Ambient temperature is known to drive changes in the thermal physiology of mammals, such as an increase in the capacity for thermogenesis in winter. Previous work has shown that acclimation to chronic cold increases the capacity for non-shivering thermogenesis (NST) and thermogenic capacity (cold-induced maximal oxygen consumption, V̇O2max) in deer mice (Peromyscus maniculatus). Photoperiod, is an important driver of seasonal changes in physiology. In the wild, animals are attuned to seasonal changes in temperature and photoperiod. However, the independent and combined effects of temperature and photoperiod on the capacity for NST in small mammals, such as the deer mouse, are still not fully understood. To address this, we acclimated deer mice to long or short photoperiods (12h or 8h light), in either thermoneutral or cold conditions (30°C or 5°C). To simulate the fall conditions in their natural habitat (Nebraska) we gradually reduced either daylength, temperature, or both over 4 weeks and acclimated mice an additional 4 weeks at those conditions. After the 8 weeks of acclimation, we determined NST and V̇O2max. We found that cold and short photoperiod were necessary to increase NST, and either short photoperiod or cold was sufficient to increase V̇O2max. There were no corresponding differences in iBAT mass, lipid droplet morphology, nor in mitochondrial content. However, there was a corresponding increase in UCP1 content per unit mitochondria. These data highlight the importance of both photoperiod and temperature as cues to prepare thermogenic responses beneficial as winter approaches. / Thesis / Master of Science (MSc)

Fire occurrence and behavior were determined by collecting and analyzing fuel , weather , and fire history ii data. Fuel plots were used to measure average fuel loading by vegetation type and integrated with weather to make worst - case fire behavior prediction s . A fire history was developed using oakbrush (Quercus qaffibelli Nutt . ) sprouts to determine age and the Global Positioning System (GPS) for mapping the burned areas . Average fuel loading was highest in the oakbrush fuel type with 16.8 t/ha , then juniper (Juniperus ost eosoerma Torrey ) with 6 . 72 t/ha , and the lowest was in sagebrush (Artemisia triden~at a Nutt. ) with 4 . 93 t/ha . Fire behavior predictions were similar for all fuel types. The fire rotation for the study area was calculated to be 30 years. The fire history showed the most hectares burned were in the oakbrush fuel type due to fuel loading and horizontal continuity. Prescribed burns and negative fuel breaks were suggested as management alternatives. The effect of fire on deer mouse (Peromyscus maniculatus) density in oakbrush was determined by using a trapping web design with distance sampling techniques . Webs were set in four pairs with one web of each pair being in 7- year- old burned oakbrush and the other web in unburned oakbrush. Variables such as shrub height and litter depth were recorded in order to reduce variance . Trapping occurred in June 1994 with each web set for two consecutive nights using 80 Museum Special snap traps spaced 6 m apart on eight lines . Density estimates were determined by using a computer program called DI STANCE and then analyzed using analysis of variance with a randomized block design . No significant differences between deer mouse densities were detected between burned and unburned oakbrush. Although litter depth and shrub height were both significantly less in burned sites, it did not affect deer mouse density . The conclusion from these result s was that after 7 years oakbrush had recovered to a point that the effect of fire on deer mouse density was negligible.

