Biology / Biologie

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Analysis of plasma immune response to clostridium difficle proteins in hospitalized patients at Health Sciences North, Sudbury, Ontario, Canada
(2017-03-03) Dubé, Kristy-Anne
Clostridium difficile infection (CDI) is the leading cause of nosocomial diarrhea worldwide. Prevention and treatment of CDI requires an intimate understanding of the immune response to the bacterium. Thus, this study investigated the plasma immune response of 79 patients at Health Sciences North in Sudbury, Ontario, Canada to identify immunodominant C. difficile proteins. Plasma samples were obtained from patients diagnosed with C. difficile infection, patients positive for C. difficile without symptoms, and patients who were negative for C. difficile, with and without symptoms. The plasma samples were tested for the presence of antibody reacting to C. difficile protein extracts using Western Blot analysis, Wes immunoblotting, and subsequent characterization by 2-D immunoblot analysis and mass spectrometry. Candidate immunodominant C. difficile proteins were found to be enolase, acetyl-coA acetyltransferase, and the 50s ribosomal protein, L7/L12. However, the presence and/or levels of antibodies that recognized these proteins in patient plasma were not statistically different between patient cohorts. Further analysis of the potential immunogenicity of these proteins could be useful to CDI treatment and prevention
Analysis of RBM5 and RBM10 expression throughout H9C2 skeletal and cardiac muscle cell differentiation.
(Laurentian University of Sudbury, 2013-07-31) Loiselle, Julie Jennifer
RNA Binding Motif (RBM) domain proteins RBM5 and RBM10 have been shown to influence apoptosis, cell cycle arrest and splicing in transformed cells. In this study, RBM5 and RBM10 were examined in non-transformed cells in order to gain a wider range of knowledge regarding their function. Expression of Rbm5 and Rbm10, as well as select splice variants, was examined at the mRNA and protein level throughout H9c2 skeletal and cardiac myoblast differentiation. Results suggest that Rbm5 and Rbm10 may (a) be involved in regulating cell cycle arrest and apoptosis during skeletal myoblast differentiation and (b) undergo post-transcriptional or translational regulation throughout myoblast differentiation. All in all, the expression profiles obtained in the course of this study will help to suggest a role for Rbm5 and Rbm10 in differentiation, as well as possible differentiation-specific target genes with which they may interact.
Analysis of the American mink (Neovison vison) harvest decline and genetic introgression
(2017-09-11) Shofstall, Krista
The American mink (Neovison vison) is endemic to North America where they have been domesticated over the course of 150 years by the fur industry. These domestic mink have been escaping from farms around the world and in North America while the harvests of wild mink across Canada are in decline. In this thesis, I used a combination of environmental data, spatial data, and genetics to better understand the declines. A multiple linear regression and a tree regression indicated that muskrat harvest growth rate, road density, and annual precipitation had the most effect on the mink harvest. To study the genetic introgression of domestic and wild mink, a 300 basepair fragment of the mitochondrial control region was used to determine regional differences between the wild and domestic populations of Nova Scotia and Ontario. Color differences and the direction of introgression were also studied. Significant differences between Nova Scotia wild, Ontario wild, and domestic mink were found. A pairwise ɸst test was used to determine directional introgression and resulted in an introgression of the hybrids towards the wild population. Together these results provide a better understanding of the decline in mink harvest although further research is needed to assess the direct impact of domestic escapees on the environment and on the wild population in North America. Prevention of domestic escapees is needed to stop hybridization which is important for the preservation of the species and to prevent further risk of outbreeding depression.
Anthropogenic effects on chronic stress and nest predation patterns in freshwater turtles
(2016-07-29) McCurdy-Adams, Hannah L.
