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Phleboviruses transmitted by phlebotomine sandflies are endemic in the Mediterranean basin. Toscana phlebovirus (TOSV), Sicilian phlebovirus (SFSV), and Naples phlebovirus (SFNV) are responsible of summer fever, with well-known pathogenic potential for humans ranging from asymptomatic to mild fever, in addition to neuro-invasive infections during summer. Although TOSV, in particular, is a significant and well-known human pathogen, SFVs remain neglected, with many gaps in the relevant knowledge. Sero-epidemiological studies and case reports recently showed a geographical wider distribution than previously considered, although the real incidence of phleboviruses infections in the Mediterranean area is still unknown. Here we retrospectively evaluated the circulation of phleboviruses during summer seasons between 2007 and 2019 in 649 patients showing neurological symptoms using both molecular and serological approaches. We found that 42/649 (6.5%) subjects experienced phlebovirus infection and only 10/42 cases were detected by molecular assays, whereas the other 32/42 were identified using serological approaches, including neutralization assays. During the 2013 summer, an outbreak in the Lombardy region is described because the prevalence of phlebovirus infection reached 37.2% (19/51 subjects). Interestingly, only 5/19 (26.5%) reported traveling in endemic areas. Of note, no cross-neutralization was observed between different strains tested, showing the possibility to be reinfected by newly discovered phlebovirus strains. In conclusion, phlebovirus infections are still inadequately considered by physicians and are generally underestimated. However, based on our results, sandfly fever viruses should be routinely included in diagnostic panels during summer period, including in Northern Italy.

Background: The service strategy (same-day transfer between the spoke hospital and the hub center with catheterization laboratory facility to perform invasive procedures) has been shown to improve the management of patients with non-ST-elevation acute coronary syndrome admitted to spoke hospitals. However, few data exist about the safety of this strategy and, in particular, the safety of retransferring patients to spoke centers immediately after successful percutaneous coronary intervention., Methods: We used data from a prospective registry to retrospectively describe the application, performance and outcome of the service strategy in the daily clinical practice in our province, organized in 5 spoke hospitals and a hub center in Reggio Emilia, Italy., Results: From January 2013 to December 2017, 1183 consecutive patients were admitted to the cath-lab in the hub center from spoke hospitals with a diagnosis of non-ST-elevation acute coronary syndrome. Mean age was 68 ± 12 years, with a mean GRACE risk score of 137 ± 3. Overall, 1063 patients (90%) were managed with a service strategy. The mean time between hospital admission and access to the cath-lab for coronary artery angiography ± percutaneous coronary intervention was 46.6 h (27.5-71.2). No major adverse events (all-cause mortality, arrhythmias, or acute myocardial infarction) were observed during the back transfer from the invasive center to the referring non-invasive center. At 30 days, acute myocardial infarction was observed in 1.3% of patients and all-cause mortality was 0.5%. At 1-year follow-up, acute myocardial infarction was reported in 10% of patients and all-cause mortality was 2.8%., Conclusions: A wide adoption of the service strategy in our provincial network in patients with non-ST-elevation acute coronary syndrome admitted to spoke centers allowed an early access to the cath-lab as recommended by international guidelines. The safety of the service strategy is confirmed in our experience, with no major adverse events occurring during the back transfer.

With several human cases reported annually since 2008 and the unapparent risk of infection of blood donors, the West Nile virus (WNV) is emerging as an important health issue in Europe. Italy, as well as other European countries, experienced a recrudescence of the virus circulation in 2018, which led to an increased number of human cases. An integrated surveillance plan was activated in the Emilia-Romagna and Lombardy regions (Northern Italy) since 2008 in order to monitor the intensity and timing of WNV circulation. A fundamental part of this plan consists in entomological surveillance. In 2018, the surveillance plan made it possible to collect 385,293 mosquitoes in 163 stations in the two Regions. In total 269,147 Culex mosquitoes were grouped into 2,337 pools and tested for WNV, which was detected in 232 pools. Circulation started in the central part of the Emilia-Romagna region in the middle of June, about one month before the previous seasons. Circulation suddenly expanded to the rest of the region and reached the Lombardy region in the middle of July. WNV circulated more intensively in the eastern part of the surveyed area, as confirmed by the highest number of human cases. A relationship between the number of mosquitoes collected and the virus incidence emerged, but the data obtained highlighted that the probability of detecting the virus in a given site was less than expected with a higher number of collected mosquitoes. A significant relationship was observed between the temperature recorded one week before the sampling and the number of collected mosquitoes, as well as between the estimated number of WNV-positive mosquitoes and the temperature recorded two weeks before the sampling. The two weeks delay in the influence of temperature on the positive mosquitoes is in line with the time of the virus extrinsic incubation in the mosquito. This finding confirms that temperature is one of the principal drivers in WNV mosquito infection. The surveillance system demonstrated the ability to detect the virus circulation early, particularly in areas where circulation was more intense. This allowed evaluating the effect of mosquito abundance and weather factors on virus circulation., (Copyright © 2020 Calzolari, Angelini, Bolzoni, Bonilauri, Cagarelli, Canziani, Cereda, Cerioli, Chiari, Galletti, Moirano, Tamba, Torri, Trogu, Albieri, Bellini and Lelli.)

Background and Aims: Hepatitis E Virus is endemic in Europe with increasing numbers of cases in recent years, also in Italy where this phenomenon has hitherto been modest. The aim of this study was to document the clinical features/natural history of locally acquired hepatitis E in our territory and explore factors which determine adverse outcome., Methods: Retrospective study of patients with locally-acquired HEV (hepatitis E virus) in Marche, Italy (2011-2019)., Results: 1189 patients were tested for HEV with 89 confirmed cases. 81 (6.8%) had locally acquired infection; 54 (66%) were male (mean age 55.5 years) and 32 (39.5%) had active co-morbidities. 41 cases were viraemic (all HEV-3 (HEV genotype 1,2,3,4)); acute infection was found in 79 and chronic infection in 2. Forty-five cases (55%) required admission to hospital, for a total of 785 days. 4 patients developed acute on-chronic liver failure, 6 developed acute kidney injury and 8 died: all had active comorbidities. Univariate analysis showed that bilirubin, INR, immunosuppression, cirrhosis and diabetes were associated with death. On multivariant analysis the only predictor of death was the presence of diabetes (p = 0.04)., Conclusions: Hepatitis E in Marche Italy is mostly locally acquired and caused by HEV-3 that impacts on the morbidity and mortality particularly for fragile patients., (Copyright © 2019 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.)

