abstract: Epidemiological theory normally does not predict host extinction from infectious disease because of a host density threshold below which pathogens cannot persist. However, host extinction can occur when a biotic or abiotic pathogen reservoir allows for density-independent transmission. Amphibians are facing global population decline and extinction from the emerging infectious disease chytridiomycosis, caused by the fungus Batrachochytrium dentrobatidis (Bd). I use the model species Eleutherodactylus coqui to assess the impact of Bd on terrestrial direct-developing frog species, a common life history in the tropics. I tested the importance of two key factors that might influence this impact and then used laboratory experiments and published field data to model population-level impacts of Bd on E. coqui. First, I assessed the ontogenetic susceptibility of E. coqui by exposing juvenile and adult frogs to the same pathogen strain and dose. Juveniles exposed to Bd had significantly lower survival rates compared with control juveniles, while adult frogs often cleared infection. Second, I conducted experiments to determine whether E. coqui can become infected with Bd indirectly from contact with zoospores shed onto vegetation by an infected frog and from direct exposure to an infected frog. Both types of transmission were observed, making this the first demonstration that amphibians can become infected indirectly in non-aquatic habitats. Third, I tested the hypothesis that artificially-maintained cultures of Bd attenuate in pathogenicity, an effect known for other fungal pathogens. Comparing two cultures of the same Bd strain with different passage histories revealed reduced zoospore production and disease-induced mortality rates for a susceptible frog species (Atelopus zeteki) but not for the less-susceptible E. coqui. Finally, I used a mathematical model to project the population-level impacts of chytridiomycosis on E. coqui. Model analysis showed that indirect transmission, combined with either a high rate of zoospore production or low rate of zoospore mortality, is required for Bd to drive E. coqui populations below an extinction threshold. High rates of transmission plus frequent re-infection could lead to poor recruitment of infected juveniles and population decline. My research adds further insight into how emerging infectious disease is contributing to the loss of amphibian biodiversity. / Dissertation/Thesis / Ph.D. Biology 2013

Este trabalho teve como objetivo propor novo método para estudar a permeabilidade de fármacos, utilizando epitélio intestinal de rã-touro (Rana catesbeiana) em método ex vivo, empregando células de Franz. Por utilizar epitélio intestinal, um material de descarte proveniente de um animal utilizado como alimento humano, pode ser considerado um método alternativo, pois não implica no sacrifício de animais. A quantidade de fármaco permeada foi determinada por método de eletroforese capilar com detecção ultravioleta e validado para os antivirais lamivudina, zidovudina e aciclovir, na presença de metoprolol e floridizina. O fármaco escolhido como modelo nos ensaios de permeabilidade foi a lamivudina. Para estabelecimento do protocolo experimental dos estudos de permeabilidade, foi proposta uma análise de variância three-way para verificar a influência na permeabilidade dos fármacos, das seguintes variáveis: secção intestinal, pH da solução de Ringer e temperatura. Foram determinados a quantidade total de fármaco permeado (Qt), o coeficiente de permeabilidade aparente (Papp) e a constante de absorção de primeira ordem (ka). A partir da análise do planejamento experimental, os efeitos das variáveis não foram significativos, exceto para a secção intestinal. Os resultados de coeficiente de permeabilidade aparente (Papp) obtidos foram de 0,09 x 10-4 cm/s para lamivudina e de 0,22 x 10-4 cm/s para o metoprolol. O valor de Papp obtido de para o metoprolol é próximo dos valores encontrados na literatura para outras técnicas. Para a lamivudina, entretanto, a diferença encontrada em comparação às células Caco-2 pode ser devida às diferentes técnicas empregadas. / This work aimed to propose a new method for studying drug permeability using frog intestinal epithelium (Rana catesbeiana) in ex vivo method, using Franz cells. By using intestinal epithelium, a disposal material from an animal used as human food, can be considered an alternative method, because it doesn\'t involve the sacrifice of animals. The amount of permeated drug was determined by capillary electrophoresis method with UV detection and validated for antiviral drugs lamivudine, zidovudine and acyclovir in the presence of metoprolol and floridizina. The drug chosen as a model in permeability studies was lamivudine. To establish the experimental protocol for the permeability studies, a three-way analysis of variance was proposed to check the influence of intestinal section, pH of Ringer\'s solution and the temperature on the permeability. Total amount of drug permeated (Qt), apparent permeability coefficient (Papp) and first-order constant absorption (ka) were determined. By the analysis of experimental design, the effects of the variables were not significant, except for intestinal section. The results of apparent permeability coefficient (Papp) obtained were 0.09 x 10-4 cm/s for lamivudine and 0.22 x 10-4 cm/s for metoprolol. The value of Papp obtained for metoprolol is quite close to the values found in literature for other methods. For lamivudine, however, the difference found in comparison to Caco-2 cells may be due to different techniques.

