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Microbiological Aspects of Pharmaceutical Aseptic Processing in the Compounding Pharmacy
Published in Graham P. Bunn, Good Manufacturing Practices for Pharmaceuticals, 2019
Media fills are performed in order to qualify the environment and the compounding process, as well as to qualify the personnel who will be performing the compounding. Media fills consist of utilizing sterile growth media to perform the most challenging compounding process that is performed in that controlled area. Trypticase soy agar (TSA) is typically utilized for this purpose. After manipulation of the media, the samples are incubated at 20°C to 25°C or 30°C to 35°C for 14 days or a combination of the two temperatures for 7 days each. Media is examined visually to determine whether contamination is present, which is evidenced by visible growth or turbidity of media. This qualification is performed initially when a new compounding area is commissioned and then annually for low- and medium-risk CSPs or twice per year for high-risk CSPs. All compounding personnel are required to qualify and requalify by performing media fills. Procedures are needed to define the media fill process. This should include rationale for what manipulations are performed, what containers and closures are utilized, and details regarding the manipulations to be performed. The duration and complexity of the media fill needs to mimic the worst-case process.
Bioremediation of Hydrocarbon-Contaminated Soils Using Indigenous Microbes
Published in Donald L. Wise, Debra J. Trantolo, Remediation of Hazardous Waste Contaminated Soils, 2018
Microbial Enumeration. One gram of soil was suspended in 10 ml of glucose salts medium minus glucose (GSM-G). The composition of this simple salts medium is given in Table 1, and preparation of this medium required that separate solutions be autoclaved and cooled prior to mixing in order to prevent precipitation of phosphate salts. One milliliter of microbial suspension cultured from the contaminated soil was serially diluted in GSM-G until approximately 10−6, where no visible particles could be detected. This dilution was pipetted onto trypticase soy agar (TSA), as well as 4-5 more serial dilutions, according to Pelczar.21 After the suspending liquid was evaporated from these plates at room temperature, the plates were inverted and incubated in an incubator at room temperature for 48 hours. The visible bacterial colonies were counted, and appropriate calculations were carried out to determine the number of microbes per gram of soil.
Airborne Microorganisms in A Domestic Waste Transfer Station
Published in Michael Muilenberg, Harriet Burge, Aerobiology, 2018
Irma Rosas, Carmen Calderón, Eva Salinas, John Lacey
Culturable airborne bacteria and fungi were sampled in a domestic waste transfer station in Mexico City. Close to where the waste was handled, the geometric mean concentration of bacteria was >6700 cfu/m3 of air, of Gramnegative bacteria >460/m3 and of fungi >4900 cfu/m3 of which 75% were Penicillium spp. Concentrations of microorganisms downwind of the waste site were greater than upwind. Salmonella was recovered on Trypticase Soy Agar from 14% of samples. The large concentrations of Gram-negative bacteria and fungi in the waste transfer station could lead to different types of pulmonary reactions, and thus constitute a respiratory hazard to workers, and possibly also to the neighboring population.
Optimization of microbial assisted phytoremediation of soils contaminated with pesticides
Published in International Journal of Phytoremediation, 2021
Asil Nurzhanova, Togzhan Mukasheva, Ramza Berzhanova, Sergey Kalugin, Anel Omirbekova, Annett Mikolasch
PDA is a general-purpose medium for micromycetes (yeasts) that was supplemented with acid to inhibit bacterial growth. TSA was used as a growth medium for the isolation and cultivation of bacteria. TSA + PDA was used for the isolation of Bacillus strains. SCA was used for the detection of Actinomycetes. The number of microorganisms recovered from the rhizosphere of X. strumarium grown in the presence of pesticides was the greatest on TSA and lowest on SCA (Table 2). The number of microorganisms recovered from C. pepo grown in the presence of pesticide also was greatest on TSA, though differences among the other media were not significant. In both cases, the numbers of microorganisms recovered on TSA were roughly two orders of magnitude greater than the numbers recovered on the other media. For both plants grown in the absence of pesticide, more microorganisms were recovered on TSA plates, and roughly one order of magnitude less colonies were recovered from the other media.
