Road expansion and increased traffic likely exacerbates barriers to amphibian migration and dispersal. Within British Columbia’s south Okanagan valley there is particular concern that the COSEWIC-listed blotched tiger salamander (Ambystoma mavortium melanostictum) and Great Basin spadefoot (Spea intermontana) are vulnerable to road effects in their annual movements from upland overwintering habitat to lowland breeding areas. My study utilizes a before after control impact approach to assess amphibian movement and population threats across this highway-bisected landscape. Throughout the spring and summer of 2010-2012, fifty two kilometers of roadways (31 km of highway, 21 km of paved backroad) were repeatedly surveyed from the Canada-USA border to north of Oliver, BC; surveys were carried out utilising vehicles and on foot. Along Highway 97, a three kilometer four-lane highway expansion project was constructed through 2010 and open to traffic use in 2011. Adjacent to a floodplain, survey effort was focused throughout this transect for informed roadkill mitigation structure placement and ongoing ecopassage effectiveness monitoring. Automated camera trap monitoring of culverts within highly concentrated amphibian road hotspots during spring and summer 2011 (three culverts) and 2012 (two culverts) resulted in over eight hundred amphibian culvert events observed. Two sample Wilcoxon tests revealed differences between years in amphibian occurrence between 2010 and 2012 (W = 4679.5, p= 0.02), and mortalities among transect areas, with the largest differences between years within the Osoyoos passing lanes transect. Amphibian mortalities within the passing lanes transect were significantly reduced with the implementation of mitigation structures (x̅2010= 13.2 ± 32.5, x̅2011= 4.7 ± 12.8, x̅2012= 2.3 ± 7.3; 2010 vs. 2012: W= 1535.5, p< 0.001). Roadkill mitigation structures proved effective in observed amphibian occurrence of the entire passing lanes stretch as well as at distances 100 m and 200 m from observed culverts. Double fenced areas resulted in a 94% reduction in amphibian road occurrence. Five species of amphibians were observed over the three survey years (4051 road incidences over 657 survey hours): Pacific chorus frog (Pseudacris regilla), Western toad (Anaxyrus boreas), long-toed salamander (Ambystoma macrodactylum) plus blotched tiger salamander and Great Basin spadefoot. This study aims to provide a better understanding of amphibian hotspots on roadways and ecopassage use within the south Okanagan. It may act as a catalyst to further wildlife-vehicle interaction studies with improved mitigation solutions for amphibian roadway fatalities.

The sudden appearance of chytridiomycosis, as the cause of amphibian deaths and population declines in several continents suggests that its etiological agent, the amphibian chytrid fungus Batrachochytrium dendrobatidis, was introduced into the affected regions. However, the origin of this virulent pathogen is unknown. Efforts were directed to determine the occurrence of chytridiomycosis in Africa, whether the disease had been introduced into South Africa in recent years and how wild frog populations were affected by infection. A chytridiomycosis survey of 2,300 archived and live specimens involving members of the Pipidae family in sub-Saharan Africa, as well as a number of unrelated frog species in South Africa was conducted by histological diagnosis of skin samples. The epidemiological evidence indicated that chytridiomycosis has been a stable endemic infection in southern Africa for 23 years before any positive specimens were found outside Africa. The occurrence of chytridiomycosis in South Africa can be described as widespread both in terms of geographical distribution and host species and generally infection is not associated with adverse effects at the individual or population level. It was proposed that the amphibian chytrid originated in Africa and that the international trade in the African clawed toad Xenopus laevis that commenced in the mid 1930s was the means of dissemination. A risk assessment of the X. laevis trade demonstrated that chytridiomycosis could spread through this pathway and culminated in the development of a management protocol to reduce the risks of spreading disease through this animate commodity. Initial comparative genetic analysis of B. dendrobatidis strains isolated from South African frogs with a global set of 35 strains, suggests that analysis of a more geographically diverse set of southern African strains is needed before this line of argument can support or reject the "out of Africa" hypothesis. / Thesis (Ph.D. (Zoology))--North-West University, Potchefstroom Campus, 2005.

