Determination of organophosphorus aromatic nitro insecticides and p-nitrophenol by microbial-cell respiratory activity

We examined the possibility of measuring the organophosphorus aromatic nitro insecticides metaphos and sumithion as well as their hydrolysis product p-nitrophenol (PNP) by the specific respiratory activity (SRA) of Pseudomonas putida C-11, P. putida BA-11, and Acinetobacter calcoaceticum A-122. The plots of cellular SRA against the two insecticides and PNP were linear over the ranges of 0.5-2.5 microM for P. putida C-11 and BA-11 and 0.5-1.0 microM for A. calcoaceticum A-122. P. putida BA-11 showed the greatest respiratory-response selectivity in the determination of the test substrates. We made comparison studies of the SRA of cells immobilised by two methods: carrier-surface adsorption and inclusion in various gels. We discuss the feasibility of developing a microbial sensor system for the determination of metaphos, sumithion, and PNP in aqueous media.     

Electrophysical characteristics of Azospirillum brasilense Sp245 during interaction with antibodies to various cell surface epitopes

This work was undertaken to examine the electrooptical characteristics of cells of Azospirillum brasilense Sp245 during their interaction with antibodies developed to various cell surface epitopes. We used the dependences of the cell suspension optical density changes induced by electroorientation on the orienting field frequency (740, 1000, 1450, 2000, and 2800kHz). Cell interactions with homologous strain-specific antibodies to the A. brasilense Sp245 O antigen and with homologous antibodies to whole bacterial cells brought about considerable changes in the electrooptical properties of the bacterial suspension. When genus-specific antibodies to the flagellin of the Azospirillum sheathed flagellum and antibodies to the serologically distinct O antigen of A. brasilense Sp7 were included in the A. brasilense Sp245 suspension, the changes caused in the electrooptical signal were slight and had values close to those for the above changes. These findings agree well with the immunochemical characteristics of the Azospirillum O antigens and with the data on the topographical distribution of the Azospirillum major cell surface antigens. The obtained results can serve as a basis for the development of a rapid test for the intraspecies detection of microorganisms.     

The electrooptical parameters of suspensions of <Emphasis Type="Italic">Escherichia coli</Emphasis> XL-1 cells interacting with helper phage M13K07

The electrophysical properties of Escherichia coli XL-1 cells interacting with helper phage M13K07 were studied as a function of the phage-to-cell ratio and the contact time. The electro-optical signal of bacterial cells changed considerably as soon as 10 min after the onset of their incubation with phage particles, presumably due to phage adsorption on the cell surface. The maximum changes in the orientational spectra of cell suspensions were observed when the phage-to-cell ratio was 20. Selectivity studies showed that E. coli XL-1 cells interacting with the helper phage M13K07 in the presence of foreign microflora, such as E. coli K-12 or Azospirillum brasilense Sp7, can be identified by using their electrophysical properties. Changes in the orientational spectra of cell suspensions are interpreted with the stage of phage-bacterium interaction taken into account. The results obtained can probably be used to devise a new rapid method for identification of microorganisms and to study the particular stages of cell infection by bacteriophages.__________Translated from Mikrobiologiya, Vol. 74, No. 2, 2005, pp. 198–203.Original Russian Text Copyright © 2005 by Bunin, Ignatov, Guliy, Zaitseva, ONeil, Ivnitskii     

Electrooptical analysis of the Escherichia coli-phage interaction

This article describes electrooptical (EO) characterization of biospecific binding between the bacterium Escherichia coli XL-1 and the phage M13K07. The electrooptical analyzer (ELUS EO), which has been developed at the State Research Center for Applied Microbiology, Obolensk, Russia, was used as the basic instrument for EO measurements. The operating principle of the analyzer is based on the polarizability of microorganisms, which depends strongly on their composition, morphology, and phenotype. The principle of analysis of the interaction of E. coli with the phage M13K07 is based on registration of changes of optical parameters of bacterial suspensions. The phage-cell interaction includes the following stages: phage adsorption on the cell surface, entry of viral DNA into the bacterial cell, amplification of phage within infected host, and phage ejection from the cell. In this work, we used M13K07, a filamentous phage of the family Inoviridae. Preliminary study had shown that combination of the EO approach with a phage as a recognition element has an excellent potential for mediator-less detection of phage-bacteria complex formation. The interaction of E. coli with phage M13K07 induces a strong and specific EO signal as a result of substantial changes of the EO properties of the E. coli XL-1 suspension infected by the phage M13K07. The signal was specific in the presence of foreign microflora (E. coli K-12 and Azospirillum brasilense Sp7). Integration of the EO approach with a phage has the following advantages: (1) bacteria from biological samples need not be purified, (2) the infection of phage to bacteria is specific, (3) exogenous substrates and mediators are not required for detection, and (4) it is suitable for any phage-bacterium system when bacteria-specific phages are available.     

