Evaluation of vector competence for West Nile virus in Italian Stegomyia albopicta (=Aedes albopictus) mosquitoes.

West Nile virus (WNV) is a zoonotic arboviral pathogen transmitted by mosquitoes in a cycle that involves wild birds as reservoir hosts. The virus is responsible for outbreaks of viral encephalitis in humans and horses. In Europe, Culex pipiens (Diptera: Culicidae) is considered to be the main vector of WNV, but other species such as Stegomyia albopicta (=Aedes albopictus) (Diptera: Culicidae) may also act as competent vectors of this virus. Since 2008 human cases of WNV disease have been reported in northeast Italy. In 2011, new areas of southern Italy became involved and a first outbreak of WNV lineage 1 occurred on the island of Sardinia. On the assumption that a potential involvement of St. albopicta in WNV transmission cannot be excluded, and in order to evaluate the competence of this species for the virus, an experimental infection of an St. albopicta laboratory colony, established from mosquitoes collected in Sardinia, was carried out. The results were compared with those obtained in a colony of the main vector Cx. pipiens. The study showed St. albopicta collected on Sardinia to be susceptible to WNV infection, which suggests this Italian mosquito species is able to act as a possible secondary vector, particularly in urban areas where the species reaches high levels of seasonal abundance.

Circulation of West Nile virus lineage 1 and 2 during an outbreak in Italy.

In 2011, from 26 September to 16 October, a small outbreak of West Nile virus (WNV) disease occurred on the island of Sardinia (Italy). According to the national case definition, six cases with acute neurological disease were confirmed in hospitalized patients, and four of them died; one of these was only 34 years old. In two case, WNV RNA was detected in urine, suggesting renal involvement. Sequence analysis showed lineage 1 and 2 circulation.

Epidemiological surveillance of West Nile neuroinvasive diseases in Italy, 2008 to 2011.

We describe the geographical and temporal distribution of West Nile neuroinvasive diseases (WNND) cases in Italy from 2008 to 2011. The increasing number of confirmed human cases from eight in 2008 to 18 in 2009 and the occurrence of the virus in a larger geographical area in 2009 (moving from east to west) prompted the Ministry of Health to publish, in spring 2010, a national programme for WNND human surveillance, comprising veterinary and vector surveillance. Subsequently, in 2011, a new national plan on integrated human surveillance of imported and autochthonous vector-borne diseases (chikungunya, dengue and West Nile disease) was issued. Between 2008 and 2011, 43 cases of WNND were reported from five regions in Italy with a case fatality rate of 16%. The incidence of WNND during the entire study period was 0.55 per 100,000 population (range: 0.06­0.23 per 100,000). During 2011, two new regions (Friuli-Venezia Giulia and Sardinia) reported confirmed cases in humans. Integrated human, entomological and animal surveillance for West Nile virus is a public health priority in Italy and will be maintained during 2012.

Serological evidence of Toscana virus infection in Portuguese patients.

Toscana virus (TOSV) is an emerging Phlebovirus of growing interest as a human pathogen in the Mediterranean Basin. In Portugal, however, little is known about the prevalence of TOSV infection. The aim of this work was to perform a seroprevalence study in patients with requests for laboratory diagnosis of vector-borne viruses. A total of 538 patients with and without neurological signs from 2004 to 2008 were studied by in-house indirect immunofluorescence assay and commercial enzyme-linked immunosorbent assays. A prevalence of 4.2% for IgG antibodies was found in the group of patients with neurological signs. Five (3%) of these had recent infections. In the group with no neurological signs, the IgG prevalence was 1.3%. Two samples, belonging to two patients, were also confirmed with plaque reduction neutralization tests with the TOSV ISS. Phl.3 Italian strain. This work showed that TOSV is present and causing disease from north to south in Portugal. The probable circulation of different phlebovirus serotypes in Portugal emphasizes the need for further studies.

Prevalence of antibodies to phleboviruses and flaviviruses in Peja, Kosovo.

In order to investigate the current and past activity of phlebovirus and flavivirus in Kosovo, a seroprevalence study among 200 blood donors was performed. Positive results were obtained for the phleboviruses TOSV and SFNV, and for a flavivirus of the Japanese Encephalitis group. No positive results for TBEV were observed.

Risk of acquiring tick-borne infections in forestry workers from Lazio, Italy.

