Cranial Nerve Involvement and Thalamic Lesions in West Nile Virus Encephalitis in a Kidney Transplant Recipient: Implications for Diagnosis and Prevention

In addition, the viral load in this study appeared to be comparatively high. These results suggest that some sapovirus genotypes are more virulent than others. Similar fi ndings were obtained with norovirus infections around the world; strains belonging to norovirus GII/4 were the most prevalent in many countries. Although several recombinant sapovirus strains have been identifi ed and found to be the cause of increased numbers of infections in some countries (1,5), they were not observed in this study. Increased sapovirus surveillance and reporting are needed to shed some more light on this poorly understood virus.

Congenital West Nile virus (WNV) infection was first described in a single case in 2002. The proportion of maternal WNV infections resulting in congenital infection and clinical consequences of such infections are unknown. In 2003 and 2004, women in the United States who acquired WNV infection during pregnancy were reported to the Centers for Disease Control and Prevention by state health departments. Data on pregnancy outcomes were collected. One of the maternal WNV infections was identified retrospectively after the infant was born. Maternal sera, placenta, umbilical cord tissue, and cord serum were tested for WNV infection by using serologic assays and reverse-transcription polymerase chain reaction. Infant health was assessed at delivery and through 12 months of age. Seventy-seven women infected with WNV during pregnancy were clinically followed in 16 states. A total of 71 women delivered 72 live infants; 4 women had miscarriages, and 2 had elective abortions. Of the 72 live infants, 67 were born at term, and 4 were preterm; gestational age was unknown for 1. Of 55 live infants from whom cord serum was available, 54 tested negative for anti-WNV IgM. One infant born with umbilical hernia and skin tags had anti-WNV IgM in cord serum but not in peripheral serum at age 1 month. An infant who had no anti-WNV IgM in cord blood, but whose mother had WNV illness 6 days prepartum, developed WNV meningitis at age 10 days. Another infant, whose mother had acute WNV illness at delivery, was born with a rash and coarctation of the aorta and had anti-WNV IgM in serum at 1 month of age; cord serum was not available. A fourth infant, whose mother had onset of WNV illness 3 weeks prepartum that was not diagnosed until after delivery, had WNV encephalitis and underlying lissencephaly detected at age 17 days and subsequently died; cord serum was not available. The following major malformations were noted among live-born infants: aortic coarctation (n = 1); cleft palate (n = 1); Down syndrome (n = 1); lissencephaly (n = 1); microcephaly (n = 2); and polydactyly (n = 1). One infant had glycogen storage disease type 1. Abnormal growth was noted in 8 infants. Of 72 infants followed to date in 2003 and 2004, almost all seemed normal, and none had conclusive laboratory evidence of congenital WNV infection. Three infants had WNV infection that could have been congenitally acquired. Seven infants had major malformations, but only 3 of these had defects that could have been caused by maternal WNV infection based on the timing of the infections and the sensitive developmental period for the specific malformations, and none had any conclusive evidence of WNV etiology. However, the sensitivity and specificity of IgM testing of cord blood to detect congenital WNV infection are currently unknown, and congenital WNV infection among newborns with IgM-negative serology cannot be ruled out. Prospective studies comparing pregnancy outcomes of WNV-infected and -uninfected women are needed to better define the outcomes of WNV infection during pregnancy.