In Italy, the first human case of West Nile Virus (WNV) infection was reported in 2008 and, since then, the number of cases has been steadily increasing. In this study, we describe the temporal and spatial pattern of WNV infection risk among humans in Italy, focusing on the human cases of West Nile Neuro-invasive Disease (WNND) observed between 2008 and 2017. Incidence rates are estimated for each year and province under study. The incidence temporal trend is estimated using Poisson regression, and a spatio-temporal cluster detection analysis is performed to detect high-risk areas. In total, 231 WNND cases were notified in Italy between 2008 and 2017. The annual incidence rates increased during the study period (annual percentage change: 11.7%; 95%CI: -0.9%; 26.1%). A geographical spread of the disease was observed during the study period throughout Northern Italy, with an increasing number of affected provinces. Provinces close to the Po River (the main river in the north of Italy) and the Oristano province (in the Sardinia Island) experienced the highest incidence rates during the study period. Our study shows a gradual, but rapid spread of WNND across Northern Italy from east to west and suggests the hypothesis that provinces close to Po River might present ecological and climatic conditions favourable to the virus circulation., (© 2019 Blackwell Verlag GmbH.)

Toscana virus (TOSV) is a Phlebovirus transmitted by phlebotomine sand flies and is an important etiological agent of summer meningitis in the Mediterranean basin. Since TOSV infection is often asymptomatic, we evaluated the seroprevalence in blood donors (BDs) in the Bologna and Ferrara provinces (Northeastern Italy)-the areas with the highest and lowest numbers of TOSV neuroinvasive cases in the region, respectively. A total of 1208 serum samples from BDs were collected in April-June 2014 and evaluated for the presence of specific TOSV-IgG by ELISA. The IgG-reactive samples were confirmed by indirect immunofluorescence assay (IIF) and by microneutralization test (MN). Serum samples were defined as positive for anti-TOSV IgG when reactive by ELISA and by at least one second-level test; TOSV seroprevalence was 6.8% in the Bologna province, while no circulation of TOSV was detected in the Ferrara province. Sand fly abundance in 2014 was also estimated by a geographic information system using a generalized linear model applied to a series of explanatory variables. TOSV seroprevalence rate was strongly associated with the sand fly abundance index in each municipality, pointing out the strong association between sand fly abundance and human exposure to TOSV., Competing Interests: The authors declare no conflicts of interest.

Background: West Nile virus (WNV) transmission was much greater in 2018 than in previous seasons in Europe. Focusing on Emilia-Romagna region (northern Italy), we analyzed detailed entomological and epidemiological data collected in 2013-2018 to quantitatively assess environmental drivers of transmission and explore hypotheses to better understand why the 2018 epidemiological season was substantially different than the previous seasons. In particular, in 2018 WNV was detected at least two weeks before the observed circulation in 2013-2017 and in a larger number of mosquito pools. Transmission resulted in 100 neuroinvasive human cases in the region, more than the total number of cases recorded between 2013 and 2017., Methodology: We used temperature-driven mathematical models calibrated through a Bayesian approach to simulate mosquito population dynamics and WNV infection rates in the avian population. We then estimated the human transmission risk as the probability, for a person living in the study area, of being bitten by an infectious mosquito in a given week. Finally, we translated such risk into reported WNV human infections., Principal Findings: The estimated prevalence of WNV in the mosquito and avian populations were significantly higher in 2018 with respect to 2013-2017 seasons, especially in the eastern part of the region. Furthermore, peak avian prevalence was estimated to have occurred earlier, corresponding to a steeper decline towards the end of summer. The high mosquito prevalence resulted in a much greater predicted risk for human transmission in 2018, which was estimated to be up to eight times higher than previous seasons. We hypothesized, on the basis of our modelling results, that such greater WNV circulation might be partially explained by exceptionally high spring temperatures, which have likely helped to amplify WNV transmission at the beginning of the 2018 season., Competing Interests: The authors have declared that no competing interests exist.

Visceral leishmaniasis (VL) caused by Leishmania (L.) infantum is a public health threat in the Emilia-Romagna region, northeastern Italy, but its epidemiology has not been fully elucidated in this area. The objective of this study was to characterize Leishmania infection in sand flies collected in a re-emerging focus of VL in the Bologna province. During the summer of 2016, 6114 sand flies were collected, identified, and tested for Leishmania detection. Of the identified sand flies, 96.5% were Phlebotomus (P.) perfiliewi and 3.5% were P. perniciosus . Detected parasites were characterized by biomolecular methods (multilocus microsatellite typing and characterization of repetitive region on chromosome 31), and quantified by real-time PCR. The prevalence of Leishmania infection in individually-tested P. perfiliewi sand flies varied from 6% to 10% with an increasing trend during the season. Promastigotes of L. infantum were isolated by dissection in one P. perfiliewi female ; the isolated strain (Lein-pw) were closely related to Leishmania parasites from VL cases in northeastern Italy, but differed from strains isolated in dogs from the same area. Our findings strongly support the vector status of P. perfiliewi for human VL in the study area., Competing Interests: The authors declare no conflict of interest.

Lineage 2 West Nile virus (WNV) caused a vast epidemic in Europe in 2018, with the highest incidence being recorded in Italy. To reconstruct the evolutionary dynamics and epidemiological history of the virus in Italy, 53 envelope gene and 26 complete genome sequences obtained from human and animal samples were characterised by means of next-generation sequencing. Phylogenetic analysis revealed two Italian strains originating between 2010 and 2012: clade A, which apparently became extinct in 2013-2014, and clade B, which was responsible for the 2018 epidemic. The mean genetic distances in clade B increased over time and with the distance between sampling locations. Bayesian birth-death and coalescent skyline plots of the clade B showed that the effective number of infections and the effective reproduction number (Re) increased between 2015 and 2018. Our data suggest that WNV-2 entered Italy in 2011 as a result of one or a few penetration events. Clade B differentiated mainly as a result of genetic drift and purifying selection, leading to the appearance of multiple locally circulating sub-clades for different times. Phylodynamic analysis showed a current expansion of the infection among reservoir birds and/or vectors.

The Trypanosoma theileri group includes several trypanosome species hardly distinguishable due to the lack of discriminating morphological characters. Trypanosomes belonging to this group have been isolated from different bovine, ovine, and cervids in Europe, Africa, Asia, and Americas. The principal vectors of the T. theileri group are considered tabanid flies; however, T. melophagium is transmitted exclusively by sheep keds. In 2016, 128 sand flies out of 2,728 trapped in Valsamoggia municipality, Italy, were individually dissected and an unknown trypanosome strain, named TrPhp1, was isolated from a female of the sand fly Phlebotomus perfiliewi . Sequence analysis placed this trypanosome in the T. theileri group with very high homology to other trypanosomes detected in European cervids. This is the first report of the T. theileri group isolation from a sand fly, and the possible role of this insect group in the trypanosome transmission cycle is discussed. Within the T. theileri group, the phylogenetic analysis distinguished several lineages, which, unfortunately, do not correspond with their host specificity and their taxonomic status remains ambiguous.

