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1.  Avian malaria parasites in the last supper: identifying encounters between parasites and the invasive Asian mosquito tiger and native mosquito species in Italy 
Malaria Journal  2015;14:32.
Background
The invasive Asian tiger mosquito Aedes albopictus has dramatically expanded its distribution range, being catalogued as one of the world’s 100 worst invasive alien species. As vectors of pathogens, Ae. albopictus may create novel epidemiological scenarios in the invaded areas.
Methods
Here, the frequency of encounters of Ae. albopictus with the avian malaria parasite Plasmodium and the related Haemoproteus was studied in an area with established populations in northeastern Italy and compared with those from four native mosquito species, Anopheles maculipennis s.l., Culex hortensis, Culex pipiens, and Ochlerotatus caspius. The abdomens of mosquitoes with a recent blood meal were used to identify both the blood meal source and the parasites harboured.
Results
Aedes albopictus had a clear antropophilic behaviour while An. maculipennis and Oc. caspius fed mainly on non-human mammals. Birds were the most common hosts of Cx. pipiens and reptiles of Cx. hortensis. Parasites were isolated from three mosquito species, with Cx. pipiens (30%) showing the highest parasite prevalence followed by Cx. hortensis (9%) and Ae. albopictus (5%).
Conclusions
These results are the first identifying the avian malaria parasites harboured by mosquitoes in Italy and represent the first evidence supporting that, although Ae. albopictus could be involved in the transmission of avian malaria parasites, the risk of avian malaria parasite spread by this invasive mosquito in Europe would be minimal.
doi:10.1186/s12936-015-0571-0
PMCID: PMC4318217  PMID: 25626918
Aedes albopictus; Avian diseases; Culex pipiens; Haemoproteus; Mosquitoes; Plasmodium; Vector; West Nile virus
2.  Evaluation of blood and bone marrow in selected canine vector-borne diseases 
Parasites & Vectors  2014;7(1):534.
Background
Bone marrow (BM) is a major hematopoietic organ that can harbour a variety of vector-borne pathogens; however, knowledge of BM pathological changes in dogs infected with vector-borne pathogens is limited. Thus, the aim of the present study was to assess the pathological changes in canine BM associated with natural infections by four vector-borne pathogens, as well as to determine the relationships between such changes and abnormalities of the peripheral blood.
Methods
Cytological disorders and pathological changes of the BM of 83 dogs naturally-infected with one or more of four vector-borne pathogens (i.e., Anaplasma platys, Leishmania infantum, Babesia vogeli and Hepatozoon canis) were evaluated and compared with the corresponding hematological findings.
Results
Dysgranulopoiesis and dysmegakaryocytopoiesis were the most frequently observed BM abnormalities in infected dogs. Erythroid suppression, and lymphocytic, monocytic and macrophage hyperplasia were also observed. Interestingly, associations between suppression and hyperplasia of specific cell lines in the marrow and corresponding changes in numbers of circulating peripheral blood cells were not observed.
Conclusions
Infections with one or more of the vector-borne pathogens examined in this study should be considered as differential diagnoses for secondary dysmyelopoiesis.
doi:10.1186/s13071-014-0534-2
PMCID: PMC4261574  PMID: 25441458
Bone marrow; Cytology; Vector-borne pathogens; Dysplasia; Secondary dysmyelopoiesis
3.  Efficacy of a slow-release imidacloprid (10%)/flumethrin (4.5%) collar for the prevention of canine leishmaniosis 
Parasites & Vectors  2014;7:327.
Background
The efficacy of a slow-release insecticidal and repellent collar containing 10% imidacloprid and 4.5% flumethrin (Seresto, Bayer Animal Health) in preventing Leishmania infantum infection was evaluated in a large population of dogs living in a hyper-endemic area of Sicily (Italy).
Methods
A total of 219 dogs, negative for L. infantum were enrolled in a multicentre, controlled study. Dogs were divided into two homogeneous groups, defined as G1 (n = 102) and G2 (n = 117). Before the start of the sand fly season, dogs in G1 were treated with the collar while animals in G2 were left untreated, serving as negative controls. Dogs were serially sampled on day D90, D180, D210 and D300 in order to assess Leishmania infection by IFAT, PCR on skin (D210-D300) and bone marrow (D300) and cytology on bone marrow aspirate (D300).
Results
Three dogs (2.9%) in G1 and 41 (40.2%) in G2 became positive for L. infantum in at least one of the diagnostic tests employed in the study. The number of seropositive dogs in G2 increased in the course of the study from 15 (D90) to 41 (D300), with some of them also positive in other diagnostic tests. Eight (19.6%) of the seropositive dogs in G2 showed an increase in antibody titers ranging from 1:160 to 1:1,280. At the last follow-up, some of dogs in G2 displayed overt clinical signs suggestive of leishmaniosis. The mean incidence density rate at the final follow-up was 4.0% for G1 and 60.7% for G2, leading to a mean efficacy of the collar in protecting dogs at both sites of 93.4%.
Conclusions
The slow-release collar tested in this study was shown to be safe and highly effective in preventing L. infantum infection in a large population of dogs. Protection conferred by a single collar (up to eight months) spanned an entire sand fly season in a hyper-endemic area of southern Italy. The regular use of collars, at least during the sand fly season, may represent a reliable and sustainable strategy for the prevention of leishmaniosis in dogs living in or travelling to an endemic area.
doi:10.1186/1756-3305-7-327
PMCID: PMC4105111  PMID: 25023573
Leishmania infantum; Dog; Canine leishmaniosis; Prevention; Control
4.  Ticks infesting humans in Italy and associated pathogens 
Parasites & Vectors  2014;7:328.
Background
Ticks may transmit a large variety of pathogens, which cause illnesses in animals and humans, commonly referred to as to tick-borne diseases (TBDs). The incidence of human TBDs in Italy is underestimated because of poor surveillance and the scant amount of studies available.
