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Human bornavirus encephalitis is a severe and often fatal infection caused by variegated squirrel bornavirus 1 (VSBV-1) and Borna disease virus 1 (BoDV-1). We conducted a prospective study of bornavirus etiology of encephalitis cases in Germany during 2018-2020 by using a serologic testing scheme applied along proposed graded case definitions for VSBV-1, BoDV-1, and unspecified bornavirus encephalitis. Of 103 encephalitis cases of unknown etiology, 4 bornavirus infections were detected serologically. One chronic case was caused by VSBV-1 after occupational-related contact of a person with exotic squirrels, and 3 acute cases were caused by BoDV-1 in virus-endemic areas. All 4 case-patients died. Bornavirus etiology could be confirmed by molecular methods. Serologic testing for these cases was virus specific, discriminatory, and a practical diagnostic option for living patients if no brain tissue samples are available. This testing should be guided by clinical and epidemiologic suspicions, such as residence in virus-endemic areas and animal exposure.

Variegated squirrel bornavirus 1 (VSBV-1) is a zoonotic virus that causes fatal encephalitis in humans who are infected after contact with exotic squirrels. We analyzed the brain lesions and the immune responses in all 4 known human cases that showed panencephalitis. Inflammatory infiltrates in areas positive for VSBV-1 RNA and antigen consisted of CD4+ and CD8+ T cells, with perivascular B-cell accumulation. Strong microglial response and bizarre astroglial expansion were present. Areas of malacia contained neutrophils and foamy microglia and macrophages. Immunopathologic examination during infection showed cleavage of caspase 3 in brain cells adjacent to CD8+ cells and widespread p53 expression, hallmarks of apoptosis. Cerebrospinal fluid analyses over time demonstrated increasing protein concentrations and cell counts, paralleled by pathologic lactate elevations in all patients. The most severe cerebrospinal fluid and histologic changes occurred in the patient with the highest viral load, shortest duration of disease, and most medical preconditions.

Limbic encephalitis is commonly regarded as an autoimmune-mediated disease. However, after the recent detection of zoonotic variegated squirrel bornavirus 1 in a Prevost's squirrel (Callosciurus prevostii) in a zoo in northern Germany, we retrospectively investigated a fatal case in an autoantibody-seronegative animal caretaker who had worked at that zoo. The virus had been discovered in 2015 as the cause of a cluster of cases of fatal encephalitis among breeders of variegated squirrels (Sciurus variegatoides) in eastern Germany. Molecular assays and immunohistochemistry detected a limbic distribution of the virus in brain tissue of the animal caretaker. Phylogenetic analyses demonstrated a spillover infection from the Prevost's squirrel. Antibodies against bornaviruses were detected in the patient's cerebrospinal fluid by immunofluorescence and newly developed ELISAs and immunoblot. The putative antigenic epitope was identified on the viral nucleoprotein. Other zoo workers were not infected; however, avoidance of direct contact with exotic squirrels and screening of squirrels are recommended.

To the Editor: Zika virus is an emerging virus that has spread to most countries in Latin America and the Caribbean (1,2). It is transmitted by mosquitoes and through sexual intercourse (3). Most persons infected with Zika virus are asymptomatic or experience mild symptoms (4). However, in a pregnant woman, infection may cause severe pregnancy and birth complications, most notably microcephaly in children infected in utero (5–7). Zika virus infection might also be associated with an increased incidence of Guillain-Barre syndrome (8). Thus, the virus represents a threat to healthcare workers who manage infected patients or diagnostic samples and researchers who work with infectious virus in laboratories.

