MELANOCYTIC NEOPLASIA IN PANTHERA SPECIES: CLINICAL PRESENTATIONS, PATHOLOGIC FINDINGS AND RESPONSES TO TREATMENT.

Neoplasia is a common cause of morbidity and mortality in captive nondomestic felids. Seven tigers (Panthera tigris), two African lions (Panthera leo), and two snow leopards (Panthera uncia) were diagnosed with melanocytic neoplasia (10 malignant melanomas, two benign melanocytomas) over a 20-yr period. Animals were 10-19 yr old and 5/7 tigers were phenotypically white. Malignant melanoma tumor location included skin (n = 4), oral mucosa (n = 2), nasal planum (n = 1), iris/uvea (n = 2), and lip margin (n = 1); melanocytomas were found in skin (n = 2). Metastasis to regional lymph nodes was seen at diagnosis in 3/7 melanoma cases. Thoracic radiography (n = 6) and/or computed tomography (n = 2) did not detect pulmonary metastasis at diagnosis but were useful for detection later in the disease course. Median survival time (MST) for all cases ranged from 1 mon - 40 mon. Seven cases with malignant melanoma underwent treatment, which included surgery, radiation therapy, and administration of the canine melanoma vaccine (Oncept®) or a combination of these treatments; MST was 5-40 mon for these cases. While multimodal therapy may provide an improved survival time, the majority of animals with malignant melanoma invariably died from neoplastic disease. Necropsy confirmed metastasis of malignant melanoma in 7/9 animals; sites included lung, liver, lymph node, kidney, mesentery, pleural cavity, heart, stomach, spleen, and adrenal gland. This case series describes the clinical and histologic findings of melanocytic neoplasia in nondomestic felids as well as multimodal treatment strategies incorporating the canine melanoma vaccine.

HISTOPLASMOSIS IN NONDOMESTIC FELIDS: A REVIEW OF SIX CASES.

One adult leopard (Panthera pardus) and five adult tigers (Panthera tigris) presented with a range of nonspecific clinical signs, including lethargy (6/6), mobility deficits (4/6), and hyporexia (3/6). Hematology and biochemistry revealed a hyperproteinemia characterized by hyperglobulinemia (4/6), hepatocellular enzyme activity increases (3/6), azotemia (3/6), leukocytosis (2/6), hyperbilirubinemia (2/6), or a combination of conditions. Further diagnostics and management varied with the presenting signs and clinicopathological findings, including supportive care, diagnostic imaging, and blastomyces urine antigen analyses. Two animals died, and four were euthanatized. Postmortem findings included granulomatous pneumonia (6/6), fibrinous pleural effusion (3/ 6), pericardial effusion (2/6), and diffuse icterus (1/6). Histopathology revealed round to oval structures with a thin clear wall and purple inclusions within cells of the mononuclear phagocyte system, consistent with Histoplasma capsulatum, in each animal. Disseminated histoplasmosis was found in five cases, with organisms present in the lung (5/5), liver (3/5), lymph nodes (3/5), spleen (2/5), bone marrow (2/5), thyroid (1/5), tongue (1/ 5), kidney (1/5), or a combination of organs. One tiger was found to have pulmonary histoplasmosis without evidence of disseminated infection. On the basis of clinical and pathological findings, histoplasmosis was diagnosed. This case series illustrates the difficulties in antemortem diagnosis of histoplasmosis on the basis of complete blood count, serum biochemistry profile, and antigen testing and underscores that histoplasmosis should be considered a differential diagnosis in any felid presenting with nonspecific clinical signs in endemic areas.

Dual purpose hafnium oxide nanoparticles offer imaging Streptococcus mutans dental biofilm and fight it In vivo via a drug free approach.

The removal of tenacious dental plaque is of paramount importance; however, early diagnosis can be a challenging task in dental clinics due to the limitations of current approaches, specifically X-ray-based techniques. We have approached this problem by integrating antibacterial properties and X-ray contrast enhancement in a single probe specific to colonies of Streptococcus mutans as the most predominant and carious oral bacteria. We report the synthesis of an inherently therapeutic polymeric silane conjugated hafnium oxide nanoparticles (Hf PS NPs). Using a high-affinity pathogen-selective peptide, the concept of molecularly targeted X-ray imaging of cariogenic pathogen S. mutans was demonstrated. Ex vivo studies using extracted human tooth demonstrated striking X-ray attenuation of NPs vs. tooth. Additionally, Hf PS NPs exhibited significant bactericidal properties against cariogenic pathogen. Electron microscopy revealed that the antibacterial activity occurred via a 'latch and kill' mechanism. Mechanistic studies determined that these NPs fragmented bacterial DNA components to exert their antimicrobial effect. Importantly, Hf PS NPs effectively inhibited the growth of a mature biofilm on an ex vivo human tooth model. Finally, the NPs were applied to the rodent model of dental biofilm. Topical administration of the Hf PS NPs for 8 days (1X daily) could effectively attenuate the S. mutans biofilm challenge. This report is the first of its kind to demonstrate that HfO2-based NPs can be used for simultaneous diagnosis and antibacterial treatment without requiring an additional drug.

Nonpathologic Infection of Macaques by an Attenuated Mycobacterial Vaccine Is Not Reactivated in the Setting of HIV Co-Infection.