To the Editor: West Nile virus (WNV) is a mosquitoborne flavivirus that is widespread in Africa, the Middle East, Asia, and southern Europe, where it causes outbreaks and sporadic cases of the disease. It has become an emergent disease in North America, where it was detected for the first time in 1999 and became epidemic shortly thereafter (1). Although WNV was initially considered to have a minor health effect in the Mediterranean basin, human and equine outbreaks reported in the last decade in different countries (2–5) have made WNV infections a public health concern. The epidemiology of WNV in Europe differs from that in America and has only been associated with nonrecurrent, sporadic outbreaks. The reasons for this difference are controversial; it may be due to environmental factors, reservoirs, or even mosquito vectors. In Spain, neither equine nor human WNV cases have been reported. However, some human serosurveys that used hemagglutination inhibition suggested that WNV or closely related flaviviruses circulated during the 1970s in the Ebro delta and areas in Spain (6,7). The Ebro delta, a wetland in Catalonia, in the northeast of Spain, is a stopping-off point for birds migrating between regions of Africa and Europe where different WNV vectors and reservoirs have been identified. The delta could be considered a high-risk area for WNV and other arthropodborne virus infections. To evaluate WNV seroprevalence in the human population of the Ebro delta, a survey was conducted in 2001. After obtaining informed consent, 992 serum samples were obtained from inhabitants of the area. The population studied was representative of the whole area and was stratified by sex and age. Anti-WNV immunoglobulin G (IgG) antibodies were determined by using an in-house indirect enzyme-linked immunosorbent assay (ELISA), as previously described (8). Results were classified as the sample absorbance/positive control absorbance ratio. Samples showing ratio values >0.2 were tested for WNV IgG and IgM by using an indirect and a μ-chain capture ELISA, respectively (Focus Technologies, Cypress, CA, USA), and an in-house microneutralization test. For the microneutralization test, samples were tested in duplicate and assayed twice. Twofold dilutions (25 μL) of the samples (1:16–1:256 dilutions) were assayed by using 100 TCID50 (50% tissue culture infectious dose) of West Nile Eg-101 reference strain in 96-well tissue culture plates with Vero cells and after 7 days of incubation at 37°C and 5% CO2. Thirty-eight samples showed IgG ratios >0.2 by the in-house ELISA. Of these, 12 showed WNV IgG, and 1 was positive for WNV IgM and IgG, according to the Focus assays. Two samples showed positive neutralizing activity, with titers of 32 and 256. The highest titer was shown by the sample that yielded positive levels of both IgM and IgG in the ELISA, which suggests recent WNV infection. Anti-WNV IgG was more often detected in participants in the 20- to 29-year age group (odds ratio [OR] 4.23, 95% confidence interval [CI] 1.04–16.02, p = 0.03) and in persons who reported frequent mosquito bites (OR 8.62, 95% CI 0.44–169, p = 0.08). IgG-positive persons were equally divided by sex. No significant differences were found between antibody-positive or antibody-negative persons with respect to their profession, place of occupation, current residence, time in current residence, outdoor activities, use of insecticides and repellents, or symptoms related to WNV infection. No symptoms related to WNV infection were reported by the IgM/IgG-positive participant, who was 31 years of age, was born in the area, worked outdoors, and was frequently bitten by mosquitoes. He also reported travel to Cuba 1 year earlier, but he had not been vaccinated against flavivirus, and serologic test results for dengue were negative. The other IgG- and neutralizing antibody–positive participant was 45 years of age and was born and works in the area. He had never traveled abroad or been vaccinated against flavivirus. He reported a 4-day fever of unknown origin during the summer 1 or 2 years before the study. He often fishes in the areas and is frequently bitten by mosquitoes. In conclusion, the study found evidence of recent WNV infections in humans living in the Ebro delta, where previous flavivirus circulation has been suggested by Lozano and Filipe (6). IgG-positive results not confirmed by neutralization could be due to cross-reactive antibodies induced by other flavivirus infections or vaccinations (9,10). The probable WNV infection described was asymptomatic, as occurs in ≈80% of cases. Other WNV infections in the area may have remained undetected, including neuroinvasive cases. Intensified research and surveillance in this area will help determine and refine thresholds for public health interventions.

Acute flaccid paralysis: the spectrum of a newly recognized complication of West Nile virus infection

West Nile virus (WNV), the causative agent of West Nile fever and West Nile neuroinvasive disease in humans, has become endemic in many countries in all continents. Concerns on long-term mobility from WNV have arisen from recent studies that reported chronic kidney disease in patients who recovered from WNV infection, supported by data from animal models that showed prolonged excretion of the virus with urine. The purpose of this review is to summarize and discuss the results of studies in the literature that investigated WNV infection of the kidney in humans and in animal models and WNV excretion with urine, the potential damage to the kidney caused by WNV infection, the risk of WNV disease in kidney transplant recipients, the significance of detecting WNV in urine and its use in the diagnosis of WNV infection, and kidney involvement by other mosquito-borne flaviviruses.