Background: Visceral leishmaniasis (VL) caused by Leishmania infantum is an ongoing health problem in southern Europe, where dogs are considered the main reservoirs of the disease. Current data point to a northward spread of VL and canine leishmaniasis (CanL) in Italy, with new foci in northern regions previously regarded as non-endemic., Methodology/principal Findings: Multilocus microsatellite typing (MLMT) was performed to investigate genetic diversity and population structure of L. infantum on 55 samples from infected humans, dogs and sand flies of the E-R region between 2013 and 2017. E-R samples were compared with 10 L. infantum samples from VL cases in other Italian regions (extra E-R) and with 52 strains within the L. donovani complex. Data displayed significant microsatellite polymorphisms with low allelic heterozygosity. Forty-one unique and eight repeated MLMT profiles were recognized among the L. infantum samples from E-R, and ten unique MLMT profiles were assigned to the extra E-R samples. Bayesian analysis assigned E-R samples to two distinct populations, with further sub-structuring within each of them; all CanL samples belonged to one population, genetically related to Mediterranean MON-1 strains, while all but one VL cases as well as the isolate from the sand fly Phlebotomus perfiliewi fell under the second population. Conversely, VL samples from other Italian regions proved to be genetically similar to strains circulating in dogs., Conclusions/significance: A peculiar epidemiological situation was observed in northeastern Italy, with the co-circulation of two distinct populations of L. infantum; one population mainly detected in dogs and the other population detected in humans and in a sand fly. While the classical cycle of CanL in Italy fits well into the data obtained for the first population, the population found in infected humans exhibits a different cycle, probably not involving a canine reservoir. This study can contribute to a better understanding of the population structure of L. infantum circulating in northeastern Italy, thus providing useful epidemiologic information for public health authorities., Competing Interests: The authors have declared that no competing interests exist.

Background: Different phleboviruses are important pathogens for humans; most of these viruses are transmitted by sand flies. An increasing number of new phleboviruses have been reported over the past decade, especially in Mediterranean countries, mainly via their detection in sand flies., Results: At least five different phleboviruses co-circulated in sand flies that were collected in three sites in Emilia-Romagna (Italy) in the summer of 2013. The well-known Toscana virus (TOSV) was isolated; three new, closely related phleboviruses differing in their M segments and tentatively named Ponticelli I, Ponticelli II and Ponticelli III virus, respectively, were isolated; a fifth putative phlebovirus, related to the sand fly fever Naples phlebovirus species, was also detected. The co-circulation, in a restricted area, of three viruses characterized by different M segments, likely resulted from reassortment events. According to the phylogenetic analysis of complete genome sequences, the TOSV belongs to clade A, together with other Italian isolates, while the Ponticelli viruses fall within the Salehabad phlebovirus species., Conclusions: Results highlight an unexpected diversity of phleboviruses that co-circulate in the same area, suggesting that interactions likely occur amongst them, that can present challenges for their correct identification. The co-circulation of different phleboviruses appears to be common, and the bionomics of sand fly populations seem to play a relevant role. Such a complex situation emphasizes the need for detailed investigations of the biology of these viruses to better characterize their pathogenic potential for mammals, including humans.

Pathogens transmitted to humans by phlebotomine sand flies are neglected, as they cause infectious diseases that are not on the priority list of national and international public health systems. However, the infections caused by protozoa of the Leishmania genus and viruses belonging to the Phlebovirus genus (family Phenuiviridae)-the most significant group of viruses transmitted by sand flies-have a relevant role for human pathology. These infections are emerging in the Mediterranean region and will likely spread in forthcoming decades, posing a complex threat to human health. Four species and 2 hybrid strains of Leishmania are pathogenic for humans in the Mediterranean Basin, with an estimated annual incidence of 239,500-393,600 cases of cutaneous leishmaniasis and 1,200-2,000 cases of visceral leishmaniasis. Among the phleboviruses, Toscana virus can cause neuroinvasive infections, while other phleboviruses are responsible for a typical "3-day fever"; the actual incidence of Phlebovirus infections in the Mediterranean area is unknown, although at least 250 million people are exposed. Here, we reviewed the current literature on epidemiology of sand fly-borne infections in the Mediterranean Basin, with a focus on humans. Our analysis indicates the need for increased public health activities directed to determine the disease burden of these infections as well as to improve their surveillance. Among the emerging challenges concerning sand fly-borne pathogens, the relationships between sand fly-borne protozoa and viruses should be considered in future studies, including epidemiological links between Leishmania and phleboviruses as well as the conditional capacity for these pathogens to be involved in interactions that may evolve towards increased virulence.

This study evaluated the vector competence of Aedes albopictus in transmitting USUV after oral infection under laboratory conditions. Ae. albopictus showed a low vector competence for USUV, although the positive body sample found with a very high number of viral copies at one week post infection indicates that a replication in the mosquito body can occur, and that USUV can escape the midgut barrier. Field data from an extensive entomological arboviruses surveillance program showed a relevant incidence of Ae. albopictus USUV positive pools in the period 2009-2012 while all pools were negative from 2013 on. No conceivable explanation regarding this field evidence was addressed, suggesting that attention must be paid to the trend of development of this vector-pathogen association, being aware of the potential rapid arbovirus' adaptation to new vectors, to prevent possible new disease's emergence., (Copyright © 2017 Elsevier B.V. All rights reserved.)

West Nile virus lineage 2 (WNV-2) was mainly confined to sub-Saharan Africa until the early 2000s, when it was identified for the first time in Central Europe causing outbreaks of human and animal infection. The aim of this study was to reconstruct the origin and dispersion of WNV-2 in Central Europe and Italy on a phylodynamic and phylogeographical basis. To this aim, discrete and continuous space phylogeographical models were applied to a total of 33 newly characterised full-length viral genomes obtained from mosquitoes, birds and humans in Northern Italy in the years 2013-2015 aligned with 64 complete sequences isolated mainly in Europe. The European isolates segregated into two highly significant clades: a small one including three sequences and a large clade including the majority of isolates obtained in Central Europe since 2004. Discrete phylogeographical analysis showed that the most probable location of the root of the largest European clade was in Hungary a mean 12.78 years ago. The European clade bifurcated into two highly supported subclades: one including most of the Central/East European isolates and the other encompassing all of the isolates obtained in Greece. The continuous space phylogeographical analysis of the Italian clade showed that WNV-2 entered Italy in about 2008, probably by crossing the Adriatic sea and reaching a central area of the Po Valley. The epidemic then spread simultaneously eastward, to reach the region of the Po delta in 2013, and westward to the border area between Lombardy and Piedmont in 2014; later, the western strain changed direction southward, and reached the central area of the Po valley once again in 2015. Over a period of about seven years, the virus spread all over an area of northern Italy by following the Po river and its main tributaries.