Human altered landscapes contribute to direct and indirect threats on wildlife populations. Chronically high stress, an indirect threat, has been associated with decreased fitness. I examined chronic stress in Midland Painted Turtles (Chrysemys picta marginata), Snapping Turtles (Chelydra serpentina), and Blanding's Turtles (Emydoidea blandingii) in areas with varying densities of anthropogenic structures in Muskoka,Ontario. Enzyme immunoassays were run, in partnership with Dr. Gabriela Mastromonaco (Toronto Zoo), on turtle claws to obtain concentrations of corticosterone (CORT). Most Snapping Turtles had above-minimum CORT, and male Midland Painted Turtles and Blanding's Turtles had relatively high CORT levels. Anthropogenic density did not seem to affect CORT levels, but more research needs to be done to understand the complex relationships. I also determined the rate and pattern of nest depredation, a direct threat, throughout egg incubation in areas of varying anthropogenic densities. Nest depredation occurred throughout egg incubation and relative nest mesopredator abundance did not differ among sites with varying anthropogenic densities. Contrary to some nest caging protocols, my findings suggest that nest cages should be kept on nests until the turtles hatch and priority should be given to caging nests in low road density areas to help increase population recruitment. Understanding indirect and direct threats to turtles can help inform mitigation strategies to improve their fitness.
Anthropogenic impacts on life history traits on Eastern Chipmunks (Tamias striatus)
(Laurentian University of Sudbury, 2015-04-20) Lyons, Jeremy
Urban centers provide unique habitats for opportunistic animal species. However, conditions in urban habitats are inherently stressful and require specialized adaptations in resident animals. Eastern chipmunks are a generalist small rodent species common in a number of different habitat types. I examined differences in behavioural traits and stress levels (chapter one), as well as body condition and parasitism rates (chapter 2) between urban and natural populations of chipmunks. I tested the hypothesis that 1), chipmunks in urban habitats would show greater levels of boldness and exploratory behaviour, 2), urban chipmunks would show greater levels of stress reflected in greater cortisol concentrations in hair and feces, 3) urban chipmunks would have a relatively poor body condition due to poor quality food and competition in urban centers, and 4) urban chipmunks would show greater parasitism rates measured in greater oocyte prevalence, richness, and abundance from fecal samples. A total of 140 individual chipmunks (80 natural and 60 urban) were sampled from natural habitats in Algonquin Provincial Park and outside of Sudbury, and urban habitats in Huntsville and Sudbury Ontario. Differences in behavioural patterns suggest urban animals may be more habituated to human presence, while body condition was significantly superior in urban animals. However, cortisol measured from hair and feces, leukocyte concentrations measured from blood and parasitism impacts did not differ between individuals from the two habitat type. Overall, chipmunks in these urban habitats have a better body condition and are more habituated to human presence than their natural conspecifics.
Antibacterial activities of Propionibacterium acnes bacteriophages against a diverse collection of P. acnes clinical isolates: prospects for novel alternative therapies for acne vulgaris
(2017-08-22) Graham, Jenna
A total of 136 chronically infected Canadian acne patients from Ottawa-Gatineau and Northeastern Ontario regions accounting for 75% of subjects (12-50 years old, with 90th percentile at the age of 30) who had suffered acne vulgaris (with various acne related scarring) for a median duration of 4 years, were sources for isolation of Propionibacterium acnes, the etiologic agent for acne vulgaris. Eighty-four percent of patients were subjected to various treatment regimens with topical and systemic agents including in combination with 1-3 different types of antibiotics (mean duration of 7 months). A diverse collection of 224 clinical P. acnes isolates from Canadian and Swedish subjects were characterized for their sensitivities to infection by a Canadian collection of 67 diverse phages belonging to siphoviridae; and multiple minimal cocktails consisting of 2-3 phages were formulated to be effective on global P. acnes isolates. Propionibacterium acnes isolates were characterized by multiplex PCR to belong to phylotypes IA, IB and II, which also showed resistance against commonly used antibiotics for treating acne vulgaris (overall resistance rate of 9.5%), were sensitive to phages regardless of their type and antibiotic resistance patterns, providing ground for phages as novel alternative therapeutics for future in vivo trials. The phage collection was diverse by virtue of their BamHI restriction patterns and full genome sequences and harboured a major tail protein (MTP) that appeared to be important in contributing to their host ranges. Three dimensional structural modeling of the N-domain of P. acnes MTPs implicated previously unreported involvement of the α1-β4 loop (C5 loop) within N-domain amino acid sequence in contributing to the expanded host range of a mutant phage to infect a naturally phage resistant P. acnes clinical isolate. Given the potential of phages for rapid mutational diversification surpassing that of their bacterial hosts and the fact that phages are generally regarded as safe (GRAS), rapid and cost-effective derivation of mutant phages with expanded host ranges provide a strong framework forimproving phage cocktails for use in future personalized medicine.