Methods
Samples (n = 561) were collected from humans in four main geographical areas of Italy (i.e., northwestern, northeastern, southern Italy, and Sicily), which represent a variety of environments. After being morphologically identified, ticks were molecularly tested with selected protocols for the presence of pathogens of the genera Rickettsia, Babesia, Theileria, Candidatus Neoehrlichia mikurensis, Borrelia and Anaplasma.
Results
Ticks belonged to 16 species of the genera Argas, Dermacentor, Haemaphysalis, Hyalomma, Ixodes and Rhipicephalus, with Ixodes ricinus (59.5%) being the species most frequently retrieved, followed by Rhipicephalus sanguineus sensu lato (21.4%). Nymphs were the life stage most frequently retrieved (41%), followed by adult females (34.6%). The overall positivity to any pathogen detected was 18%. Detected microorganisms were Rickettsia spp. (17.0%), Anaplasma phagocytophilum (0.8%), Borrelia afzelii (0.5%), Borrelia valaisiana (0.3%), C. N. mikurensis (0.5%) and Babesia venatorum (0.6%).
Conclusions
Results indicate that people living in the Italian peninsula are at risk of being bitten by different tick species, which may transmit a plethora of TBD causing pathogens and that co-infections may also occur.
doi:10.1186/1756-3305-7-328
PMCID: PMC4223688  PMID: 25023709
Ticks; Pathogens; Humans; Tick-borne diseases; Italy; Distribution
5.  Indirect versus direct detection methods of Trichinella spp. infection in wild boar (Sus scrofa) 
Parasites & Vectors  2014;7:171.
Background
Trichinella spp. infections in wild boar (Sus scrofa), one of the main sources of human trichinellosis, continue to represent a public health problem. The detection of Trichinella spp. larvae in muscles of wild boar by digestion can prevent the occurrence of clinical trichinellosis in humans. However, the analytical sensitivity of digestion in the detection process is dependent on the quantity of tested muscle. Consequently, large quantities of muscle have to be digested to warrant surveillance programs, or more sensitive tests need to be employed. The use of indirect detection methods, such as the ELISA to detect Trichinella spp. infections in wild boar has limitations due to its low specificity. The aim of the study was to implement serological detection of anti-Trichinella spp. antibodies in meat juices from hunted wild boar for the surveillance of Trichinella spp. infections.
Methods
Two tests were used, ELISA for the initial screening test, and a specific and sensitive Western blot (Wb) as a confirmatory test. The circulation of anti-Trichinella IgG was determined in hunted wild boar muscle juice samples in 9 provinces of 5 Italian regions.
Results
From 1,462 muscle fluid samples, 315 (21.5%, 95% C.I. 19.51-23.73) were tested positive by ELISA. The 315 ELISA-positive muscle fluid samples were further tested by Wb and 32 (10.1%, 95% C.I. 7.29-13.99) of these were positive with a final seroprevalence of 2.2% (95% C.I 1.55-3.07; 32/1,462). Trichinella britovi larvae were detected by artificial digestion in muscle tissues of one (0.07%, 95%C.I. 0.01-0.39) out of the 1,462 hunted wild boars. No Trichinella spp. larvae were detected in Wb-negative wild boar. From 2006 to 2012, a prevalence of 0.017% was detected by muscle digestion in wild boar hunted in the whole Italian territory.
Conclusions
The combined use of both serological methods had a sensitivity 31.4 times higher than that of the digestion (32/1,462 versus 1/1,462), suggesting their potential use for the surveillance of the Trichinella spp. infection in wild boar populations.
doi:10.1186/1756-3305-7-171
PMCID: PMC3995759  PMID: 24708795
Trichinella spp; ELISA; Western blot; Wild boar; Meat juice; Chemiluminescence; Prevalence; Surveillance; Italy
6.  Human and entomological surveillance of West Nile fever, dengue and chikungunya in Veneto Region, Italy, 2010-2012 
Background
Since 2010 Veneto region (North-Eastern Italy) planned a special integrated surveillance of summer fevers to promptly identify cases of West Nile Fever (WNF), dengue (DENV) and chikungunya (CHIKV). The objectives of this study were (i) To increase the detection rate of imported CHIKV and DENV cases in travellers from endemic areas and promptly identify potential autochthonous cases.(ii) To detect autochthonous cases of WNF, besides those of West Nile Neuroinvasive Disease (WNND) that were already included in a national surveillance.
Methods
Human surveillance: a traveler who had returned within the previous 15 days from endemic countries, with fever >38°C, absence of leucocytosis (leukocyte count <10,000 μL), and absence of other obvious causes of fever, after ruling out malaria, was considered a possible case of CHIKV or DENV. A possible autochthonous case of WNF was defined as a patient with fever >38°C for <7 days, no recent travel history and absence of other obvious causes of fever. Entomologic surveillance: for West Nile (WNV) it was carried out from May through November placing CDC-CO2 traps in five provinces of Veneto Region, while for DENV and CHIKV it was also performed around residences of viremic cases.
Results
Human surveillance: between 2010 and 2012, 234 patients with fever after travelling were screened, of which 27 (11,5%) were found infected (24 with DENV and 3 with CHIKV). No autochthonous case of DENV or CHIKV was detected. Autochthonous patients screened for WNF were 408, and 24 (5,9%) were confirmed cases. Entomologic surveillance: the WNV was found in 10, 2 and 11 pools of Culex pipiens from 2010 to 2012 respectively, in sites of Rovigo, Verona, Venezia and Treviso provinces). No infected Aedes albopictus with DENV or CHIKV was found.
