The Toll-like receptor (TLR) 7 response represents a vital host-defence mechanism in a murine model of systemic West Nile virus (WNV) infection. Here, we investigated the role of the TLR7-induced immune response following cutaneous WNV infection. We found that there was no difference in susceptibility to WNV encephalitis between wild-type and TLR7"/" mice upon intradermal injection or infected mosquito feeding. Viral load analysis revealed similar levels of WNV RNA in the peripheral tissues and brains of these two groups of mice following intradermal infection. There was a higher level of cytokines in the blood of wild-type mice at early stages of infection; however, this difference was diminished in the blood and brains at later stages. Langerhans cells (LCs) are permissive to WNV infection and migrate from the skin to draining lymph nodes upon intradermal challenge. Our data showed that WNV infection of TLR7"/"keratinocytes was significantly higher than that of wild-type keratinocytes. Infection of wild-type keratinocytes induced higher levels of alpha interferon and interleukin-1b (IL-1b), IL-6 and IL-12, which might promote LC migration from the skin. Co-culture of naïve LCs of wild-type mice with WNV-infected wild-type keratinocytes resulted in the production of more IL-6 and IL-12 than with TLR7 "/" keratinocytes or by cultured LCs alone. Moreover, LCs in the epidermis were reduced in wild-type mice, but not in TLR7 "/" mice, following intradermal WNV infection. Overall, our results suggest that the TLR7 response following cutaneous infection promotes LC migration from the skin, which might compromise its protective effect in systemic infection. INTRODUCTIONWest Nile virus (WNV), a single-stranded (ss)RNA flavivirus, has been endemic in Africa, Asia, Europe and, more recently, North America (Granwehr et al., 2004; Pletnev et al., 2006). The virus is maintained in an enzootic cycle that involves mosquitoes and birds. Human infection results primarily from mosquito bites, and symptoms include fever, headache, myalgia, meningitis and encephalitis. Neurological disease (including encephalitis) has been observed in .30 % of confirmed WNV cases, with a higher frequency in the elderly and immunocompromised (Campbell et al., 2002; Pletnev et al., 2006). Human vaccines are not yet available. Treatment is mostly nonspecific and supportive.Murine infection via subcutaneous or intraperitoneal injection has been an effective in vivo experimental model for human infections to investigate WNV pathogenesis and host immune response (Beasley et al., 2002;. Work from the murine model has demonstrated that type I interferons (IFNs), cd T cells and humoral immunity are critical in controlling the dissemination of WNV (Anderson & Rahal, 2002;Diamond et al., 2003;Fredericksen et al., 2008;Klein et al., 2005;Lucas et al., 2003;Roehrig et al., 2001;Wang et al., 2003a Intradermal WNV infection, which presumably mimics natural infection in humans, has also been studied in mice (Johnston et al., 1996(Johnston et al., , 2000. Follow...

A One Health approach to the epidemiology, management, surveillance, and control of leptospirosis relies on accessible and accurate diagnostics that can be applied to humans and companion animals and livestock. Diagnosis should be multifaceted and take into account exposure risk, clinical presentation, and multiple direct and/or indirect diagnostic approaches. Methods of direct detection of Leptospira spp. include culture, histopathology and immunostaining of tissues or clinical specimens, and nucleic acid amplification tests (NAATs). Indirect serologic methods to detect leptospiral antibodies include the microscopic agglutination test (MAT), the enzyme-linked immunosorbent assay (ELISA), and lateral flow methods. Rapid diagnostics that can be applied at the point-of-care; NAAT and lateral flow serologic tests are essential for management of acute infection and control of outbreaks. Culture is essential to an understanding of regional knowledge of circulating strains, and we discuss recent improvements in methods for cultivation, genomic sequencing, and serotyping. We review the limitations of NAATs, MAT, and other diagnostic approaches in the context of our expanding understanding of the diversity of pathogenic Leptospira spp. Novel approaches are needed, such as loop mediated isothermal amplification (LAMP) and clustered regularly interspaced short palindromic repeats (CRISPR)-based approaches to leptospiral nucleic acid detection.

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BackgroundMosquito salivary proteins (MSPs) modulate the host immune response, leading to enhancement of arboviral infections. Identification of proteins in saliva responsible for immunomodulation and counteracting their effects on host immune response is a potential strategy to protect against arboviral disease. We selected a member of the D7 protein family, which are among the most abundant and immunogenic in mosquito saliva, as a vaccine candidate with the aim of neutralizing effects on the mammalian immune response normally elicited by mosquito saliva components during arbovirus transmission.Methodology/Principal FindingsWe identified D7 salivary proteins of Culex tarsalis, a West Nile virus (WNV) vector in North America, and expressed 36 kDa recombinant D7 (rD7) protein for use as a vaccine. Vaccinated mice exhibited enhanced interferon-γ and decreased interleukin-10 expression after uninfected mosquito bite; however, we found unexpectedly that rD7 vaccination resulted in enhanced pathogenesis from mosquito-transmitted WNV infection. Passive transfer of vaccinated mice sera to naïve mice also resulted in increased mortality rates from subsequent mosquito-transmitted WNV infection, implicating the humoral immune response to the vaccine in enhancement of viral pathogenesis. Vaccinated mice showed decreases in interferon-γ and increases in splenocytes producing the regulatory cytokine IL-10 after WNV infection by mosquito bite.Conclusions/SignificanceVector saliva vaccines have successfully protected against other blood-feeding arthropod-transmitted diseases. Nevertheless, the rD7 salivary protein vaccine was not a good candidate for protection against WNV disease since immunized mice infected via an infected mosquito bite exhibited enhanced mortality. Selection of salivary protein vaccines on the bases of abundance and immunogenicity does not predict efficacy.

Background Factors associated with outcome in dogs diagnosed with infective endocarditis (IE) are not well characterized. Objectives Evaluate outcome and prognostic factors in dogs with IE. Animals One hundred and thirteen dogs with IE. Methods Medical records for dogs that fulfilled the modified Duke criteria between 2005 and 2020 were retrospectively reviewed. Signalment, preexisting conditions, clinicopathologic findings, treatment regimen, and outcomes were recorded. Univariate logistic regression was performed to identify categorical factors associated with mortality, and then multivariate analysis was performed. Results Dogs were categorized as survivors (n = 47), non‐survivors (n = 57), or lost to follow‐up (n = 9). Survival to discharge and at 1 month was documented in 79 (70%) of 113 and 56 (54%) of 104 dogs, respectively, with median survival time (MST) of 72 days. Risk factors associated with mortality included development of congestive heart failure (odds ratio [OR], 11.8; 95% confidence interval [CI], 1.4‐97.8), thromboembolic events (OR, 5.7; 95% CI, 2.3‐14.4), and acute kidney injury (OR, 6.2; 95% CI, 2.0‐18.8). Administration of antithrombotic medications was associated with survival (OR, 0.35; 95% CI, 0.13‐0.97). Dogs that were not treated with antithrombotics had MST of 92 days, whereas dogs treated with antithrombotics did not reach MST during the study period. The heart valves involved and etiologic agent identified did not correlate with outcome. Conclusion and Clinical Importance Dogs with IE that had thromboembolic events, acute kidney injury, or congestive heart failure had higher risk of mortality. Administration of antithrombotics was associated with prolonged survival time.