We describe four solid-organ transplant recipients with donor-derived West Nile virus (WNV) infection (encephalitis 3, asymptomatic 1) from a common donor residing in a region of increased WNV activity. All four transplant recipients had molecular evidence of WNV infection in their serum and/or cerebrospinal fluid (CSF) by reverse transcription polymerase chain reaction (RT-PCR) testing. Serum from the organ donor was positive for WNV IgM but negative for WNV RNA, whereas his lymph node and spleen tissues tested positive for WNV by RT-PCR. Combination therapy included intravenous immunoglobulin (4 cases), interferon (3 cases), fresh frozen plasma with WNV IgG (2 cases), and ribavirin (1 case). Two of the four transplant recipients survived.Review of the 20 published cases of organ-derived WNV infection found that this infection is associated with a high incidence of neuroinvasive disease (70%) and severe morbidity and mortality (30%). Median time to onset of symptomatic WNV infection was 13 days after transplantation (range 5-37 days). Initial unexplained fever unresponsive to antibiotic therapy followed by rapid onset of neurologic deficits was the most common clinical presentation. Confirmation of infection was made by testing serum and CSF for both WNV RNA by RT-PCR and WNV IgM by serological assays. Treatment usually included supportive care, reduction of immunosuppression, and frequent intravenous immunoglobulin. The often negative results for WNV by current RT-PCR and serological assays and the absence of clinical signs of acute infection in donors contribute to the sporadic occurrence of donor-derived WNV infection. Potential organ donors should be assessed for unexplained fever and neurological symptoms, particularly if they reside in areas of increased WNV activity.

In addition to neuroinvasive disease, West Nile virus (WNV) infection is frequently associated with self-limiting chorioretinitis and vitritis. However, the mechanisms of ophthalmic WNV infection are rarely investigated, in part because of the lack of reliable in vitro models. The authors therefore established the first model of ocular WNV infection and investigated interaction of WNV with IFN signal-transduction mechanisms. Human retinal pigment epithelial (RPE) cells were infected with WNV strain NY385-99 at a multiplicity of infection of 5. Virus replication was evaluated by virus titers at different times after infection. The susceptibility of RPE cells to WNV infection was confirmed by transmission electron microscopy. IFN-beta expression was assessed by quantitative real-time PCR and by measurements of antiviral activity in cell culture supernatants. IFN signaling was evaluated by phosphorylation of transducer and activator of transcription 1 and 2 (STAT1/2) proteins, with immunoblot analysis. RPE cells appeared to be highly sensitive to WNV infection. Maximum viral titers were found 24 hours after infection, followed by a continuous decline during the course of infection. WNV infection of RPE cells was followed by increased IFN-beta expression associated with IFN signaling and subsequent inhibition of WNV replication. In this study, the first cell culture model of ophthalmic WNV infection was developed and characterized in RPE cells, and the molecular mechanisms of WNV infection were studied. The data suggest that WNV induces a general antiviral state in RPE cells. This general antiviral state correlates with WNV-induced IFN signaling in retinal cells.

To describe the prevalence of West Nile virus (WNV) infection and evaluate factors associated with positive IgM capture ELISA results in equids with clinical signs compatible with WNV infection. Retrospective case series. Laboratory submission forms from 1,104 equids tested for WNV in Colorado in 2003. Submission forms accompanying samples submitted for detection of WNV via IgM capture ELISA were obtained from the Colorado state veterinarian and diagnostic laboratories performing the tests. Data on signalment, clinical signs, history of vaccination against WNV, and assay results were collected from laboratory submission forms. Equids with clinical signs compatible with WNV infection in which IgM capture ELISA results were positive were considered as case equids. 1,104 equids were tested for WNV; 1,017 (92.1%) had clinical signs compatible with WNV infection. Among equids with clinical signs compatible with WNV infection, the odds of testing positive for WNV via IgM capture ELISA were lower in males and in vaccinated equids and higher in equids with moderate and severe illness, compared with females, unvaccinated equids, and equids with mild illness. Among equids with clinical signs compatible with WNV infection, vaccination against WNV, severity of clinical signs, duration of illness, and region in Colorado were associated with increased risk of having a positive IgM capture ELISA result.