Hypoxia encountered at high altitude (HA) can limit energy production via aerobic metabolism in animals. Carbohydrate oxidation (CHO) has a greater ATP yield/mole O2 than fat oxidation, and HA-native deer mice show an increased reliance on CHO during submaximal exercise after hypoxia acclimation as an O2-saving strategy. However, hypoxia acclimation does not increase glycolytic capacity in muscle. We therefore tested the hypothesis that altered metabolic regulation of the CHO pathway allows HA mice to achieve higher rates of CHO during submaximal exercise. The objective of our study was to identify the effects of hypoxia acclimation on the regulation of two key proteins in the CHO pathway and their activation with exercise. Using first generation (G1) laboratory born and raised HA deer mice acclimated to normoxia or chronic hypoxia, we examined the metabolic regulation of muscle glucose uptake by glucose transporter (GLUT) 4 and of pyruvate oxidation by pyruvate dehydrogenase (PDH). The gastrocnemius was electrically stimulated in situ under anaesthesia and acute normoxia at two submaximal workloads relative to maximal force production, which was measured using a force transducer. In frozen gastrocnemius following stimulation or rest, GLUT4 protein content was measured via Western blotting of the sarcolemmal membrane fraction and PDH activity was measured using a radiolabelled assay. We found no differences in sarcolemmal GLUT4 content with stimulation, but PDH activity was increased in hypoxia, indicating increased rates of carbohydrate breakdown at similar workloads after acclimation. These data were compared to data from wild HA deer mice sampled at their native altitude. In support of our hypothesis, these data show that the metabolic regulation of the carbohydrate oxidative pathway changes with acclimation to support higher CHO rates during submaximal exercise. These data will help uncover the mechanistic underpinnings responsible for the exercise fuel use strategies observed exclusively in HA-native mice. / Thesis / Master of Science (MSc) / At high altitude, oxygen availability is low and can be challenging for active animals. Preferential carbohydrate oxidation is a metabolic strategy used by high altitude-native deer mice to fuel exercise because of its high energy yield per oxygen consumed. Despite the increase in carbohydrate breakdown, the capacity for muscles to use carbohydrates did not change, suggesting that the regulation of this metabolic pathway may be changing instead. We measured the contributions of two proteins involved in carbohydrate metabolism in active muscle, pyruvate dehydrogenase (PDH) and glucose transporter 4 (GLUT4), at different muscle workloads and after acclimation to high altitude conditions. We found no differences in GLUT4 content, but PDH activity was higher in hypoxia-acclimated mice at similar intensities, indicating increased rates of carbohydrate breakdown after acclimation. These data suggest that the regulation of the carbohydrate metabolic pathway changes with acclimation to support higher rates of carbohydrate oxidation during exercise.

Cryptosporidium is a protozoan parasite infecting the intestines of its hosts, leading to acute diarrheal disease. Out of 26 recognized species, 14 are known to infect humans. Of most importance, from a human perspective are Cryptosporidium parvum and Cryptosporidium hominis, of which the former is known to have zoonotic potential. Globally, cryptosporidiosis affect people with lowered immune status particularly hard; among children under five it is the most important parasitic cause of gastroenteritis. In the region of KwaZulu-Natal, on the east coast of South Africa, Cryptosporidium is considered endemic. Drinking water is frequently collected from river systems and as Cryptosporidium spp. can be transmitted via contaminated water, this may be one source of infection. Research on the species distribution is important for outbreak investigations and prevention efforts. In water and wastewater such speciation is commonly performed using immunomagnetic separation, an antibody dependent method. There is however a suspicion that these antibodies have less affinity to some species and hence contorts the detected species distribution. An alternative approach is therefore of interest.   In the present study, Cryptosporidium diversity in wastewater collected from four different wastewater treatment plants in KwaZulu-Natal, is evaluated with an optimized antibody-free workflow and a single cell platform. It was shown that the workflow is suitable for complex samples, such as wastewater. Furthermore, diversity was assessed with amplicon sequencing, revealing four different species and genotypes. Further modifications of the methods used could benefit the field of Cryptosporidium research, along with improving global health and preventing disease outbreaks.