Conclusions
Veneto is the only Italian region reporting WNV human cases every year since 2008. WNV is likely to cause sporadic cases and unforeseeable outbreaks for decades. Including WNF in surveillance provides additional information and possibly an early alert system. Timely detection of DENV and CHIKV should prompt vector control measures to prevent local outbreaks.
doi:10.1186/1471-2334-14-60
PMCID: PMC3922982  PMID: 24499011
Dengue; Chikungunya; West Nile; Surveillance
7.  Determinants of the population growth of the West Nile virus mosquito vector Culex pipiens in a repeatedly affected area in Italy 
Parasites & Vectors  2014;7:26.
Background
The recent spread of West Nile Virus in temperate countries has raised concern. Predicting the likelihood of transmission is crucial to ascertain the threat to Public and Veterinary Health. However, accurate models of West Nile Virus (WNV) expansion in Europe may be hampered by limited understanding of the population dynamics of their primary mosquito vectors and their response to environmental changes.
Methods
We used data collected in north-eastern Italy (2009–2011) to analyze the determinants of the population growth rate of the primary WNV vector Culex pipiens. A series of alternative growth models were fitted to longitudinal data on mosquito abundance to evaluate the strength of evidence for regulation by intrinsic density-dependent and/or extrinsic environmental factors. Model-averaging algorithms were then used to estimate the relative importance of intrinsic and extrinsic variables in describing the variations of per-capita growth rates.
Results
Results indicate a much greater contribution of density-dependence in regulating vector population growth rates than of any environmental factor on its own. Analysis of an average model of Cx. pipiens growth revealed that the most significant predictors of their population dynamics was the length of daylight, estimated population size and temperature conditions in the 15 day period prior to sampling. Other extrinsic variables (including measures of precipitation, number of rainy days, and humidity) had only a minor influence on Cx. pipiens growth rates.
Conclusions
These results indicate the need to incorporate density dependence in combination with key environmental factors for robust prediction of Cx. pipiens population expansion and WNV transmission risk. We hypothesize that detailed analysis of the determinants of mosquito vector growth rate as conducted here can help identify when and where an increase in vector population size and associated WNV transmission risk should be expected.
doi:10.1186/1756-3305-7-26
PMCID: PMC3896690  PMID: 24428887
Culex pipiens; Culicidae; Mosquito population dynamics; Density dependence; West nile virus; Disease vectors
8.  Distribution and habitat characterization of the recently introduced invasive mosquito Aedes koreicus [Hulecoeteomyia koreica], a new potential vector and pest in north-eastern Italy 
Parasites & Vectors  2013;6:292.
Background
The container breeding species belonging to the genus Aedes (Meigen) are frequently recorded out of their place of origin. Invasive Aedes species are proven or potential vectors of important Arboviruses and their establishment in new areas pose a threat for human and animal health. A new species of exotic mosquito was recorded in 2011 in north-eastern Italy: Aedes (Finlaya) koreicus [Hulecoeteomyia koreica]. The aim of this study was to characterize the biology, the environment and the current distribution of this mosquito in north-eastern Italy. Morphological details useful to discriminate this species from other invasive Aedes mosquitoes are also given (see Additional files).
Methods
All possible breeding sites for larval development were monitored. In addition, ovitraps and traps for adults were used to collect eggs and adults. The mosquitoes (larvae and adults) were identified morphologically and molecularly. Environmental data and climatic variables during the period of mosquito activity (from April to October) were considered.
Results
Aedes koreicus was found in 37 municipalities (39.4%) and was detected in 40.2% of places and in 37.3% of larval habitats monitored, in a range of altitude from 173 to 1250 m.a.s.l.. Garden centres were the most common locations (66.7%), followed by streets/squares (57.1%), private gardens (46.4%) and cemeteries (21.1%) (p < 0.01). The main larval habitats were catch basins (48.5%) and artificial water containers (41.8%). As for Aedes albopictus [Stegomyia albopicta], ovitraps were attractive for adult females resulting in the higher rate of positivity (15/21; 71.4%) among breeding sites. The period of Ae. koreicus activity ranged from March 29 to October 29.
Conclusion
The species is clearly established in the area and is now overlapping with other vectors such as Ae. albopictus and colonizing areas over 800 m.a.s.l, not yet or sporadically reached by the tiger mosquito. The data collected are essential to assess the risk of colonization of other parts of Italy and Europe, as well as the risk of spreading of pathogens transmitted. These findings stress the importance of implementing entomological surveillance for early detection of invasive species, which is necessary for eradication or limitation of its further spread.
doi:10.1186/1756-3305-6-292
PMCID: PMC3852218  PMID: 24457085
Aedes koreicus; Invasive mosquito species; Entomological surveillance; North-eastern Italy
9.  European Surveillance for West Nile Virus in Mosquito Populations 
A wide range of arthropod-borne viruses threaten both human and animal health either through their presence in Europe or through risk of introduction. Prominent among these is West Nile virus (WNV), primarily an avian virus, which has caused multiple outbreaks associated with human and equine mortality. Endemic outbreaks of West Nile fever have been reported in Italy, Greece, France, Romania, Hungary, Russia and Spain, with further spread expected. Most outbreaks in Western Europe have been due to infection with WNV Lineage 1. In Eastern Europe WNV Lineage 2 has been responsible for human and bird mortality, particularly in Greece, which has experienced extensive outbreaks over three consecutive years. Italy has experienced co-circulation with both virus lineages. The ability to manage this threat in a cost-effective way is dependent on early detection. Targeted surveillance for pathogens within mosquito populations offers the ability to detect viruses prior to their emergence in livestock, equine species or human populations. In addition, it can establish a baseline of mosquito-borne virus activity and allow monitoring of change to this over time. Early detection offers the opportunity to raise disease awareness, initiate vector control and preventative vaccination, now available for horses, and encourage personal protection against mosquito bites. This would have major benefits through financial savings and reduction in equid morbidity/mortality. However, effective surveillance that predicts virus outbreaks is challenged by a range of factors including limited resources, variation in mosquito capture rates (too few or too many), difficulties in mosquito identification, often reliant on specialist entomologists, and the sensitive, rapid detection of viruses in mosquito pools. Surveillance for WNV and other arboviruses within mosquito populations varies between European countries in the extent and focus of the surveillance. This study reviews the current status of WNV in mosquito populations across Europe and how this is informing our understanding of virus epidemiology. Key findings such as detection of virus, presence of vector species and invasive mosquito species are summarized, and some of the difficulties encountered when applying a cost-effective surveillance programme are highlighted.
doi:10.3390/ijerph10104869
PMCID: PMC3823308  PMID: 24157510
West Nile virus; mosquito; surveillance; vector; invasive species
10.  Efficacy of an imidacloprid/flumethrin collar against fleas, ticks and tick-borne pathogens in dogs 
Parasites & Vectors  2013;6:245.