Usutu virus is an arbovirus closely related to West Nile virus (genus Flavivirus), which circulates between mosquitoes and wild birds. This virus has been increasingly reported in Europe, raising concerns for its possible pathogenic potential for wild birds and humans. This study reports the whole genome sequences of 15 strains of USUV, isolated between 2010 and 2014 from mosquitoes and wild birds in the course of West Nile virus surveillance in the Emilia-Romagna and Lombardy regions of Northern Italy. Both whole and 656 partial genome sequences, obtained from isolated viruses and field samples (mosquitoes and wild birds), were analyzed to describe the temporal and geographical spread of USUV in the surveyed area. The detected sequences belonged to two groups, with one circulating primarily in the northwestern part of the area and the other in the southeastern part. This pattern is likely the result of different routes of introduction from the North (over the Alps) and from the East, respectively. The phylogenetic analysis of obtained sequences and other European sequences demonstrated that the majority of European strains belonged to one main clade, while less common strains, mainly from Western Europe, fell in other two clades. This analysis strongly suggested an autochthonous evolution process of strains of the main clade from a common ancestor with an estimated time of arrival in Europe at the beginning of the 1990s. In addition to causing mass mortality in wild birds, Usutu virus can infect humans and can sporadically cause disease. These factors and the endemization of the Usutu virus in a large area of Europe, sustained by the obtained data, strongly support the need to adequately survey Usutu virus in areas in which its circulation is detected., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Usutu (USUV) and West Nile (WNV) are mosquito-borne Flavivirus emerged in Italy in 1996 and 1998, respectively, and reappeared 10 years later. The aim of this work is to review the Italian mosquito species found positive for WNV and USUV between 2008 and 2014. Moreover, the role of mosquitoes in promoting the overwintering of these viruses is discussed, as a result of the mosquito collections performed in Molise region between September 2010 and April 2011. Overall 99,000 mosquitoes were collected: 337 and 457 mosquito pools tested positive by real time reverse transcriptase polymerase chain reaction (real time RT-PCR) for WNV and USUV, respectively. West Nile virus was detected in pools of Culex pipiens s.l. (329), Ochlerotatus caspius (4), Culex modestus (2), and Culex spp. (2). Positive USUV pools were from Cx. pipiens s.l. (435), Aedes albopictus (12), Oc. caspius (5), Culex spp. (2), Anopheles maculipennis s.l. (1), Culiseta annulata (1), and Ochlerotatus detritus (1). In Molise region, 1,694 mosquitoes were collected, and USUV was identi ed in Cx. pipiens s.l., Cs. annulata, and Oc. detritus pools. This paper shows that Cx. pipiens s.l. is the mosquito species most involved in the WNV and USUV circulation in Italy, although other species would also support the spread of both the viruses during Winter.

Human leishmaniasis is an emerging problem in Italy and is on the increase in the Emilia-Romagna region, northeastern part of the country. Nevertheless, studies dealing with the molecular characterization of Leishmania spp. circulating in these areas are limited. In the present work, we explored the genetic polymorphism of Leishmania isolates from 28 cases of canine leishmaniasis and three cases of human visceral leishmaniasis (VL), which occurred in 2013-2014 in the Emilia-Romagna region. The characterization was carried out in comparison with nine human isolates of Leishmania from other VL endemic Italian regions and two reference strains. Nucleic acid from 31 Leishmania-positive phlebotomine sandfly pools, sampled in 2012-2013 in the Emilia-Romagna region, were also evaluated. DNA amplification and sequencing of the ribosomal internal transcribed spacer-1 and of a repetitive nuclear region on chromosome 31 were carried out for genotyping. Two size polymorphic targets were also analyzed by PCR, the cpb E/F-gene and the k26-gene. Altogether, the analysis showed the circulation of different Leishmania infantum genotypes in the Emilia-Romagna region: two genotypes found in dogs from public kennels were similar to VL isolates from other Italian regions, whereas a third genotype was detected in VL cases of the Emilia-Romagna region and in all but one of the sandfly pools. The combined molecular tools applied in this study can constitute a helpful support for parasite tracking (e.g., in outbreak investigations) and for a better understanding of the epidemiological evolution of leishmaniasis in northeastern Italy.

From 1 May 2015 to 31 October 2015 over 20 million visitors from all over the world visited the Universal Exhibition (EXPO) hosted by Milan (Lombardy region, Italy), raising concerns about the possible introduction of mosquito-borne diseases from endemic countries. The entomological surveillance protocol performed in Lombardy over the last three years was implemented in the EXPO area and in the two major regional airports using both Center for Disease Control CO₂ and Biogents Sentinel traps. This surveillance aimed to estimate the presence and densities of putative vectors, and also to support investigations, including the vector species involved and area of diffusion, on the local spread of Chikungunya, Dengue and West Nile viruses (WNV) by competent vectors. From 3544 mosquitoes belonging to five different species, 28 pools of Culex spp. and 45 pools of Aedes spp. were screened for the presence of WNV, and for both Chikungunya and flaviviruses, respectively. The entomological surveillance highlighted a low density of potential vectors in the surveyed areas and did not reveal the presence of Chikungunya or Dengue viruses in the local competent vectors inside the EXPO area or in the two airports. In addition, the surveillance reported a low density of Culex spp. mosquitoes, which all tested negative for WNV.

Several flaviviruses are important pathogens for humans and animals (Dengue viruses, Japanese encephalitis virus, Yellow-fever virus, Tick-borne encephalitis virus, West Nile virus). In recent years, numerous novel and related flaviviruses without known pathogenic capacity have been isolated worldwide in the natural mosquito population. However, phylogenetic studies have shown that genomic sequences of these viruses diverge from other flaviviruses. Moreover, these viruses seem to be exclusive of insects (they do not seem to grow on vertebrate cell lines), and were already defined as mosquito-only flaviviruses or insect-specific flaviviruses. At least eleven of these viruses were isolated worldwide, and sequences ascribable to other eleven putative viruses were detected in several mosquito species. A large part of the cycle of these viruses is not well known, and their persistence in the environment is poorly understood. These viruses are detected in a wide variety of distinct mosquito species and also in sandflies and chironomids worldwide; a single virus, or the genetic material ascribable to a virus, was detected in several mosquito species in different countries, often in different continents. Furthermore, some of these viruses are carried by invasive mosquitoes, and do not seem to have a depressive action on their fitness. The global distribution and the continuous detection of new viruses in this group point out the likely underestimation of their number, and raise interesting issues about their possible interactions with the pathogenic flaviviruses, and their influence on the bionomics of arthropod hosts. Some enigmatic features, as their integration in the mosquito genome, the recognition of their genetic material in DNA forms in field-collected mosquitoes, or the detection of the same virus in both mosquitoes and sandflies, indicate that the cycle of these viruses has unknown characteristics that could be of use to reach a deeper understanding of the cycle of related pathogenic flaviviruses., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Background: The invasive species Aedes (Finlaya) koreicus was first identified in north-eastern Italy in 2011, during the ongoing surveillance activity of Aedes albopictus. Following this finding, a more intensive monitoring was carried out to assess the distribution of the species and to collect biological data. Herein, we report the new records obtained by four years of surveillance., Findings: The presence of Ae. koreicus was checked using ovitraps, adults traps and by larval collections in all possible breeding sites from May 2011 to July 2015. The monitoring started in the site of the first detection (Province of Belluno) and was then extended in the neighbouring Provinces belonging to four Regions. Aedes koreicus was found in 73 municipalities out of 155 monitored (47.1 %), including 23 municipalities (14.8 %) previously not infested. The area of first detection of Ae. koreicus (Province of Belluno) was also the most infested (68 %). However the mosquito has also been found to the west (Province of Trento) and to the south and south-west (Provinces of Vicenza and Treviso) of the initially infested area., Conclusions: The spread of Ae. koreicus is directed towards south and west from the original infested area, likely due to the dense road connections and the habitat suitability of the new areas. According to these records, northern Italy has a high probability to be invaded by Ae. koreicus in the next decade. These data can be useful to validate predictive models of potential distribution and dispersal of this species in Italy or in Europe.

