Cardiovascular disease in central bearded dragons (Pogona vitticeps): 54 cases (2007-2022).

OBJECTIVE: To describe the clinical features, histopathologic lesions, and outcome of cardiovascular disease in central bearded dragons. ANIMALS: 54 bearded dragons. METHODS: Retrospective evaluation of captive bearded dragons with antemortem imaging or postmortem diagnosis of cardiovascular disease from 2007 to 2022 from 6 hospitals. RESULTS: The total prevalence of cardiovascular disease was 3.3% (54/1,655). Physical examination findings were available in 46 cases with change in mentation being the most common finding (n = 28/46 [60.9%]), followed by dehydration (17/46 [37%]), palpable coelomic mass (13/46 [28.3%]), dyspnea (10/46 [21.7%]), and sunken eyes (10/46 [21.7%)]. Doppler auscultation revealed an arrhythmia in 5/34 (14.7%) animals. Diagnostic imaging was only performed on 21 animals, and 10 (47.6%) had cardiovascular abnormalities described. In total, 84 cardiovascular diagnoses were found in 54 animals. The most common diagnosis was myocarditis (n = 14) followed by aneurysms (11), pericardial effusion (9), atherosclerosis (7), epicarditis (7), and myocardial degeneration/necrosis (7). Overall, 62 causes of death were identified in 52 cases, with cardiovascular disease being the most common (n = 18/52 [34.5%]). Only 3/54 animals were diagnosed with congestive heart failure. Animals with aneurysms were more likely to die to due cardiovascular disease compared to other types of cardiovascular diagnoses (OR, 43.75; 95% CI, 4.88 to 392.65; P < .001). CLINICAL RELEVANCE: Diagnosis of cardiovascular disease in bearded dragons is challenging given the inconsistent clinical presentation; however, it should remain a differential in animals with nonspecific signs of illness. Antemortem diagnostics are recommended in suspected cases, including diagnostic imaging. Of the cardiovascular diseases described, aneurysms most often contributed to clinical demise.

Characterization of an Injectable Chitosan Hydrogel for the Tunable, Localized Delivery of Immunotherapeutics.

Localized delivery of immunotherapeutics within a tumor has the potential to reduce systemic toxicities and improve treatment outcomes in cancer patients. Unfortunately, local retention of therapeutics following intratumoral injection is problematic and is insufficiently considered. Dense tumor architectures and high interstitial pressures rapidly exclude injections of saline and other low-viscosity solutions. Hydrogel-based delivery systems, on the other hand, can resist shear forces that cause tumor leakage and thus stand to improve the local retention of coformulated therapeutics. The goal of the present work was to construct a novel, injectable hydrogel that could be tuned for localized immunotherapy delivery. A chitosan-based hydrogel, called XCSgel, was developed and subsequently characterized. Nuclear magnetic resonance studies were performed to describe the chemical properties of the new entity, while cryo-scanning electron microscopy allowed for visualization of the hydrogel's cross-linked network. Rheology experiments demonstrated that XCSgel was shear-thinning and self-healing. Biocompatibility studies, both in vitro and in vivo, showed that XCSgel was nontoxic and induced transient mild-to-moderate inflammation. Release studies revealed that coformulated immunotherapeutics were released over days to weeks in a charge-dependent manner. Overall, XCSgel displayed several clinically important features, including injectability, biocompatibility, and imageability. Furthermore, the properties of XCSgel could also be controlled to tune the release of coformulated immunotherapeutics.

Safety and Pharmacokinetics of Intravesical Chitosan/Interleukin-12 Immunotherapy in Murine Bladders.