Antibacterial activity of green microalgae from stressed environmental conditions
(2020-10-28) Little, Shannon
Bacterial resistance to antibiotics is a growing problem in worldwide health care. The discovery of new antibiotics has been less successful in recent years which have so far not proven to be as successful or safe as natural products. Photosynthetic fresh (non-saline) water green microalgae have, however, been recently shown to be a promising source of compounds with antibacterial activity. Extracts from microalgae from a variety of environments have inhibited growth of both gram-negative and gram-positive bacteria. By investigating untapped microalgal species that thrive in stressed environments, most likely through the production of protective secondary metabolites, there is a greater likelihood of discovering strains capable of producing bioactive compounds with antibacterial activity. In this study, microalgal species obtained from environments with low pH, high metal concentrations or municipal wastewater were tested against a variety of bacterial species. The results confirm that microalgae from stressed environments are a promising source of compounds exhibiting antibacterial activity.
Applicability of remote sensing technologies for use in Ontario forest fire management protocols
(2020-08-20) Hendel, Isabelle-Gabriele
The use of remote sensing in forest fire detection could allow earlier mitigation of active fires resulting in a significant decrease in fire related health effects. Infrared detection can provide more detailed and accurate information on fire size, spatial accuracy and temperature. Hyperspectral sensors can allow the detection and earlier mitigation of high risk areas. Few studies have concentrated on the efficiency of multiple types of sensors surch as infrared, hyperspectral and multispectral being used in combination. A literature review was done to highlight health effects on humans and the environment in order to determine the need for changes in current forest fire detection practices. The infrared portion of this research focused on sensors abilities to detect heat signatures and compared the specifications of each sensor with the others to ensure a cost-effective solution to current practices using off-the-shelf technologies. The hyperspectral portion of this research focused on using vegetation indices to examine potential use of such sensors in the detection of high risk areas.
Aquatic microbial community structure and function across a gradient of logging, fire, and industrial watershed disturbance.
(2014-12-16) Emilson, Caroline.
The role of microbial communities in the recovery of aquatic ecosystems from watershed disturbance has received little attention despite their important role in energy and nutrient cycling. This study investigates the structure and function of microbial communities on a standardized substrate (alder leaves) in small streams across a wide gradient of watershed disturbances. Microbial communities exhibited variation with disturbance regime with lower hydrolase enzyme activities at all disturbed streams compared to undisturbed streams, and the lowest rates of microbial decomposition, fungal biomass, and differences in microbial community composition at the most severely disturbed streams. Forest and wetland cover were identified as important watershed features that provide DOC to fuel microbial activity in aquatic ecosystems. Increasing road density within the watershed was identified as having a negative impact or association on microbial activity that appeared to be linked to inputs of inorganic solutes that were measured through increased levels of specific conductance in stream water samples. This study is one of the first of its kind and it provides some important evidence that leaf litter associated microbial communities can be influenced by factors linked to watershed disturbance and as such may be useful as indicators of watershed disturbance and potentially the state of recovery of aquatic ecosystems.
