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Recombinant Escherichia coli displaying organophosphorus hydrolase (OPH) was used to overcome the diffusion barrier limitation of organophosphorus pesticides. A new anchor system derived from the N-terminal domain of ice-nucleation protein from Pseudomonas syringae InaV (InaV-N) was used to display OPH onto the surface. The designed sequence was cloned in the vector pET-28a(+) and then was expressed in E. coli. Tracing of the expression location of the recombinant protein using SDS-PAGE showed the presentation of OPH by InaV-N on the outer membrane, and the ability of recombinant E. coli to utilize diazinon as the sole source of energy, without growth inhibition, indicated its significant activity. The location of OPH was detected by comparing the activity of the outer membrane fraction with the inner membrane and cytoplasm fractions. Studies revealed that recombinant E. coli can degrade 50% of 2 mM chlorpyrifos in 2 min. It can be concluded that InaV-N can be used efficiently to display foreign functional protein, and these results highlight the high potential of an engineered bacterium to be used in bioremediation of pesticide-contaminated sources in the environment.

Pasteurella haemolytica isolates from cattle and sheep, including representatives of different serotypes and untypable strains, were examined for leukotoxin (Lkt) production at the end of the log phase of growth in brain heart infusion broth. There were marked differences in leukotoxic activity in culture supernate samples, as measured by chemiluminescence-inhibition assays with bovine and ovine neutrophils, even between strains of the same serotype. There was also some variation in the amount and mol.wt of the Lkt protein produced by different strains, as judged by SDS-PAGE, immunoblotting and ELISA. Some strains produced normal amounts of Lkt protein which had only low leukotoxic activity. Most strains produced Lkt of 105 kDa whereas four strains produced a higher mol. wt form of c. 108 kDa, including two of the five serotype A2 strains examined. Thus, the R haemolytica isolates showed considerable heterogeneity in terms of leukotoxin production, mol. wt and activity, even within a given serotype.

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Background: Final elimination of some intracellular bacterial agents, such as Brucella, is often a complex issue and impossible to achieve, primarily due to the presence and survival of the bacteria within phagocytic cells. By penetrating into the cell membrane, drug delivery nanosystems can reduce the number of intracellular bacteria. The aim of this study was to assess the efficacy of chitosan nanoparticles on the delivery of gentamicin into Brucella infected J774A.1 murine cells in vitro. Materials and Methods: Chitosan nanoparticles (NPs) were synthesized using ionic gelation technique. The shape, size and charge of NPs, loading rate and release of the drug were investigated. Finally, the effects of gentamicin-loaded chitosan NPs (Gen-Cs) and free gentamicin on J774A.1 murine cells infected with these bacteria were examined. Results: The mean size and charge of NPs were computed as 100 nm and +28mV, respectively. The loading capacity of NPs was 22%. About 70% of the drug was released from NPs during the first 8 hours. Antimicrobial activity of the two formulations showed that MIC (minimum inhibitory concentration) of the Gen-Cs and free drug was 3.1 and 6.25 µg, respectively. The minimum bactericidal concentration of the NPs-loaded drug and free drug was 6.25 and 12.5 µg, respectively. Cell culture analysis revealed that there was a significant reduction in the load of the intercellular bacteria in J774A.1 murine cells in both formulations. Conclusion: Our results showed the Gen-Cs have a proper potential for optimal treatment of intracellular bacterial agents. [GMJ.2019;8:e1296]

1IntroductionFoodborne infections are an important public health concern worldwide. Escherichia coli O157 :H7i saparto ft he enterohemorrhagic group of E. coli (EHEC). Clinical symptoms of patients infected with E. coli O157 :H7h ave aw ide range and diagnosis of the symptoms is difficult. Thep athogen causes diarrhea and hemolytic uremic syndrome (HUS) in human beings.H emolytic uremic syndrome (HUS), al ife threatening complication that causes kidney failure,i sd evelopedb ya bout 10 %o fp atients, mostly in elderly people and children. So the exploitation of new methods for rapid and sensitived etection of such microorganism predicates significance to the safety in food or water supplies [1,2].C onventional detection methods for bacterialp athogens are the culture and colony counting,i mmunological techniquesa nd polymerase chain reaction( PCR). In addition to culture based methods, other techniques such as immunoassays, enzyme assays,a nd nucleic acid probe assays have been exploited for the identification of some foodborne pathogens.U nfortunately,d espite being powerful and errorproof approaches,m ost of them are laborious,c omplex, and time consuming,a nd they are not specific enough towards theirt argets [3,4].E nrichment, immunomagnetic separation, plating on selective media, and the agglutination of O157 antigenf ollowedb yP CR detection of virulence genes for confirmation, are the main strategies which are generally used for the isolation and identification of E. coli O157 in food and feces [5].T he nanobiosensors offers everala dvantages in comparison to the mentioned methods;l imited hands-on time,i mproved detectability,h igh-throughput screening, the possibility of real-time analysis,l abel-freed etection,s impler nucleic acid assays, and finally,t he faster and cheaper process has triggered ag reat interest toward nanobiosensor techniques [6].Adiagnostic label-free,P CR-free nanobiosensor could be an extremely powerful and rapid tool for detecting microorganisms in environmental and food samples,a sw ell as for clinical diagnosis of microbial infectionsa nd genetic diseases.Due to its tremendous molecular recognition potential, DNAi se specially well suited to biosensing applications [7].D NA biosensors could detect the DNAs equences via the hybridization to the complementary single stranded sequences,a nd offer ap romising approach for rapid, sen-Abstract:Asimple and sensitiven anobiosensor was developedf or diagnosingl abel-free,P CR-free of rfbE gene in Escherichia coli O157 :H7r eal sample.T he Au nanoparticles modified electrode (Au nano ME) was prepared by electrodepositiont echnique.T he single-strandedD NA probeswere immobilized through the chemisorptions process on Au nano ME (ssDNA/Au nano ME). Electrochemical impedance spectroscopy (EIS) techniqueh as been used to investigate the hybridization and sensing properties. Using electrochemical impedance spectroscopy (EIS) technique,t he charge-transferr esistance (R ct )v alues were raised by immobilization of the DNAp robea nd after the hyb...

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Platinum oxide nanoparticles were prepared by a simple hydrothermal route and chemical reduction using carbohydrates (fructose and sucrose) as the reducing and stabilizing agents. In comparison with other metals, platinum oxide has less environmental pollution. Therefore, Pt is considered an appropriate candidate to deal with environmental pathogens. The crystallite size of these nanoparticles was evaluated from X-ray diffraction, atomic force microscopy, and transmission electron microscopy (TEM) and was found to be 10 nm, which is the demonstration of EM bright field and transmission electron microscopy. The effect of carbohydrates on the morphology of the nanoparticles was studied using TEM. The nanoparticles were administered to the Pseudomonas stutzeri and Lactobacillus cultures, and the incubation was done at 37°C for 24 h. The nanocomposites exhibited interesting inhibitory as well as bactericidal activity against P. stutzeri and Lactobacillus species. Incorporation of nanoparticles also increased the thermal stability of the carbohydrates. The results of this paper showed that carbohydrates can serve as a carrier for platinum oxide nanoparticles, and nanocomposites can have potential biological applications.