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Microbiological Aspects
Published in Héctor A. Videla, Manual of Biocorrosion, 2018
Among the variant cellular components, the cellular wall is a compact layer surrounding the cytoplasmatic membrane that gives rigid structure and shape to the cell. The prokaryotic cell wall is chemically different than that of the eukaryotic cell, being an important differential aspect between the two types of organisms. Although it is difficult to observe with the optical microscope, it can be seen clearly in thin sections through the TEM. The cellular wall is the structural element which is differentially stained by the Gram method, the most important differential staining method used in microbiology. Under the electron microscope, the Gram negative cell wall presents a complex and multilayer structure, whereas the Gram positive cell wall is characterized by a single and thicker layer.5
Modern Microscopic Methods of Bioaerosol Analysis
Published in Christopher S. Cox, Christopher M. Wathes, Bioaerosols Handbook, 2020
Of the many bacterial stains in common use, methylene blue serves for simple examination. The Gram stain is the most important differential staining procedure, where Gram-negative bacteria lose the violet stain and take a counter stain. The staining response reflects important differences in the cell wall structure of the two cellular classes, for example, the bactericidal action of Penicillin and the enzyme lysozyme, found in tears and egg white, is much more effective against Gram-positive cells. Very few species have a variable Gram stain response.
Wetlands Microbiology: Form, Function, Processes
Published in Donald A. Hammer, Constructed Wetlands for Wastewater Treatment, 2020
Ralph J. Portier, Stephen J. Palmer
Bacteria are divided into two groups on the basis of a differential staining technique called the gram stain. They differ mainly in whether their cell walls retain the stain (gram-positive) or not (gram-negative). Gram-negative bacteria have short, filamentous fibers composed of protein called pili or fimbriae attached to their walls. Such bacteria tend to stick to each other because pili apparently are used for attachment to surfaces.
Formation mechanisms of viable but nonculturable bacteria through induction by light-based disinfection and their antibiotic resistance gene transfer risk: A review
Published in Critical Reviews in Environmental Science and Technology, 2021
Yiwei Cai, Jianying Liu, Guiying Li, Po Keung Wong, Taicheng An
A common method to detect total viable cells is through differential staining, which stains all viable and dead cells separately so they can be counted via fluorescence microscopy or flow cytometry. A commonly used method is LIVE/DEAD Baclight assay, which relies on SYTO 9 and propidium iodide fluorescent probes (Ding et al., 2017; Foddai & Grant, 2020). The green fluorescent probe SYTO 9 penetrates both intact and damaged cell membranes, so that all cells fluoresce green. The red fluorescent stain propidium iodide only penetrates damaged cell membranes and is used to mark membrane-permeable cells (Salive et al., 2020). The number of cells with a red fluorescent signal is subtracted from the total number of cells with green fluorescence to obtain the total number of viable cells. Although this method has the advantages of sensitivity, intuitive results and a short processing time, it also has disadvantages such as requiring expensive dyes and equipment. Furthermore, when using flow cytometry for quantification, the operation is more complicated and requires both positive and negative controls. In addition, this method requires a combined plate count method to calculate VBNC bacteria (Table 1S).
Groundwater inhabited Bacillus and Paenibacillus strains alleviate arsenic-induced phytotoxicity of rice plant
Published in International Journal of Phytoremediation, 2020
Adrita Banerjee, Anjan Hazra, Sauren Das, Chandan Sengupta
For detection of antioxidant enzyme activities, leaf samples (1 g) from nine experimental set up were taken and homogenized in extraction buffer (0.05 M Na phosphate pH 7.2, 20% v/v glycerol, 0.05% v/v Triton X-100 and 14 mM mercaptoethanol) with the help of pre-chilled mortar-pestle placing on ice tray following earlier reported methodology (Hazra et al. 2018). The homogenates were centrifuged at 10,000 rpm for 10 min at 4 °C and the supernatant was used as enzyme source. On 10% native polyacrylamide gels, 100 µl of each sample from the stock solutions was loaded. Electrophoresis was done at 60 V for 3 h with 0.025 M tris/glycine running buffer, pH 8.8. Quantification of SOD, CAT and POD enzymes was performed following the method described by Maksimović and Živanović (2012) and Uarrota et al. (2016). The differential staining of the isozymes (SOD, POD, and CAT [catalase]) was performed by the following methods.
Novel spray-dried PHA microparticles for antitumor drug release
Published in Drying Technology, 2018
Anna Shershneva, Anastasiya Murueva, Elena Nikolaeva, Ekaterina Shishatskaya, Tatiana Volova
The cell viability in HeLa culture was investigated by the differential staining techniques using DNA-intercalate fluorescent dyes such as acridine orange (AO) and ethidium bromide (EB). Cell staining was performed according to procedure, the previously described by Liu et al.[37] The cells were initially fixed with a 1% formalin solution at room temperature. The culture was washed with Dulbecco’s phosphate-buffered saline (DPBS) solution, 70% alcohol. The degradation of RNA was achieved using 0.2 mL of RNAse which was added to the cells culture and incubated at 37°C for 30 min. Furthermore, solution mixture containing AO (100 µg/mL) and EB (100 µg/mL) was added in a ratio of 1:1. The cell morphological analysis and counting were performed on the first and third day using a fluorescence microscope Leica DM6000B (Leica Camera AG, Wetzlar, Germany) at a lens of 20 × in 10 fields of vision. The staining with 4′,6-diamidino-2-phenylindole (DAPI) and fluorescein isothiocyanate (FITC) (Sigma-Aldrich, USA) was also performed to study cell morphology and the results of MTT. Visualization and counting of cells were carried out in a similar way.