Assessing differential expression of genes associated with nickel resistance in jack pine (Pinus banksiana) and eastern white pine (Pinus strobus): effects of nickel toxicity
(2021-06-28) Moarefi, Nastaran
Eastern white pine (Pinus strobus) and jack pine (P. banksiana) represent two of the major coniferous species planted in the metal contaminated region of Northern Ontario. Studies on coping mechanisms of these two species to soil metal contamination are limited. The main objectives of the present project were to 1) determine how these two species respond to soil nickel (Ni) contamination and 2) assess the effects of different doses of nickel on the expression of six genes associated with Ni resistance. These genes include high affinity Ni transporter family (AT2G16800), 1-aminocyclopropane-1-carboxylic acid deaminase (ACC deaminase), Natural resistance-associated macrophage proteins (NRAMP3), and serine acetyltransferase (SAT), Nicotianamine synthase (NAS) and Glutathione reductase (GR). Seedlings were grown in soils treated with increasing concentrations of Ni(NO3)2. Potassium nitrate (KNO3) was used to control for potential toxicity of nitrate ions. Gene expression was assessed using RT-qPCR. Damage ratings reveal that P. banksiana is sensitive to Ni(NO3)2 doses of 1,600 and 3,200 mg/kg as well as 3,200 mg/kg KNO3. However, P. strobus was tolerant to all treatment. Soil Ni treatments induced a downregulation of NAS in P. strobus as well as AT2G16800 and NRAMP3 in both species with the severity increasing at high doses of Ni. An upregulation of ACC in both species consistent with an inverted-U hormetic effect was observed. Ni also triggered a hormetic U-effect downregulation of SAT in P. strobus with the lowest dose showing the highest repression. An opposite trend was observed in P. banksiana where Ni induced an upregulation of SAT. Moreover, GR induced an inverted-U hormetic upregulation in P. banksiana. Knowing the coping mechanisms of these conifers to metal contamination is essential for ensuring their longterm sustainability.
Assessing patterns and biogeochemical impacts of shrubification in sedge fens of the Hudson Bay subarctic tundra
(2021-09-30) Robinson, Chantae Ucaeci
Mean annual air temperatures in Arctic regions have increased by about 2–3°C over the past 50 years. As a result, the abundance and maximum height of woody shrub species has increased (shrubification) at sites around the circumpolar Arctic. This paper examines the extent and impact of shrubification on relevant nutrient cycling, soil microbial community and GHG production potential at a fen environment in the Hudson Bay Lowlands near Churchill, MB. Greening during 1984-2017 was analyzed using Google Earth Engine, a 10-year litterbag study was conducted to compare nutrient release patterns between a sedge and shrub dominated site, soil microbial DNA was extracted from sites and compared, and active layer samples were incubated to determine potential GHG production. We found that greening was extensive in the area, methane production was lower in soils from shrub-dominated sites and shrub litters decomposed significantly slower than sedge litters in the first year. These results suggest that it is unlikely that permafrost is sustainable where shrubs encroach, yet the effects of permafrost thaw on carbon cycling could be in part offset by lower microbial methane production associated with shrubs in near surface soils.