West Nile virus (WNV) is a recently re-emerged health problem in Europe. In Italy, an increasing number of outbreaks of West Nile disease, with occurrences of human cases, have been reported since 2008. This is particularly true in northern Italy, where entomological surveillance systems have been implemented at a regional level. The aim of this study was to use, for the first time, all the entomological data collected in the five regions undergoing surveillance for WNV in northern Italy to characterize the viral circulation (at a spatial and temporal scale), identify potential mosquito vectors, and specify relationships between virus circulation and meteorological conditions. In 2013, 286 sites covering the entire Pianura Padana area were monitored. A total of 757,461 mosquitoes were sampled. Of these, 562,079 were tested by real-time PCR in 9,268 pools, of which 180 (1.9%) were positive for WNV. The largest part of the detected WNV sequences belonged to lineage II, demonstrating that, unlike those in the past, the 2013 outbreak was mainly sustained by this WNV lineage. This surveillance also detected the Usutu virus, a WNV-related flavivirus, in 241 (2.6%) pools. The WNV surveillance systems precisely identified the area affected by the virus and detected the viral circulation approximately two weeks before the occurrence of onset of human cases. Ninety percent of the sampled mosquitoes were Culex pipiens, and 178/180 WNV-positive pools were composed of only this species, suggesting this mosquito is the main WNV vector in northern Italy. A significantly higher abundance of the vector was recorded in the WNV circulation area, which was characterized by warmer and less rainy conditions and greater evapotranspiration compared to the rest of the Pianura Padana, suggesting that areas exposed to these conditions are more suitable for WNV circulation. This observation highlights warmer and less rainy conditions as factors able to enhance WNV circulation and cause virus spillover outside the sylvatic cycle.

In the summer of 2013, an outbreak of West Nile virus (WNV) infection occurred in the Lombardy, a region of northern Italy to the west of districts affected by WNV in previous years. Eighteen cases of human WNV infection were diagnosed--10 cases of acute WNV neuroinvasive disease and eight of WNV fever. In the same period, WNV was detected in birds (one crow) in horses (11 cases) and from mosquitoes (six pools).

The impact of weather variables on Culex pipiens L. (Diptera: Culicidae) population dynamics in the Po Valley, Northern Italy, a densely populated region containing the largest industrial and agricultural areas in Italy, was investigated. Monitoring of mosquitoes was carried out by using CO(2)-baited traps without light, collecting data weekly from 1700 to 0900 hours during the period May-September, from 1997 to 2011. Daily minimum, average, and maximum relative humidity; daily minimum, maximum, and average temperature; rainfall; and hydroclimatic balance (rainfall-potential evapotranspiration) were obtained from three weather stations within the surveillance zone. The average population dynamic trend over the 15-yr period showed a bell-shaped curve with a major peak in June and a secondary peak at the end of August in the rural areas, whereas bimodality was not evidenced in the urban areas. The correlation analyses showed that the mosquito seasonal population and the population in the period of maximum West Nile virus circulation (August-September) was mostly affected by the relative humidity registered from March to July, particularly in May, and, to a lower extent, also by hydroclimatic balance registered in April-July, and by the rainfall occurred in June-July. In addition, the rate of increase of the population during the spring months influenced the development of the mosquito population of the following months.

Since 2007, a canine leishmaniasis (CanL) surveillance program has been carried out in public kennels of the Emilia-Romagna region with the aim of providing health guarantees for dog adoptions. According to this program, monitoring activities were performed to verify the presence of sandflies and infected dogs, and a specific CanL risk class was assigned to each kennel, resulting in different control approaches (entomological and/or serological monitoring, clinical surveillance, therapeutic treatment of infected dogs, protections against vector bites). From 2007 to 2012, 20,931 dogs, 89.8% of which were identified by microchip and housed in 73 kennels, were examined using an indirect fluorescent antibody test. In all, 528 (2.8%) dogs tested positive, and 43.0% of these were asymptomatic. The authors used monitoring results, in particular serological tests performed on dogs at admittance to the kennel and annual controls of sentinel dogs, to estimate CanL risk in the whole region and to evaluate the efficacy of the preventive measures adopted. CanL seroprevalence in dogs tested at the admittance in kennels increased significantly from 2010 (1.0%; 29/2858) to 2012 (2.4%; 69/2841). In contrast, the number of seroconversions in sentinel dogs was stable in 2010 (1.2%; 11/896) and 2011 (1.6%; 13/825) and decreased in 2012 (0.9%; 8/850), suggesting the efficacy of the preventive measures applied.

The purpose of the study described here was to prospectively evaluate the significance of embolization of the spleen in patients with definite left-sided infective endocarditis (IE) using contrast-enhanced ultrasound (CEUS). From March through October 2012, 6 consecutive patients (4 females and 2 males, aged 27 to 83 years) with definite left-sided IE according to the revised Duke criteria were enrolled. All patients gave informed written consent, and the study was performed in conformity with the ethical guidelines of the Declaration of Helsinki. All patients underwent CEUS of the spleen within 1 week of the definite diagnosis of IE. A blood pool second-generation contrast agent and an ultrasound machine with contrast harmonic imaging technology were used for CEUS. Fifteen consecutive patients (7 females and 8 males, aged 39 to 88 years) who underwent CEUS from October through November 2012 for the study of focal liver lesions constituted the control group. The number of patients did not permit statistical analysis. Splenic CEUS revealed infarctions in 5 patients and an infarcted area in the only patient with negative echocardiography. All splenic CEUS procedures in the control group were negative. In this study, CEUS of the spleen, a repeatable and low-cost imaging technique, easily allowed bedside detection of asymptomatic and even tiny infarctions, and revealed a high rate of embolization in patients with definite left-sided IE. Indeed, splenic CEUS, if applied to the workup of patients with suspect IE, has the potential to accelerate or upgrade the diagnosis itself., (Copyright © 2013 World Federation for Ultrasound in Medicine & Biology. Published by Elsevier Inc. All rights reserved.)