The seroprevalence of antibodies to Borrelia burgdorferi and tick-borne encephalitis (TBE) virus was evaluated in a group of forestry rangers in the Lazio region of Italy. One hundred and forty-five forestry rangers and 282 blood donors were examined by two-tiered serological tests for B. burgdorferi and TBE virus. Information on occupation, residence, tick bites, outdoor leisure activities and other risk factors was obtained. The prevalence of IgG/IgM antibodies to B. burgdorferi showed no statistical difference between the two groups, but there was a higher occurrence of IgM antibodies. There were significant differences between indoor and outdoor, urban and rural workplaces among the 145 exposed workers (χ² test: p < 0.001), and a higher risk for outdoor rural than urban tasks was detected among the ten Western blot-tested forestry rangers positive to B. burgdorferi (χ² test: p < 0.1). No seropositivity was observed for the TBE virus. Forestry rangers from the Lazio region did not have a higher risk of Borrelia infection than the blood donors, though an increase in the risk for outdoor tasks in a rural environment was observed.

West Nile virus transmission with human cases in Italy, August - September 2009.

In 2009, to date 16 human cases of West Nile neuroinvasive disease (WNND) have been reported in Italy, in three regions: Veneto, Emilia-Romagna and Lombardia. The number of cases is higher compared with last year when nine cases were identified (eight cases of WNND and one case of West Nile fever) and the geographical distribution indicates spread from east to west.

Arboviruses in Italy.

A brief review of the arboviruses isolated In Italy is presented and discussed. Some of the Arboviruses considered in this paper are endemic in the country and are transmitted by arthropods that play actually a role of vectors, (sand flyes, hard ticks and mosquitoes); other arboviruses, sporadically isolated, are potential agents of emerging human or zoonotic diseases.

Infection with chikungunya virus in Italy: an outbreak in a temperate region.

BACKGROUND: Chikungunya virus (CHIKV), which is transmitted by Aedes spp mosquitoes, has recently caused several outbreaks on islands in the Indian Ocean and on the Indian subcontinent. We report on an outbreak in Italy. METHODS: After reports of a large number of cases of febrile illness of unknown origin in two contiguous villages in northeastern Italy, an outbreak investigation was done to identify the primary source of infection and modes of transmission. An active surveillance system was also implemented. The clinical case definition was presentation with fever and joint pain. Blood samples were gathered and analysed by PCR and serological assays to identify the causal agent. Locally captured mosquitoes were also tested by PCR. Phylogenetic analysis of the CHIKV E1 region was done. FINDINGS: Analysis of samples from human beings and from mosquitoes showed that the outbreak was caused by CHIKV. We identified 205 cases of infection with CHIKV between July 4 and Sept 27, 2007. The presumed index case was a man from India who developed symptoms while visiting relatives in one of the villages. Phylogenetic analysis showed a high similarity between the strains found in Italy and those identified during an earlier outbreak on islands in the Indian Ocean. The disease was fairly mild in nearly all cases, with only one reported death. INTERPRETATION: This outbreak of CHIKV disease in a non-tropical area was to some extent unexpected and emphasises the need for preparedness and response to emerging infectious threats in the era of globalisation.

Prevalence and incidence of antibodies to Borrelia burgdorferi and to tick-borne encephalitis virus in agricultural and forestry workers from Tuscany, Italy.

The ticks Ixodes persulcatus and Ixodes ricinus are the main vectors of both Borrelia burgdorferi sensu lato and tick-borne encephalitis (TBE) virus in Eurasia. Borrelia burgdorferi is the cause of Lyme borreliosis, and TBE is a biphasic meningoencephalitis induced by an arbovirus belonging to the flavivirus family. The principal aims of the current investigation were (i) to determine the frequency of serological evidence of Borrelia burgdorferi sensu lato and TBE infections in healthy agricultural and forestry workers, (ii) to determine the incidence of seroconversion for antibodies against Borrelia burgdorferi sensu lato and TBE virus in Tuscan workers during a 1-year survey; and (iii) to assess the occupational risk for agricultural and forestry activities in a defined area (Tuscany, Italy). A total of 412 blood samples were taken from agricultural and forestry workers, and information on age, duration of employment, and history of tick bites was collected in a questionnaire to establish the risk factors for the diseases. Three hundred sixty-five blood donors from the same region served as controls. To estimate the rate of seroconversion, 176 of the agricultural and forestry workers were tested 1 year later. IgG and IgM antibodies against Borrelia burgdorferi sensu lato and TBE virus were detected in serum by an enzyme-linked immunosorbent assay and confirmed by Western blot analysis for Borrelia burgdorferi and by a test for inhibition of hemagglutination for TBE. Antibodies against Borrelia burgdorferi were more frequent among the workers than in the control group (7.8% vs. 4.9% in the IgG-IgM enzyme-linked immunosorbent assay and 7.03% vs. 3.56% in the confirmatory test). No seropositivity was observed for TBE virus. Eighteen of 176 subjects who underwent a second blood test developed specific antibodies against Borrelia burgdorferi within 1 year.