Coronavirus disease 2019 (COVID-19) has had a major effect on kidney and other solid organ transplant recipients.1 In addition to public health measures, improved access to testing, and therapeutic developments, vaccination has emerged as a key tool for controlling the ongoing pandemic. In December 2020, multiple regulatory agencies worldwide authorized the use of two mRNA vaccines for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and several other vaccine platforms are in advanced-stage clinical trials.2,3 Individuals who have received a transplanted kidney or other solid organ have been identified as high-risk populations and prioritized for vaccination in public health guidelines, but unfortunately have been excluded from major SARS-CoV-2 vaccine clinical trials.1,2 Thus, studies are urgently needed to characterize the safety, immunogenicity, and ultimately, efficacy of SARS-CoV-2 vaccines for such patients. Below, we provide an overview of SARS-CoV-2 vaccines and highlight key concepts that should be considered in evaluating their safety in solid organ transplant recipients. Despite the theoretical concerns described below, we emphasize that available evidence from studies demonstrates safety and efficacy in the general population. Because of the known substantial risks of COVID-19–associated morbidity and mortality in recipients of kidney and other solid organs, and the long track record of safety of other vaccinations in such recipients, we anticipate the benefits of selected SARS-CoV-2 vaccines will far outweigh risks of vaccination. Accordingly, current guidance from multiple professional organizations recommend vaccination for all eligible organ transplant recipients.2,4,5 Each vaccine platform has distinct safety considerations for kidney transplant recipients. Live (replication-competent) vaccines are generally contraindicated in immunocompromised individuals because of a risk of vaccine-acquired disease.6 The SARS-CoV-2 candidate vaccines that are furthest along in development do not contain replication-competent SARS-CoV-2 virus, and therefore do not carry risk of SARS-CoV-2 infection (Table 1). Table 1. - Major SARS-CoV-2 platforms in developmenta Vaccine Platform Vaccine Name (Manufacturer) Vehicle Phase of Development Adjuvant Safety and Efficacy in the General Population Specific Considerations for Kidney Transplant Recipients mRNA BNTb162b2 (Pfizer/BioNTech) mRNA encapsulated in lipid nanoparticles Authorized for emergency use in the United States and other countries Unadjuvanted, but lipid nanoparticles possess natural adjuvant activity7 95% efficacy in phase 3 trials.1 Anaphylaxis has been reported. Avoid in patients with a known allergy to a vaccine component (e.g., polyethylene glycol). Close monitoring after administration for patients with a history of anaphylaxis to any food or drug.3 Does not contain live virus. No evidence of vaccine-induced off-target immune responses in large phase 3 clinical trials.2,3 mRNA-1273 (Moderna) Replication-defective viral vectors AZD122 (Oxford/AstraZeneca) Human-chimpanzee adenovirus (ChAdOx1) Phase 3 Unadjuvanted 70%–90% efficacy depending on dose in phase 3 trials.8 Transverse myelitis reported.8 Removal of genes necessary for replication reduces risk of vaccine-associated AdV disease.9 Theoretical risk of emergence of new AdV type with replicative potential through homologous recombination, although this has never been demonstrated to occur with AdV-vectored vaccines.9 JNJ78436735/Ad26.COV2.S (Janssen) Human adenovirus (Ad26) Phase 3 Unadjuvanted Unknown Convidecia (Ad5-nCov) Human adenovirus (Ad5) Approved for limited use in China Unadjuvanted Unknown Sputnik V (Gamaleya) Human adenovirus (Ad5 and Ad26 in consecutive doses) Early use in Russia, Belarus, and Argentina Unadjuvanted Unknown Protein subunit NVX-CoV2373 (Novavax) Recombinant spike glycoprotein Phase 3 Matrix-M1 system plus an additional, unnamed adjuvant Unknown Does not contain live virus. Matrix-M1 contains the same QS21 saponin as the AS01B adjuvant system contained in the recombinant varicella zoster vaccine.7 SARS-CoV-2 recombinant protein formulation (GSK/Sanofi) Recombinant spike protein Phase 2 AS03 adjuvant Unknown High incidence of anti-HLA antibodies in KTR vaccinated with AS03-adjuvanted influenza vaccines, but no association between AS03 exposure and rejection.3,10 EpiVacCorona (Vector Institute) Peptide epitope Early use in Russia Unknown Limited data available Whole-inactivated (killed) BBIBP-CoV (Sinopharm) CoronaVac (SinoVac) Whole-inactivated SARS-CoV-2 viral particles Limited use in China and other countries Unknown Unknown Does not contain live virus. Limited data available in peer-reviewed literature. GSK, GlaskoSmithKline; KTR, kidney transplant recipients.aDoes not include all candidate vaccines or platforms under investigation; limited to platforms in advanced stages of clinical development or authorized for use as of December 31, 2020. However, viral vector–based vaccines that incorporate viruses other than SARS-CoV-2 are in advanced-phase studies, including adenovirus (AdV) vector–based vaccines that have been licensed in Europe. These vaccines consist of intact virions that are engineered to include the gene encoding the SARS-CoV-2 spike protein, a technique that leverages the viral vector's ability to efficiently infect cells and enhances spike gene delivery. Vaccines that use viral vectors contain either replication-deficient or replication-competent viruses (Table 1). The majority of viral-vectored vaccines in the most advanced phases of development have been rendered replication-deficient through deletion of genes essential for replication.8 By limiting vector replication, the potential for vaccine-associated AdV disease is greatly diminished. There are, however, theoretical mechanisms by which replication-deficient viral vector–based vaccines could become replication competent and cause disease, especially in immunocompromised individuals. For example, in cells concurrently infected with two different AdVes, homologous recombination of genetic elements could occur and result in the emergence of new, pathogenic, replication-competent AdV types.9 This has been observed in patients with advanced HIV disease during natural AdV infections, and is theoretically possible with AdV vector–based vaccines in patients who are immunocompromised with a concurrent wild-type AdV infection.9 Although infrequent, severe AdV infections, including allograft nephritis, can occur in kidney transplant recipients during natural infection.9 Notably, vaccine-associated AdV disease has not been reported, albeit there is little experience in immunocompromised populations. It should be emphasized that, despite the theoretical concerns with replication-deficient viral vector–based vaccines, immunosuppression is not considered a contraindication to their use.11 Replication-competent viral-vectored vaccines carry a greater risk of vaccine-derived vector infection in patients who are immunocompromised and should only be administered under carefully controlled circumstances (specifically, clinical trials). Other vaccine candidates that are in advanced stages of development, including mRNA, protein subunit, or whole virus–inactivated SARS-CoV-2 vaccines, do not contain intact virus and thus do not carry a risk of vaccine-associated infection.3 Induction of generalized systemic inflammation by either the vaccine antigen or an associated adjuvant, or by more specific cellular and humoral crossreactivity between vaccine epitopes and allograft antigens, theoretically could promote undesired allograft-directed immune responses. AdV vectors elicit potent innate immune responses through complement activation and induce a diverse cytokine repertoire.10 Although this phenomenon is most prominent at the site of AdV-vector inoculation, systemic inflammation could promote vaccine-associated allograft rejection. Concern for autoimmunity related to the SARS-CoV-2 vaccine on the basis of a modified chimpanzee AdV vector (ChAdOx1) arose after two vaccine recipients developed transverse myelitis, although the possibility of an unrecognized preexisting demyelinating