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Browsing by Author "Eisen, Rebecca J., committee member"

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    Identifying blood meals in cat fleas (Ctenocephalides felis) from a plague-endemic region of Uganda using a SYBR Green real-time polymerase chain reaction-based assay
    (Colorado State University. Libraries, 2012) Graham, Christine B., author; Black, William C., advisor; Eisen, Rebecca J., committee member; Karkhoff-Schweizer, RoxAnn R., committee member; Huyvaert, Kathryn P., committee member
    A zoonotic disease that has killed millions over the course of at least three pandemics, plague remains a threat in regions where the etiologic agent, Yersinia pestis, persists in natural cycles involving small mammals and their fleas. Numerous flea species have been implicated as Y. pestis vectors, and some provide a "bridge" from zoonotic hosts to humans, particularly during the epizootics that decimate susceptible small mammal populations. In order to serve as a bridging vector, a flea species must be able to transmit Y. pestis, it must feed on infectious zoonotic hosts, and it must feed on humans. Identifying bridging vector species in plague-endemic regions can aid in the development of vector-control activities aimed at reducing the incidence of human plague. The West Nile region is an established plague focus in northwest Uganda. Since 1999, more than 2400 suspect human plague cases have been reported from Vurra and Okoro counties. The most likely source of infection for humans in this region is the black rat, Rattus rattus, which commonly infests human habitations and is highly susceptible to Y. pestis infection. Other potential zoonotic hosts include other rodent and shrew species that predominate in the peridomestic environment and occasionally enter huts. Two rat flea species, Xenopsylla cheopis and X. brasiliensis, both among the most efficient flea vectors of Y. pestis, are very likely to serve as bridging vectors to humans in Vurra and Okoro counties. Recent investigations, however, have found that the cat flea, Ctenocephalides felis, comprises more than 88% of host-seeking (off-host) fleas captured in huts in this region. Though an inefficient vector, this species is capable of transmitting Y. pestis. Given its dominance in human habitations and its catholic feeding habits in other regions, we hypothesized that C. felis might serve as a secondary bridging vector in Vurra and Okoro counties. In order to address this hypothesis, we sought to determine what proportion of blood meals in off-host cat fleas collected in huts in this region come from humans, and what proportion come from potentially-infectious small mammal species. Blood meal assays have long been used to examine the feeding behavior of a wide variety of disease vectors, but existing blood meal assays were deemed inadequate for our purposes because they were either not sensitive enough to detect the very small amounts of host DNA in field-collected fleas, or they were unable to capture the wide range of potential cat flea hosts in the West Nile region. Therefore, we developed a blood meal assay that takes advantage of the exquisite sensitivity of SYBR Green I-based real-time polymerase chain reaction (PCR) and combines it with the specificity and flexibility afforded by sequencing. We found that this highly-sensitive assay was subject to human DNA contamination, so we analyzed vertebrate DNA detection in artificially-fed and unfed fleas to establish a threshold cycle (Ct) cutoff that would optimize specificity without completely sacrificing sensitivity. Specifically, we identified a Ct cutoff that maximized positive predictive value. Using the established cutoff, our assay was 94 percent specific, detecting contaminating human DNA in 3 of 50 unfed fleas, and it detected and correctly identified the source of human and rat blood meals in 100 percent of artificially fed fleas held alive for up to 4 hours post feeding. Assay sensitivity declined as the time between feeding and collection increased, but we were able to detect and identify human and rat DNA in a proportion of artificially-fed fleas held alive for up to 72 hours post feeding. Using this assay, we detected and identified vertebrate DNA in 148 off-host C. felis collected in human habitations in Vurra and Okoro counties, none of it from wild rodents or shrews. Our findings indicate that cat fleas infesting huts in the West Nile region probably feed on humans, but the majority of off-host C. felis blood meals came from domesticated species that are unlikely to play a significant role in perpetuating transmission of Y. pestis. We concluded that C. felis is unlikely to serve as a bridging vector for Y. pestis in the West Nile region.
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    The ecological and evolutionary mechanisms behind the persistence of highly virulent pathogens: plague as a case study
    (Colorado State University. Libraries, 2013) Buhnerkempe, Michael G., author; Webb, Colleen T., advisor; Poff, N. LeRoy, committee member; Eisen, Rebecca J., committee member; Hoeting, Jennifer A., committee member
    The persistence of emerging infectious diseases is the result of eco-evolutionary feedbacks between a pathogen and its novel host. Spatial structure both within and between host populations (i.e., a metapopulation) in particular can have a large effect on the establishment and subsequent coevolution of a host and pathogen. Here, my colleagues and I explore how differing metapopulation structures in a host and pathogen affect the coevolutionary maintenance of high virulence and low resistance in an emerging infectious disease. We use the relatively recent emergence of plague, caused by the bacterium Yersinia pestis, in North America as a case study to both understand how spatial structure in the pathogen may differ from that of its host and how these differences may affect coevolutionary trajectories. Host responses to Y. pestis infection are highly variable with some species, like black-tailed prairie dogs (Cynomys ludovicianus), experiencing massive population declines upon introduction of the plague bacterium (i.e., epizootics), while others, like the California ground squirrel (Spermophilus beecheyi), exhibit enzootic maintenance of Y. pestis. These species in particular have markedly different spatial structures, but it is unclear how regional transmission of plague may structure the pathogen population. To understand transmission more fully, we developed a mechanistic model of plague infection in a single population that incorporated multiple routes of transmission and parameterized the model for the two species mentioned above. We found that transmission in the epizootic system is driven largely through on-host cycling of fleas (i.e., a booster-feed infection cycle). In contrast, enzootics are driven by an off-host, questing flea reservoir. The potential for off-host fleas to drive plague dynamics reveals the potential for non-overlapping host and pathogen metapopulation structures. The effect of such a structure on coevolution is not well-understood, particularly for quantitative traits where no theoretical methods exist to study coevolution in a metapopulation. Consequently, we also developed a novel theoretical framework for studying quantitative trait coevolution in a metapopulation. This new framework reveals that coevolutionary outcomes for resistance and virulence depend on the interaction between host and pathogen dispersal strategies with local reproduction and transmission dynamics favoring a diversity of resistance-virulence combinations. Host-pathogen coevolution is also affected by the shape of life-history trade-offs for both the host and the pathogen. We predicted coevolutionary outcomes under different host and pathogen dispersals assuming three different trade-off functions when resistance comes at the cost of reproduction and virulence increases transmission while decreasing the infectious period: accelerating, linear, and decelerating costs. We found that selection on resistance is most sensitive to concave trade-off functions, and selection on virulence was most sensitive to convex functions, although coevolutionarily stable strategies were only predicted when both resistance and virulence had accelerating cost trade-off functions. Predictions from the model also differ from those observed in well-mixed and spatially structured single populations indicating that eco-evolutionary dynamics do not scale directly with space. Implications for future models of plague coevolution are also discussed.