and type IV restriction enzymes and belong to the family of DNA-mimic proteins because their three-dimensional structure is similar to the double-helical B-form DNA. It is proposed that the DNA-mimic proteins are able to bind nucleoid protein H-NS and alleviate H-NS-silencing of the transcription of bacterial genes. Escherichia coli lux biosensors were constructed by inserting H-NS-dependent promoters into a vector, thereby placing each fragment upstream of the promoterless Photorhabdus luminescens luxCDABE operon...
November 2016: Microbiological Research
Krupa Naran, Atica Moosa, Clifton E Barry, Helena I M Boshoff, Valerie Mizrahi, Digby F Warner
The tuberculosis (TB) drug discovery pipeline is fueled by compounds identified in whole-cell screens against the causative agent, Mycobacterium tuberculosis Phenotypic screening enables the selection of molecules that inhibit essential cellular functions in live, intact bacilli grown under a chosen in vitro condition. However, deducing the mechanism of action (MOA), which is important to avoid promiscuous targets, often requires significant biological resources in a lengthy process that risks decoupling medicinal chemistry and biology efforts...
November 2016: Antimicrobial Agents and Chemotherapy
S Bartkova, B Kokotovic, I Dalsgaard
Recent development of imaging tools has facilitated studies of pathogen infections in vivo in real time. This trend can be exemplified by advances in bioluminescence imaging (BLI), an approach that helps to visualize dissemination of pathogens within the same animal over several time points. Here, we employ bacterial BLI for examining routes of entry and spread of Aeromonas salmonicida susbp. salmonicida in rainbow trout. A virulent Danish A. salmonicida strain was tagged with pAKgfplux1, a dual-labelled plasmid vector containing the mutated gfpmut3a gene from Aequorea victoria and the luxCDABE genes from the bacterium Photorhabdus luminescens...
January 2017: Journal of Fish Diseases
Bo Jiang, Wei E Huang, Guanghe Li
Here, we constructed an Escherichia coli recA::luxCDABE bioreporter for genotoxicity assessments. The recA promoter was cloned from the marine bacterium Vibrio natriegens. This bioreporter showed a dose-response relationship following induction by mitomycin C, and other pollutants or environmental samples could be calculated as mitomycin C equivalents, which provided a way to quantitatively compare the genotoxicities of different environmental samples. This bioreporter was used to evaluate the genotoxicity under a wide range of external environmental conditions, like temperatures ranging from 15 °C to 42 °C, pH between 4...
June 15, 2016: Environmental Science. Processes & Impacts
Ryan R Chaparian, Stephen G Olney, Christine M Hustmyer, Dean A Rowe-Magnus, Julia C van Kessel
The cell-cell signaling process called quorum sensing allows bacteria to control behaviors in response to changes in population density. In Vibrio harveyi, the master quorum-sensing transcription factor LuxR is a member of the TetR family of transcription factors that both activates and represses genes to coordinate group behaviors, including bioluminescence. Here, we show that integration host factor (IHF) is a key coactivator of the luxCDABE bioluminescence genes that is required together with LuxR for precise timing and expression levels of bioluminescence during quorum sensing...
September 2016: Molecular Microbiology
Elin Verbrugghe, Alexander Van Parys, Roel Haesendonck, Bregje Leyman, Filip Boyen, Freddy Haesebrouck, Frank Pasmans
OBJECTIVES: Antibiotics are among the most frequently prescribed drugs in human and animal medicine. With antibiotic resistance being a serious threat to veterinary and public health, the prudent use of antibiotics receives much attention. Less well known is that incorrect use of antimicrobial agents may also lead to increased bacterial virulence with the potential of a more severe clinical course of infection. Therefore, the aim of this study was to investigate the effect of subtherapeutic doses of tetracyclines on htpG virulence gene expression in Salmonella Typhimurium and on the course of salmonellosis...
August 2016: Journal of Antimicrobial Chemotherapy
Nathalie Franche, Manon Vinay, Mireille Ansaldi
Water quality is a major safety consideration in environments that are impacted by human activity. The key challenge of the COMBITOX project is to develop a unique instrument that can accommodate several biodetector systems (see the accompanying COMBITOX papers) able to detect different pollutants such as bacteria, toxins, and heavy metals. The output signal chosen by our consortium is based on luminescence detection. Our group recently developed phage-based biosensors using gfp as a reporter gene to detect enteric bacteria in complex environments such as sea water, and the main challenge we faced was to adapt our biodetector to a luminescent signal that could fit the COMBITOX project requirements...