condition has raised questions about the significance of one of these events.8In vitro reactivity between spike protein antibodies and human collagen has been demonstrated, but molecular mimicry has not been identified as a primary mechanism of kidney injury in COVID-19.12 Available data suggest acute allograft rejection is uncommon during COVID-19, despite frequent reduction in immunosuppression as a therapeutic strategy.1 In the absence of an observed association between natural SARS-CoV-2 infection and acute allograft rejection in kidney transplant recipients, it is unlikely that vaccine antigens would precipitate clinically significant immune responses to renal allografts. In general, adjuvants used to enhance vaccine immunogenicity also elicit nonspecific inflammatory responses, and thus have the potential to induce acute allograft rejection. Concern about adjuvant safety in organ transplant recipients arose from observations of an unusually high incidence of anti-HLA antibodies in kidney transplant recipients who received the 2009 influenza A(H1Na1)pdm09 vaccine, which contained the squalene-based AS03 adjuvant system.6,7 However, only a fraction of these anti-HLA antibodies were donor specific, and a subsequent investigation of >10,000 solid organ transplant recipients found no definitive association between the AS03 adjuvant system and acute allograft rejection.6 The AS01B adjuvant used in the recombinant varicella zoster virus vaccine contains a combination of monophosphoryl lipids and QS21, a saponin.13 This adjuvant induces a potent innate immune response and associated concerns for precipitating acute allograft rejection in kidney transplant recipients. Several recombinant spike protein SARS-CoV-2 vaccines contain adjuvants, such as AS03 and the novel Matrix M1 adjuvant, which contains the same QS21 saponin found in the recombinant varicella zoster vaccine.3 Viral-vectored and mRNA vaccines do not generally contain adjuvants, although lipid nanoparticle delivery devices used in the mRNA vaccines have natural adjuvant activity.13 Postmarketing surveillance will be essential to assess any potential association between SARS-CoV-2 vaccine components and acute allograft rejection. In the interim, theoretical concerns associated with any vaccine must be weighed against the benefits of preventing or mitigating the severity of a life-threatening infection in a vulnerable population. We emphasize that a broad range of vaccines have a substantial track record of safety in kidney and other solid organ transplant recipients. Furthermore, no definitive association between any vaccine or adjuvant and allograft rejection has been identified to date.2,6 Immunosuppression in kidney transplant recipients is anticipated to reduce the immunogenicity of SARS-CoV-2 vaccines, and immunogenicity may vary by vaccine platform. Available data across a broad range of vaccines in solid organ transplant recipients suggest they have relative humoral response rates that are approximately 50%–70% of those seen in nontransplant populations.6,7 Patients with ESRD may have more a robust response to vaccines before rather than after kidney transplant,6 and when possible, SARS-CoV-2 vaccines should be given before transplantation.2 In the post-transplant setting, age >65 years, more recent transplantation, use of mycophenolate and mammalian target of rapamycin inhibitors, and lower graft function are associated with decreased serologic responses to influenza vaccines.6,7 Despite the effects of lymphocyte-depleting immunosuppression in the early period after transplant or treatment for rejection, the benefits of even modest SARS-CoV-2 protection might outweigh the risk of delaying immunization during a pandemic.2 In general, vaccines are not recommended immediately post-transplant due to a presumed decrease in immunogenicity after recent high-level immunosuppression. Expert opinion advises that delaying SARS-CoV-2 vaccination of vaccine-naive transplant recipients until 3 months after transplant or receipt of T cell or B cell ablative therapies may be appropriate; for patients who received a first dose before transplant, administration of the second dose should be delayed until at least 4 weeks post-transplant.2 Higher doses, booster doses, adjuvants, and intradermal delivery have all been used with variable success to improve immunogenicity of commonly administered vaccines in solid organ transplant recipients.6,7 If immunogenicity of standard regimens in kidney transplant recipients is suboptimal, these alternative approaches should be considered. The diversity of vaccine platforms and phased vaccine allocation present unique challenges for assessing the safety, immunogenicity, and efficacy of SARS-CoV-2 vaccination in kidney transplant recipients. Surveillance for adverse events related to each specific formulation through prospective multicenter registries of vaccinated solid organ transplant recipients is one potential approach to assessing safety, especially given that large population-specific studies of kidney transplant recipients may not be feasible. Prospective clinical trials should directly compare different SARS-CoV-2 vaccine platforms, assess the magnitude and durability of humoral and cellular responses, and utilize the same functional laboratory immunogenicity assays as the vaccine trials to facilitate direct comparisons between transplant recipients and general populations. It is hoped these investigations will identify relevant differences among the various vaccine platforms to guide future studies of alternative vaccination strategies, if warranted. Because immunosuppression may increase SARS-CoV-2 viral load and prolong the duration of SARS-CoV-2 viral shedding and transmissibility, studies to monitor both symptomatic and asymptomatic infection in vaccinated kidney transplant recipients—with quantitation of viral loads, viral culture, or both—would complement studies of safety and immunogenicity. Large prospective studies of vaccine efficacy in kidney transplant recipients that include a placebo arm are likely not feasible and may not be ethically appropriate, depending on the final results of ongoing phase 3 studies. Case-control trials of kidney transplant recipients with COVID-19 that examine the effect of prior vaccination on disease severity, viral load, and duration of viral persistence, although less definitive, may offer a more practical approach. SARS-CoV-2 vaccines have significant potential to reduce COVID-19–associated morbidity and mortality among recipients of solid organ transplants, including kidney transplants. Because transplant recipients' responses to vaccines may be suboptimal, continued emphasis on nonvaccine preventive measures—use of face covers, hand hygiene, and physical distancing—will be needed, even after vaccination.2 Although the vaccines' ability to disrupt viral transmission in either immunocompetent or immunocompromised individuals is not yet established, vaccination of caregivers and close contacts of kidney transplant recipients, recommended for influenza vaccination, would be an important strategy to reduce the risk of infection.6 Assessment of vaccine efficacy against emerging SARS-CoV-2 variants is necessary to establish optimal vaccine strategies for both immunocompetent and immunocompromised populations. Future evaluations of SARS-CoV-2 vaccine platforms in kidney transplant recipients are imperative to confirm safety and immunogenicity, but the expectation is that SARS-CoV-2 vaccines will add to the armamentarium of vaccines that have safely protected transplant recipients from serious infectious diseases for decades. Disclosures A. Limaye reports having consultancy agreements with AlloVir, Amplyx, GSK, Merck, NovaNordisk, Novartis, and Sana Biotech and being a scientific advisor or member with NobelPharma and Novartis. M.R. Heldman reports receiving speaking honoraria from CignaLife Source, outside the scope of the submitted work. Funding This work was supported by the National Institute of Allergy and Infectious Diseases (T32AI118690 to M.R. Heldman). The content of this work is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

