Selection and analytical applications of aptamers binding microbial pathogens

DNA aptamers specifically recognizing microbial cells and viruses have a range of analytical and therapeutic applications. This article describes recent advances in the development of aptamers targeting specific pathogens (e.g., live bacteria, whole viral particles, and virally-infected mammalian cells). Specific aptamers against pathogens have been used as affinity reagents to develop sandwich assays, to label and to image cells, to bind with cells for flow-cytometry analysis, and to act as probes for development of whole-cell biosensors. Future applications of aptamers to pathogens will benefit from recent advances in improved selection and new aptamers containing modified nucleotides, particularly slow off-rate modified aptamers (SOMAmers).     

50 Hz magnetic field effect on the morphology of bacteria

Atomic force microscopy was used to distinguish changes in morphology of bacteria induced by 50 Hz 10 mT magnetic field exposure. It is known that alternating magnetic field exposure causes decrease of viability of different bacterial strains. Previously we found that the viability of rod-like bacteria exposed to magnetic field decreased twice more in comparison with the spherical ones. Motivated by this fact we carried out this study with bacterial cells of both shapes. We used Escherichia coli (rod-like) and Paracoccus denitrificans (spherical) bacteria. As a result we have not observed any change in bacterial morphology neither of rod-like nor of spherical bacteria after 1 h, 50 Hz and 10 mT magnetic field exposure.     

Screening of biosorption bacteria tolerance towards copper and cadmium from oil sludge pond

Certain bacterial strains can be employed in the removal of heavy metals from the environment. The aim of this study was to screen potential bacteria that were tolerant towards Cu and Cd and instrumental neutron activation analysis (INAA) was used to determine the concentrations of heavy metals in the sludge samples. The sludge samples from oil refinery plant in Malaysia contained Cr, Cs, Cu, Eu, Fe, Hg, Mn, Sb, Sc, Th, U and Zn. Seven bacterial isolates were identified to be tolerant to 100 mg/kg of copper and cadmium.     

Ag/SBA-15复合材料的制备及其抗菌性质

本文采用二维六方结构的介孔硅SBA-15作为主体, 先将其浸渍在葡萄糖溶液中, 利用土伦试剂在原位发生氧化-还原反应, 成功地在介孔孔道中制备出分散的银纳米粒子, 并以金黄葡萄球菌为研究对象, 对Ag/SBA-15的抑菌性能及持续抑菌能力进行了检测, 结果表明, 样品对金黄葡萄球菌有明显的抑菌作用并具有持续的抑菌能力.     

Rapid differentiation among bacteria that cause gastroenteritis by use of low-resolution Raman spectroscopy and PLS discriminant analysis

Use of classical microbiological methods to differentiate bacteria that cause gastroenteritis is cumbersome but usually very efficient. The high cost of reagents and the time required for such identifications, approximately four days, could have serious consequences, however, mainly when the patients are children, the elderly, or adults with low resistance. The search for new methods enabling rapid and reagentless differentiation of these microorganisms is, therefore, extremely relevant. In this work the main microorganisms responsible for gastroenteritis, Escherichia coli, Salmonella choleraesuis, and Shigella flexneri, were studied. For each microorganism sixty different dispersions were prepared in physiological solution. The Raman spectra of these dispersions were recorded using a diode laser operating in the near infrared region. Partial least-squares (PLS) discriminant analysis was used to differentiate among the bacteria by use of their respective Raman spectra. This approach enabled correct classification of 100% of the bacteria evaluated and unknown samples from the clinical environment, in less time (∼10 h), by use of a low-cost, portable Raman spectrometer, which can be easily used in intensive care units and clinical environments.     

Optical imaging: bacteria,viruses, and mammalian cells encoding light-emitting proteins reveal the locations of primary tumors and metastases in animals

Early detection of tumors and their metastases is crucial for the prognosis of cancer treatment. Traditionally, tumor detection is achieved by various methods, including magnetic resonance imaging and computerized tomography. With the recent cloning, cellular expression, and real-time imaging of light-emitting proteins, such as Renilla luciferase (Ruc), bacterial luciferase (Lux), firefly luciferase (Luc), green fluorescent protein (GFP), or Ruc-GFP fusion protein, significant efforts have been focused on using these marker proteins for tumor detection. It has also been demonstrated that certain bacteria, viruses, and mammalian cells (BVMC), when administered systemically, are able to gain entry and replicate selectively in tumors. In addition, many tissue/tumor specific promoters have been cloned which allow transgene expression specifically in tumor tissues. Therefore, when light-emitting protein encoded BVMC are injected systemically into rodents, tumor-specific marker gene expression is achieved and is detected in real time based on light emission. Consequently, the locations of primary tumors and previously unknown metastases in animals are revealed in vivo. In the future it will likely be feasible to use engineered light-emitting BVMC as probes for tumor detection and as gene-delivery vehicles in vivo for cancer therapy.     

Gram-negative bacteria viable in ultrapure water: identification of bacteria isolated from ultrapure water and effect of temperature on their behavior

This study describes the effect of temperature on the behavior of bacteria viable in ultrapure water and the contamination of ultrapure water by bacteria. Three species of bacteria were isolated from ultrapure water (total organic carbon, 60 ppb and 5 ppb; effluent resistivity > 18 MΩ cm at 25°C) and identified by morphological and physiological characteristics. The three isolates were incubated in water for injection and PYG broth to check the growth profile at various temperatures. In PYG broth, temperature influenced the behavior of bacteria directly; however, it did not in water for injection. By checking both viable and non-viable bacterial numbers and endotoxin concentration in pure water, the water was found to be contaminated with non-viable bacteria and newly generated endotoxins besides viable bacteria. A column treatment, a mixed bed of fully regenerated strong acid cation exchange resin (SACER) and strong base anion exchange resin (SBAER), was used to remove bacteria from pure water. Bacteria could not grow on the surfaces of ion exchange resins in the mixed bed. The removal of bacteria was more effective as pure water was circulated through the mixed bed more rapidly.     

Development of a LC-MS/MS method for the analysis of volatile primary and secondary amines as NIT (naphthylisothiocyanate) derivatives

High-performance liquid chromatography with mass spectrometric detection was used for the structure elucidation of eighteen primary and secondary amines and ammonia derivatised with naphthylisothiocyanate (NIT). A fragmentation scheme was established using reference compounds and the scheme was applied to real air samples from a tyre repair shop and from the air above a bacterial culture. The sampling was performed using a solid sorbent, XAD-2, impregnated with NIT, and the derivatives were extracted with acetonitrile and analysed with LC-MS/MS. A three-step process was developed for screening and identifying of volatile amines. The first step, selected reaction monitoring; SRM was applied in order to screen the samples for NIT derivatives. In the second step, a precursor ion scan gave the [M+H]⁺ ion, and in the third step a product ion scan gave the fragments needed for identification. The detection limits varied between 0.12 and 0.25 ng L^–1 when screening for unknown derivatised amines. It was possible to separate and identify all the amines with the structural information obtained and the method proved to be general, sensitive and well suited for sampling and analysis of complex environmental samples.