Effect of river water exposition on adhesion and invasion abilities of Salmonella Oranienburg and Saintpaul
Published in International Journal of Environmental Health Research, 2018
Mitzi Dayanira Estrada-Acosta, Karina Ramirez, José Andrés Medrano-Félix, Nohelia Castro-Del Campo, Hector S. López-Moreno, Maribel Jimenez Edeza, Jaime Martínez-Urtaza, Cristóbal Chaidez
Bacterial adhesion and gentamicin-protection invasion assays were performed as previously described by Steele-Mortimer (2008) and Dostal et al. (2014), with some modifications. Briefly, HEp-2 cells were seeded in 24-well tissue-culture plates (Corning, U.S.A) at 5×104 cell/mL per well 18 h prior infection. A multiplicity of infection (MOI) of 50–100 bacteria per cell was used for laboratory and river water conditions. Cell monolayers were inoculated with 500 μL of the bacterial suspension and incubated for 1 h at 37 °C. For the adhesion assay, non-adherent bacteria were removed by two washes with sterile Phosphate-buffered saline solution (PBS), and the cells were treated with 1 mL of lysis buffer [0.1 % Triton X-100 and 0.1 % of sodium dodecyl sulfate (SDS, Sigma, U.S.A) in PBS]. Adhered bacteria were enumerated by serial dilutions on TSA (Difco, U.S.A). The percentages of adhered bacteria were calculated dividing the number of CFU of adhered bacteria by the number of CFU of the inoculum. For the invasion assay, after 1 h of infection, the cell monolayers were washed twice with PBS and treated with 500 μL of gentamicin (100 μg/mL), and incubated 1 h at 37 °C. Then, the cells were washed with PBS and treated with 1 ml of lysis buffer. Invasive bacteria were enumerated by plating appropriate dilutions on TSA. Bacterial death by gentamicin was confirmed by negative growth in samples from supernatants of infected cultures treated with gentamicin before cell lysis. A non-invasive E. coli strain was used as a control. Invasion percentages were calculated dividing the number of CFU of invasive bacteria by the number of CFU of the inoculum. Invasion efficiency was calculated as follows: (% of invasive bacteria / % of adhered bacteria) × 100. All steps were incubated at 37 °C in 5 % CO2 humidified atmosphere. The assays were performed as replicate cultures of three independent experiments.
Isolation, characterization and growth kinetics of phenol hyper-tolerant bacteria from sewage-fed aquaculture system
Published in Journal of Environmental Science and Health, Part A, 2020
Lucky Nandi, Ashis Kumar Panigrahi, Nilanjan Maitra, Asoke Prasun Chattopadhyay, Sanjib Kumar Manna
Phenol acts both as a substrate and an inhibitor of bacteria and thus determination of the highest specific growth rate along with growth kinetics of the phenol degrading bacteria is essential in development of bioremediation technology. Growth kinetics of the bacteria was studied in MM with added phenol (concentration: 200-1200 mg L−1) as sole carbon source. The test bacteria were grown on Tryptic Soy Agar (TSA). Single colony from culture plate was taken in a 1.5 mL centrifuge tube and washed in normal saline (0.85%) by repeated centrifugation. The cell pellet was then suspended in saline solution, bacteria concentration in the cell suspension was determined by further plating and 9.18 x 109 - 1.04 x 1010 CFU bacteria were inoculated in MM containing aforesaid different concentrations of phenol. The culture flasks were incubated in a shaker (120 rpm) at 37 °C and samples were taken at 12-hour intervals for cell concentration measurement. The specific growth rate (μ) of cells in the batch culture was determined at the exponential phase of the growth curve using the logarithm of bacterial population determined from absorbance at 600 nm. The data were fitted to standard models described by Haldane,[24] Aiba,[25] Yano,[26] Edwards[27] and Webb[28] to describe kinetics of bacteria in presence of phenol. In all cases, specific growth rate was a function of initial substrate concentration S (mg L−1). Parameters such as µmax, KS and KI (and for Yano model, K in addition) varied. Initial variations were coarse grained, over a wide range of values (e.g. 0-106 for KS, KI and K values, 0-10 for µmax). For each set of values, correlation coefficient, given as where SSe = regression (or explained) sum of squares, and SSt = total sum of squares (or experimental variance), are maximized. Systematically, the grain size is lowered, and iterations are stopped at a consistency of 10−7 in R2 and standard deviation values. All calculations were carried out in a Dell Power Edge R815 server, using Fortran programs written specifically for this purpose.