Background
Tick-borne diseases comprise a group of maladies that are of substantial medical and veterinary significance. A range of tick-borne pathogens, including diverse species of bacteria and protozoa, can infect both dogs and humans. Hence, the control of tick infestations is pivotal to decrease or prevent tick-borne pathogen transmission. Therefore, different commercial products with insecticidal, repellent or both properties have been developed for use on dogs. Recently, a collar containing a combination of imidacloprid 10% and flumethrin 4.5% has proven effective to prevent tick and flea infestations in dogs under field conditions and the infection by some vector-borne pathogens they transmit under laboratory-controlled conditions.
Methods
From March 2011 to April 2012, a field study was conducted in a private shelter in southern Italy to assess the efficacy of the imidacloprid/flumethrin collar against tick and flea infestations and to determine if this strategy would decrease tick-borne pathogen transmission in young dogs. A total of 122 animals were enrolled in the study and randomly assigned to group A (n = 64; collared) or group B (n = 58; untreated controls). Dogs were examined monthly for ticks and fleas and systematically tested for selected tick-borne pathogens.
Results
Compared to controls, the collar provided overall efficacies of 99.7% and 100% against tick and flea infestation, respectively. The overall efficacy for the prevention of tick-borne pathogens (i.e., Anaplasma platys and Babesia vogeli) was 91.6%.
Conclusions
This study demonstrates that the imidacloprid/flumethrin collar is efficacious against flea and tick infestation as well as tick-borne pathogen transmission to dogs under field conditions.
doi:10.1186/1756-3305-6-245
PMCID: PMC3766024  PMID: 23972013
Canine vector-borne diseases; Anaplasma platys; Babesia vogeli; Hepatozoon canis; Prevention; Dog; Imidacloprid; Flumethrin; Tick
11.  Risk of canine and human exposure to Dirofilaria immitis infected mosquitoes in endemic areas of Italy 
Parasites & Vectors  2013;6:60.
Background
The occurrence of infections by Dirofilaria immitis in canine and human populations depends on several factors linked to both the definitive and intermediate hosts. Little data are available on the risk of human and dog exposure to D. immitis in endemic areas. Data collected on dog- and human-bait traps in endemic areas of north-eastern Italy were used to estimate the likelihood of a receptive host coming into contact with an infected vector.
Methods
From 1997 to 1999, mosquitoes were collected from three sampling sites of north-eastern Italy on D. immitis microfilaraemic dogs and on human baits. The bite/night/host rates were determined based on the number of feeding and probing mosquitoes on dogs and humans, respectively. The survival/mortality rates of different species of mosquitoes following the blood meal, and the rate of natural Dirofilaria infection in unfed specimens were estimated. The risk of exposure of dogs and humans to infected mosquito species was determined by combining the bite/host/night and the mosquito infection rates.
Results
A total of 1,165 mosquitoes were collected on human (n = 815) and dog (n = 350) baits with varying species composition (i.e., Culex pipiens, 87.3% and Ochlerotatus caspius, 11.6%). Overall, dogs were more attractive to Cx pipiens than humans (feeding rate 70.2% vs probing rate 25.9%). The highest bite/night/host rate was 84.0 for dogs and 26.5 for humans. Cx pipiens displayed a mortality rate of 76.3% within 13 days and Oc. caspius of 100% within two days following the infective blood meal. In addition, D. immitis DNA was detected in unfed Cx pipiens (infection rate of 0.26%-2.07%). The infection rate adjusted for mosquito mortality was 0.38%. Based on data collected, the contact between an infected mosquito and a host can occur as often as every four nights for D. immitis infected-mosquitoes in dogs and within two weeks for humans.
Conclusions
Cx pipiens was confirmed as the most efficient natural vector of D. immitis in the studied area. In endemic areas, the risk of transmission can be very high for dogs and relevant for humans. Despite the increased awareness of veterinarians and owners on canine dirofilarioses, dogs from rural areas still maintain the natural life cycle of Dirofilaria spp., therefore acting as a source of infection to humans through vector bites.
doi:10.1186/1756-3305-6-60
PMCID: PMC3599681  PMID: 23510597
Dirofilaria immitis; Mosquito vectors; Dogs; Humans; Risk of exposure
12.  Prevention of Canine Leishmaniosis in a Hyper-Endemic Area Using a Combination of 10% Imidacloprid/4.5% Flumethrin 
PLoS ONE  2013;8(2):e56374.
Background
Dogs are the main reservoir hosts of Leishmania infantum, the agent of human zoonotic visceral leishmaniosis. This study investigated the efficacy of a polymer matrix collar containing a combination of 10% imidacloprid and 4.5% flumethrin as a novel prophylactic measure to prevent L. infantum infections in young dogs from a hyper-endemic area of southern Italy, with a view towards enhancing current control strategies against both human and canine leishmaniosis.