West Nile virus (WNV) infections were first detected in Germany in 2018, but information about WNV seroprevalence in horses is limited. The study’s overall goal was to gather information that would help veterinarians, horse owners, and veterinary-, and public health- authorities understand the spread of WNV in Germany and direct protective measures. For this purpose, WNV seroprevalence was determined in counties with and without previously registered WNV infections in horses, and risk factors for seropositivity were estimated. The cohort consisted of privately owned horses from nine counties in Eastern Germany. A total of 940 serum samples was tested by competitive panflavivirus ELISA (cELISA), and reactive samples were further tested by WNV IgM capture ELISA and confirmed by virus neutralization test (VNT). Information about potential risk factors was recorded by questionnaire and analyzed by logistic regression. A total of 106 serum samples showed antibodies against flaviviruses by cELISA, of which six tested positive for WNV IgM. The VNT verified a WNV infection for 54 samples (50.9%), while 35 sera neutralized tick-borne encephalitis virus (33.0%), and eight sera neutralized Usutu virus (7.5%). Hence, seroprevalence for WNV infection was 5.8% on average and was significantly higher in counties with previously registered infections (p = 0.005). The risk factor analysis showed breed type (pony), housing in counties with previously registered infections, housing type (24 h turn-out), and presence of outdoor shelter as the main significant risk factors for seropositivity. In conclusion, we estimated the extent of WNV infection in the resident horse population in Eastern Germany and showed that seroprevalence was higher in counties with previously registered equine WNV infections.