To the Editor: Mayaro virus (MAYV), a mosquitoborne alphavirus endemic to South America, causes a self-limiting febrile arthralgia syndrome closely resembling Chikungunya fever (1). MAYV has been detected increasingly as imported infections in international travelers returning to Europe and North America (2–9). Joint pain, the most prominent symptom, is often long-lasting (several months), sometimes incapacitating (4,6,7,9), and may recur (8). Arthralgia develops during the acute phase and symmetrically affects the wrists, ankles, and small joints of hands and feet. Joint swelling may occur initially, but permanent joint damage has not been described (5). The clinical disease and diagnostic procedures have been described (1–9), but immunologic parameters and their possible role in the clinical follow-up of patients (i.e., during the postacute long-lasting arthralgia period) remain to be investigated. To further our knowledge of MAYV infection, we analyzed cytokine levels in serum samples from 6 travelers to South America who returned to Europe with Mayaro fever (MF). Two of the cases occurred during 2014; 4 occurred during 2011–2013 (2–5). The 6 travelers comprised 2 men and 4 women who were 20–54 (median 36) years of age (Technical Appendix Table). The 2 most recent cases occurred in spring 2014 in a 28-year-old female student and a 54-year-old male physician. Serologic testing was performed for both patients at the Bernhard Nocht Institute and confirmed by virus neutralization testing (4). The student had traveled for 3 weeks in Ecuador, visiting rainforest villages and hiking in the jungle. During her stay, she had experienced myalgia of the forearms, arthralgia of fingers and toes, subfebrile but elevated body temperatures, and maculopapular exanthema. On examination in Germany, the student had no clinical signs of disease, but she reported arthralgia of the ankles and hands. Laboratory test results showed a slightly increased C-reactive protein level (6.2 mg/L, reference value 12 months. In the examined patients, the prolonged arthralgia recovery phase was paralleled by significantly increased proinflammatory cytokine levels, indicating ongoing inflammation, probably related to arthritis. Elevated levels of RANTES and IP-10 suggest T-cell recruitment, possibly reflecting virus persistence and replication, as described for the related Chikungunya virus (10). Thus, cytokine measurements may be helpful for monitoring patient symptoms, especially when signs of arthritis (swellings and redness) and elevated standard serum inflammatory parameters are no longer present. In our study, the picture of MF cytokine elevations paralleled those described for Chikungunya virus infection (10); these elevated levels may help to elucidate the pathogenesis of MAYV-induced arthralgia. More immunology data are required to complete this evolving picture of viral arthralgia syndromes. Technical Appendix: International travelers who were recently diagnosed with Mayaro fever and were included in this study. Click here to view.(151K, pdf)

To the Editor: Usutu virus (USUV) is an arthropod-borne flavivirus that belongs to the Japanese encephalitis serocomplex (1). USUV circulates between ornithophilic mosquito vectors (mainly Culex spp. mosquitoes) and avian amplification hosts (2). Migratory birds play a key role in the introduction of USUV into new areas (3). USUV has recently been introduced from Africa into Europe, causing epizootics among wild birds and Usutu fever in humans (4–6). The detection and isolation of USUV from different bird and mammalophilic mosquitoes during the epizootic in Germany raise questions regarding the USUV host range (2,3). Bats have been considered natural reservoir hosts of a wide diversity of viruses, including several flaviviruses (7,8). Their ability to fly and their social behavior enable efficient maintenance, spread, and evolution of viruses.