Assessment of the recovery of littoral zone food-webs in Lake Charr Lakes in Killarney Provincial Park affected by acidification and contemporary stressors
(2023-12-12) Moskal, Haley
Killarney Provincial Park is a site of significant environmental damage from acidic deposition resulting in the extreme loss of biodiversity. As many lakes in Killarney Park acidified to pH< 6, many populations of fish and sensitive species were lost from Killarney’s lakes and streams. In recent decades, effective legislation, and massive emission reductions programs have resulted in substantial chemical recovery, however biological recovery has lagged considerably behind. The objective of my study was to assess the recovery of littoral zone food- webs in Killarney’s acid damaged lake charr (Salvelinus namaycush) lakes by resampling 20 lakes previously studied 25-years earlier, with a focus on assessing change in water quality, and key prey and indicator species (crayfish, mayflies, amphipods, littoral fish). Dramatic water quality shifts have occurred in these lakes including increases in pH and DOC and decreases in SO4 2-, Ca2+ and metals and increase in surface water temperatures. The substantial rise in DOC may represent a significant increase in nutrient conditions, but more importantly appears to have affected the thermal structure and cold-water habitat for charr in some ultra-clear lakes. In contrast to the physicochemical changes in the lakes, particularly the substantial pH rise to above toxic levels, the biotic changes were not all as positive. There is evidence that emerging contemporary stressors, including declining Ca levels and shifts in littoral fish communities to centrarchid-dominated may be inhibiting recovery potential in the littoral zones. My results demonstrate the importance of the interactions between historical and contemporary stressors, and that contemporary stressors may now be controlling variables in determining littoral food web recovery. Overall, my study provides evidence that many of Killarney Park’s historically acid-damaged lakes are no longer limited by acidic conditions but contemporary stressors, such as the arrival of warm water centrarchids (e.g. smallmouth bass) have emerged as key factors shaping littoral zone recovery. In particular, the increase in concentration of dissolved organic carbon may be a potential driver of littoral zone recovery, providing food source additions to the nearshore environment and increasing the productivity capacity of food webs.
Assessments of genetic and reproductive health in Canada’s endangered Oregon Spotted Frog (Rana pretiosa)
(2023-08-04) Hunter, Briar
Zoological institutions are increasingly relied upon for ex situ management of species at risk, with conservation breeding and reintroduction programs providing both assurance against extinction and a reliable source of offspring to reinforce wild population sizes. Ex situ efforts face many challenges, however. One often overlooked challenge is balancing genetic priorities, like retaining genetic diversity and reducing inbreeding, with reproductive priorities, like mate compatibility and reliable breeding, both of which are required for a successful program. Like many amphibians, the Oregon Spotted Frog (Rana pretiosa) is highly threatened in its native habitat and ongoing conservation breeding and reintroduction programs are experiencing limited success; the genetic sustainability of these populations remains unassessed. In this study, I evaluated the genetic health of both zoo and wild populations of R. pretiosa in Canada and investigated some potential causes of the ongoing ex situ reproductive failures associated with egg binding. I found that zoos have maintained stable genetic metrics relative to their wild sources, but ongoing collections from wild populations should be reassessed due to low genetic diversity available therein. No clear causes of egg binding were elucidated but I found older female frogs (> 3 years old) who became egg bound generally had a higher body mass than conspecifics, while body mass did not differ in females becoming egg bound in their first breeding season (2-3 years old). This suggests frogs should be monitored for egg binding and changes in body condition differently depending on their age. Overall, there were no significant differences in genetic or reproductive health across the three zoo conservation breeding populations, but scaled mass index was significantly lower in the zoo with larger holding tanks and less genetic management: a reminder of the importance of husbandry and environment in conservation breeding program outcomes. The costs and benefits of strict genetic management vs. a more communal breeding approach should be carefully considered in light of these results. Along with more cohesion, communication is required between all involved institutions to have an effective impact on the conservation of Oregon Spotted Frogs in Canada, the recommendations discussed here have applicability to amphibian ex situ programs worldwide.
Bacteria against bacteria: isolation, identification and characterization of bacteria with antimicrobial activities against Gardnerella vaginalis
(2018-06-13) Ross, Megan
Gardnerella vaginalis is the etiological agent of bacterial vaginosis (BV), a chronic vaginal infection that affects women globally. Increasing antibiotic resistance and failing treatment options has resulted in chronic BV, demonstrating a need for novel alternative therapeutics. A total of 34 environmental bacterial isolates with antimicrobial properties against a G. vaginalis collection (n=17) were isolated and characterized for their antagonistic effects against G. vaginalis isolates and for their production of antimicrobial active agents. The G. vaginalis collection was shown to be diverse by genotyping and by their antibiotic susceptibility profiles. Anti-G. vaginalis (αGV) isolates were identified and phylogenetically clustered following full genome sequencing and their antiGV target range specificities were investigated. A subset of the αGV collection (n=20) were shown to produce and export active antimicrobial agents with molecular sizes of 4.6-35kD, which showed specific lethal effect on G. vaginalis isolates (target ranges of 5-100%). Cocktail formulations consisted of promising representatives from αGV clusters, with overlapping complimentary properties and specificity to G. vaginalis were identified for further study as the antimicrobial candidates for BV.
Bioelectrical and biophysical interactions across spatio-temporal domains: identifying the conduit for the emergence of material from the immaterial
(2016-01-15) Carniello, Trevor N.
The emergence of biophysical and bioelectrical dynamics at the level of the organism are explored in an attempt to identify the physiological correlates of the onset of senescence in plants. Furthermore, the ability of phylogenically disparate entities to demonstrate the potential to simultaneously display correlated electro-dynamic processes akin to sharing the "same space" or spatial parameters is investigated. The underlying, aforementioned electrophysiological changes associated with senescence and the potential for phylogenically disparate systems to display markedly correlated processes may share water as a common source of variance. The imperative investigations on the physical nature of water and its interactions with applied electromagnetic fields with respect to enhanced latency of the onset of deviations in pH and enhanced electro-dynamic correlations are experimentally explored. A hypothesis, supplemented by marked quantitative convergence, suggests that water is the central conduit necessary for the emergence of material from the immaterial.
Biological recovery in an urban industrial stream: using the reference condition approach to assess the current state of Junction Creek, Sudbury, Ontario
(2018-06-22) Wittmann, Amanda
Benthic macroinvertebrate communities in Junction Creek, Sudbury, have recovered remarkably since the 1970s, following government initiatives to lower atmospheric pollution, the implementation of mine wastewater treatment, and Greater Sudbury’s Regreening Program. To understand contemporary temporal and spatial patterns of biological condition in this stream system, I employed a number of benthic community metrics and the Reference Condition Approach. There was little evidence of temporal trends in the benthic macroinvertebrate communities across the 2003-2015 study period, however there was strong evidence of community composition, water and sediment quality differences among study sites. This urban industrial stream is affected by multiple stressors (straightening, culverts, urban and mining pollutants, overflow from sewer outfalls, etc.), which accumulate as the water flows downstream. Lime-treated metal mining effluent forms the headwaters, which receive cold groundwater from the upper reaches and is further diluted by the Maley tributary before reaching the heavily urbanized city. In these upper reaches, community metrics that are indicative of sensitive organisms and higher diversity are elevated and metal levels are lowest within the study area. Biological and chemical conditions decrease slightly heading downstream, but are much better following the millions of dollars spent in 2001 on diverting acid mine drainage underground to be treated kilometers away before being discharged into the creek below the study area. Treated surface runoff from a historic mine site, and a portion of each the Clarabelle Mill property and slag storage area from one of the world’s largest metal mining and smelting complexes in the world feed Nolin Creek, which then enters the stream through concrete box culverts that run beneath the city’s downtown core before re-surfacing and flowing South of the city. Despite the recent appearance of fish within Nolin Creek, it is still a major source of contaminants to Junction Creek, with elevated water and sediment metal levels and low benthic macroinvertebrate community diversity and abundance. This research showed that use of select benthic macroinvertebrate community metrics served as a better tool to assess biological conditions within the stream than comparison to near-pristine reference sites, but that local reference sites would serve as a best practise since they take account for naturally high regional metal levels, and the effects from decades of atmospheric pollution have had on Sudbury soil, lakes and streams. Additionally, test site benthic macroinvertebrate metric scores were surprisingly similar to those of pristine reference sites, but abundance was much higher at all reference sites. The apparent similarities between test and reference biotic conditions may be due to high nutrients and ions found in urban environments, inappropriate matching of reference and test sites or numerous cumulative effects. It was therefore recommended that ecosystem processes and functioning be studied in order to obtain a more accurate view of the current biological condition of Junction Creek. Finally, aqueous metal levels have decreased substantially and benthic macroinvertebrate community abundance and richness have increased substantially in the last 50 years, but appear to be at a relatively stable point currently, although the variation in both metrics is generally higher than those of reference sites. Biological and chemical conditions will likely not improve until the major current stressors (residual contamination of sediments and soils, and addition of mining effluent) are removed and habitat is improved.