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.

West Nile virus (WNV) is one of the most serious public health threats that Europe and the Mediterranean countries are currently facing. In Italy, WNV emerged in 1998 and has been circulating since 2008. To tackle its continuous incursions, Italian national and regional institutions set up a surveillance program, which includes the serological screening of sentinel horses, sentinel-chickens and backyard poultry flocks and the surveillance on all equine neurological cases, resident captured and wild dead birds, and vectors. This communication aims to assess the importance of the entomological surveillance program as an early warning system for WNV circulation. In the province of Modena, the circulation of WNV lineage 2 strains was first detected in pools of Culex pipiens on July the 3rd, 42 days prior to the onset of the first 2013 human WNV neuroinvasive case reported in the same province. Similarly in Veneto, WNV was first detected on July 3rd in a pool of Cx. pipiens collected in the province of Venezia. The first human neuroinvasive case in this region occurred in the Rovigo province on July the 24th, seven days after the detection of WNV lineage 2 in a mosquito pool collected in the same province. Up to the end of July 2013, WNV circulation was further detected in several other pools of Cx. pipiens mosquitoes collected in Emilia-Romagna, Veneto and Lombardia. According to the NS3 partial sequence alignments including all recent European and Italian Lineage 2 strains, the new circulating WNV lineage 2 strains share high nt homology with the Hungarian and with the previous lineage 2 strains isolated in Veneto and Sardegna in 2011 and 2012. These data provide a clear and practical demonstration of the relevance of a reliable entomological surveillance program to early detect WNV in Italy.

Background: The circulation of West Nile virus and Usutu virus was detected in the Emilia-Romagna region in 2008 and 2009. To evaluate the extent of circulation of both viruses, environmental surveillance, based on bird and mosquito testing, was conducted in 2008 and gradually improved over the years., Methods: In February-March 2009-2011, 5,993 hibernating mosquitoes were manually sampled, out of which 80.1% were Culex pipiens; none tested positive for the viruses. From 2008 to 2011, 946,213 mosquitoes, sampled between May and October, were tested; 86.5% were Cx. pipiens. West Nile virus was detected in 32 Cx. pipiens pools, and Usutu virus was detected in 229 mosquito pools (217 Cx. pipiens, 10 Aedes albopictus, one Anopheles maculipennis s.l., and one Aedes caspius). From 2009 to 2011, of 4,546 birds collected, 42 tested positive for West Nile virus and 48 for Usutu virus. West Nile virus and Usutu virus showed different patterns of activity during the 2008-2011 surveillance period. West Nile virus was detected in 2008, 2009, and 2010, but not in 2011. Usutu virus, however, was continuously active throughout 2009, 2010, and 2011., Conclusions: The data strongly suggest that both viruses overwinter in the surveyed area rather than being continually reintroduced every season. The lack of hibernating mosquitoes testing positive for the viruses and the presence of positive birds sampled early in the season support the hypothesis that the viruses overwinter in birds rather than in mosquitoes. Herd immunity in key bird species could explain the decline of West Nile virus observed in 2011, while the persistence of Usutu virus may be explained by not yet identified reservoirs. Reported results are comparable with a peri-Mediterranean circulation of the West Nile virus lineage 1 related strain, which became undetectable in the environment after two to three years of obvious circulation.

Background: In 2011, a new orthobunyavirus, named the Schmallenberg virus (SBV), was discovered in Europe. Like the related Shamonda virus, SBV is an arbovirus (arthropod-borne virus). After its discovery, the virus was detected in a wide area in north-western Europe, an unexpected finding in a territory where climatic conditions would not seem ideal for arbovirus transmission. This sudden expansion suggests the effect of 2011 drought as a key factor that may have triggered SBV circulation. The possible influence of drought, recorded in north-western Europe in early 2011, on virus circulation was evaluated., Methods and Results: The locations of SBV detections in Europe until April 2012 were obtained, and area of virus circulation was evaluated by kernel density estimation. Precipitation data in SBV circulation area, summarized by the 3 month precipitation indexes of May, were compared with precipitation data outside that area, confirming driest conditions in that area., Conclusions: The onset of drought conditions recorded in the SBV detection area in early 2011 may have promoted the circulation of this virus. A correlation between circulation of some arboviruses and drought has been reported elsewhere. This was mainly explained by an effect of water deficit on the environment, which altered the relationships between vectors and reservoirs, but this correlation might be also the result of unknown effects of drought on the vectors. The effect of drought conditions on arbovirus circulation is most likely underestimated and should be considered, since it could promote expansion of arboviruses into new areas in a global warming scenario.

The genus Flavivirus, family Flaviviridae, includes a number of important arthropod-transmitted human pathogens such as dengue viruses, West Nile virus, Japanese encephalitis virus and yellow fever virus. In addition, the genus includes flaviviruses without a known vertebrate reservoir, which have been detected only in insects, particularly in mosquitoes, such as cell fusing agent virus, Kamiti River virus, Culex flavivirus, Aedes flavivirus, Quang Binh virus, Nakiwogo virus and Calbertado virus. Reports of the detection of these viruses with no recognized pathogenic role in humans are increasing in mosquitoes collected around the world, particularly in those sampled in entomological surveys targeting pathogenic flaviviruses. The presence of six potential flaviviruses, detected from independent European arbovirus surveys undertaken in the Czech Republic, Italy, Portugal, Spain and the UK between 2007 and 2010, is reported in this work. Whilst the Aedes flaviviruses, detected in Italy from Aedes albopictus mosquitoes, had already been isolated in Japan, the remaining five viruses have not been reported previously: one was detected in Italy, Portugal and Spain from Aedes mosquitoes (particularly from Aedes caspius), one in Portugal and Spain from Culex theileri mosquitoes, one in the Czech Republic and Italy from Aedes vexans, one in the Czech Republic from Aedes vexans and the last in the UK from Aedes cinereus. Phylogenetic analysis confirmed the close relationship of these putative viruses to other insect-only flaviviruses.

