Prevalence and Distribution of Ranavirus in Amphibians From

Several infectious diseases are threatening amphibian species worldwide and have resulted in massmortality events across the globe. An emerging group of viral pathogens (ranaviruses) are documented to cause die-offs in amphibian populations worldwide, including in several regions of the U.S. Unfortunately, large gaps remain in our understanding of the distribution of this systemic pathogen in the U.S., including within the state of Oklahoma. To address this gap in our understanding, we carried out surveys of this infectious pathogen across 14 sites in seven southeastern Oklahoma counties in spring 2015, screening 17 amphibian species from this region. Using liver and tail tissue samples collected from individual amphibians, we screened for the presence and infection load of ranavirus. Of the 390 samples, 84 (21.5%) tested positive for ranavirus, with infection prevalence varying among species surveyed. Notably, the family Bufonidae had no samples that tested positive for ranavirus, whereas the remaining families had an infection prevalence ranging from 14–50%. Despite an overall infection prevalence of 21.5%, we detected no clinical signs of ranavirosis and all sampled individuals appeared outwardly healthy. These results provide data on the geographic and host distribution of ranavirus in southeastern Oklahoma, as well as the first documented cases of the pathogen in three species of anurans: Gastrophryne carolinensis (Eastern Narrow-mouthed Toad), G. olivacea (Western Narrow-mouthed Toad), and Pseudacris fouquettei (Cajun Chorus Frog). With widespread ranavirus infection, there is potential for transmission from abundant, widespread species to more vulnerable, state-threatened amphibians

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival