Thesis (Ph.D.) -- University of Adelaide, Dept. of Microbiology, 1974. / Reprints to two articles published by the author held in pocket in back of publication.

Thesis (Ph. D.)--University of Adelaide, 1985. / Includes bibliographical references (leaves 267-287).

Master of Science / Geology / Saugata Datta / The province (upazila) of Matlab in SE Bangladesh is highly affected with elevated concentrations of dissolved As content and widespread fecal contamination in untreated drinking waters. The study area is sedimentologically composed of thick floodplain deposits of Holocene age overlying Plio-Pleistocene grey fine to coarse sands with considerable clays (consisting of Dupi Tila formation). The goal of the current study is to understand the possible impact of co-occurrence of dissolved organic carbon (along with As release) and fecal indicator bacteria (e.g., E.coli) in aquifers from shallow to deep groundwater quality in this area. Nineteen groundwater (spanning a depth range of 14 to 240 m) samples and nine surface water samples (eight ponds and one canal in proximity to the piezometer nests) were collected from four different piezometric nests within north and south Matlab Upazila in Bangladesh during the monsoonal season (Jun-Jul 2014). The analyses of dissolved organic carbon (DOC) and its fluorescence properties indicate that the chemical character of DOC from shallow to intermediate groundwaters (<150 m) and surface water is dominated by more aromatic and humic materials than deeper groundwaters. Both groundwaters and surface waters may receive humic substances leached from soil and/or from the cellular constituents and exudates of indigenous aquatic organisms. Dissolved organic carbons in groundwater and surface waters are composed of predominantly UVA and UVC-humic like along with tryptophan like components. Only 15% of total C is modern carbon at shallowest depths (<30 m) in groundwaters. The recharge source of groundwaters is from local precipitation, with or without some evaporation before infiltration as depicted by the δ2H and δ¹⁸O variations and the water is infiltrating through mostly terrestrially derived weathered sediments into the aquifers. The type of water in the study area is Ca-Na-HCO₃⁻ type. More toxic and soluble As (III) is present in shallow groundwaters (<30 m). High concentrations of As (V) and As[subscript (t)] are observed high in shallow and intermediate depth wells (<150 m). The most probable number based on the Colilert test and qPCR result for E.coli suggest that unprotected surface waters are harbingers for high microbial population compared to hand pumped wells. However, the very low observed concentrations of cultured E. coli (<1-10 MPN/100 mL) and E. coli DNA (<40 Copies/100 mL) in the wells indicates that the abundance of E.coli cells decrease rapidly with residence time in oligotrophic aquifers. Thus, it may be suggested that more humic DOC in shallow and intermediate groundwaters may be involved in complexation or other biogeochemical reactions that may mobilize As in groundwater. The non-indigenous bacteria can be the primary producers of DOC in the aquifers which can be utilizing surface derived DOC.

Cells dedicate tremendous amounts of energy to express essential genes for survival. During transcription, RNA polymerase (RNAP) actively scans the template strand of DNA, stalling when it meets DNA damages. Stalled RNAP prevents repair by the nucleotide excision repair pathway (NER); a sub-pathway of NER named transcription-coupled repair (TCR) resolve this problem by removing RNAP and recruiting repair proteins. In Escherichia coli, a TCR protein named “Mutation Frequency Decline” (Mfd) couples removal of RNAP through its motor activity with recruitment of the NER repair proteins. Mfd can be divided into two functional halves; the N-terminal region (MfdN, domains 1-3) is essential for NER protein recruitment, and the C-terminal region (MfdC, domains 4-7) is responsible for RNAP-interaction and motor activity. Data suggest Mfd undergoes large conformational movement to activate RNAP removal and repair protein recruitment. To study the activation mechanism of Mfd, we created several full-length “hyperactive” mutants by perturbing interactions between MfdN and MfdC. In all mutants, residue 79 in domain 1 is changed from aspartic acid to arginine (D79R), disrupting a key salt bridge interaction with arginine 804 in domain 6. The linker connecting MfdN and MfdC was made cleavable to allow separation of MfdN and MfdC, which enable us to study activities in equal molar concentration. We have studied the effect of the D79R mutation in vivo (cytotoxicity and UV sensitivity) and in vitro (enzyme activity and thermal stability), and demonstrate that this single residues change render the enzyme “hyperactive”. This confirms our model of activation: activation of Mfd results from breaking communication between MfdN and MfdC

The Glycine and Proline Reduction systems are two of the best characterized selenoenzymes in bacteria and have been found to occur in a wide variety of clostridia [1-5]. These enzymes are utilized to reduce glycine or D-proline to obtain energy via substrate level phosporylation or membrane gradients, respectively [6, 7]. This includes the pathogens C. difficile and C. botulinum [5, 8]. Strains of C. difficile are activate toxigenic pathways whenever either of these pathways is active within the cell [5, 8]. Though evolutionary studies have been conducted on ammonia producing bacteria [9] none has been done to directly characterize these two system by themselves. This includes an understanding of whether or not this system is transferred between organisms, as many of the clostridia that are to be studied are known to have an “open genome.” [8, 10] With this information we were able to generate a phylogenic model of the proline and glycine reduction systems. Through this analysis, we were able to account for many clostridial organisms that contain the system, but also many other organisms as well. These included enterobacteriaceae including a strain of the model organism, Escherichia coli. It was further concluded that Glycine Reductase was a much less centralized system and included a wide range of taxa while Proline Reductase was much more centralized to being within the phyla of firmicutes. It was also concluded that the strain of E. coli has a fully functional operon for Glycine Reductase.