Potential vectors of West Nile virus following an equine disease outbreak in Italy.

In the late summer of 1998, an outbreak of equine encephalomyelitis due to West Nile virus (WNV) occurred in the Tuscany region of central Italy. The disease was detected in 14 race horses from nine localities in four Provinces: Firenze, Lucca, Pisa and Pistoia. The outbreak area included Fucecchio wetlands (1800 ha), the largest inland marsh in Italy, and the adjacent hilly Cerbaie woodlands with farms breeding horses. To detect potential vectors of WNV, entomological surveys of Fucecchio and Cerbaie were undertaken during 1999-2002 by collecting mosquito larvae from breeding sites and adult mosquitoes by several methods of sampling. Among 6023 mosquitoes (Diptera: Culicidae) collected, 11 species were identified: Aedes albopictus (Skuse), Ae. vexans (Meigen), Anopheles atroparvus Van Thiel, An. maculipennis Meigen s.s., An. plumbeus Stephens, Culex impudicus Ficalbi, Cx. pipiens L., Culiseta longiareolata Macquart), Ochlerotatus caspius (Pallas), Oc. detritus (Haliday) and Oc. geniculatus (Olivier). In Fucecchio marshes, Cx. impudicus predominated with seasonal peak densities in spring and autumn: its greatest abundance during early spring coincides with arrival of migratory birds from Africa. In Cerbaie hills, Cx. pipiens predominated with peak population density in late summer. No viruses were isolated from 665 mosquitoes processed. These findings, plus other data on Italian mosquito bionomics, suggest a possible mode of WNV transmission involving the most abundant Culex in the Fucecchio-Cerbaie areas. Culex impudicus, being partly ornithophilic, might transmit WNV from migratory to non-migratory birds during springtime; Cx. pipiens, having a broader host range, would be more likely to transmit WNV from birds to horses and, perhaps, to humans by late summer.

Evidence of Toscana virus circulation in Umbria: first report.

To evaluate, for the first time, circulation and clinical expression of Toscana virus (ToSv) in Umbria region we studied: (1) 93 cases of aseptic meningitis and meningoencephalitis admitted to our Department from 1989 to 2001 with negative results for common neurotropic virus; (2) 50 healthy subjects. Specific antibodies (IgM and IgG) anti-TOSv were found in 36.6% of aseptic meningitis, in 6.06% of meningoencephalitis and (IgG) in 16% of healthy subjects.

Encephalitis without meningitis due to sandfly fever virus serotype toscana.

The role of Toscana (TOS) virus in producing encephalitis without meningitis is uncertain. We studied 2 cases of TOS virus encephalitis without meningitis by means of nested polymerase chain reaction assay and DNA sequencing. Findings confirm that TOS virus may directly cause encephalitis and suggest the usefulness of DNA sequencing in investigating relationships between TOS virus molecular patterns and the spectrum of neurological involvement.

Detection of Toscana virus-specific immunoglobulins G and M by an enzyme-linked immunosorbent assay based on recombinant viral nucleoprotein.

An enzyme-linked immunosorbent assay (ELISA) based on the recombinant Toscana virus nucleoprotein (rN) has been developed. Its sensitivity and specificity for the detection of virus-specific immunoglobulins G and M in human sera were similar to those of the ELISA that is based on an antigen extracted from infected mouse brain and that is routinely used for serodiagnosis.

Toscana virus infections of the central nervous system in children: a report of 14 cases.