Background: In 2008, after the first West Nile virus (WNV) detection in the Emilia-Romagna region, a surveillance system, including mosquito- and bird-based surveillance, was established to evaluate the virus presence. Surveillance was improved in following years by extending the monitoring to larger areas and increasing the numbers of mosquitoes and birds tested., Methodology/principal Findings: A network of mosquito traps, evenly distributed and regularly activated, was set up within the surveyed area. A total of 438,558 mosquitoes, grouped in 3,111 pools and 1,276 birds (1,130 actively sampled and 146 from passive surveillance), were tested by biomolecular analysis. The survey detected WNV in 3 Culex pipiens pools while Usutu virus (USUV) was found in 89 Cx. pipiens pools and in 2 Aedes albopictus pools. Two birds were WNV-positive and 12 were USUV-positive. Furthermore, 30 human cases of acute meningoencephalitis, possibly caused by WNV or USUV, were evaluated for both viruses and 1,053 blood bags were tested for WNV, without any positive result., Conclusions/significance: Despite not finding symptomatic human WNV infections during 2010, the persistence of the virus, probably due to overwintering, was confirmed through viral circulation in mosquitoes and birds, as well as for USUV. In 2010, circulation of the two viruses was lower and more delayed than in 2009, but this decrease was not explained by the relative abundance of Cx. pipiens mosquito, which was greater in 2010. The USUV detection in mosquito species confirms the role of Cx. pipiens as the main vector and the possible involvement of Ae. albopictus in the virus cycle. The effects of meteorological conditions on the presence of USUV-positive mosquito pools were considered finding an association with drought conditions and a wide temperature range. The output produced by the surveillance system demonstrated its usefulness and reliability in terms of planning public health policies.

We investigated the impact of CHIKV strains on some Aedes albopictus (Skuse) reproductive parameters and the possibility of vertical transmission. Two strains were collected in the area where the epidemic occurred in 2007, one isolated from mosquitoes, the other one isolated from a viraemic patient. Different types of blood meals, either infected or non-infected, were offered to Ae. albopictus females, that were then analyzed at increasing time post infection. The virus titre, measured by two RT-PCR methods in the blood meals, influenced the rate of infection and the rate of dissemination of CHIKV in Ae. albopictus body. We found individual variability with respect to the infection/dissemination rates and their latency both considering the female's body and appendages. The hatching rate was significantly lower for the eggs laid by the infected females than for the control eggs, while the mortality during the larval development (from first instar larva to adult emergence) was similar among the progeny of infected and non-infected female groups. Our findings seem to support the hypothesis that the vertical transmission is a rare event under our conditions, and that a certain time period is required in order to get the ovarioles infected. Field observations conducted during the Spring 2008 showed no evidence of the presence of infected overwintering progeny produced by Ae. albopictus females infected during the 2007 outbreak.

Usutu virus (USUV) is a mosquito-borne flavivirus belonging to the Japanese encephalitis serocomplex, recently related to neurological disease in immunosuppressed patients. In the same area of Northern Italy where USUV human cases occurred in 2009, a regional West Nile virus (WNV) surveillance program based on mosquito monitoring and wild birds screening has been implemented since 2008. Mosquito pools and wild birds were tested using three different polymerase chain reactions (Flavivirus, WNV, and USUV). During summer 2009, 56 pools (54 consisting of Culex pipiens and 2 of Aedes albopictus) and 27 pools (Cx. pipiens) out of 1789 mosquito pools were, respectively, USUV and WNV positive. Moreover, out of 1218 wild birds tested, 44 were WNV positive, whereas only 11 birds were USUV positive by polymerase chain reaction. Data collected during 2009 prove a cocirculation of USUV and WNV in Northern Italy, but these two viruses show different incidence values in both mosquitoes and birds, suggesting involvement of different animals (other bird species or mammals) in their natural cycles. The cocirculation of WNV and USUV poses a new potential threat to human health in this area. The extent of WNV surveillance to other Flaviviruses will require new diagnostic procedures able to process a large number of samples in a limited period of time and highlights the importance of developing more specific serological tests that could be used in field.

Background: In recent years human diseases due to mosquito-borne viruses were increasingly reported in Emilia-Romagna region (Italy), from the chikungunya virus in 2007 to the West Nile virus (WNV) in 2008. An extensive entomological survey was performed in 2009 to establish the presence and distribution of mosquito arboviruses in this region, with particular reference to flaviviruses., Methodology/principal Findings: From May 6 to October 31, a total of 190,516 mosquitoes were sampled in georeferenced stations, grouped in 1,789 pools according date of collection, location, and species, and analyzed by reverse transcription polymerase chain reaction (RT-PCR) to detect the presence of RNA belong to Flavivirus genus. WNV was detected in 27 mosquito pools, producing sequences similar to those of birds and human strains obtained in 2008 outbreak, pointed out the probable virus overwintering. Isolation of WNV was achieved from one of these pools. Moreover 56 pools of mosquitoes tested positive for Usutu virus (USUV). Most PCR positive pools consisted of Culex pipiens, which also was the most analyzed mosquito species (81.4% of specimens); interestingly, USUV RNA was also found in two Aedes albopictus mosquito pools. Simultaneous circulation of WNV and USUV in the survey area was highlighted by occurrence of 8 mosquito WNV- and USUV-positive pools and by the overlaying of the viruses "hot spots", obtained by kernel density estimation (KDE) analysis. Land use of sampled stations pointed out a higher proportion of WNV-positive Cx. pipiens pool in rural environments respect the provenience of total sampled pool, while the USUV-positive pools were uniformly captured in the different environments., Conclusions/significance: Obtained data highlighting the possible role of Cx. pipiens mosquito as the main vector for WNV and USUV in Northern Italy, and the possible involvement of Ae. albopictus mosquito in USUV cycle. The described mosquito-based surveillance could constitute the foundation for a public health alert system targeting mosquito borne arboviruses.

Recently, Italy-particularly the Emilia-Romagna region-was the location of consecutive outbreaks of human diseases caused by the arboviruses chikungunya virus and West Nile virus. The two outbreaks, spread by different species of mosquitoes, were not related, but pointed out the lack of an arboviral surveillance program in this region. Beginning in 2007 entomological surveillance was initiated in the Emilia-Romagna region, and in 2008 the program was improved and extended at Lombardia region. Using CO(2)-baited traps, 65,292 mosquitoes were collected; pooled by date of collection, location, and species; macerated manually; and tested by reverse transcription (RT)-polymerase chain reaction for the presence of alphaviruses, orthobunyaviruses, and flaviviruses. Amplicons were sequenced and employed for identification of viral RNA by basic local alignment search tool search in GenBank. Results of these assays showed (1) the presence of West Nile virus in two pools of Culex pipiens mosquitoes, (2) the presence of RNA of two orthobunyaviruses, Tahyna virus in a pool of Ochlerotatus caspius mosquitoes and Batai virus in a pool of Anopheles maculipennis mosquitoes, and (3) the presence of flavivirus RNAs in pools of Oc. caspius, Aedes albopictus, and Aedes vexans mosquitoes; the sequences of these amplicons were most closely related to flaviviruses that have been detected only in mosquitoes and had no recognized vertebrate host (Aedes flavivirus, Culex flavivirus, and Kamiti River virus).

