Investigating the Symbiotic Fungi and Bacteria on the Skin of Colorado Boreal Toads (Anaxyrus boreas boreas) and Their Interactions with The Amphibian Pathogen Batrachochytrium dendrobatidis

Fungal emerging infectious diseases (EID&rsquo;s) can affect a broad range of host species, have rapid global dispersal, are fast evolving, and can be highly lethal to hosts. Batrachochytrium dendrobatidis (Bd) is a fungal EID that threatens amphibians globally. One affected species is the Colorado boreal toad (Anaxyrus boreas boreas), a high-elevation, long-lived toad that is locally endangered in Colorado due to the Bd pandemic. The fungal pathogen infects and destroys amphibian skin cells. The skin environment of amphibians involves a mucous matrix rich with symbiotic microbes and past studies have found that Bd interacts with amphibian skin microbes in ways that affect the trajectory of disease. A single-strain anti-Bd probiotic, Janthinobacterium lividum, is currently being field tested in wild Colorado boreal toads, but large knowledge gaps remain pertaining to other microbial domains and microbe-microbe interactions that affect the host in context of disease. My first chapter characterizes the diversity of the host-associated fungal community on boreal toad skin, across the host&rsquo;s different life stages. I found that there is a diverse fungal community on boreal toad skin that changes over the course of the host&rsquo;s life stages, and I identified eight putatively host-associated fungi. In my second chapter, I investigated whether the fungal community harbors Bd-inhibitory strains and how host-associated fungal strains respond to Bd. Five out of the 16 tested fungi were Bd-inhibitive. In addition, the growth patterns of some isolates were affected by Bd, implying that fungi-pathogen interactions are also important. Lastly, my third chapter screened over 100 boreal toad skin bacteria for Bd-inhibition and tested whether multi-strain probiotic cocktails are stronger Bd inhibitors than single strains. I found very few multi-strain combinations that worked better than their single-strain components. Probiotic applications remain a strong tool against fungal EID&rsquo;s, but such therapies cannot succeed without knowledge about the whole microbial community, across domains and including multiple species&rsquo; interactions. My findings contribute to this knowledge gap by characterizing a previously unstudied domain of life in this host and investigating the anti-Bd functions in the bacterial community in the context of single- and multi-strain probiotic targets.</p

Bacterial Community Sequences of Submerged Aquatic Vegetation in the Potomac River.

Here, we report results from PCR and sequencing of bacterial 16S rRNA genes from leaf and root surfaces from nine submerged aquatic vegetation (SAV) samples comprising five species. Samples were from four sites along the Potomac River

Identifying fungal-host associations in an amphibian host system

Host-associated microbes can interact with macro-organisms in a number of ways that affect host health. Few studies of host-associated microbiomes, however, focus on fungi. In addition, it is difficult to discern whether a fungal organism found in or on an ectotherm host is associating with it in a durable, symbiotic interaction versus a transient one, and to what extent the habitat and host share microbes. We seek to identify these host-microbe interactions on an amphibian, the Colorado boreal toad (Anaxyrus boreas boreas). We sequenced the ITS1 region of the fungal community on the skin of wild toads (n = 124) from four sites in the Colorado Rocky Mountains, across its physiologically dynamic developmental life stages. We also sampled the common habitats used by boreal toads: water from their natal wetland and aquatic pond sediment. We then examined diversity patterns within different life stages, between host and habitat, and identified fungal taxa that could be putatively host-associated with toads by using an indicator species analysis on toad versus environmental samples. Host and habitat were strikingly similar, with the exception of toad eggs. Post-hatching toad life stages were distinct in their various fungal diversity measures. We identified eight fungal taxa that were significantly associated with eggs, but no other fungal taxa were associated with other toad life stages compared with their environmental habitat. This suggests that although pre- and post-metamorphic toad life stages differ from each other, the habitat and host fungal communities are so similar that identifying obligate host symbionts is difficult with the techniques used here. This approach does, however, leverage sequence data from host and habitat samples to predict which microbial taxa are host-associated versus transient microbes, thereby condensing a large set of sequence data into a smaller list of potential targets for further consideration. &nbsp;</p

MOLECULAR VIROLOGICAL ANALYSIS OF THE TRANSMISSION CLUSTERS AND RESISTANCE MUTATIONS OF HIV-1 SUBTYPE B IN BULGARIA (2012-2020)

HIV-1 infection in Bulgaria is known for its high level of genetic diversity. Previous studies have indicated that subtype B is the most common strain in Bulgaria, particularly among men who have sex with men, who are at a high risk of transmission. The primary objective of this study was to identify any transmission clusters and transmission resistance in individuals newly diagnosed with HIV-1 who have not yet received antiretroviral therapy (ART). To this end, we sequenced the HIV-1 pol gene in the samples from the study participants using either the Viroseq HIV-1 Genotyping Test (Abbott) and the Applied Biosystems 3130xl genetic analyzer or the TruGene DNA Sequencing System (Siemens Healthcare) and an OpenGene DNA sequencing system. We then subtyped the HIV-1 pol sequences, and further analyzed those that met the criteria for subtype B. The study included a total of 595 HIV-1 subtype B sequences. Our analysis revealed that the majority of those diagnosed with HIV-1 subtype B were male and lived in Sofia region. The most common transmission mode was through sexual intercourse among men who have sex with men, followed by heterosexual transmission. We also observed the presence of multiple transmission clusters , and a low percentage of transmitted drug resistance mutations (TDRMs). Overall, our study confirms that HIV-1 subtype B remains the most dominant strain in Bulgaria

Structured Operational Semantics for Graph Rewriting

Process calculi and graph transformation systems provide models of reactive systems with labelled transition semantics. While the semantics for process calculi is compositional, this is not the case for graph transformation systems, in general. Hence, the goal of this article is to obtain a compositional semantics for graph transformation system in analogy to the structural operational semantics (SOS) for Milner's Calculus of Communicating Systems (CCS). The paper introduces an SOS style axiomatization of the standard labelled transition semantics for graph transformation systems. The first result is its equivalence with the so-called Borrowed Context technique. Unfortunately, the axiomatization is not compositional in the expected manner as no rule captures "internal" communication of sub-systems. The main result states that such a rule is derivable if the given graph transformation system enjoys a certain property, which we call "complementarity of actions". Archetypal examples of such systems are interaction nets. We also discuss problems that arise if "complementarity of actions" is violated.Comment: In Proceedings ICE 2011, arXiv:1108.014