Satu Kurkela

Characterization of a novel flavivirus from mosquitoes in northern europe that is related to mosquito-borne flaviviruses of the tropics.

ABSTRACT A novel flavivirus was isolated from mosquitoes in Finland, representing the first mosquito-borne flavivirus from Northern Europe. The isolate, designated Lammi virus (LAMV), was antigenically cross-reactive with other flaviviruses and exhibited typical flavivirus morphology as determined by electron microscopy. The genomic sequence of LAMV was highly divergent from the recognized flaviviruses, and yet the polyprotein properties resembled those of mosquito-borne flaviviruses. Phylogenetic analysis of the complete coding sequence showed that LAMV represented a distinct lineage related to the Aedes sp.-transmitted human pathogenic flaviviruses, similarly to the newly described Nounané virus (NOUV), a flavivirus from Africa (S. Junglen et al., J. Virol. 83:4462-4468, 2009). Despite the low sequence homology, LAMV and NOUV were phylogenetically grouped closely, likely representing separate species of a novel group of flaviviruses. Despite the biological properties preferring replication in mosquito cells, the genetic relatedness of LAMV to viruses associated with vertebrate hosts warrants a search for disease associations.

Causative agent of Pogosta disease isolated from blood and skin lesions.

Pogosta disease is a mosquito-borne viral disease in Finland, which is clinically manifested by rash and arthritis; larger outbreaks occur in 7-year intervals. The causative agent of the disease has been suspected of being closely related to Sindbis virus (SINV). We isolated SINV from five patients with acute Pogosta disease during an outbreak in fall 2002 in Finland. One virus strain was recovered from a whole blood sample and four other strains from skin lesions. The etiology of Pogosta disease was confirmed by these first Finnish SINV strains, which also represent the first human SINV isolates from Europe. Phylogenetic analysis indicates that the Finnish SINV strains are closely related to the viral agents isolated from mosquitoes and that cause clinically similar diseases in nearby geographic areas.

Sindbis Virus Infection in Resident Birds, Migratory Birds, and Humans, Finland

Resident grouse may be involved in the epidemiology of SINV in humans.

Novel insect-specific flavivirus isolated from northern Europe

Mosquitoes collected in Finland were screened for flaviviral RNA leading to the discovery and isolation of a novel flavivirus designated Hanko virus (HANKV). Virus characterization, including phylogenetic analysis of the complete coding sequence, confirmed HANKV as a member of the “insect-specific” flavivirus (ISF) group. HANKV is the first member of this group isolated from northern Europe, and therefore the first northern European ISF for which the complete coding sequence has been determined. HANKV was not transcribed as DNA in mosquito cell culture, which appears atypical for an ISF. HANKV shared highest sequence homology with the partial NS5 sequence available for the recently discovered Spanish Ochlerotatus flavivirus (SOcFV). Retrospective analysis of mitochondrial sequences from the virus-positive mosquito pool suggested an Ochlerotatus mosquito species as the most likely host for HANKV. HANKV and SOcFV may therefore represent a novel group of Ochlerotatus-hosted insect-specific flaviviruses in Europe and further afield.

Sindbis virus as a human pathogen-epidemiology, clinical picture and pathogenesis.

Sindbis virus (SINV; family Togaviridae, genus Alphavirus) is an enveloped RNA virus widely distributed in Eurasia, Africa, Oceania and Australia. SINV is transmitted among its natural bird hosts via mosquitoes. Human disease caused by SINV infection has been reported mainly in South Africa and in Northern Europe. Vector mosquito abundance affects the annual incidence of SINV infections with occasional outbreaks of up to 1500 patients. Symptoms include fever, malaise, rash and musculoskeletal pain. In a significant portion of patients the debilitating musculoskeletal symptoms persist for years. Chronic disease after SINV infection shares many features with autoimmune diseases. Currently there is no specific treatment available. Recently SINV infections have been detected outside the previously known distribution range. In this article we will summarize the current knowledge on epidemiology, clinical disease and pathogenesis of SINV infection in man. Copyright

Novel flaviviruses from mosquitoes: mosquito-specific evolutionary lineages within the phylogenetic group of mosquito-borne flaviviruses.

Novel flaviviruses that are genetically related to pathogenic mosquito-borne flaviviruses (MBFV) have been isolated from mosquitoes in various geographical locations, including Finland. We isolated and characterized another novel virus of this group from Finnish mosquitoes collected in 2007, designated as Ilomantsi virus (ILOV). Unlike the MBFV that infect both vertebrates and mosquitoes, the MBFV-related viruses appear to be specific to mosquitoes similar to the insect-specific flaviviruses (ISFs). In this overview of MBFV-related viruses we conclude that they differ from the ISFs genetically and antigenically. Phylogenetic analyses separated the MBFV-related viruses isolated in Africa, the Middle East and South America from those isolated in Europe and Asia. Serological cross-reactions of MBFV-related viruses with other flaviviruses and their potential for vector-borne transmission require further characterization. The divergent MBFV-related viruses are probably significantly under sampled to date and provide new information on the variety, properties and evolution of vector-borne flaviviruses.