BACKGROUND: Intravesical administration of interleukin 12 (IL-12) co-formulated with the biopolymer, chitosan (CS/IL-12), has demonstrated remarkable antitumor activity against preclinical models of bladder cancer. However, given historical concerns regarding severe toxicities associated with systemic IL-12 administration in clinical trials, it is important to evaluate the safety of intravesical CS/IL-12 prior to clinical translation. OBJECTIVE: To evaluate the pharmacokinetics as well as the local and systemic toxicities of intravesical CS/IL-12 immunotherapy in laboratory mice. METHODS: Local inflammatory responses in mouse bladders treated with intravesical IL-12 or CS/IL-12 were assessed via histopathology. Serum cytokine levels following intravesical and subcutaneous (s.c.) administrations of IL-12 or CS/IL-12 in laboratory mice were compared. Systemic toxicities were evaluated via body weight and liver enzyme levels. RESULTS: Intravesical IL-12 and CS/IL-12 treatments did not induce significant local or systemic toxicity. IL-12 dissemination and exposure from intravesical administration was significantly lower compared to s.c. injections. Weekly intravesical CS/IL-12 treatments were well-tolerated and did not result in blunted immune responses. CONCLUSIONS: Intravesical CS/IL-12 is safe and well-tolerated in mice. In particular, the lack of cystitis and acute inflammation justifies continued investigation of intravesical CS/IL-12 immunotherapy in larger animals and patients with bladder cancer.

Evaluating the Effects of Various Decalcification Protocols on Immunohistochemical Staining in Zebrafish (Danio rerio).

Fixation and decalcification can alter protein structure in tissues, influencing the efficacy of primary antibodies routinely used in immunohistochemical (IHC) staining. Histologic examination of zebrafish requires both processes, making staining and analysis potentially challenging. Here, we investigated the effects of common fixation and decalcification protocols on IHC staining in zebrafish. We also identified zebrafish-reactive and -specific antibodies for use in research and diagnostics. For several of the antibodies, time spent in Dietrich's fixative containing 2% glacial acetic acid or 3.4% formaldehyde followed by decalcification with ethylenediaminetetraacetic acid (EDTA) significantly impacted IHC staining quality, particularly regarding staining intensity. Protocols utilizing shorter fixation times produced higher-quality stains. In addition, individual markers were variably affected by the type of fixative. Dietrich's fixative significantly reduced staining quality for the "neural" markers: glial fibrillar acidic protein, chromogranin A, S100. A negative time-dependent effect of fixation on staining quality was found for several antibodies: muscle actin (Dietrich's only), cytokeratin AE1/AE3, chromogranin, and S100. Neither decalcification protocol had a statistically significant negative time-dependent effect on staining quality. Based on our results, we suggest shorter fixation and decalcification protocols to best preserve IHC staining quality as well as recommend deliberate selection of the fixative used depending on the protein of interest.

Aspergillus Section Nigri-Associated Calcium Oxalate Crystals in an Eurasian Eagle Owl (Bubo bubo).

An adult male Eurasian eagle owl (Bubo bubo) housed at a wildlife rehabilitation facility in southern Oregon died after a short period of progressive ill-thrift. Radiographs taken prior to death demonstrated abnormal radiopaque material in the coelom and the owl was submitted for postmortem examination. Black pigmented fungus was noted grossly, particularly in the respiratory tissues, with abundant oxalate crystal deposition associated with and without hyphal elements subsequently observed histologically. Aspergillus section Nigri was cultured from the lesions. Although there have been a few reports of aspergillosis caused by Aspergillus niger in avian species, the severity and wide tissue distribution of oxalates in this case are highly unusual.

Development of Quantitative Real-Time PCR Assays for Postmortem Detection of Mycobacterium spp. Common in Zebrafish (Danio rerio) Research Colonies.

Mycobacterium spp. infections are common in zebrafish kept in research facilities. These comorbidities can substantially modulate the responses of these fish to external and internal stimuli. Therefore, diagnostic tests to detect Mycobacterium spp. infections in zebrafish colonies prove essential. Here, we outline the development of quantitative simplex real-time PCR assays to detect the 3 Mycobacterium species most commonly identified in laboratory zebrafish. The assays targeted the heatshock protein 65 gene of M. marinum, M. chelonae, and M. haemophilum. The assays are both highly specific and sensitive for fresh-frozen samples and highly specific and moderately sensitive for formalin-fixed paraffin-embedded (FFPE) samples. Two sampling techniques for FFPE samples of sagittally sectioned zebrafish were evaluated. Both paraffin cores targeting granulomas containing bacteria and scrolls from the entire fish yielded DNA of equivalent quantity and purity. The diagnostic sensitivity of cores was superior to that of scrolls for M. chelonae and M. haemophilum but not M. marinum. The assays are cost-effective and ideally suited to diagnosing common Mycobacterium spp. infections in zebrafish.