A New Species of Myxidium (Myxosporea: Myxidiidae), from the Western Chorus Frog, Pseudacris triseriata triseriata, and Blanchard's Cricket Frog, Acris crepitans blanchardi (Hylidae), from Eastern Nebraska: Morphology, Phylogeny, and Critical Comments on Amphibian Myxidium Taxonomy

During March 2001-April 2004, 164 adult anurans of 6 species (47 Rana blairi, 35 Rana catesbeiana, 31 Hyla chrysoscelis, 31 Pseudacris triseriata triseriata, 11 Bufo woodhousii, and 9 Acris crepitans blanchardi) from Pawnee Lake, Lancaster County, Nebraska, were surveyed for myxozoan parasites. Of these, 20 of 31 (65%) P. triseriata triseriata and 1 of 9 (11%) A. crepitans blanchardi were infected with a new species of Myxidium. Myxidium melleni n. sp. (Myxosporea) is described from the gallbladder of the western chorus frog, P. triseriata triseriata (Hylidae). This is the second species of Myxidium described from North American amphibians. Mature plasmodia are disc-shaped or elliptical 691 (400-1,375) × 499 (230-1,200) × 23 (16-35) μm, polysporic, producing many disporic pansporoblasts. The mature spores, 12.3 (12.0-13.5) × 7.6 (7.0-9.0) × 6.6 (6.0-8.0) μm, containing a single binucleated sporoplasm, are broadly elliptical, with 2-5 transverse grooves on each valve, and contain two equal polar capsules 5.2 (4.8-5.5) × 4.2 (3.8-4.5) μm positioned at opposite ends of the spore. Myxidium melleni n. sp. is morphologically consistent with other members of Myxidium. However, M. melleni n. sp. was phylogenetically distinct from other Myxidium species for which DNA sequences are available. Only with improved morphological analyses, accompanied by molecular data, and the deposit of type specimens, can the ambiguous nature of Myxidium be resolved. Guidelines for descriptions of new species of Myxidium are provided.

Development of an Electrochemical Insulin Sensor Based on a High Affinity DNA Sequence Found in the Insulin-linked Polymorphic Region

The detection of pertinent biomarkers has the potential provide an early indication of disease progression before considerable damage has been incurred. A decrease in an individual’s sensitivity to insulin, which may be quantified as the ratio of insulin to glucose in the blood after a glucose pulse, has recently been reported as an early predictor of insulin-dependent diabetes mellitus. Routine measurement of insulin levels is therefore desirable in the care of diabetes-prone individuals. A rapid, simple, and reagentless method for insulin detection would allow for wide-spread screenings that provide earlier signs of diabetes onset. The aim of this thesis is to develop a folding-base electrochemical sensor for the detection of insulin. The sensor described herein consists of a DNA probe immobilized on a gold disc electrode via an alkanethiol linker and embedded in an alkanethiol self-assembled monolayer. The probe is labeled with a redox reporter, which readily transfers electrons to the gold electrode in the absence of insulin. In the presence of insulin, electron transfer is inhibited, presumably due to a binding-induced conformational or dynamic change in the DNA probe that significantly alters the electron-tunneling pathway. A 28-base segment of the insulin-linked polymorphic region that has been reported to bind insulin with high affinity serves as the capture element of the DNA probe. Three probe constructs that vary in their secondary structure and position of the redox label are evaluated for their utility as insulin-sensing elements on the electrochemical platform. The effects of probe modification on secondary structure are also evaluated using circular dichroism spectroscopy.