January 2017: Environmental Science and Pollution Research International
Alberto Hernández-Eligio, Ángel Andrade, Lizeth Soto, Enrique Morett, Katy Juárez
In Geobacter sulfurreducens, metal reduction and generation of bioelectricity require the participation of several elements, and among them, the type IV pili has an essential role. The pilus is composed of multiple PilA monomers. Expression of pilA gene depends mainly on the σ54 factor and the response regulator protein PilR. In this work, we characterized the role of the PilS-PilR two-component system in the regulation of the pilA gene expression. Experimental evidence indicates that PilS is autophosphorylated at the His-334 residue, which in turn is transferred to the conserved Asp-53 in PilR...
February 18, 2016: Environmental Science and Pollution Research International
Xiwen Wang, Zhiping Li, Bo Li, Hang Chi, Jiakuan Li, Hongchao Fan, Ruizhi Yao, Qianxue Li, Xiaolin Dong, Man Chen, Han Qu, Yuanyuan Wang, Weicun Gao, Yutian Wang, Yu Sun, Rui Sun, Jun Qian, Zhiping Xia
PURPOSE: The goal of this study was to develop a plasmid-based lux bio-reporter for use to obtain in vivo images of Brucella suis vaccine strain 2 (B.suis S2) infection with high resolution and good definition. PROCEDURES: The pBBR-lux (pBBR1MCS-2-lxCDABE) plasmid that carries the luxCDABE operon was introduced into B. suis S2 by electroporation yielding B. suis S2-lux. The spatial and temporal transit of B. suis S2 in mice and guinea pigs was monitored by bioluminescence imaging...
August 2016: Molecular Imaging and Biology: MIB: the Official Publication of the Academy of Molecular Imaging
Aaron C Liu, Nikhil A Thomas
Vibrio parahaemolyticus (Vp) is a marine halophilic bacterium that is commonly associated with oysters and shrimp. Human consumption of contaminated shellfish can result in Vp mediated gastroenteritis and severe diarrheal disease. Vp encodes two type 3 secretion systems (T3SS-1 and T3SS2) that have been functionally implicated in cytotoxicity and enterotoxicity respectively. In this study, we profiled protein secretion and temporal promoter activities associated with exsA and exsB gene expression. exsA is an AraC-like transcriptional activator that is critical for activating multiple operons that encode T3SS-1 genes, whereas exsB is thought to encode an outer membrane pilotin component for T3SS-1...
2015: Frontiers in Microbiology
Yong-Zhi Wang, Dan Li, Miao He
Mercury and its organic compounds have been of severe concern worldwide due to their damage to the ecosystem and human health. The development of effective and affordable technology to monitor and signal the presence of bioavailable mercury is an urgent need. The Mer gene is a mercury-responsive resistant gene, and a mercury-sensing recombinant luminescent bacterium using the Mer gene was constructed in this study. The mer operon from marine Pseudomonas putida strain SP1 was amplified and fused with prompterless luxCDABE in the pUCD615 plasmid within Escherichia coli cells, resulting in pTHE30-E...
September 1, 2015: Journal of Environmental Sciences (China)
Irina V Gryazeva, Оlga K Davydova, Dmitrii G Deryabin
The antioxidant activities of five alkylresorcinol (AR) homologs with alkyl chains of 1, 3, 5 6 and 12 carbon atoms were studied using molecular and cellular assays for superoxide anions (O2.-). The effect of ARs as superoxide anion scavengers was assessed using the photochemical reaction of spontaneous photo-reduced flavin re-oxidation. In this system, ARs reaction with O2.- produced dye derivatives, as C6- and C12-AR prevented the O2.--induced conversion of nitroblue tetrazolium into formazan in AR-containing mixtures...