To the Editor: Dirofilariosis caused by infection with the filarial nematode Dirofilaria repens is considered an emerging zoonosis in Europe (1). The main reservoirs for this parasite are dogs and other carnivores. As is the case for other filarial species, mosquitoes transmit infectious third-stage larvae, which develop into fertile adults in their definitive vertebrate hosts. Humans may become infected as aberrant hosts, and, in most cases, these worms remain infertile (1,2). Infections in humans usually manifest as subcutaneous nodules, which are caused by developing worms that are trapped by immune mechanisms (2). Subcutaneous migration of a worm may result in local swelling. Severe clinical manifestations have also been reported and may affect various organs, including the brain, lung, and eye (2,3). Eye infections are found in particular during the migratory phase of the parasite. Transmission of D. repens occurs in various regions of the Old World, including Europe, Africa, and Asia. The primary areas in Europe to which D. repens is endemic are countries of the Mediterranean region, where warm temperatures enable development of infectious third-stage larvae in mosquitoes. However, during the past decade, several autochthonous cases of canine and human dirofilariasis have been reported from countries farther north, including Austria, the Czech Republic, and Poland (4–6). Several factors might be responsible for this spread to the north, including climate change and increased translocation of dogs from southern to central Europe. Until recently, central Europe, including Germany, was not considered a region to which D. repens was endemic. However, a survey in 44 hunting dogs from the Upper Rhine region showed that 3 animals were positive for D. repens microfilariae (7). In addition, testing of a kennel of sled dogs located in the federal state of Brandenburg, near Berlin, found 8 of 28 animals were infected with the parasite (8). To determine whether local transmission of D. repens is taking place in Germany, we retrospectively analyzed 74,547 mosquitoes from a large-scale German mosquito-borne virus surveillance program for the presence of filarial DNA. Mosquitoes were collected during the main animal-trapping seasons (May–September) 2011 and 2012 by using carbon dioxide–baited encephalitis virus surveillance or gravid traps (9). Mosquitoes were collected from 55 trapping sites located in 9 federal states in the southwest and northeastern parts of Germany (Figure), frozen at −70°C, and transported to the laboratory at the Bernhard Nocht Institute, where they were classified morphologically to the species level. Up to 25 mosquitoes of the same species from individual collection sites were subsequently combined to create a total of 4,113 pools. These pools comprised a broad range of ornithophilic and mammalophilic mosquitoes common in Germany; Culex spp. and Aedes vexans were the most abundant species. Figure Origin of mosquito samples analyzed for infection with Dirofilaria repens, by federal state, Germany, May–September 2011 and 2012 (n = 74,547). Black dots indicate collection locations of mosquitoes that tested positive for D. repens DNA. Nucleic acids were extracted from each mosquito pool, and the presence of filarial DNA was determined by PCR using a set of generic primers that amplify DNA from a broad range of filarial species, including those affecting birds and other animals (9). A total of 1,050 (25.5%) of the 4,113 pools were found to be positive for filarial DNA. To further determine if any of these pools contained DNA from D. repens, we analyzed all 1,050 pools by PCR using the D. repens–specific primers 5′-GAGATGGCGTTTCCTCGTG-3′ and 5′-GACCATCAACACTTAAAG-3′. Four pools were positive for D. repens DNA; all of these pools consisted of mammalophilic mosquitoes collected at 2 trapping sites in the federal state of Brandenburg, near Oder Valley, at a latitude of 52°N (global positioning satellite coordinates 52°47′N, 14°14′E, and 52°51′N, 14°7′E). One pool, consisting of Culiseta annulata mosquitoes, was collected in August 2011; the other 3 pools, 2 of Anopheles maculipennis sensu lato and 1 of Ae. vexans mosquitoes, were trapped in July and August 2012. The identification of D. repens DNA in all 4 pools was confirmed by DNA sequencing of a 538-bp fragment of the gene encoding cytochrome oxidase 1 (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"KF410864","term_id":"556527619","term_text":"KF410864"}}KF410864). Sequence comparisons indicated 99.8% identity to the same gene in a D. repens nematode isolate from Italy (accession no. {"type":"entrez-nucleotide","attrs":{"text":"AM749234","term_id":"183579534","term_text":"AM749234"}}AM749234). In conclusion, we found mosquitoes that were positive for D. repens DNA at a latitude of 52°N in northern Germany; these mosquitoes, from 3 species, were collected at 2 locations in 2011 and 2012. Our findings support recent projections that have suggested increases in average temperatures could affect climatic conditions during summer sufficiently to enable development of D. repens larvae in central Europe at latitudes of up to 56°N (10). In the federal state of Brandenburg in Germany, conditions for larval development were predicted to be sufficient in each of the past 12 years. These projections, together with our finding of mosquitoes carrying D. repens in Brandenburg in 2 successive years and the recurrent detection of infected dogs in this area (8), present a strong case for the existence of stable local transmission of D. repens in this region of Germany. Technical Appendix: PCR analyses of mosquitoes from various federal states of Germany for the presence of filarial DNA. Click here to view.(114K, pdf)