Electrooptical parameters of kanamycin-treated <Emphasis Type="Italic">E. coli</Emphasis> cell suspensions

The effect of kanamycin on the electrophysical parameters of cell suspensions of Escherichia coli K-12 and pMMB33 was investigated. Incubation of the sensitive K-12 strain with kanamycin resulted in significant changes in the orientation spectra (OS) of the cell suspensions; these changes were not revealed in the case of the resistant pMMB33 strain. In the case of the sensitive K-12 strain incubated with different kanamycin concentrations, changes in the OS of the cell suspensions occurred within the 10–1000 kHz frequency range of the orienting electrical field. The most pronounced change in the electrooptical signal was observed at 10 μg/ml of kanamycin. Control experiments were carried out by standard plating on nutrient media. Thus, the OS changes of suspensions in the presence of antibiotics may be used as a test for microbial resistance to such antibiotics.     

An acoustic method for the analysis of bacterial cells

The application of a biological electroacoustic sensor based on a lateral electric-field-excited piezoelectric resonator for the study of bacterial cells that interact with specific bacteriophages, mini-antibodies, and polyclonal antibodies was successfully demonstrated. The determined lower limit of microbialcell detection was approximately of 10³ to 10⁴ cells/mL for the duration of the assay of 10 min. The possibility of bacterial-cell detection via interaction with specific agents in the presence of extraneous microbiota was shown. The method allowed us to determine the spectrum of lytic activity of bacteriophages and the sensitivity of microbial cells to bacteriophages. The results of the study showed that application of a sensor piezoelectric lateral-field resonator is a promising technique for the detection and identification of microbial cells and determination of their phage resistance in microbiology, medicine, and veterinary medicine. Furthermore, the results of the experiments made it possible to understand the mechanisms of the processes that occur in a suspension of bacterial cells in the presence of various biological agents. The method also may provide useful information regarding biophysical mechanisms of interactions that occur in microbial populations.     

The effect of ampicillin on the electrophysical properties of Escherichia coli cells

The study of the effect of ampicillin on the electrophysical properties of Escherichia coli cells showed that this antibiotic influences the orientational spectra (OSs) of the ampicillin-susceptible E. coli strains K-12 and XL-1 within the frequency range 10–1000 kHz of the orienting electric field and does not affect the OSs of the ampicillin-resistant strains K-12(pUC-18) and XL-1(pHEN1). The change in the electrooptical signal of the ampicillin-susceptible cells was maximum at an ampicillin concentration of 50 µg/ml and did not depend on the exposure time. The conclusion is drawn that changes in the OSs of cells can be used to evaluate their resistance to ampicillin.Translated from Mikrobiologiya, Vol. 74, No. 1, 2005, pp. 126–131.Original Russian Text Copyright © 2005 by Guliy, Markina, Ignatov, Shchegolev, Zaitseva, Bunin, Ignatov.     

Biosensor Systems for Antibiotic Detection

Antibiotics are widely used in medicine, veterinary medicine, and the food industry. However, the active use of antibacterial drugs leads to environmental pollution. In this regard, there is a great need for monitoring and determining antibiotics in various environments such as drinking water, food, drinks, waste water from pharmaceutical factories, etc. A number of methods, including those based on biosensors, have been developed to determine antibiotics. Biosensor methods of analysis are widely used and are an integral part of environmental monitoring. Electrochemical, optical, acoustic, microbial biosensors, immuno- and aptasensors, as well as sensors based on molecularly imprinted polymers are in the most demand for the analysis of antibiotics. This article provides a brief overview of biosensor methods and approaches for the determination of antibiotics. The most promising biosensor systems for determining antibacterial drugs were analyzed.

     

The adjuvant effect of selenium nanoparticles,Triton X-114 detergent micelles,and lecithin liposomes for <Emphasis Type="Italic">Escherichia coli</Emphasis> antigens

We studied the adjuvant properties of micelles from nonionogenic detergents, liposome, and selenium nanoparticles containing extracellular and intracellular vaccine antigens of a weakly virulent α-hemolytic Escherichia coli B-5 strain used for the immunization of experimental animals. Triton X-100 was used as a nonionogenic detergent for micelle preparation. The liposomes were obtained on the basis of lecithin from a chicken egg and E. coli B-5 membrane lipids. Native lipoproteins of E. coli B-5 cells and peptides for the proteolytic hydrolysis of toxin-containing culture liquid were used as antigens for micelles and liposomes. The obtained data suggested that micelles, liposomes, and selenium nanoparticles can be used for immunization with cellular and extracellular E. coli antigens.