March 2015: Cellular & Molecular Biology Letters
Sharon Yagur-Kroll, Eden Amiel, Rachel Rosen, Shimshon Belkin
The use of bacterial bioreporters for the detection of buried landmines and other explosive devices has been promoted for over 25 years, and several bacterial sensor strains capable of detecting traces of either 2,4,6-trinitrotoluene (2,4,6-TNT) or 2,4-dinitrotoluene (2,4-DNT) have since been reported. In all cases, however, detection sensitivity failed to reach the levels required to reliably sense the minute concentrations of 2,4-DNT vapors expected to exist in the soil above buried landmines. Here, we report on the application of a directed evolution process to enhance the performance of a previously described E...
September 2015: Applied Microbiology and Biotechnology
D G Deryabin, I V Gryazeva, O K Davydova, G I El'-Registan
Molecular and cellular luminescent biotests were used to reveal the effects of five alkylresorcinol homologues (C7-, C9-, C11-, C12-, and C18-AR) on the thermally induced denaturation and refolding ofbac- terial luciferases, as well as on the synthesis of heat shock proteins. The ARs activities were found to depend on their fine structure and concentration. Direct heat-protective effect of short-chain C7- and C9-AR on the chromatographically pure Photobactrium leiognathii luciferase/oxidoreductase was shown within broad range of concentration (10(-6)-10(-3) M)...
November 2014: Mikrobiologiia
Anthony Vocat, Ruben C Hartkoorn, Benoit Lechartier, Ming Zhang, Neeraj Dhar, Stewart T Cole, Claudia Sala
Targeting dormant Mycobacterium tuberculosis represents a challenge to antituberculosis drug discovery programs. We previously reported and validated the use of the streptomycin (STR)-dependent M. tuberculosis 18b strain as a tool for assessing drug potency against nonreplicating bacteria both in vitro and in vivo. In this study, we generated a luminescent 18b strain, named 18b-Lux, by transforming the bacteria with a vector expressing the luxCDABE operon from Photorhabdus luminescens. Luciferase expression was demonstrated under replicating conditions, and, more importantly, luminescence levels significantly above background were detected following STR removal...
July 2015: Antimicrobial Agents and Chemotherapy
Keila Martín-Betancor, Ismael Rodea-Palomares, M A Muñoz-Martín, Francisco Leganés, Francisca Fernández-Piñas
A self-luminescent bioreporter strain of the unicellular cyanobacterium Synechococcus sp. PCC 7942 was constructed by fusing the promoter region of the smt locus (encoding the transcriptional repressor SmtB and the metallothionein SmtA) to luxCDABE from Photorhabdus luminescens; the sensor smtB gene controlling the expression of smtA was cloned in the same vector. The bioreporter performance was tested with a range of heavy metals and was shown to respond linearly to divalent Zn, Cd, Cu, Co, Hg, and monovalent Ag...
2015: Frontiers in Microbiology
Bo Jiang, Yizhi Song, Dayi Zhang, Wei E Huang, Xu Zhang, Guanghe Li
Bacterial whole-cell bioreporters are practical and reliable analytical tools to assess the toxicity and bioavailability of environmental contaminants, yet evidence has shown that their performance could be affected by different carbon sources. This paper evaluated the influence of carbon sources on the recA gene (ACIAD1385) in a DNA damage-inducible recA::luxCDABE Acinetobacter bioreporter and optimized the induction conditions for its practical application in environmental monitoring. Different carbon sources, including LB, potassium acetate (MMA), sodium citrate (MMC), sodium pyruvate (MMP), and sodium succinate (MMS), significantly influenced (p < 0...
April 2015: Environmental Science. Processes & Impacts
Shu-Jing Sun, Yu-Chen Liu, Jiao Sun, Hu Zhu
Bacteria communicate with one another using chemical signal molecules. This phenomenon termed quorum sensing enables the bacteria to monitor the environment for other bacteria and to alter behavior on a population-wide scale in response to cell density. Serratia marcescens JG, a quorum sensing bacterium, can secrete a furanosyl borate diester autoinducer (AI-2) in the exponential phase of growth. In this study, to further investigate the regulation of AI-2 production in S. marcescens JG, the pfs and luxS promoter fusions to an operon luxCDABE reporter were constructed in a low-copy-number vector pBR322K, which allows an examination of transcription of the genes in the pathway for signal synthesis...
June 2015: Archives of Microbiology
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