Glyphosate-based herbicides are the dominant pesticide on the market and are utilized worldwide in both the agricultural and forestry industries. Their prevalence comes at a time when concern over the potential effects of pesticide application in amphibian spawning grounds is growing. The primary goal of this thesis was to determine if the glyphosate-based herbicide WeatherMax® has the potential to disrupt sexual differentiation and growth in the wood frog (Lithobates sylvaticus) in a pulse exposure at the predicted maximal environmental concentration (PMEC) of 2.88 mg acid equivalent per liter. This was carried out in laboratory, mesocosm and in-situ field exposures, in an attempt to determine how a potential disruption might vary between experimental environments. In this study, tadpoles from three split-wetlands targeted at the PMEC for WeatherMax were found to display no significant change in survival or growth, however gene expression of several genes involved in steroidogenesis during sexual differentiation (cyp19, cyp17, star, foxl2) were found to be affected. The effects on these genes appeared to be dependant on the exposure concentration of WeatherMax in each wetland, which varied even though all three wetlands were meant to target the PMEC. The wetland that was measured as having the highest herbicide concentration (PMEC 13) was found to have a female biased sex ratio. The results found in the field varied from those found in the more artificial exposures. In the laboratory the PMEC of WeatherMax experienced complete mortality, whereas in the mesocosms survival was not significantly affected. Sex ratios were unaffected in the laboratory, however at the PMEC there was a significant male bias in the mesocosms. The discrepancies in the results obtained from the different exposure types highlights the importance of real world exposures. That the same concentration that caused complete mortality in the laboratory caused sublethal effects in the field is of importance as it denotes that these endpoints may not be easily investigated in these synthetic exposures. This project is a part of the Long Term Experimental Wetlands Area (LEWA) and contributes to the body of information amassed therein on the impact of a glyphosate-based herbicide on amphibians in a wetland ecosystem.

Nanoparticles (NPs) are engineered in the nanoscale (<100nm) to have unique physico-chemical properties from their bulk counterparts. Nanosilver (nAg) is the most prevalent nanoparticle in consumer products due to its strong antimicrobial action and can be released to the environment during product manufacture, usage and disposal. The predicted environmental concentrations are within the North American guidelines for the protection of aquatic life and in drinking water. While nAg toxicity at high concentrations has been well described, the sublethal effects at environmentally-relevant concentrations are relatively unknown. Initial screening in our lab showed nAg was a potential endocrine disrupting chemical (EDC). Amphibian metamorphosis is mediated by thyroid hormone (TH), and nAg perturbed TH-dependent transcriptional responses in the tailfin of bullfrog (Rana catesbeiana) tadpoles. The primary objective of this thesis was to further investigate and characterize the effects of low, environmentally relevant concentrations of nAg on TH-dependent metamorphosis in R. catesbeiana and Xenopus laevis. Two chronic, 28 day in vivo exposures at 0.06 and 6µg/L nAg were conducted with premetamorphic R. catesbeiana tadpoles using TH to induce precocious metamorphosis. Ionic silver (iAg) was also examined to control for the complete dissolution of Ag. Analysis of metamorphic stage progression demonstrated nAg-induced acceleration of hindlimb growth and development. After 6 days of nAg exposure, analysis with quantitative real-time polymerase chain reaction (QPCR) demonstrated nAg-induced disruption of TH-responsive transcripts in a tissue-specific manner. Furthermore, the nAg effects could not be fully explained by iAg, indicating NP-specific disruption. Two chronic, 28 day exposures to 0.018-1.8 µg/L nAg were conducted on X. laevis premetamorphic and prometamorphic tadpoles. nAg was found to significantly bioaccumulate in tadpole tissue after 28 days. Furthermore, nAg increased the hindlimb length during early premetamorphosis and in post-metamorphic juvenile tadpoles. Using an in-house MAGEX microarray and QPCR transcriptional analysis, 7 biomarkers of nAg exposure were validated. Five of these targets showed disruption to their TH-response. Furthermore, the increased mRNA abundance of two peroxidases indicated that nAg generated reactive oxygen species (ROS) even at low, environmental concentrations. This thesis demonstrates that nAg has consistent EDC actions across two distinct amphibian species, and the data suggest that regulatory guidelines for silver may need revision. A X. laevis derived fibroblast-like TH-responsive cell line, XTC-2, was used in conjunction with the 7 biomarkers of nAg exposure to gain mechanistic insight into the role of ROS in TH signaling disruption. Monocultures were created and validated to increase the specificity of TH-response. While the monocultures were successfully created, the biomarkers were not responsive to nAg in this cell line. Additional investigations were made into the relationship between genetic sex and responsiveness to TH. Genetic sexing methods were used to investigate transcriptional differences between males and females during natural and TH-induced metamorphosis. The sexing protocol was optimized and validated successfully. The genetic sex was determined for premetamorphic and prometamorphic X. laevis tadpoles exposed to TH for 48 h. QPCR and microarray analysis were used to identify three markers that demonstrated transcriptional sex-bias during early gonadal differentiation stages. / Graduate / 0307 / 0383 / 0487 / amanda_carew14@yahoo.ca

This study examined the effects of temperature and chronic hypoxia (CH) on pH/CO2- sensitive fictive breathing, and central pH/CO2 chemosensitivity, in cane toads (Bufo marinus). Toads were exposed to CH (10% or 15% O2) or control conditions (21% O2) for 10 days at either room temperature (controls), 10°C or 30°C following which in vitro brainstem-spinal cord preparations were used to examine central pH/CO2-sensitive fictive breathing (i.e., motor output from respiratory nerves which is the neural correlate of breathing). A reduction in artificial cerebral spinal fluid (aCSF) pH increased fictive breathing frequency (fR) and total fictive ventilation (TFV). Cold temperature reduced and hot temperature increased fR and TFV under control conditions. CH attenuated fictive breathing independently of temperature. Additional experiments in which the aCSF temperature was varied indicate that the effects of temperature acclimation result from neural plastic changes within respiratory control centres in the brain.