Failure to replace Bacille Calmette-Guerin vaccines with efficacious anti-tuberculosis (TB) vaccines have prompted outside-the-box thinking, including pulmonary vaccination to elicit local immunity. Inhalational MtbΔsigH, a stress-response-attenuated strain, protected against lethal TB in macaques. While live mycobacterial vaccines show promising efficacy, HIV co-infection and the resulting immunodeficiency prompts safety concerns about their use. We assessed the persistence and safety of MtbΔsigH, delivered directly to the lungs, in the setting of HIV co-infection. Macaques were aerosol-vaccinated with ΔsigH and subsequently challenged with SIVmac239. Bronchoalveolar lavage and tissues were sampled for mycobacterial persistence, pathology, and immune correlates. Only 35% and 3.5% of lung samples were positive for live bacilli and granulomas, respectively. Our results therefore suggest that the nonpathologic infection of macaque lungs by ΔsigH was not reactivated by simian immunodeficiency virus, despite high viral levels and massive ablation of pulmonary CD4+ T cells. Protective pulmonary responses were retained, including vaccine-induced bronchus-associated lymphoid tissue and CD8+ effector memory T cells. Despite acute simian immunodeficiency virus infection, all animals remained asymptomatic of pulmonary TB. These findings highlight the efficacy of mucosal vaccination via this attenuated strain and will guide its further development to potentially combat TB in HIV-endemic areas. Our results also suggest that a lack of pulmonary pathology is a key correlate of the safety of live mycobacterial vaccines.

Hypoxia Sensing and Persistence Genes Are Expressed during the Intragranulomatous Survival of Mycobacterium tuberculosis.

Although it is accepted that the environment within the granuloma profoundly affects Mycobacterium tuberculosis (Mtb) and infection outcome, our ability to understand Mtb gene expression in these niches has been limited. We determined intragranulomatous gene expression in human-like lung lesions derived from nonhuman primates with both active tuberculosis (ATB) and latent TB infection (LTBI). We employed a non-laser-based approach to microdissect individual lung lesions and interrogate the global transcriptome of Mtb within granulomas. Mtb genes expressed in classical granulomas with central, caseous necrosis, as well as within the caseum itself, were identified and compared with other Mtb lesions in animals with ATB (n = 7) or LTBI (n = 7). Results were validated using both an oligonucleotide approach and RT-PCR on macaque samples and by using human TB samples. We detected approximately 2,900 and 1,850 statistically significant genes in ATB and LTBI lesions, respectively (linear models for microarray analysis, Bonferroni corrected, P < 0.05). Of these genes, the expression of approximately 1,300 (ATB) and 900 (LTBI) was positively induced. We identified the induction of key regulons and compared our results to genes previously determined to be required for Mtb growth. Our results indicate pathways that Mtb uses to ensure its survival in a highly stressful environment in vivo. A large number of genes is commonly expressed in granulomas with ATB and LTBI. In addition, the enhanced expression of the dormancy survival regulon was a key feature of lesions in animals with LTBI, stressing its importance in the persistence of Mtb during the chronic phase of infection.

CD4+ T-cell-independent mechanisms suppress reactivation of latent tuberculosis in a macaque model of HIV coinfection.

The synergy between Mycobacterium tuberculosis (Mtb) and HIV in coinfected patients has profoundly impacted global mortality because of tuberculosis (TB) and AIDS. HIV significantly increases rates of reactivation of latent TB infection (LTBI) to active disease, with the decline in CD4(+) T cells believed to be the major causality. In this study, nonhuman primates were coinfected with Mtb and simian immunodeficiency virus (SIV), recapitulating human coinfection. A majority of animals exhibited rapid reactivation of Mtb replication, progressing to disseminated TB and increased SIV-associated pathology. Although a severe loss of pulmonary CD4(+) T cells was observed in all coinfected macaques, a subpopulation of the animals was still able to prevent reactivation and maintain LTBI. Investigation of pulmonary immune responses and pathology in this cohort demonstrated that increased CD8(+) memory T-cell proliferation, higher granzyme B production, and expanded B-cell follicles correlated with protection from reactivation. Our findings reveal mechanisms that control SIV- and TB-associated pathology. These CD4-independent protective immune responses warrant further studies in HIV coinfected humans able to control their TB infection. Moreover, these findings will provide insight into natural immunity to Mtb and will guide development of novel vaccine strategies and immunotherapies.

Treatment with aspirin or clopidogrel does not affect equine platelet expression of P selectin or platelet-neutrophil aggregates.

Inflammation-induced P-selectin (CD62P) expression on platelets and endothelial cells facilitates interactions among platelets and polymorphonuclear leukocytes (PMN), and can also promote coagulation. The effects of clopidogrel and aspirin (ASA) on equine platelet CD62P expression were investigated. Six horses were treated in a cross-over design with clopidogrel (2mg/kg PO q 24) or ASA (5mg/kg PO q 24h) for 5 days. Platelets collected at 24, 72, 96, 120, and 168 h after the initiation of therapy were stimulated using 0.1 µg/mL thrombin, followed by flow cytometric analysis using anti-CD41/61 and anti-equine CD62P antibodies. Platelet-PMN aggregates were also enumerated. Baseline CD62P positive platelet numbers were not different between groups (mean ± SD): 4254 ± 1785 (clopidogrel) and 3600 ± 1780 (ASA, P=0. 435). Although expression tended to decrease, there were no significant changes in CD62P+platelets after treatment with either drug (clopidogrel P=0.139, ASA P=0.161). There was also no difference in platelet-PMN aggregates during or after treatment with ASA (P=0.513) or clopidogrel (P=0.543). Due to small numbers of horses, this study may have been underpowered to detect a true decrease in expression, and differences between therapies may have been more pronounced if this study had evaluated horses with systemic inflammation.