Biomonitoring under changing climate conditions: assessing seasonal variability of benthic macroinvertebrate communities and stream characteristics in two ecozones in Northern Ontario, Canada.
(2017-05-12) Bourne, Vanessa A.
The vast “Ring of Fire” region, at the interface of the Boreal Shield and Hudson Plains ecozones in the far north of Ontario, Canada, is considered to be one of the most promising mineral exploration areas in Ontario in almost a century. The region is undeveloped and remote, accessible only by air, water or winter road. Multiple stresses to the natural ecosystems are anticipated with mining development, in a time of climate change which is predicted to significantly impact hydrological and temperature regimes (intensified spring freshet, more high discharge events, warmer temperatures for longer). Preserving and protecting the aquatic ecosystems in this pristine region will require effective baseline environmental monitoring in advance of development. Quantifying seasonal variation of habitat characteristics and benthic macroinvertebrate (BMI) communities, and assessing the seasonal difference between ecozones are considered key challenges in designing effective monitoring programs. In this study, I examined seasonal variability in habitat characteristics and BMI communities by sampling 43 stream sites distributed across the two ecozones. In 2015 each site was sampled three times during the open water season: post-freshet, mid-summer and fall. BMI communities varied among sampling seasons, but less variation was observed between the post-freshet and summer sampling periods than either of these compared to the fall. Ordination analyses of BMI communities identified to the family level indicated that sites from the same sampling season were more likely to group together. Water temperature, stream velocity and canopy coverage were the key factors associated with seasonal differences in benthic macroinvertebrates. Differences in location of the sampling sites across the two ecozones did not significantly impact seasonal variability, but differences may have been constrained by the common habitat features used because of the sampling protocol. Conducting sampling during the post-freshet or summer seasons is recommended for future stream bioassessments to reduce the influence of seasonal variability and thus ensure comparability over time.
Cellular mechanisms associated with chemotherapy induced ribosomal RNA disruption in ovarian cancer cells
(2019-03-15) Fluke, Amanda
The degradation of RNA is a ubiquitous process prevalent in all cells; employed to maintain nucleotide turnover, maturation, and quality control of RNA. A recent breast cancer clinical trial demonstrated that the degradation of ribosomal RNA (rRNA) in tumour biopsies, referred to as RNA disruption, could serve as a potential biomarker for patients receiving chemotherapy treatment. Given that most, if not all, chemotherapy treatments are associated with toxic side effects, the ability to differentiate early in treatment between patients whose tumours respond to chemotherapy versus patients whose tumours do not would be a very useful tool to prevent needless toxicity in nonresponding patients. Further research, in vitro, demonstrated that this phenomenon was reproducible using a number of different chemotherapy agents in a variety of cancer cell lines. While studies have indicated measuring RNA disruption can be a useful tool in clinical diagnosis, the mechanism(s) of RNA disruption in response to chemotherapy treatment are not well understood. The aim of this study was to investigate the role of Nonfunctional RNA Decay (NRD), RNase L, autophagy, and apoptosis in chemotherapyinduced RNA disruption. Treatment of A280 ovarian carcinoma cells with a proteasome inhibitor did not reduce chemotherapy-induced RNA disruption, suggesting that NRD does not play a role. However, treating A2780 cells with the synthetic dsRNA Poly I:C and a small molecule RNase L activator (RLA), induced an rRNA disruption pattern comparable to that of docetaxel (DXL). Furthermore, A2780 cells transfected with the RNase L inhibitor ABCE1 reduced rRNA disruption in response to DXL or RLA treatment, demonstrating a potential link between RNase L and RNA disruption. Immunoblot experiments revealed that DXL and RLA induced elevated levels of protein associated with the activation of autophagy (LC3-II) at 12, 24, and 48 hours after treatment. DXL and RLA also induces increased levels of proteolytic products associated with apoptosis (PARP) at 12 and 24 hours post-treatment. DXL and RLA-induced RNA disruption was strongest at 48 hours, suggesting that that DXL and RLA-induced RNA disruption are not concurrent with autophagy and apoptosis. The results of the study provide greater insight into the mechanisms associated with chemotherapy-dependent RNA disruption, which could be helpful in the future in improving the accuracy of the RNA disruption assay as a chemotherapy response biomarker.