Methodology/Principal Findings
The study was carried out on 124 young dogs, of which 63 were collared (Group A) while 61 were left untreated (Group B), from March-April 2011 until March 2012. Blood and skin samples were collected at baseline (April 2011) and at the first, second, third and fourth follow-up time points (July, September 2011 and November 2011, and March 2012, respectively). Bone marrow and conjunctiva were sampled at baseline and at the fourth follow-up. Serological, cytological and molecular tests were performed to detect the presence of L. infantum in the different tissues collected. At the end of the trial, no dog from Group A proved positive for L. infantum at any follow-up, whereas 22 dogs from Group B were infected (incidence density rate = 45.1%); therefore, the combination of 10% imidacloprid and 4.5% flumethrin was 100% efficacious for the prevention of L. infantum infection in young dogs prior to their first exposure to the parasite in a hyper-endemic area for CanL.
Conclusions
The use of collars containing 10% imidacloprid and 4.5% flumethrin conferred long-term protection against infection by L. infantum to dogs located in a hyper-endemic area, thus representing a reliable and sustainable strategy to decrease the frequency and spread of this disease among the canine population which will ultimately result in the reduction of associated risks to human health.
doi:10.1371/journal.pone.0056374
PMCID: PMC3581506  PMID: 23451043
13.  Vector-borne helminths of dogs and humans in Europe 
Parasites & Vectors  2013;6:16.
Presently, 45% of the total human population of Europe, as well as their domestic and companion animals, are exposed to the risk of vector-borne helminths (VBH) causing diseases. A plethora of intrinsic biological and extrinsic factors affect the relationship among helminths, vectors and animal hosts, in a constantly changing environment. Although canine dirofilarioses by Dirofilaria immitis and Dirofilaria repens are key examples of the success of VBH spreading into non-endemic areas, another example is represented by Thelazia callipaeda eyeworm, an emergent pathogen of dogs, cats and humans in several regions of Europe. The recent finding of Onchocerca lupi causing canine and human infestation in Europe and overseas renders the picture of VBH even more complicated. Similarly, tick-transmitted filarioids of the genus Cercopithifilaria infesting the skin of dogs were recently shown to be widespread in Europe. Although for most of the VBH above there is an increasing accumulation of research data on their distribution at national level, the overall impact of the diseases they cause in dogs and humans is not fully recognised in many aspects. This review investigates the reasons underlying the increasing trend in distribution of VBH in Europe and discusses the diagnostic and control strategies currently available. In addition, this article provides the authors’ opinion on some topics related to VBH that would deserve further scientific investigation.
doi:10.1186/1756-3305-6-16
PMCID: PMC3564894  PMID: 23324440
Zoonosis; Dirofilaria immitis; Dirofilaria repens; Onchocerca lupi; Cercopithifilaria; Thelazia callipaeda; Europe; Risk; Mosquito; Tick; Vector; Treatment; Control
14.  Intestinal parasite infections in immigrant children in the city of Rome, related risk factors and possible impact on nutritional status 
Parasites & Vectors  2012;5:265.
Background
Parasitic diseases can represent a social and economic problem among disadvantaged people - even in developed countries. Due to the limited data available concerning Europe, the aims of the present study were to evaluate the presence of parasites in immigrant children and the risk factors favouring the spread of parasites. Subsequently, the possible correlation between nutritional status and parasitic infections was also investigated.
Findings
A convenience sample of two hundred and forty seven immigrant children (aged 0–15) attending the Poliambulatorio della Medicina Solidale in Rome was examined. Data were collected using structured questionnaires, and parasitological and anthropometric tests were applied. Chi-squared test and binary logistic multiple-regression models were used for statistical analysis.
Thirty-seven children (15%) tested positive to parasites of the following species: Blastocystis hominis, Entamoeba coli, Giardia duodenalis, Enterobius vermicularis, Ascaris lumbricoides and Strongyloides stercoralis. A monospecific infection was detected in 30 (81%) out of 37 parasitized children, while the others (19%) presented a polyparasitism. The major risk factors were housing, i.e. living in shacks, and cohabitation with other families (p<0.01). Children classified in the lower height Z-scores had a significantly greater prevalence of parasites (30.9%) than the others (p<0.01).
Conclusions
This study shows that parasite infection in children is still quite common, even in a developed country and that children’s growth and parasitism may be related. Extensive improvements in the living, social and economic conditions of immigrants are urgently needed in order to overcome these problems.
doi:10.1186/1756-3305-5-265
PMCID: PMC3519717  PMID: 23168023
Intestinal parasites; Risk factors; Nutritional status; Immigrant children; Developed country
15.  Molecular xenomonitoring of Dirofilaria immitis and Dirofilaria repens in mosquitoes from north-eastern Italy by real-time PCR coupled with melting curve analysis 
Parasites & Vectors  2012;5:76.
Background
Dirofilaria immitis and Dirofilaria repens are transmitted by bloodsucking culicid mosquitoes belonging to Culex, Aedes, Ochlerotatus, Anopheles and Mansonia genera.
The detection of filarioids in mosquitoes for assessing distribution of vectors and/or of pathogens in a given area (also known as “xenomonitoring”), when based on individual dissection of wild-caught female mosquitoes is time consuming and hardly applicable in large epidemiological surveys.
Our study aimed to evaluate the recently developed duplex real-time PCR for screening large number of culicids and to assess their positivity for D. immitis and D. repens in an area where both species are endemic.
Methods
A duplex real-time PCR was used to detect and differentiate D. immitis and D. repens in mosquitoes collected in six provinces of the Veneto region using 43 carbon dioxide-baited traps under the frame of an entomological surveillance program to monitor the vectors of West Nile disease. From early May till October 2010, unfed female mosquitoes (n = 40,892) were captured in 20 selected sites.