Obsessive–compulsive disorder (OCD) is a psychiatric condition that is characterized by two main symptom cohorts, namely recurrent inappropriate thoughts (obsessions) and seemingly purposeless repetitive motor actions (compulsions). In 70% of cases, the condition only re–sponds to chronic, but not sub–chronic, high dose treatment with the selective serotonin reup–take inhibitors (SSRIs), such as fluoxetine and escitalopram. This indicates a role for hyposero–tonergic functioning in the primary brain areas involved in OCD, namely the components of the cortico–striatal–thalamic–cortical (CSTC) circuit which include the prefrontal cortex, the basal ganglia, and the thalamus. A number of studies have demonstrated a lower serotonin trans–porter (SERT) availability in OCD patients compared with healthy controls, supporting the hy–pothesis of a hyposerotonergic state in OCD. The current study focuses on the validation of the deer mouse (Peromyscus maniculatus bairdii) model of OCD and builds on previous work done in our laboratory. Deer mice that are bred and housed in confinement naturally develop two main forms of stereotypical behaviour, namely vertical jumping and pattern running. Furthermore, these behaviours can be catego–rized into various levels of severity, namely high (HSB), low (LSB) and non–stereotypic (NSB) cohorts. The seemingly purposeless and repetitive nature of these behaviours mimics the com–pulsions that characterize human OCD and constitutes the basis for the face validity of the model. However, although these two forms of stereotypy seem equally repetitive and persis–tent, stereotypical pattern runners do not complete the required number of cage revolutions per 30 minutes compared to the amount of jumps executed by stereotypical vertical jumpers. As only one set of criteria for the appraisal of the different topographies of deer mouse stereotypy has been applied in previous studies, the matter of whether pattern runners do in fact generate stereotypical behaviour of the same persistent and severe nature as opposed to the behaviour expressed by vertical jumpers, is problematic. Therefore, the first objective of the current study was to develop a new classification system for the appraisal of the different forms of behavioural topographies of deer mice and subse–quently to evaluate whether pattern runners can indeed be categorized into non–, low– and high stereotypical cohorts. After an eight–week behavioural assessment period, deer mice express–ing the two different behavioural topographies could be classified into non–, low– and high stereotypical cohorts (NSB, LSB, and HSB respectively), applying different criteria for each be–havioural topography. Based on the weekly mean stereotypy count generated during three 30–minute intervals of highest stereotypical behaviour over the course of a 12–hour assessment period, HSB pattern runners were found to execute on average 296 cage revolutions per 30 minutes, while HSB vertical jumpers executed an average of 3063 jumps per 30 minutes. This discrepancy between the generated numbers of the different topographies of stereotypy indi–cates that one classification system for the appraisal of both behavioural topographies is indeed inappropriate, and hence requires re–evaluation and validation. As patients with OCD present with a lower central SERT availability compared to healthy controls, the second objective of the study was to determine whether a decrease in SERT den–sity could be demonstrated in HSB animals compared to the NSB and LSB controls. After eight weeks of behavioural assessment, animals were sacrificed and frontal–cortical and striatal SERT binding was performed. HSB deer mice presented with significantly lower striatal, but not fron–tal–cortical SERT availability compared to the [NSB/LSB] control animals (p = 0.0009). As far as it concerns a lower SERT availability in HSB animals and involvement of the CSTC circuitry, this data is congruent with that demonstrated in human OCD and strengthens the construct validity of the model. Although previous studies undertaken in our laboratory demonstrated that deer mouse stereotypy is attenuated after chronic (21–day) fluoxetine administration, OCD only responds to chronic, but not sub–chronic treatment with the SSRIs. The lack of response of deer mouse stereotypy to sub–chronic treatment has not been established and therefore the third study ob–jective was to assess the behavioural effects of sub–chronic (7–day) and chronic (28–day) SSRI treatment on expression of deer mouse stereotypy. Chronic, but not sub–chronic treatment with oral escitalopram (50 mg/kg/day) significantly increased the number of intervals over a 12–hour assessment period during which no stereotypical behaviour were expressed by HSB deer mice (p = 0.0241) and decreased the number of intervals during which high–stereotypical be–haviour were executed (p = 0.0054). Neither chronic, nor sub–chronic treatment significantly affected the behaviour of animals in the [NSB/LSB] cohort. The fact that the model demon–strates a lack of response to sub–chronic treatment with high dose SSRIs, positively contributes to the predictive validity of the deer mouse model of OCD. The results from the current study therefore strengthens the construct and predictive valid–ity of the deer mouse model of OCD and confirm the model’s status as a prominent animal model of OCD. Not only is hyposerotonergic functioning in the CSTC circuitry implicated in the behaviour of HSB animals, but the model also demonstrates selective response to chronic SSRI–treatment - two core characteristics of human OCD. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2012.