Characterization of microbial communities associated with the rhizosphere of wetland plants from the Sudbury Region
(Laurentian University of Sudbury, 2015-07-27) Yücel, Çağdaş Kera
The role of microbial communities within the plant rhizosphere is a rapidly developing area of research. Specificity of microbial community structure to plant species and environmental drivers of this relationship are not yet well understood in natural communities, particularly in wetlands. In the present thesis, species-specific differences in those communities as well as environmental influences on those differences were examined separately. In an experiment, six different wetland species taken from field sites around Sudbury, Ontario were grown in mesocosms in a wetland garden for two growing seasons. The species included two species from two genera of the family Cyperaceae each, one species of Poaceae and one species of Ericaceae. Mesocosms were inoculated with a mixture of field rhizosphere soils from all the collected plants. A field study was simultaneously conducted on the rhizospheres of two of those species from wetlands along an industrial disturbance gradient. The microbial community structures of the rhizospheres of the selected wetland plants were determined using next generation 454-pyrosequencing techniques. Microbial community structure in the garden experiment showed specificity to plant taxa which was related to the phylogeny of the host plant, the differences increasing with decreasing taxonomic relatedness of the plants. Differences in the microbial community structure between the investigated plant species were also found in the field, but were secondary to site-specific effects. I conclude that the microbial community structure of the rhizosphere does differ between plant species and that environmental conditions were stronger than plant-microbe interactions in the scale of influence over microbial community structure.
Characterization of secreted acid phosphatase activity of a streptomyces albus clinical isolate
(Laurentian University of Sudbury, 2015-07-27) Alsalmi, Fatin
Due to limitations in the efficacy of the current vaccines against Tuberculosis, there are efforts currently underway exploring the possibility of using the phylogenetically related Streptomycetes as heterologous vaccine vehicles against Tuberculosis. Ideal candidates for such a vaccine should be somewhat pathogenic and physiologically similar to M. tuberculosis so as to induce an immune response capable of targeting and inactivating this pathogen. Towards establishing such a physiological relatedness, previous work in our laboratory has established a similarity profile between M. tuberculosis and a clinical isolate of Streptomyces albus at the level of secreted enzymes. Amongst these was a strong secreted acid phosphatase activity. This work reports on the characterization of the secreted acid phosphatase activity in this clinical isolate of Streptomyces albus. In this work, two enzymes with sequence homology to those encoding a protein tyrosine phosphatase and an inorganic pyrophosphatase were purified from the culture supernatant of S. albus. The calculated molecular masses of these two putative phosphatases were approximately 18 and 30 kDa, respectively. The S. albus protein tyrosine phosphatase has 50% amino acid sequence identity to the protein tyrosine phosphatase of M. tuberculosis (gi|686037535). The inorganic pyrophosphatase has 68% amino acid sequence identity with M. tuberculosis inorganic pyrophosphatase (gi|625006479). Based on our knowledge of the role of acid phosphatases in pathogenic bacteria, presence of this activity in the supernatant of S. albus further supports the physiological relatedness of M. tuberculosis and S. albus and supports the proposition of using S. albus as a heterologous vaccine against Tuberculosis.

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