Sindbis virus as a human pathogenepidemiology, clinical picture and pathogenesis

Sindbis virus (SINV; family Togaviridae, genus Alphavirus) is an enveloped RNA virus widely distributed in Eurasia, Africa, Oceania and Australia. SINV is transmitted among its natural bird hosts via mosquitoes. Human disease caused by SINV infection has been reported mainly in South Africa and in Northern Europe. Vector mosquito abundance affects the annual incidence of SINV infections with occasional outbreaks of up to 1500 patients. Symptoms include fever, malaise, rash and musculoskeletal pain. In a significant portion of patients the debilitating musculoskeletal symptoms persist for years. Chronic disease after SINV infection shares many features with autoimmune diseases. Currently there is no specific treatment available. Recently SINV infections have been detected outside the previously known distribution range. In this article we will summarize the current knowledge on epidemiology, clinical disease and pathogenesis of SINV infection in man. Copyright

Diagnostics of Pogosta disease: antigenic properties and evaluation of Sindbis virus IgM and IgG enzyme immunoassays.

Sindbis virus (SINV) is a mosquito-borne causative agent of a fever-rash arthritis, Pogosta disease, as verified recently by virus isolation from acutely ill patients. Pogosta disease occurs annually, but it emerges as unique epidemics every 7 years in Finland; over 10,000 patient samples have been analyzed for SINV antibodies, with over 2000 diagnosed acute SINV infections. However, the performance of these serological tests with a large number of samples has not been described before. The aim of the present study was to characterize and evaluate methods developed for the serodiagnostics of SINV infection, suitable for large sample numbers, and to examine the protein-specific responses to the antigen used. We developed SINV IgM and IgG enzyme immunoassays (EIA) using highly purified SINV. The EIAs were compared to hemagglutination inhibition (HI) and neutralization tests. We studied paired samples from 46 acutely ill patients taken at approximately 2-week intervals, with a verified SINV infection confirmed by a fourfold rise in HI antibody titer. The assay cut-off values and specificity were determined with confirmed negative sera. Protein-specific antibody responses were examined with immunoblot assay. The optical density values of the EIAs correlated well with the HI titers. The sensitivities of the IgM and IgG EIAs were 97.6% and 100%, and specificities were 95.2% and 97.6%, respectively. We consider that a verified serological diagnosis of acute SINV infection requires (1) in addition to a positive IgM result at least a fourfold increase in HI (or IgG) titer between paired sera or (2) a positive IgM result and a negative/borderline IgG result (which excludes old immunity) and specific reaction in HI. Both E1 and E2 glycoproteins of SINV were shown to be recognized by IgM and IgG antibodies early in infection.

Prolonged Myalgia in Sindbis Virus Infection: Case Description and In Vitro Infection of Myotubes and Myoblasts

BACKGROUND Sindbis virus (SINV) is a mosquito-borne alphavirus found in Eurasia, Africa, and Oceania. Clinical SINV infection is characterized by febrile rash and arthritis and sometimes prolonged arthralgia and myalgia. The pathophysiological mechanisms of musculoskeletal and rheumatic disease caused by SINV are inadequately understood. METHODS We studied the muscle pathology of SINV infection ex vivo by examining a unique muscle biopsy obtained from a patient with chronic myalgia and arthralgia 6 months after acute SINV infection and assessed potential genetic predisposing factors by determining the human leukocyte antigen (HLA) and complement factor C4 genes and proteins. In addition, we performed in vitro SINV infections of primary human myoblasts and myotubes. RESULTS In the muscle biopsy we found evidence of muscle regeneration due to previous necrotic lesions likely caused by earlier SINV infection. We showed that human myoblasts and myotubes were susceptible in vitro for SINV infection as the cells became immunoreactive for viral antigens and cytopathic effect was observed. The patient was homozygous for HLA-B*35 alleles and heterozygous for HLA-DRB1*01 and HLA-DRB1*03 alleles and had total deficiency of C4B protein. CONCLUSIONS This study provides new insights concerning pathological processes leading to chronic symptoms in SINV infection and demonstrates for the first time the susceptibility of human myogenic cells to SINV infection.

Complete coding sequence and molecular epidemiological analysis of Sindbis virus isolates from mosquitoes and humans, Finland

Sindbis virus (SINV) is an arthropod-borne alphavirus, which causes rash-arthritis, particularly in Finland. SINV is transmitted by mosquitoes in Finland but thus far no virus has been isolated from mosquitoes. In this study, we report the isolation of the first SINV strain from mosquitoes in Finland and its full-length protein-coding sequence. We furthermore describe the full-length coding sequence of six SINV strains previously isolated from humans in Finland and from a mosquito in Russia. The strain isolated from mosquitoes (Ilomantsi-2005M) was very closely related to all the other Northern European SINV strains. We found 9 aa positions, of which five in the nsP3 protein C terminus, to be distinctive signatures for the Northern European strains that may be associated with vector or host species adaptation. Phylogenetic analyses further indicate that SINV has a local circulation in endemic regions in Northern Europe and no novel strains are frequently being introduced.