Results
Mosquitoes identified as Culex pipiens, Ochlerotatus caspius, Aedes vexans and Culex modestus were grouped into 995 pools according to species, day and site of collection (from minimum of 1 to maximum of 57). Out of 955 pools, 23 (2.41 %) scored positive for Dirofilaria spp. of which, 21 (2.2 %) for D. immitis and two (0.21 %) for D. repens. An overall Estimated Rate of Infection (ERI) of 0.06 % was recorded, being higher in Och. caspius and Ae. vexans (i.e., 0.18 % and 0.14 %, respectively). At least one mosquito pool was positive for Dirofilaria spp. in each province with the highest ERI recorded in Vicenza and Padova provinces (i.e., 0.42% and 0.16 %, respectively). Mosquitoes collected in all provinces were positive for D. immitis whereas, only two (i.e., Padova and Rovigo) provinces scored positive for D. repens. All mosquito species, except for Cx. modestus, were positive for D. immitis, whereas D. repens was only found in Cx. pipiens.
Conclusions
The results suggest that both Dirofilaria species are endemic and may occur in sympatry in the examined area. The molecular approach herein used represents a powerful tool for surveillance programs of D. immitis and D. repens in the culicid vectors towards a better understanding of the epidemiology of the infections they cause and their seasonal transmission patterns.
doi:10.1186/1756-3305-5-76
PMCID: PMC3438040  PMID: 22520170
Dirofilaria immitis; Dirofilaria repens; Real-time PCR; Mosquitoes; Vector; Surveillance
16.  Surveillance for West Nile, Dengue, and Chikungunya Virus Infections, Veneto Region, Italy, 2010 
Emerging Infectious Diseases  2012;18(4):671-673.
In 2010, in Veneto Region, Italy, surveillance of summer fevers was conducted to promptly identify autochthonous cases of West Nile fever and increase detection of imported dengue and chikungunya in travelers. Surveillance highlighted the need to modify case definitions, train physicians, and when a case is identified, implement vector control measures
doi:10.3201/eid1804.110753
PMCID: PMC3309689  PMID: 22469230
West Nile virus; dengue virus; chikungunya virus; summer fever; surveillance; Italy; viruses; vector-borne infections; arboviruses
17.  Occurrence and identification of risk areas of Ixodes ricinus-borne pathogens: a cost-effectiveness analysis in north-eastern Italy 
Parasites & Vectors  2012;5:61.
Background
Ixodes ricinus, a competent vector of several pathogens, is the tick species most frequently reported to bite humans in Europe. The majority of human cases of Lyme borreliosis (LB) and tick-borne encephalitis (TBE) occur in the north-eastern region of Italy. The aims of this study were to detect the occurrence of endemic and emergent pathogens in north-eastern Italy using adult tick screening, and to identify areas at risk of pathogen transmission. Based on our results, different strategies for tick collection and pathogen screening and their relative costs were evaluated and discussed.
Methods
From 2006 to 2008 adult ticks were collected in 31 sites and molecularly screened for the detection of pathogens previously reported in the same area (i.e., LB agents, TBE virus, Anaplasma phagocytophilum, Rickettsia spp., Babesia spp., "Candidatus Neoehrlichia mikurensis"). Based on the results of this survey, three sampling strategies were evaluated a-posteriori, and the impact of each strategy on the final results and the overall cost reductions were analyzed. The strategies were as follows: tick collection throughout the year and testing of female ticks only (strategy A); collection from April to June and testing of all adult ticks (strategy B); collection from April to June and testing of female ticks only (strategy C).
Results
Eleven pathogens were detected in 77 out of 193 ticks collected in 14 sites. The most common microorganisms detected were Borrelia burgdorferi sensu lato (17.6%), Rickettsia helvetica (13.1%), and "Ca. N. mikurensis" (10.5%). Within the B. burgdorferi complex, four genotypes (i.e., B. valaisiana, B. garinii, B. afzelii, and B. burgdorferi sensu stricto) were found. Less prevalent pathogens included R. monacensis (3.7%), TBE virus (2.1%), A. phagocytophilum (1.5%), Bartonella spp. (1%), and Babesia EU1 (0.5%). Co-infections by more than one pathogen were diagnosed in 22% of infected ticks. The prevalences of infection assessed using the three alternative strategies were in accordance with the initial results, with 13, 11, and 10 out of 14 sites showing occurrence of at least one pathogen, respectively. The strategies A, B, and C proposed herein would allow to reduce the original costs of sampling and laboratory analyses by one third, half, and two thirds, respectively. Strategy B was demonstrated to represent the most cost-effective choice, offering a substantial reduction of costs, as well as reliable results.
Conclusions
Monitoring of tick-borne diseases is expensive, particularly in areas where several zoonotic pathogens co-occur. Cost-effectiveness studies can support the choice of the best monitoring strategy, which should take into account the ecology of the area under investigation, as well as the available budget.
doi:10.1186/1756-3305-5-61
PMCID: PMC3337281  PMID: 22452970
Ixodes ricinus; tick-borne diseases; surveillance; economic evaluation; Italy.
18.  Vector-Borne Diseases - constant challenge for practicing veterinarians: recommendations from the CVBD World Forum 
Parasites & Vectors  2012;5:55.
The human-animal bond has been a fundamental feature of mankind's history for millennia. The first, and strongest of these, man's relationship with the dog, is believed to pre-date even agriculture, going back as far as 30,000 years. It remains at least as powerful today. Fed by the changing nature of the interactions between people and their dogs worldwide and the increasing tendency towards close domesticity, the health of dogs has never played a more important role in family life. Thanks to developments in scientific understanding and diagnostic techniques, as well as changing priorities of pet owners, veterinarians are now able, and indeed expected, to play a fundamental role in the prevention and treatment of canine disease, including canine vector-borne diseases (CVBDs).
The CVBDs represent a varied and complex group of diseases, including anaplasmosis, babesiosis, bartonellosis, borreliosis, dirofilariosis, ehrlichiosis, leishmaniosis, rickettsiosis and thelaziosis, with new syndromes being uncovered every year. Many of these diseases can cause serious, even life-threatening clinical conditions in dogs, with a number having zoonotic potential, affecting the human population.