The cross-situational word learning paradigm argues that word meanings can be approximated by word-object associations, computed from co-occurrence statistics between words and entities in the world. Lexicon acquisition involves simultaneously guessing (1) which objects are being talked about (the ”meaning”) and (2) which words relate to those objects. However, most modeling work focuses on acquiring meanings for isolated words, largely neglecting relationships between words or physical entities, which can play an important role in learning. Semantic parsing, on the other hand, aims to learn a mapping between entire utterances and compositional meaning representations where such relations are central. The focus is the mapping between meaning and words, while utterance meanings are treated as observed quantities. Here, we extend the joint inference problem of word learning to account for compositional meanings by incorporating a semantic parsing model for relating utterances to non-linguistic context. Integrating semantic parsing and word learning permits us to explore the impact of word-word and concept-concept relations. The result is a joint-inference problem inherited from the word learning setting where we must simultaneously learn utterance-level and individual word meanings, only now we also contend with the many possible relationships between concepts in the meaning and words in the sentence. To simplify design, we factorize the model into separate modules, one for each of the world, the meaning, and the words, and merge them into a single synchronous grammar for joint inference. There are three main contributions. First, we introduce a novel word learning model and accompanying semantic parser. Second, we produce a corpus which allows us to demonstrate the importance of structure in word learning. Finally, we also present a number of technical innovations required for implementing such a model.

A high proportion of amphibian species are threatened with extinction globally, and habitat loss and degradation are the most frequently implicated causes. Rapid deforestation for the establishment of agricultural production is a primary driver of habitat loss in tropical zones where amphibian diversity is highest. Land-cover change affects native assemblages, in part, through the reduction of habitat area and the reduction of movement among remnant populations. Decreased gene flow contributes to loss of genetic diversity, which limits the ability of local populations to respond to further environmental changes. The focus of this dissertation is on the degree to which common land uses in Sarapiquí, Costa Rica impede the movement of two common amphibian species. First, I used field experiments, including displacement trials, and a behavioral landscape ecology framework to investigate the resistance of pastures to movement of Oophaga pumilio. Results from experiments demonstrate that pastures do impede movement of O. pumilio relative to forest. Microclimatic effects on movement performance as well as limited perceptual ranges likely contribute to reduced return rates through pastures. Next, I linked local processes to landscape scale estimates of resistance. I conducted experiments to measure habitat-specific costs to movement for O. pumilio and Craugastor bransfodrii, and then used experimental results to parameterize connectivity models. Model validation indicated highest support for resistance estimates generated from responses to land-use specific microclimates for both species and to predator encounters for O. pumilio. Finally, I used abundance and experiment-derived resistance estimates to analyze the effects of prevalent land uses on population genetic structure of the two focal species. While O. pumilio did not exhibit a strong response to landscape heterogeneity and was primarily structured by distances among sites, C. bransfordii genetic variation was explained by resistance estimates from abundance and experiment data. Collectivity, this work demonstrates that common land uses can offer different levels of resistance to amphibian movements in Sarapiquí and illustrates the value of investigating local scales processes to inform interpretation of landscape-scale patterns.

The release of chemical transmitter from nerve terminals is critically dependent on a transient increase in intracellular Ca2+.6,25 The increase in Ca2+ may be due to influx of Ca2+ from the extracellular fluid15 or release of Ca2+ from intracellular stores such as mitochondria.1,8,18 Whether Ca2+ utilized in transmitter release is liberated from organelles other than mitochondria is uncertain. Smooth endoplasmic reticulum is known to release Ca2+, e.g., on activation by inositol trisphosphate or cyclic adenosine diphosphate-ribose,2 so the possibility exists that Ca2+ from this source may be involved in the events leading to exocytosis. We examined this hypothesis by testing whether inositol trisphosphate and cyclic adenosine diphosphate-ribose modified transmitter release. We used liposomes to deliver these agents into the cytoplasmic compartment and binomial analysis to determine their effects on the quantal components of transmitter release. Administration of inositol trisphosphate (10-4M) caused a rapid, 25% increase in the number of quanta released. This was due to an increase in the number of functional release sites, as the other quantal parameters were unaffected. The effect was reversed with 40min of wash. Virtually identical results were obtained with cyclic adenosine diphosphate-ribose (10-4M). Inositol trisphosphate caused a 10% increase in quantal size, whereas cyclic adenosine diphosphate-ribose had no effect. The results suggest that quantal transmitter release can be increased by Ca2+ released from smooth endoplasmic reticulum upon stimulation by inositol trisphosphate or cyclic adenosine diphosphate-ribose. This may involve priming of synaptic vesicles at the release sites or mobilization of vesicles to the active zone. Inositol trisphosphate may have an additional action to increase the content of transmitter within the vesicles. These findings raise the possibility of a role of endogenous inositol phosphate and smooth endoplasmic reticulum in the regulation of cytoplasmic Ca2+ and transmitter release.