To the Editor: Zika virus (ZIKV), a member of the family Flaviviridae, is a mosquito-borne virus that is endemic to Africa and Southeast Asia. ZIKV causes illness that is similar to dengue fever, characterized by joint pain, myalgia, headache, and rash (1). ZIKV has caused several recent outbreaks, including one in Micronesia in 2007 (2) and one in French Polynesia (≈30,000 cases) ongoing since October 2013 (3) and spreading to New Caledonia and Easter Island (4). We report the clinical and laboratory findings for a patient with ZIKV infection imported from Tahiti, French Polynesia. The previously healthy 31-year-old woman from Norway was admitted to the Oslo University Hospital, Norway, on December 13, 2013. Six days earlier, she had returned from a 14-day vacation to Tahiti, where she mainly stayed in the capital, Pape'ete, and took a short trip to the island of Mo'orea. One day after her return to Norway, she experienced fever, intense joint pain, and myalgia. Subsequently, a maculopapular rash developed. At the time of admission, her temperature was 37.7°C, and she had enlarged nuchal lymph nodes; injected conjunctivae; and a maculopapular rash on her trunk, extremities, and face (Technical Appendix). Clinical examination findings were otherwise unremarkable. Laboratory tests showed leukopenia of 2.7 × 109 cells/L (reference range 3.5–10 × 109/L), with mild lymphopenia of 1.0 × 109 cells/L (reference range 1.5.–4.0 × 109/L) and neutropenia of 1.4 ×109 cells/L (reference range 1.5–7.3 × 109/L). No thrombocytopenia or elevated liver enzyme levels were detected. C-reactive protein levels (1.4 mg/L) were within reference range. Because of the patient’s clinical picture and travel history, an acute ZIKV infection was suspected and several diagnostic tests were ordered. In a serum sample taken 5 days after symptom onset, no IgM or IgG against ZIKV, dengue virus (DENV), Japanese encephalitis virus, yellow fever virus, or chikungunya virus was detected by in-house indirect immunofluorescence (5,6). Only a weak IgG titer of 1:20 (and no IgM) against tick-borne encephalitis virus was found (cutoff

To the Editor: Mayaro virus (MAYV), a mosquito-borne New World alphavirus of the family Togaviridae, causes a febrile arthralgia syndrome resembling dengue and chikungunya fever. The virus is maintained in a natural cycle involving nonhuman primates and Haemagogus spp. mosquitoes in tropical rainforest areas of South America (1). After an incubation time of 7–12 days following an infectious mosquito bite, rash, fever, headache, and arthralgia develop in patients, followed by restoration to their original conditions after several weeks (1). Outbreaks of Mayaro fever have been reported from the Amazon region (1,2). There are increasing reports of travel-related infections imported from South America to Europe and the United States (3–7). We describe an acute MAYV infection in a German traveler who returned from French Guiana. Full-length MAYV genome amplification was performed on virus obtained from a serum sample of the patient. In August 2013, a 44-year-old woman (bookkeeper) came to an outpatient clinic with fever (temperature ≤38.7°C), chills, a mild headache, severe fatigue, highly painful swelling of small finger joints, and pain in both feet. Symptoms appeared 2 days before when she experienced aches in her wrists and left forefoot. Four days before, the patient had returned from a 2.5-week visit to French Guiana, where she traveled with her partner and caught butterflies. She had conducted these activities during her holidays for the past 5 years, mostly in spring or autumn. In July 2013 at the end of the rainy season, she had many mosquito bites, especially on her hands, despite use of repellents and bed nets. Physical examination showed a body temperature of 38°C, throat enanthema, generalized macular exanthema, and slightly swollen and tender interphalangeal joints of the hands and feet. Her medical history was unremarkable, and her partner was asymptomatic. Laboratory tests showed reference values for hemoglobin concentration; platelet count; and levels of liver enzymes, creatinine, and anti-nuclear and anti–citrulline peptide antibodies. C-reactive protein level was increased (24.2 mg/L; reference value

We present the molecular identification of Apodemus agrarius (striped field mouse) as reservoir host of the Dobrava-Belgrade virus (DOBV) lineage DOBV-Aa in 3 federal states of Germany. Phylogenetic analyses provided evidence for multiple spillover of DOBV-Aa to A. flavicollis, a crucial prerequisite for host switch and genetic reassortment.

To the Editor: Since 2005, new parvoviruses forming a novel genus of the proposed name Partetravirus, within the subfamily Parvovirinae, have been described (1). Human parvovirus 4 (PARV4) with 3 different genotypes globally infects humans (2). A related porcine virus, hokovirus (HoV or porcine partetravirus), was found in wild boar and domestic pig populations in Germany, Romania, China, and the United States, with prevalences of 12%–47%, forming 1 common genotype (3–6). Prevalence figures from sub-Saharan Africa are not available. Furthermore, no information about possibly region-associated genotypes is available for porcine HoV, although it is for human PARV4 from the same genus. We therefore used samples (collected during February–March 2012) from a study investigating hepatitis E virus (HEV) in pigs from Cameroon (7) to analyze the occurrence of porcine HoV in pigs in Africa and to determine the respective genotype.

