Spontaneous Lesions of Endangered Geriatric Julia Creek Dunnarts (Sminthopsis douglasi, Archer 1979) with Emphasis in Reproductive Pathology.

Julia Creek dunnarts are an endangered species of carnivorous marsupials and the focus of multiple conservation strategies involving significant resources such as captive breeding programs. Despite the relevance for conservation, no study to date has focused on evaluating geriatric diseases in dunnarts. This study describes the pathology findings in a group of one wild and thirty-five captive-born, mostly geriatric Julia Creek dunnarts that failed to produce offspring over multiple breeding periods. A total of 20 females and 16 males were submitted for a postmortem examination, with ages ranging from 9 to 42 and 12 to 42 months for females and males, respectively. Of these, 10 had unremarkable findings. The most common condition in females was cystic glandular hyperplasia (n = 8), typical of hormonal dysregulation profiles in senescence, particularly hyperestrogenism. Rarely, cutaneous disease represented by unidentified dermal round cell infiltrates was observed in females (n = 2). Primary reproductive hormonal dysregulation was also suspected in males diagnosed with testicular degeneration, aspermatogenesis and/or atrophy (n = 3). Cutaneous round cell infiltrates, possibly compatible with epitheliotropic lymphomas, were seen in males (n = 3), and 2/3 affected males also had concurrent testicular degeneration or atrophy, indicating male sex could be a predictor for lymphoid neoplasia in aged dunnarts, especially in individuals with concurrent testosterone-luteinizing hormone dysregulation as it occurs in gonadectomized animals. The role of an underlying viral etiology is also explored. This study is the first to describe major spontaneous diseases in endangered aged Julia Creek dunnarts, providing an important understanding of senescence and geriatric diseases within a conservation context.

Development of body, head and brain features in the Australian fat-tailed dunnart (Sminthopsis crassicaudata; Marsupialia: Dasyuridae); A postnatal model of forebrain formation.

Most of our understanding of forebrain development comes from research of eutherian mammals, such as rodents, primates, and carnivores. However, as the cerebral cortex forms largely prenatally, observation and manipulation of its development has required invasive and/or ex vivo procedures. Marsupials, on the other hand, are born at comparatively earlier stages of development and most events of forebrain formation occur once attached to the teat, thereby permitting continuous and non-invasive experimental access. Here, we take advantage of this aspect of marsupial biology to establish and characterise a resourceful laboratory model of forebrain development: the fat-tailed dunnart (Sminthopsis crassicaudata), a mouse-sized carnivorous Australian marsupial. We present an anatomical description of the postnatal development of the body, head and brain in dunnarts, and provide a staging system compatible with human and mouse developmental stages. As compared to eutherians, the orofacial region develops earlier in dunnarts, while forebrain development is largely protracted, extending for more than 40 days versus ca. 15 days in mice. We discuss the benefits of fat-tailed dunnarts as laboratory animals in studies of developmental biology, with an emphasis on how their accessibility in the pouch can help address new experimental questions, especially regarding mechanisms of brain development and evolution.

Teaching structure: student use of software tools for understanding macromolecular structure in an undergraduate biochemistry course.

Because understanding the structure of biological macromolecules is critical to understanding their function, students of biochemistry should become familiar not only with viewing, but also with generating and manipulating structural representations. We report a strategy from a one-semester undergraduate biochemistry course to integrate use of structural representation tools into both laboratory and homework activities. First, early in the course we introduce the use of readily available open-source software for visualizing protein structure, coincident with modules on amino acid and peptide bond properties. Second, we use these same software tools in lectures and incorporate images and other structure representations in homework tasks. Third, we require a capstone project in which teams of students examine a protein-nucleic acid complex and then use the software tools to illustrate for their classmates the salient features of the structure, relating how the structure helps explain biological function. To ensure engagement with a range of software and database features, we generated a detailed template file that can be used to explore any structure, and that guides students through specific applications of many of the software tools. In presentations, students demonstrate that they are successfully interpreting structural information, and using representations to illustrate particular points relevant to function. Thus, over the semester students integrate information about structural features of biological macromolecules into the larger discussion of the chemical basis of function. Together these assignments provide an accessible introduction to structural representation tools, allowing students to add these methods to their biochemical toolboxes early in their scientific development.

Genetic and fluorescence studies of affinity maturation in related antibodies.

Affinity maturation, the process by which an organism's response to infection becomes more specific and more effective over time, occurs after somatic hypermutation of antibody genes in B-cells. This increase in affinity might be a result of the evolution of either specific interactions between antigen and antibody over time (enthalpic factors) or antibody binding site rigidification (entropic factors) or both. Here, monoclonal antibodies, derived from antibodies elicited at different points in the murine immune response after inoculation with the same diketone hapten, have been characterized both genetically and functionally. Though this hapten has previously been shown to produce the catalytic aldolase antibody 38C2, antibodies described here are not catalytic and unlike 38C2, form no covalent enzyme-substrate complex. Thus, they provide a system in which to assess contributions to the evolution of binding affinity. The genes for these non-catalytic antibodies have been sequenced and analyzed both with regard to their relationships to germ line genes, to each other, and to two commercially available catalytic aldolase antibodies. Consequences of particular mutations for antigen binding behavior are discussed. The protein products of these genes have been expressed, purified, and binding properties measured by two complementary techniques: the hapten-induced quenching of the native antibody fluorescence and the changes in the anisotropy of Prodan (6-propionyl-2-(dimethylamino)naphthalene), a fluorescent hapten analogue. Differences in binding affinity are related back to differences in the lengths and amino acid sequences of the complementary determining region 3 (CDR3) binding loop. Taken together with our earlier results on binding site heterogeneity from tryptophan lifetime analysis [Mohan, G.S., Chiu, P.T., Southern, C.A., O'Hara, P.B., 2004. Steady-state and multifrequency phase fluorometry studies of binding site flexibility in related antibodies. J. Phys. Chem. A 108, 7871-7877], affinity appears to be modulated by a combination of entropic and enthalpic factors, and not dominated by one or the other. Because these antibodies are not related to the same germ line gene, however, these results do not provide evidence for the dominance of enthalpy or entropy in evolving binding affinity in this system.