Browsing by Author "Wadhawan, Tanush"

Filter results by typing the first few letters
Now showing 1 - 2 of 2
  • No Thumbnail Available
    Item
    In situ Quantification of Hydrogel Entrapped Microbial Cells
    (North Dakota State University, 2010) Wadhawan, Tanush
    Entrapped bacteria are used in several applications including food and beverage production, antibiotic production, and wastewater treatment. To date in order to determine the viability of entrapped bacteria, they have to be de-entrapped from the matrix first. However, cell de-entrapment procedures, such as matrix dissolution by acid or heating at high temperatures, may affect the viability of the cells. In this study, three viability assays were assessed for in situ estimation of the number of entrapped cells. Also, a new method was developed to determine the effect of entrapment procedures on bacterial cell viability using one of the three assays, LIVE/DEAD® BacLight™ Bacterial Viability Kit. The other two quantitative assays used in this study were the bioreducible tetrazolium salt (XTT) assay and the adenosine triphosphate (ATP) based assay. The applications of the assays and the new method were performed on two cell entrapment techniques widely used in environmental applications, phosphorylated-polyvinyl alcohol (PPV A) and calcium alginate (CA). The data from the XTT and ATP assays showed linearity and strong correlations between the viability signals and number of beads in which each bead contained a similar number of live cells. An application of the XTT assay on the PPV A entrapped bacterial beads was an exception to these results. Effects of the acid and heat dissolution deentrapment procedures on cell viability were also evaluated by using both assays and a traditional plate count method. The heating process showed the greatest reduction in bacterial viability when compared to the other de-entrapment procedures. The ATP assay is a more sensitive and less time consuming approach for viability estimation when compared to the XTT assay and traditional plate count method. Both XTT and ATP assays have potential for use in quantifying the viability of entrapped bacteria. The new method developed for determining the effect of entrapment procedures on bacterial cell viability involved entrapping bacteria directly onto glass slides. This new method was compared with traditional approaches which require dissolution of the entrapment matrix using chelating agents and heat. Both the developed and traditional methods require labeling with fluorescent dyes from the LIVE/DEAD® assay and observing and quantifying live and dead cells under fluorescence illumination. The viability of entrapped cells was compared to the viability of free cells prior to the entrapment. The developed method was applicable to both PPV A and CA entrapped cells. Both methods indicated that the entrapment procedures resulted in reductions in cell viability, but the new method showed less viability reduction than the previously used method. This suggests that the matrix dissolution prescribed in the traditional method negatively affected cell viability and the new method is therefore more reliable. The percent of live bacterial cells before the entrapment ranged from 54 to 74%, while the percent of live cells following the entrapment based on the new method was 39 to 62%. The approach used in the method could potentially be adopted for other cell entrapment techniques.
  • No Thumbnail Available
    Item
    Investigating Biodegradability of Dissolved Organic Nitrogen in Oligotrophic and Eutrophic Systems
    (North Dakota State University, 2014) Wadhawan, Tanush
    Dissolved organic nitrogen (DON) in water and wastewater is a major public concern. In drinking water treatment plants (WTP), DON and biodegradable DON (BDON) may form carcinogenic by-products during disinfection and might also serve as a nutrient for microbiological growth in distribution systems. BDON in treated wastewater can promote algal growth in receiving water bodies. Understanding biodegradability of DON is important to develop strategies and processes capable of minimizing DON impact on the wastewater effluent receiving water bodies and drinking water. WTPs are nutrient-poor oligotrophic systems that receive source water with DON of about ≤2 mg N/L. Wastewater treatment plants (WWTPs) are nutrient-rich eutrophic systems which receive raw wastewater with DON of ≥8 mg N/L. At WWTPs, sidestream deammonification is a highly eutrophic system employed to treat highly concentrated streams of DON (≥100 mg N/L) and ammonia (≥1,500 mg N/L) generated from filtrate from anaerobically digested sludge dewatering. DON characteristics including biodegradability for different trophic levels could differ. The main goal of this dissertation is to investigate biodegradability of DON in these oligotrophic and eutrophic systems. Three research tasks were performed. In the first task, a method to measure BDON in oligotrophic systems was developed and applied to determine the fate of BDON along four treatment stages of a WTP with ozonation prior to filtration. Optimum dose of inocula and incubation time were identified for the BDON measurement. The Moorhead WTP, Moorhead, MN on average removed 30% of DON and 68% of BDON. The second task involved investigating the role of four biological wastewater treatment processes in removing DON from eutrophic systems. Nitrification process biodegraded 70, 54, and 57% of DON in influent, primary effluent, and secondary effluent, respectively. Heterotrophic DON removal was less (1.7 to 38%) while denitrification and deammonification did not remove DON. For the third task, BDON biodegradability in highly eutrophic system was investigated using nitrifying sludge. About 45 to 90% of DON in sidestream effluent was biodegradable. Information from this dissertation provides a better understanding on DON and BDON fate through water and wastewater treatment processes representing different trophic levels.