In Hamburg, Northern Germany, no locally acquired human Tick-borne encephalitis virus (TBEV) or West Nile Virus (WNV) infections have been recorded as of to date. Since 2018, several WNV infections have been diagnosed in birds/horses. As a majority of TBEV and WNV infections are asymptomatic, diagnoses may be missed. Our objectives were to investigate TBEV- and WNV-antibody prevalence and TBE-protection rates through vaccination in Hamburg. In May 2023, we conducted a TBEV- and WNV-seroprevalence study in 1,321 blood donors at a large donation service; additionally, the percentage of TBE vaccination-induced seroprotection rates were assessed. Laboratory analyses were conducted at the National TBEV Consultant Laboratory. Geographical patterns of TBEV and WNV infections in humans and WNV infections in birds/horses were evaluated. The median age of donors was 43 years (IQR 32-56), 59% were male. TBEV-prevalence was 0.30%. In 24% of blood donors, we detected TBEV-vaccination-induced antibodies; WNV-seroprevalence was 0.30%. No geographical pattern or proximity between TBEV infections was observed. Three human WNV-infections were detected in individuals living in relative proximity. In the non-endemic area Hamburg, a low TBEV and WNV-prevalence could be detected in blood donors. For the first time, TBEV vaccination rates in this non-TBEV-risk area were assessed. Due to the healthy donor effect, vaccination rates may be overestimated. Despite the low seroprevalence of TBEV and WNV in our study, they highlight the need for continuous disease surveillance and local physicians should be aware of possible TBEV and WNV infections, especially in patients with neurological symptoms.