I investigated the value of under-road tunnels as a conservation strategy to protect a long-toed salamander population, in south-west Alberta, whose overwintering sites and breeding habitat (Linnet Lake) are separated by a road. I conducted a mark-recapture study from 2008-2009, capturing salamanders using roadside fences and pitfall traps. Four tunnels were monitored in 2009 using traps and cameras. A 2008 estimate indicated that the population declined by 60% since 1994, however, road mortality was dramatically reduced following installation of fences and tunnels. Camera and trap data documented 130 salamanders navigating tunnels in 2009. I found little evidence of juvenile recruitment from Linnet Lake, likely because of predation by lake chub. Experiments showed that lake chub consumed salamander larvae, and fish presence altered larval behaviour. Continued monitoring is needed to determine if reduced road mortality translates into population gains, and whether fish predation threatens the persistence of the long-toed salamander population. / Ecology

Amphibian populations are declining on a global scale and although disease outbreaks are a commonly accepted hypothesis they are not the only one. My aims for my thesis were to study the home range of Leiopelma hamiltoni, to determine whether a photographic database could be used to individual identified them and monitor the health status of the population. Habitat loss is a possible cause. For this reason monitoring an animals' home range is a possible method to detect early impacts the population is facing. By tracking 12 L. hamiltoni within a 12 m x 6 m grid on Maud Island, it was shown that the home range size can vary from 0.5 m2 to 25 m2 based on the minimum convex polygon method. However, to track multiple individuals it is important to be able to distinguish among frogs. The commonly used methods of identification, such as toe clipping, pose potentially detrimental effects. Therefore, non-invasive methods based on natural markings need to be established. Through the use of the dark pigmented patterns found on the skin of L. hamiltoni individuals can be identified on recapture with a mean accuracy of 93%. By developing a database to maintain the photographs used for individual identification, the database can also be used to monitor the status of the population. During 2003 numerous L. hamiltoni were observed with denuded patches predominantly on the facial region. By monitoring five individuals within the captive facility at the University of Canterbury it was discovered that frogs appear to be able to cure themselves. Through researching the home range requirements and developing a photographic database to monitor the population status of L. hamiltoni, it will aid in the management of ensuring the long-term survival of this archaic species of frog.

Thesis (Ph. D.)--Oregon State University, 2009. / Printout. Includes bibliographical references (leaves 95-104). Also available on the World Wide Web.

Thesis (Ph. D.)--Ohio State University, 2009. / Title from first page of PDF file. Includes vita. Includes bibliographical references (p. 82-92).

abstract: Infectious diseases have emerged as a significant threat to wildlife. Environmental change is often implicated as an underlying factor driving this emergence. With this recent rise in disease emergence and the acceleration of environmental change, it is important to identify the environmental factors that alter host-pathogen dynamics and their underlying mechanisms. The emerging pathogen Batrachochytrium dendrobatidis (Bd) is a clear example of the negative effects infectious diseases can have on wildlife. Bd is linked to global declines in amphibian diversity and abundance. However, there is considerable variation in population-level responses to Bd, with some hosts experiencing marked declines while others persist. Environmental factors may play a role in this variation. This research used populations of pond-breeding chorus frogs (Pseudacris maculata) in Arizona to test if three rapidly changing environmental factors nitrogen (N), phosphorus (P), and temperature influence the presence, prevalence, and severity of Bd infections. I evaluated the reliability of a new technique for detecting Bd in water samples and combined this technique with animal sampling to monitor Bd in wild chorus frogs. Monitoring from 20 frog populations found high Bd presence and prevalence during breeding. A laboratory experiment found 85% adult mortality as a result of Bd infection; however, estimated chorus frog densities in wild populations increased significantly over two years of sampling despite high Bd prevalence. Presence, prevalence, and severity of Bd infections were not correlated with aqueous concentrations of N or P. There was, however, support for an annual temperature-induced reduction in Bd prevalence in newly metamorphosed larvae. A simple mathematical model suggests that this annual temperature-induced reduction of Bd infections in larvae in combination with rapid host maturation may help chorus frog populations persist despite high adult mortality. These results demonstrate that Bd can persist across a wide range of environmental conditions, providing little support for the influence of N and P on Bd dynamics, and show that water temperature may play an important role in altering Bd dynamics, enabling chorus frogs to persist with this pathogen. These findings demonstrate the importance of environmental context and host life history for the outcome of host-pathogen interactions. / Dissertation/Thesis / Ph.D. Biology 2012