To the Editor: West Nile virus (WNV) is a single-stranded RNA virus in the family Flaviviridae that is transmitted to humans by mosquitoes. Approximately 80% of WNV infections in humans are asymptomatic, whereas ≈20% of infected persons experience fever, often accompanied by a rash. Less than 1% of infections are manifested as neuroinvasive disease, such as meningoencephalitis, polyradiculoneuritis, and polio-like flaccid paralysis (1). WNV is endemic in Africa, southern Asia, and northern Australia, and only sporadic cases or small epidemics are seen in Europe (2). In 1999, WNV emerged in North America. By 2010, ≈1.8 million persons had become infected, with 12,852 reported cases of meningoencephalitis and 1,308 deaths (2). In Europe, the last notable outbreak of WNV infection occurred in Greece in 2010; 197 persons were infected, and 33 died (3). The Czech Republic, Denmark, France, and the Netherlands reported laboratory-confirmed WNV infections in travelers returning from North America (1). We report a case of WNV meningoencephalitis in a 28-year-old German woman, who sought treatment the emergency department of a hospital in Potsdam, Germany, on September 7, 2011. She had a 3-day history of fever of up to 40°C and mental confusion. Six days before admission, she had returned from a 2-week holiday trip to Ottawa, Ontario, Canada. She had spent most of her time in the city of Ottawa. The patient’s medical history was unremarkable. She was in a reduced general condition because of a severe encephalitic syndrome characterized by somnolence, meningism, fever, and mental confusion. Laboratory investigations revealed leukocytosis with 15.000 leukocytes/µL (reference range 4,400–11,300 leukocytes/µL) and elevated C-reactive protein of 14.8 mg/L (reference

To the Editor: O’nyong-nyong virus (ONNV) is a mosquitoborne RNA virus of the Togaviridae family. The virus was first isolated in June 1959 from serum samples from febrile patients in the northern province of Uganda (1). Unlike ONNV is primarily transmitted by anopheline mosquitoes (2). ONNV is genetically and serologically related to chikungunya virus (CHIKV) (1), but is restricted to the African continent. The clinical picture resembles CHIKV infection, i.e., a self-limited febrile illness characterized by headache, rash, and joint pain. In contrast to CHIKV, ONNV is reported to cause lymphadenopathy more often and affected joints do not show effusions (3).