West Nile virus-induced pancreatitis

Lipocalin 2 (Lcn2) is a bacteriostatic factor produced during the innate immune response to bacterial infection. Whether Lcn2 has a function in viral infection is unknown. We investigated the regulation and function of Lcn2 in the central nervous system (CNS) of mice during West Nile virus (WNV) encephalitis. Lcn2 mRNA and protein were induced in the brain by day 5, and this induction increased further by day 7 postinfection but was delayed compared with the induction of the toll-like receptor 3 (TLR3) gene, retinoic acid-inducible gene 1 (RIG-I), and melanoma differentiation-associated protein 5 (MDA5) gene. The Lcn2 mRNA and protein were both found at high levels in the choroid plexus, vascular endothelium, macrophage/microglia, and astrocytes. However, some neuronal subsets contained Lcn2 protein but no detectable mRNA. In Lcn2 knockout (KO) mice, with the exception of CXC motif chemokine 5 (CXCL5), which was significantly more downregulated than in wild-type (WT) mice, expression levels of a number of other host response genes were similar in the two genotypes. The brain from Lcn2 and WT mice with WNV encephalitis contained similar numbers of infiltrating macrophages, granulocytes, and T cells. Lcn2 KO and WT mice had no significant difference in tissue viral loads or survival after infection with different doses of WNV. We conclude that Lcn2 gene expression is induced to high levels in a time-dependent fashion in a variety of cells and regions of the CNS of mice with WNV encephalitis. The function of Lcn2 in the host response to WNV infection remains largely unknown, but our data indicate that it is dispensable as an antiviral or immunoregulatory factor in WNV encephalitis.

A Cluster Study of Predictors of Severe West Nile Virus Infection

SummaryThe Toll-like receptor (TLR) 7 response represents a vital host defense 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 wild-type keratinocytes. Infection of wild-type keratinocytes induced higher levels of IFN-α, IL-1β, IL-6 and IL-12, which might promote LC migration from the skin. Coculture of naive 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 LCs migration from the skin, which might compromise its protective effect in systemic infection.