Obsessive–compulsive disorder (OCD) is a psychiatric condition that is characterized by two main symptom cohorts, namely recurrent inappropriate thoughts (obsessions) and seemingly purposeless repetitive motor actions (compulsions). In 70% of cases, the condition only re–sponds to chronic, but not sub–chronic, high dose treatment with the selective serotonin reup–take inhibitors (SSRIs), such as fluoxetine and escitalopram. This indicates a role for hyposero–tonergic functioning in the primary brain areas involved in OCD, namely the components of the cortico–striatal–thalamic–cortical (CSTC) circuit which include the prefrontal cortex, the basal ganglia, and the thalamus. A number of studies have demonstrated a lower serotonin trans–porter (SERT) availability in OCD patients compared with healthy controls, supporting the hy–pothesis of a hyposerotonergic state in OCD. The current study focuses on the validation of the deer mouse (Peromyscus maniculatus bairdii) model of OCD and builds on previous work done in our laboratory. Deer mice that are bred and housed in confinement naturally develop two main forms of stereotypical behaviour, namely vertical jumping and pattern running. Furthermore, these behaviours can be catego–rized into various levels of severity, namely high (HSB), low (LSB) and non–stereotypic (NSB) cohorts. The seemingly purposeless and repetitive nature of these behaviours mimics the com–pulsions that characterize human OCD and constitutes the basis for the face validity of the model. However, although these two forms of stereotypy seem equally repetitive and persis–tent, stereotypical pattern runners do not complete the required number of cage revolutions per 30 minutes compared to the amount of jumps executed by stereotypical vertical jumpers. As only one set of criteria for the appraisal of the different topographies of deer mouse stereotypy has been applied in previous studies, the matter of whether pattern runners do in fact generate stereotypical behaviour of the same persistent and severe nature as opposed to the behaviour expressed by vertical jumpers, is problematic. Therefore, the first objective of the current study was to develop a new classification system for the appraisal of the different forms of behavioural topographies of deer mice and subse–quently to evaluate whether pattern runners can indeed be categorized into non–, low– and high stereotypical cohorts. After an eight–week behavioural assessment period, deer mice express–ing the two different behavioural topographies could be classified into non–, low– and high stereotypical cohorts (NSB, LSB, and HSB respectively), applying different criteria for each be–havioural topography. Based on the weekly mean stereotypy count generated during three 30–minute intervals of highest stereotypical behaviour over the course of a 12–hour assessment period, HSB pattern runners were found to execute on average 296 cage revolutions per 30 minutes, while HSB vertical jumpers executed an average of 3063 jumps per 30 minutes. This discrepancy between the generated numbers of the different topographies of stereotypy indi–cates that one classification system for the appraisal of both behavioural topographies is indeed inappropriate, and hence requires re–evaluation and validation. As patients with OCD present with a lower central SERT availability compared to healthy controls, the second objective of the study was to determine whether a decrease in SERT den–sity could be demonstrated in HSB animals compared to the NSB and LSB controls. After eight weeks of behavioural assessment, animals were sacrificed and frontal–cortical and striatal SERT binding was performed. HSB deer mice presented with significantly lower striatal, but not fron–tal–cortical SERT availability compared to the [NSB/LSB] control animals (p = 0.0009). As far as it concerns a lower SERT availability in HSB animals and involvement of the CSTC circuitry, this data is congruent with that demonstrated in human OCD and strengthens the construct validity of the model. Although previous studies undertaken in our laboratory demonstrated that deer mouse stereotypy is attenuated after chronic (21–day) fluoxetine administration, OCD only responds to chronic, but not sub–chronic treatment with the SSRIs. The lack of response of deer mouse stereotypy to sub–chronic treatment has not been established and therefore the third study ob–jective was to assess the behavioural effects of sub–chronic (7–day) and chronic (28–day) SSRI treatment on expression of deer mouse stereotypy. Chronic, but not sub–chronic treatment with oral escitalopram (50 mg/kg/day) significantly increased the number of intervals over a 12–hour assessment period during which no stereotypical behaviour were expressed by HSB deer mice (p = 0.0241) and decreased the number of intervals during which high–stereotypical be–haviour were executed (p = 0.0054). Neither chronic, nor sub–chronic treatment significantly affected the behaviour of animals in the [NSB/LSB] cohort. The fact that the model demon–strates a lack of response to sub–chronic treatment with high dose SSRIs, positively contributes to the predictive validity of the deer mouse model of OCD. The results from the current study therefore strengthens the construct and predictive valid–ity of the deer mouse model of OCD and confirm the model’s status as a prominent animal model of OCD. Not only is hyposerotonergic functioning in the CSTC circuitry implicated in the behaviour of HSB animals, but the model also demonstrates selective response to chronic SSRI–treatment - two core characteristics of human OCD. / Thesis (M.Sc. (Pharmacology))--North-West University, Potchefstroom Campus, 2012.