Today, CVBDs pose a growing global threat as they continue their spread far from their traditional geographical and temporal restraints as a result of changes in both climatic conditions and pet dog travel patterns, exposing new populations to previously unknown infectious agents and posing unprecedented challenges to veterinarians.
In response to this growing threat, the CVBD World Forum, a multidisciplinary group of experts in CVBDs from around the world which meets on an annual basis, gathered in Nice (France) in 2011 to share the latest research on CVBDs and discuss the best approaches to managing these diseases around the world.
As a result of these discussions, we, the members of the CVBD Forum have developed the following recommendations to veterinarians for the management of CVBDs.
doi:10.1186/1756-3305-5-55
PMCID: PMC3359183  PMID: 22433172
19.  First report in italy of the exotic mosquito species Aedes (Finlaya) koreicus, a potential vector of arboviruses and filariae 
Parasites & Vectors  2011;4:188.
Background
In the Veneto region (north-eastern Italy) an entomological surveillance system has been implemented since the introduction of the Asian tiger mosquito (Aedes albopictus) in 1991. During the routine monitoring activity in a tiger mosquito-free area, an unexpected mosquito was noticed, which clearly did not belong to the recorded Italian fauna.
Findings
At the end of May 2011, twelve larvae and pupae were collected in a small village in Belluno province (Veneto region) from a single manhole. Ten adults reared in the laboratory were morphologically and genetically identified as Aedes (Finlaya) koreicus (Edwards, 1917), a species native to Southeast Asia. The subsequent investigations carried out in the following months in the same village provided evidence that this species had become established locally. Entomological and epidemiological investigations are currently ongoing in the surrounding area, to verify the eventual extension of the species outside the village and to trace back the route of entry into Italy.
Conclusions
This is the first report in Italy of the introduction of the exotic mosquito Ae. koreicus. This species has been shown experimentally to be competent in the transmission of the Japanese encephalitis virus and of the dog heartworm Dirofilaria immitis and is considered a potential vector of other arboviruses. Thus, the establishment of this species may increase the current risk or pose new potential threats, for human and animal health. This finding considerably complicates the entomological monitoring of the Asian tiger mosquito Ae. albopictus in Italy and stresses the importance of implementing the entomological surveillance for the early detection of and the rapid response against invasive mosquito species.
doi:10.1186/1756-3305-4-188
PMCID: PMC3203849  PMID: 21951867
Aedes koreicus; Italy; exotic mosquito; invasive species; entomological monitoring
20.  Diagnosis of Hepatozoon canis in young dogs by cytology and PCR 
Parasites & Vectors  2011;4:55.
Background
Hepatozoon canis is a widespread tick-borne protozoan affecting dogs. The diagnosis of H. canis infection is usually performed by cytology of blood or buffy coat smears, but this method may not be sensitive. Our study aimed to evaluate the best method to achieve a parasitological diagnosis of H. canis infection in a population of receptive young dogs, previously negative by cytology and exposed to tick infestation for one summer season.
Results
A total of 73 mongrel dogs and ten beagles younger than 18 months of age, living in an animal shelter in southern Italy where dogs are highly infested by Rhipicephalus sanguineus, were included in this study. In March-April 2009 and in October 2009, blood and bone marrow were sampled from each dog. Blood, buffy coat and bone marrow were examined by cytology only (at the first sampling) and also by PCR for H. canis (second sampling). In March-April 2009, only one dog was positive for H. canis by cytological examination, whereas in October 2009 (after the summer season), the overall incidence of H. canis infection by cytological examinations was 43.9%. Molecular tests carried out on samples taken in October 2009 showed a considerably higher number of dogs positive by PCR (from 27.7% up to 51.2% on skin and buffy coat tissues, respectively), with an overall positivity of 57.8%. All animals, but one, which were positive by cytology were also PCR-positive. PCR on blood or buffy coat detected the highest number of H. canis-positive dogs displaying a sensitivity of 85.7% for both tissues that increased up to 98% when used in parallel. Twenty-six (74.8%) out of the 28 H. canis-positive dogs presented hematological abnormalities, eosinophilia being the commonest alteration observed.
Conclusions
The results suggest that PCR on buffy coat and blood is the best diagnostic assay for detecting H. canis infection in dogs, although when PCR is not available, cytology on buffy coat should be preferred to blood smear evaluation. This study has also demonstrated that H. canis infection can spread among young dogs infested by R. sanguineus and be present in the majority of the exposed population within 6 months.
doi:10.1186/1756-3305-4-55
PMCID: PMC3083373  PMID: 21489247
21.  Diagnosis of Canine Vector-Borne Diseases in Young Dogs: a Longitudinal Study▿ †  
Journal of Clinical Microbiology  2010;48(9):3316-3324.
Canine vector-borne diseases (CVBDs) pose a diagnostic challenge, particularly when a dog is coinfected with more than one pathogen. The purpose of this study was to generate information about the diagnosis of CVBDs in young dogs following their first exposure to flea, tick, sand fly, louse, and mosquito vectors. From March 2008 to May 2009, 10 purpose-bred young naive beagle dogs and a cohort of 48 mixed-breed dogs living in an area to which CVBD is endemic in southern Italy were monitored using different diagnostic tests (cytology, serology, and PCR). Overall, PCR detected the highest number of dogs infected with Anaplasma platys, Babesia vogeli, and Ehrlichia canis, whereas seroconversion was a more sensitive indicator of exposure to Leishmania infantum. For A. platys infection, combining blood and buffy coat cytology in parallel enhanced the relative sensitivity (SErel) (87.3%). For B. vogeli, the best diagnostic combination was buffy coat cytology and serology used in parallel (SErel, 67.5%), whereas serology and PCR used in parallel (SErel, 100%) was the best combination for L. infantum. Overall, 12 (20.7%) dogs were coinfected; however, the percentage of new coinfections decreased from baseline (50%) to the first (33.3%) and second (16.6%) follow-up time points. Numbers of coinfections with A. platys and B. vogeli were significantly higher (P < 0.05) than coinfections with other pathogen combinations. The data generated in this study provide insights on the incidence of certain pathogens infecting young dogs in southern Italy, highlight important diagnostic testing limitations, and support the use of multiple diagnostic modalities when attempting to confirm a tick-borne infection in an individual dog or in a canine population.
doi:10.1128/JCM.00379-10
PMCID: PMC2937705  PMID: 20660218
22.  Occurrence of Rickettsia felis in dog and cat fleas (Ctenocephalides felis) from Italy 
Parasites & Vectors  2009;2(Suppl 1):S8.