OBJECTIVE: To evaluate the pathogenicity of a recently discovered arthropod-transmitted bunyavirus (Toscana virus) on the CNS in children and to provide information on the epidemiologic and clinical aspects of Toscana virus infection. STUDY DESIGN: Case-series analysis of children hospitalized with clinical and cerebrospinal fluid examination compatible with a CNS disease of viral origin. METHODS: Cerebrospinal fluid, acute, and convalescent sera were investigated for conventional neurotropic viruses and for Toscana and tickborne encephalitis viruses. A clinical-epidemiologic analysis was carried out on confirmed Toscana virus cases to clarify the profile of Toscana virus infection in children. RESULTS: The study indicates that (1) Toscana virus has been endemic in the Siena province for at least 15 years; (2) the virus is responsible for at least 80% of acute viral infections of the CNS in children throughout the summertime; (3) the clinical signs and symptoms range from aseptic meningitis to meningoencephalitis; (4) infected children resided habitually or temporarily in rural or suburban areas of the Siena province, where ecological characteristics allow arthropods to be peridomestic in human settlements. CONCLUSIONS: Toscana virus is the most common viral agent involved in acute infections of CNS in children in central Italy.

Phlebotomus-transmitted toscana virus infections of the central nervous system: a seven-year experience in Tuscany.

Toscana virus (TOSv) is a recently discovered Phlebotomus-transmitted human pathogen involved in acute infections of the central nervous system (CNS) occurring during the summer in natural foci in Italy. The purpose of this prospective study was to investigate the role of this virus in 170 patients with meningitis-meningoencephalitis of suspected viral origin, admitted to the Departments of Infectious Diseases at the Siena Hospital from 1990 to 1996. Infections caused by tick-borne encephalitis virus (TBEv) and TOSv or other neurotropic viruses were routinely diagnosed by means of conventional virological methods. 89 cases were attributed to TOSv, about 10% of which were Europeans on vacation in Tuscany. All of the TOSv-positive cases were observed during the summer and were residents of hilly areas in Siena and its province at an altitude not above 500 m. An increase in the number of cases was observed over the years, with a higher incidence among younger people. The clinical picture was similar to that observed in other viral infections of the CNS. Evolution was benign in all cases; in 2 subjects symptoms and signs of encephalitis were present.

Evidence of Toscana virus infections without central nervous system involvement: a serological study.

In central Italy, acute lymphocytic meningitis and meningoencephalitis due to a Phlebotomus transmitted virus (Phlebovirus Toscana, TOSv) occurring throughout the summer are frequently observed. Several serum specimens of patients hospitalized with a clinical picture of viral meningitis/meningoencephalitis showed anti TOS-IgG reactivity suggestive of a previous infection occurring at an unknown time in the past. This observation led us to design a serological investigation of 83 household contacts of 46 summertime CNS infection patients (index cases) with the purpose of evaluating the percentage of both IgG and IgM in seropositive healthy individuals, living, like the index cases, in areas at high risk of phlebotomine sandfly bites. The serological study was carried out using an ELISA method: 22% of the sera showed a reactivity for anti-TOS IgG antibodies and 6% resulted IgG/IgM positive; none of them reported having had any symptoms of CNS involvement; the distribution of seropositive cases was similar in contacts of both TOS-confirmed and TOS-negative cases. Our results indicate: (1) that phlebovirus Toscana is frequently implicated in cases with CNS disease as well as in infections occurring without neurological involvement; (2) in our country the ecological requirements encompass the conditions in and around the human settlements for phlebotomine sandflies to become peridomestic thus amplifying the risk of TOSv infections, which are in fact widespread and frequent in Siena and its surroundings.

Sandfly fever viruses in Italy.

Two serologically distinct agents, the sandfly fever Sicilian and the sandfly fever Naples viruses, were isolated by Sabin from blood samples taken during an Italian epidemic of febrile illness. Since then, several different viruses have been isolated from sandflies and/or humans in both the Old and New World. Toscana virus, a new virus closely antigenically related to sandfly fever Naples virus, was isolated in 1971 from the sandfly Phlebotomus perniciosus in Italy. Extensive studies on the importance of Toscana virus as a human pathogen demonstrated its association with acute neurologic diseases. A serosurvey for the presence of antibodies to sandfly fever Sicilian, sandfly fever Naples and Toscana viruses indicated that, as in other Mediterranean areas, both sandfly fever Sicilian and sandfly fever Naples viral infections decreased or disappeared after the 1940s in countries performing insecticide-spraying malaria eradication campaigns. In contrast, clinical cases of aseptic meningitis or meningoencephalitis caused by Toscana virus are observed annually in Central Italy during the summer. Toscana virus may be present in other Mediterranean countries where sandflies of the genus Phlebotomus are present.