Francesco Severini

COLD ACCLIMATION AND OVERWINTERING OF FEMALE AEDES ALBOPICTUS IN ROMA

ABSTRACT Eight years after the introduction and establishment of Aedes albopictus in Roma, females of the species extended their trophic activity to the coldest months of the year. Winter monitoring carried out from December 2003 to March 2004 in the urban area of the capital city of Italy recorded a weekly rate of positive ovitraps constantly around 30%, for the entire period of surveillance (14 wk). Eggs from ovitraps did not hatch when stimulated in laboratory with a wet and dry procedure. The hypothesis that long-lived female Ae. albopictus, belonging to the last seasonal generation, could have continued their trophic activity for 3 months, or that 1 or more cycles of reproduction have occurred in peculiar breeding sites, where a favorable microclimate may have allowed the development of at least 1 larval generation, are discussed and compared with the existing literature.

Experimental studies of susceptibility of Italian Aedes albopictus to Zika virus.

We report a study on vector competence of an Italian population of Aedes albopictus for Zika virus (ZIKV). Ae. albopictus was susceptible to ZIKV infection (infection rate: 10%), and the virus could disseminate and was secreted in the mosquitos saliva (dissemination rate: 29%; transmission rate: 29%) after an extrinsic incubation period of 11 days. The observed vector competence was lower than that of an Ae. aegypti colony tested in parallel.

Experimental investigation of the susceptibility of Italian Culex pipiens mosquitoes to Zika virus infection.

We investigated the susceptibility of an Italian population of Culex pipiens mosquitoes to Zika virus (ZIKV) infection, tested in parallel with Aedes aegypti, as a positive control. We analysed mosquitoes at 0, 3, 7, 10, 14, 20 and 24 days after an infectious blood meal. Viral RNA was detected in the body of Cx. pipiens up to three days post-infection, but not at later time points. Our results indicate that Cx. pipiens is not susceptible to ZIKV infection.

Detection of a chikungunya outbreak in Central Italy, August to September 2017

An autochthonous chikungunya outbreak is ongoing near Anzio, a coastal town in the province of Rome. The virus isolated from one patient and mosquitoes lacks the A226V mutation and belongs to an East Central South African strain. As of 20 September, 86 cases are laboratory-confirmed. The outbreak proximity to the capital, its late summer occurrence, and diagnostic delays, are favouring transmission. Vector control, enhanced surveillance and restricted blood donations are being implemented in affected areas.

Assessment of the risk of malaria re-introduction in the Maremma plain (Central Italy) using a multi-factorial approach

AbstractKeywords: Mosquito-borne diseases, Residual anophelism, Anopheles labranchiae, Vectorial capacity, Climatechange, Plasmodium falciparum, Experimental infectionBackgroundIn recent years, the increase in globalization [1], the risein the average temperature of the earth together with anincreasing frequency and intensity of extreme weatherevents, as storms, floods and droughts [2,3], and theenvironmental changes induced by human activities [4],have raised the concern about the possible introductionor reintroduction of Vector Borne Diseases in Countrieswhere these were absent or eradicated [5]. These con-siderations, coupled with the recent spread of somemosquito vector borne diseases in Europe [6,7] and theincreasing number of imported malaria cases recordedin the Continent [8] have renewed interest in the possi-ble reintroduction of malaria in Southern Europe [7-9],particularly in the countries facing the Western Medi-terranean Basin, where potential Anopheline vectors arestill present [10-13]. Moreover, in recent years auto-chthonous malaria cases have been sporadically reportedin Italy, France, Spain and Greece [14-20].In 2005, a five-year study was implemented in Italy, aswell as in other South European countries, with the aim toassess the status of the local potential malaria vectors andthe possible re-introduction of malaria transmission[21-25]. In Italy, the selected study area was the Maremmaplain, a region that was hyperendemic for malaria until60 years ago [26-28] and that more recently was recog-nized as the major “at risk” area for the malaria reintro-duction into Italy [14,29,30].In Maremma, after the malaria eradication campaign(1947-1951), Anopheles labranchiae, the main endophilicvector of the Anopheles maculipennis complex was dra-matically reduced in abundance. However, in subsequentyears, the species has progressively re-colonized most ofthe area coming back to substantial densities [31-33].This was mainly due to the introduction of intensive ricecultivation in the early 1970s. Since then, Maremma hasbeen subjected to continuous entomological surveillancethat was intensified after1997, when an autochthonousPlasmodium vivax malaria case, transmitted byAn. lab-ranchiae, occurred in the Province of Grosseto [14]. Thestudies carried out in the area since eradication, providesa database that allowed a follow-up the history of malariaand its vectors in Maremma over the past 60 years. Start-ing from the findings of the most recent entomologicaland environmental studies [23,34], the present study waschosen to evaluate the malariogenic potential of the areausing a multifactorial approach.Methods

Ecological Niche Modelling of Potential West Nile Virus Vector Mosquito Species and Their Geographical Association with Equine Epizootics in Italy

In Italy, West Nile virus (WNV) equine outbreaks have occurred annually since 2008. Characterizing WNV vector habitat requirements allows for the identification of areas at risk of viral amplification and transmission. Maxent-based ecological niche models were developed using literature records of 13 potential WNV Italian vector mosquito species to predict their habitat suitability range and to investigate possible geographical associations with WNV equine outbreak occurrence in Italy from 2008 to 2010. The contribution of different environmental variables to the niche models was also assessed. Suitable habitats for Culex pipiens, Aedes albopictus, and Anopheles maculipennis were widely distributed; Culex modestus, Ochlerotatus geniculatus, Ochlerotatus caspius, Coquillettidia richiardii, Aedes vexans, and Anopheles plumbeus were concentrated in north-central Italy; Aedes cinereus, Culex theileri, Ochlerotatus dorsalis, and Culiseta longiareolata were restricted to coastal/southern areas. Elevation, temperature, and precipitation variables showed the highest predictive power. Host population and landscape variables provided minor contributions. WNV equine outbreaks had a significantly higher probability to occur in habitats suitable for Cx. modestus and Cx. pipiens, providing circumstantial evidence that the potential distribution of these two species coincides geographically with the observed distribution of the disease in equines.