Rickettsia felis is an obligate intracellular bacterium belonging to the spotted fever group, suspected to cause a murine typhus-like illness in humans, with a cosmopolitan distribution. This study was designed to estimate presence and occurrence of this pathogen in fleas collected from dogs and cats in different areas of Italy. Two species of fleas were identified, Ctenocephalides felis (80.3%) and Ctenocephalides canis (19.7%).
Overall, 320 fleas (257 C. felis and 63 C. canis) collected from 117 animals (73 dogs and 44 cats) were tested. Thirty-eight (11.9%) C. felis fleas, 13 from cats (17.6%) and 25 from dogs (10.2%) were positive for R. felis. No C. canis was positive. Fleas from cats showed a tendency to be more positive than fleas from dogs. Prevalence of R. felis among areas and within provinces of the same area was extremely variable, ranging from 0 to 35.3%. Overall, prevalence in north-eastern Italy (23.2%) was significantly higher than in south-western Italy (7.1%). This study confirmed the occurrence of R. felis in cat and dog fleas (C. felis) from Italy, similar to other European countries. The results also suggest that R. felis should be considered in the human differential diagnosis of any spotted-like fever in Italy, especially if the patient is known to have been exposed to flea bites.
doi:10.1186/1756-3305-2-S1-S8
PMCID: PMC2679400  PMID: 19426447
23.  Changing distribution patterns of canine vector borne diseases in Italy: leishmaniosis vs. dirofilariosis 
Parasites & Vectors  2009;2(Suppl 1):S2.
Ecological and climatic changes, human and animal population dynamics are among the several factors that have favoured the spread or the (re)introduction and establishment of "novel" vector species and pathogens they transmit in previously disease-free geographical areas. As key examples of the changing pattern of distribution of canine vector borne diseases (CVBDs), the current distribution of canine leishmaniosis (CanL) by Leishmania infantum and dirofilariosis by Dirofilaria immitis causing heart worm disease (HW) in Italy is discussed on the basis of retrospective historical reports until the 90's and later on until 2009. For long time, D. immitis has been considered mainly present along the Po River Valley and northward areas, while L. infantum in south-central Italy and Sicily and Sardinia. Comparison of current available and historical data (up to 1989) confirms that HW and CanL, although with different prevalence rates, have been changing their distribution patterns in Italy as a result of many biological and ecological factors, including those related to vector distribution and introduction of new species (e.g. the Asian tiger mosquito Aedes albopictus, a competent vector of D. immitis). New autochthonous foci of HW in southern Italy (i.e. Apulia and Calabria regions) have recently been reported.
Although analysing retrospective data may represent a difficult task, the "paradigm" about the dual distribution of HW and CanL in northern and southern Italy cannot yet be considered valid. The research needs for managing HW and CanL in previously uninfected areas are discussed.
doi:10.1186/1756-3305-2-S1-S2
PMCID: PMC2679394  PMID: 19426441
24.  Evaluation of Enzyme-Linked Immunosorbent Assays, an Immunofluorescent-Antibody Test, and Two Rapid Tests (Immunochromatographic-Dipstick and Gel Tests) for Serological Diagnosis of Symptomatic and Asymptomatic Leishmania Infections in Dogs 
Journal of Clinical Microbiology  2005;43(11):5515-5519.
Enzyme-linked immunosorbent assays (ELISAs) based on soluble antigens derived from promastigote or amastigote-like stages of Leishmania infantum and on the recombinant rK39 antigen, each in combination with different conjugates [anti-immunoglobulin G1 [IgG1], anti-IgG2, anti-IgG(γ), and anti-IgG heavy plus light chains], were compared to an immunofluorescent-antibody test (IFAT) and two commercially available rapid test systems (DiaMed-Vet-IT Leish and ID-PaGIA canine leishmaniasis antibody test) for the detection of specific anti-Leishmania antibodies in symptomatic and asymptomatic dogs with proven L. infantum infections. ELISAs based on soluble promastigote and amastigote antigens had very high sensitivities in symptomatic (n = 30; 100%) and asymptomatic dogs (n = 17; 94.1 to 100%), except when combined with the anti-IgG1 conjugate (41.2 to 82.4%). Specificities were high for all combinations (n = 50; 96 to 100%). The rK39 ELISA detected fewer asymptomatic cases (sensitivities, 52.9 to 64.7%) but was highly specific (96 to 100%). The IFAT was 90% sensitive in symptomatic dogs but was significantly less sensitive in asymptomatic cases (29.4%). However, it had an excellent specificity (100%). Test performances of the rapid tests based on the rK39 antigen were comparable to the ELISAs based on the same antigen. ELISAs based on soluble promastigote or amastigote antigens seem to be most suited for the serological diagnosis of canine Leishmania infections in both symptomatic and asymptomatic dogs. IFAT and the rK39 ELISA lack sensitivity in asymptomatic cases but are highly specific. Rapid tests like the rK39 dipstick test or the ID-PaGIA are helpful for confirming clinically suspected cases because of their high specificities in symptomatic animals.
doi:10.1128/JCM.43.11.5515-5519.2005
PMCID: PMC1287801  PMID: 16272479

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