The greenhouse frog, Eleutherodactylus planirostris, native to Cuba and the Bahamas, was recently introduced to Hawaii. Studies from other invaded habitats suggest that it may impact Hawaiian ecosystems by consuming and potentially reducing endemic invertebrates. However, there have been no studies on the greenhouse frog in Hawaii. The first component of this study was to conduct a diet analysis. We conducted a stomach content analysis of 427 frogs from 10 study sites on the island of Hawaii. At each site, we also collected invertebrates using two different sampling methods: leaf litter collection and sticky traps to characterize available resources. Greenhouse frogs consumed predominantly leaf litter invertebrates. Dominant prey items consisted of Hymenoptera: Formicidae (32.4%), Acari (19.2%), and Collembola (17.4%). Greenhouse frogs consumed more Formicidae than was measured in the environment. At one study site, we estimated there were 12,500 frogs ha-1 using mark-recapture methods and greenhouse frogs consumed 129,000 invertebrates ha-1 night-1 at this site. The second component of this study was to determine the distribution of the greenhouse frog on the island of Hawaii, with a male breeding call presence/absence survey at 446 points along the major road network. The greenhouse frog was detected at 61 sites (14%), and found mostly in lowland areas, in habitats of native shrublands and forests, nonnative forests, agricultural lands, and pastures on the southwestern and eastern sides of the island. We determined detection probabilities of the greenhouse frog and the invasive coqui frog, E. coqui. Detection probability of the greenhouse frog was low on the first two surveys and improved by the third survey. Detection probability of the coqui was higher than the greenhouse frog, but overall site occupancy estimates were similar for both species. Because the greenhouse frog appears to be as widespread as the coqui, we recommend that research be conducted to investigate its impacts ecologically to determine whether control efforts should also be aimed at this species.

This study is a multimodal discourse analysis of how involuntary celibates, commonly known as incels, use memes in their discourse to create a group identity. The study focuses on memes used on the internet forum Incels.is. Through an examination of 100 threads on the forum the study has picked out the 7 most frequently used memes for analysis. The study concludes that the memes allude to themes of humor, anger, hate, fixation with appearance, hopelessness and dejection. These themes appeal to feelings that help create a sense of community and in-group-think, which excludes the “other” and creates an echo-chamber where the group's ideas and opinions aren’t challenged and therefore become more extreme.

<p>Perfluoroalkyl substances are synthetic organic chemicals of environmental concern because they have been associated with adverse effects in both human epidemiological studies and standard laboratory animals. In the environment, PFAS occur as mixtures, especially in areas with a history of PFAS application, such as aqueous film forming foam (AFFF) sites. Among the PFAS, perfluorooctanesulfonic acid (PFOS) and perfluorohexanesulfonic acid (PFHxS) are the most common, and occur at the highest concentrations. Thus, amphibian populations at or near AFFF sites are at risk of exposure to known bioaccumulative and persistent chemicals, likely compromising the physiology and body condition of the animals. Here, we exposed northern leopard frogs to environmentally relevant concentrations of 0.5 and 1 ppb PFOS and PFHxS, alone or as a mixture comprised of 0.5 ppb PFOS and 0.5 ppb PFHxS. Univariate analyses showed that in the larval stages, tadpoles exposed to PFAS had significantly reduced scaled mass indexes (SMI’s) relative to the control, and only the organisms exposed to PFHxS 0.5 ppb were significantly larger. Sex did not significantly influence toxicity in the later stages (GS 42 & 46), indicating no sex-related effects. Altered body condition (i.e., fat stores) in the larval stages indicate potential effects to energy balance. There is a need to assess fitness-related effects as amphibians’ transition into the terrestrial environment, and include endpoints such as: reproductive, developmental, immunological, mating, feeding, competition, and survival. Early developmental effects in the larval stages also suggests that earlier developmental endpoints may be of interest. Establishing ecological risk assessments for PFAS are necessary, as they are toxic, persistent, and bioaccumulative.</p>

The Mississippi gopher frog (MGF, Lithobates sevosa) is a critically endangered amphibian which requires the use of assisted reproductive technologies (ART) to successfully reproduce in captivity. Several challenges to successfully breeding MGFs were investigated to improve reproduction in this species. Accurate identification of biological sex is a necessary first step to reproducing species, thus several non-invasive techniques for sex identification were compared in the weakly dimorphic MGF. Reliably obtaining eggs from female MGFs has also proved difficult, therefore, four exogenous hormone therapies were compared for their success to induce ovulation. Ultrasound imaging was used alongside hormone treatments to improve understanding of ovarian response to hormone treatment. Lastly, short-term cold storage of spermic urine was studied to determine the feasibility of this technique for MGF gamete storage. Findings from these research projects are important for the successful conservation of the MGF by aiding in the production offspring for future reintroduction programs.

No description available.