A 2-year entomological study of potential malaria vectors in Central Italy.

Europe was officially declared free from malaria in 1975; nevertheless, this disease remains a potential problem related to the presence of former vectors, belonging to the Anopheles maculipennis complex. Autochthonous-introduced malaria cases, recently reported in European countries, together with the predicted climatic and environmental changes, have increased the concern of health authorities over the possible resurgence of this disease in the Mediterranean Basin. In Italy, to study the distribution and bionomics of indigenous anopheline populations and to assess environmental parameters that could influence their dynamics, an entomological study was carried out in 2005-2006 in an at-risk study area. This model area is represented by the geographical region named the Maremma, a Tyrrhenian costal plain in Central Italy, where malaria was hyperendemic up to the 1950s. Fortnightly, entomological surveys (April-October) were carried out in four selected sites with different ecological features. Morphological and molecular characterization, blood meal identification, and parity rate assessment of the anophelines were performed. In total, 8274 mosquitoes were collected, 7691 of which were anophelines. Six Anopheles species were recorded, the most abundant of which were Anopheles labranchiae and An. maculipennis s.s. An. labranchiae is predominant in the coastal plain, where it is present in scattered foci. However, this species exhibits a wider than expected range: in fact it has been recorded, for the first time, inland where An. maculipennis s.s. is the most abundant species. Both species fed on a wide range of animal hosts, also showing a marked aggressiveness on humans, when available. Our findings demonstrated the high receptivity of the Maremma area, where the former malaria vector, An. labranchiae, occurs at different densities related to the kind of environment, climatic parameters, and anthropic activities.

Ecological Distribution and CQ11 Genetic Structure of Culex pipiens Complex (Diptera: Culicidae) in Italy.

Mosquitoes in the Culex pipiens complex are considered to be involved in the transmission of a range of pathogens, including West Nile virus (WNV). Although its taxonomic status is still debated, the complex includes species, both globally distributed or with a more limited distribution, morphologically similar and characterised by different physiological and behavioural traits, which affect their ability as vectors. In many European countries, Cx. pipiens and its sibling species Culex torrentium occur in sympatry, exhibiting similar bionomic and morphological characters, but only Cx. pipiens appears to play a vector role in WNV transmission. This species consists of two biotypes, pipiens and molestus, which can interbreed when in sympatry, and their hybrids can act as WNV-bridge vectors, due to intermediate ecological features. Considering the yearly WNV outbreaks since 2008 and given the morphological difficulties in recognising species and biotypes, our aim was to molecularly identify and characterised Cx. pipiens and Cx. torrentium in Italy, using recently developed molecular assays. Culex torrentium was not detected; as in other European countries, the pipiens and molestus biotypes were widely found in sympatry with hybrids in most environments. The UPGMA cluster analysis applied to CQ11 genotypic frequencies mainly revealed two groups of Cx. pipiens populations that differed in ecological features. The high propensity of the molestus biotype to exist in hypogean environments, where the habitat’s physical characteristics hinder and preclude the gene flow, was shown. These results confirmed the CQ11 assay as a reliable diagnostic method, consistent with the ecological and physiological aspects of the populations analysed. Since the assessment of the actual role of three biotypes in the WNV circulation remains a crucial point to be elucidated, this extensive molecular screening of Cx. pipiens populations can provide new insights into the ecology of the species and may give useful indications to plan and implement WNV surveillance activities in Italy.

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.

Impact of Environmental Changes and Human-Related Factors on the Potential Malaria Vector, Anopheles labranchiae (Diptera: Culicidae), in Maremma, Central Italy

ABSTRACT The Maremma Plain (central Italy) was hyper-endemic for malaria until the mid-20th century, when a national campaign for malaria elimination drastically reduced the presence of the main vector Anopheles labranchiae Falleroni. However, the introduction of rice cultivation over 30 yr ago has led to an increase in the An. labranchiae population and concern over possible malaria reemergence. We studied the impact of anthropogenic environmental changes on the abundance and distribution of An. labranchiae in Maremma, focusing on rice fields, the main breeding sites. Adults and larvae were collected in three main areas with diverse ecological characteristics. Data were collected on human activity, land use, and seasonal climatic and demographic variations. We also interviewed residents and tourists regarding their knowledge of malaria. Our findings showed that the most important environmental changes have occurred along the coast; An. labranchiae foci are present throughout the area, with massive reproduction strictly related to rice cultivation in coastal areas. Although the abundance of this species has drastically decreased over the past 30 yr, it remains high and, together with climatic conditions and the potential introduction of gametocyte carriers, it may represent a threat for the occurrence of autochthonous malaria cases. Our findings suggest the need for the continuous monitoring of An. labranchiae in the study area. In addition to entomological surveillance, more detailed knowledge of human-induced environmental changes is needed, so as to have a more complete database that can be used for vector-control plans and for properly managing emergencies related to autochthonous introduced cases.