To the Editor: We report the case of a 27-year-old male Swiss tourist who spent 3.5 weeks (July 6–30, 2011) vacationing in the vicinity of Tarapoto, a small city located in the rainforests of the Amazon Basin in northern Peru. An acute febrile illness developed in the man during the second week of his stay. Signs and symptoms of illness were chills, malaise, frontal headache, generalized myalgia, a self-limiting painful cervical and inguinal lymphadenopathy (lasting ≈1 week), slowly progressive and pronounced polyarthralgic pains of the peripheral joints, and a transient nonpruritic maculopapular rash (starting at the forearms ≈1 week after onset of fever and spreading to the trunk and later to the neck and face before fading after 3 days). The traveler sought care at the local hospital, where physicians diagnosed suspected dengue fever on the basis of the clinical signs and symptoms, and received symptomatic treatment with paracetamol. The fever and other signs subsided within 1 week, except the arthralgia, which did not improve. The polyarthritis initially was accompanied by swelling of the affected joints and showed a symmetric pattern, mainly affecting the small joints of the hands and feet as well as the wrists, ankles, and knees. After returning home to Switzerland, the patient consulted his general practitioner (August 1, 2011) because of persisting, incapacitating joint pains. The patient reported stiffness of the affected joints, mainly in the morning and after immobility. Physical examination of the affected joints did not reveal visible clinical signs of inflammation (swelling, redness, effusion). Laboratory tests were performed for complete blood count, liver and kidney function, C-reactive protein, and serologic testing for dengue virus, chikungunya virus, Borrelia burgdorferi, Chlamydia trachomatis, Epstein-Barr virus, parvovirus B19, Salmonella Typhi, and S. Paratyphi, but none revealed a cause for the symptoms. Over almost 2 more months, the joint pains did not improve; thus, the patient was referred to a rheumatologic clinic and subsequently to the Swiss Tropical and Public Health Institute, Basel, Switzerland, for evaluation of a putative travel-related cause of the polyarthralgia. Because of the patient’s travel history, the course of the illness and clinical signs and symptoms experienced during the journey, and the evolution and characteristics of the persisting joint pains, we suspected an underlying Mayaro virus (MAYV) infection. Serologic testing (indirect immunofluorescence and virus neutralization assays) for several alphaviruses were performed as described (1), and the results (Table) confirmed our presumptive diagnosis. Table Results of serologic testing for a Mayaro virus–infected Swiss traveler returning from the Amazon Basin, northern Peru, 2011* Several viral infections (e.g., dengue, rubella, parvovirus B19, hepatitis B, hepatitis C, HIV, and human T-lymphotropic virus type 1) can be accompanied by arthralgia. However, the most prominent and long-lasting polyarthritic symptoms occur in patients infected by alphaviruses (family Togaviridae). Alphaviruses are arthropod-borne viruses (arboviruses) that circulate among a wide variety of wild animals in relative mosquito vector–specific and host-specific enzootic cycles; infection in humans (dead-end hosts) is almost exclusively incidental. Clinical cases and virus isolation have been reported only from northern South America, where MAYV circulates in an enzootic sylvatic cycle (similar to that for yellow fever) involving forest-dwelling Haemagogus spp. mosquitoes as vectors and nonhuman primates as natural hosts (2). Infections in humans mostly occur sporadically, are strongly associated with occupational or recreational exposure in rainforest environments, and are assumed to represent spillover from the enzootic cycle (2). MAYV was first isolated in Trinidad in 1954; since then, sporadic cases, clusters, outbreaks, and small epidemics of Mayaro fever have been reported from Brazil, Bolivia, Columbia, French Guiana, Guyana, Peru, Venezuela, and Surinam (3). In addition to the clinical cases and virus isolates reported from northern South America, serologic survey findings suggest the presence of MAYV in Costa Rica, Guatemala, and Panama (4) (Figure A1). Clinical signs of this acute, dengue-like, febrile illness last 3–7 days and typically include chills, headache, retro-orbital and epigastric pain, myalgia, arthralgia, nausea, vomiting, diarrhea, and a maculopapular rash (sometimes followed by desquamation) (7). However, as with other alphavirus infections (i.e., chikungunya [Africa, Asia], o’nyong-nyong [Africa], Ross River [Australia, Oceania], Barmah Forest [Australia], and Sindbis [Africa, Europe, Asia, Australia]), the hallmark of MAYV infection is the highly debilitating arthralgia. Permanent damage of the affected joints has not been reported. Hemorrhagic manifestations of MAYV infections are rare but have been described (3). Concerns over the potential emergence of urban transmission of MAYV were raised after a laboratory study showed vector competence of Aedes aegypti mosquitoes (8). International travelers are rarely given a diagnosis of MAYV infection (9,10). This might be attributed to the overall low frequency of the infection; the dengue-like signs and symptoms, which may lead to a misdiagnosis; and the fact that the disease is not well known outside MAYV-endemic regions. Physicians treating patients with signs and symptoms of a dengue-like illness and a recent history of travel to MAYV-endemic areas should consider MAYV infection in the differential diagnosis, especially if arthralgia is prominent and prolonged and dominates the clinical picture.

To the Editor: Old World hantaviruses (family Bunyaviridae) are rodentborne pathogens that can cause hemorrhagic fever with renal syndrome (HFRS) (1). At least 3 different pathogenic hantavirus species have been detected in Europe: Dobrava-Belgrade virus (DOBV), Tula virus, and Puumala virus (PUUV) (1–3). Most human hantavirus infections in Europe are assigned to PUUV transmitted by bank voles (Myodes glareolus, formerly Clethrionomys glareolus). Although PUUV is thought to cause a mild form of HFRS, designated as nephropathia epidemica (NE), severe courses have been described with a case-fatality ratio of up to 0.6% (3).