Spongy or noncompacted myocardium is a rare congenital cardiomyopathy characterized by parietal myocardial structural alterations secondary to intrauterine arrest of myocardial fiber compaction. It can be associated with other congenital cardiovascular malformations or manifest as an isolated disease. It is characterized by a wide variety of clinical presentations that range from complete absence of symptoms to thromboembolic manifestations, ventricular arrhythmias and congestive heart failure; the short- and mid-term prognosis is quite severe. We here describe a casual finding of an advanced form of noncompacted myocardium associated with patent ductus arteriosus in a young asymptomatic athlete during a routine visit for agonistic sports eligibility. Although the patient had a normal functional capacity, transthoracic echocardiography showed left ventricular dilation with a moderately depressed contractile function and the presence of numerous prominent trabeculae and deep intertrabecular recesses which communicated with the left ventricular cavity. These findings which were confirmed by further echocardiographic examinations, and the absence of other cardiovascular disorders associated with this disease, led us to the diagnosis of noncompacted myocardium. The patient was not allowed to perform sports activities and underwent percutaneous closure of the patient ductus arteriosus. He was asymptomatic and because in the recent literature there are no data regarding pharmacological therapy in these patients, the subject was submitted to routine clinical follow-up evaluation with no medical therapy.

Aims and Background: General practitioners could play a key role in preventive programs against tobacco-related diseases. However, they seldom take action in the office even with minimal advice counselling. Such behaviour might reflect the lack of academic teaching and the lack of practice with motivational and dependence questionnaires, considered basic tools to help smokers to quit successfully. The study was aimed to investigate the awareness of a sample of Italian family doctors as regards tobacco epidemiology and smoking cessation strategies., Methods: A total of 428 family doctors were administered a questionnaire with a set of questions on their personal smoking habits and on personal initiatives in the office towards smokers. Another set of questions regarded their knowledge on tobacco issues, with special attention to carbon monoxide, which is widely perceived as a very dangerous poison and works as a motivational tool on smokers and adolescents. Carbon monoxide measurement was carried out on all participants to obtain objective data on smoking and to show the feasibility of the test., Results: The percentage of self-reported current smokers among general practitioners was 24%, with a high prevalence of ex-smokers (46%), and 29% of never smokers. Family doctors were more keen to counsel adolescents than adults about tobacco, and they were very interested in continuing medical education on the issue. The doctors who took part in our study showed a surprising limited knowledge of all the issues associated with smoking cessation and prevention such as epidemiology, cigarette characteristics, success rate of smoking cessation programs, Fagerström's tolerance questionnaire, safety of nicotine replacement therapy and the knowledge of carbon monoxide as a product of cigarette smoke., Conclusions: The scenario depicted by our survey underscores the necessity to improve the knowledge and performance of primary care physicians on tobacco-related issues in order to implement primary and secondary prevention in clinical practice.

Several studies on spontaneously hypertensive rats (SHR) have demonstrated increased activity of the sympathetic nervous system (SNS). Using microdialysis, we have observed a greater release of norepinephrine (NE) into the interstitia of striated muscle, than that observed in control Wistar-Kyoto (WKY) rats in the prehypertensive phase. We confirmed these results in the subcutaneous adipose tissue where the sympathetic output controls metabolism. This study was carried out in order to evaluate SNS activity in two district tissue types conducted during both the prehypertensive phase (4-5 weeks of age) and the established hypertensive phase (15-16 weeks of age). Interstitial concentrations of NE were measured by microdialysis in striated muscle and subcutaneous adipose tissue. Two groups of rats were studied. Each group was made up of 8 subjects, SHR and WKY, males of 4-5 weeks of age with a mean body weight of 80 and 75 g respectively. Arterial systolic pressure (tail-cuff) values were 106 mmHg (standard deviation +/-8.2) in SHR and 101 mmHg (standard deviation +/-6.9) in WKY rats (NS). Two microdialysis probes were positioned in the subcutaneous fatty tissue and in the striated muscle of the parascapular region and perfused with Ringers' solution. The dialysate was collected every 30 min for 150 min and analyzed in high-performance liquid chromatography-every day. The content of NE and other catecholamines was determined. The same animals in both groups were reevaluated at 15-16 weeks of age. The mean body weight at this time was 246 g for the SHR and 289 g for the WKY rats. Arterial systolic pressure was 161 mmHg (standard deviation +/-13.3) and 108 mmHg (standard deviation +/-15.6) respectively (p < 0.01, Student's t test). Interstitial levels of NE were higher in SHR than in WKY rats in both tissues examined in the prehypertensive phase and in the established hypertensive phase. Mean NE values from subcutaneous adipose tissue in 4-5 week-old SHR were 1362.1 +/- 181.3 pg/ml compared to 479.0 +/- 162.3 pg/ml in WKY rats (p < 0.001, Student's t test). Muscle tissue NE levels in SHR were 1292.7 +/- 319.1 vs 536.3 +/- 146.7 pg/ml in WKY rats (p < 0.001, Student's t test). Values from the same rats at 15-16 weeks of age were 1405.0 +/- 148.3 pg/ml in SHR compared to 501.6 +/- 131.2 pg/ml in fatty tissue from WKY rats and 1893.7 +/- 214.6 vs 502.0 +/- 118.8 pg/ml in muscle tissue from the respective groups (p < 0.001, Student's t test). Significant differences (p < 0.01, Student's t test) were also observed in mean NE values in striated muscle tissue during the developing phase of hypertension. These findings document SNS hyperactivity in SHR when compared to WKY normotensive controls. This increase in SNS activity was observed in both the prehypertensive phase and in the established hypertensive phase indicating a complete disassociation from regional components of regulation (baroreceptor control and metabolic control), at least in the prehypertensive phase. These results may suggest as alteration in primitive sympathetic central outflow. Higher interstitial NE concentrations in the muscle tissue from SHR during the hypertensive phase compared to levels of young animals that are still normotensive, reveal an interesting pathophysiological aspect for the development of arterial hypertension.

We report a case of a young burkinabian, who presented with the disseminated african form of histoplasmosis, Histoplasma duboisii. There was pulmonary mediastinal disease with superficial lymphadenopathy and also hepatic involvement. The diagnosis was achieved by a histological examination of the peripheral glands and micrological examination of the expectorate. A clinical cure (despite the radiological persistence of enlarged but stable left hilar glands) with the follow up 10 months was achieved after 15 months of treatment with ketoconazole in a dose which was increased to 10 mg/kg/day.

The pharmacokinetic behaviour and metabolism of ursodeoxycholic acid (UDCA) have been studied in the rat. After oral administration of both 3H-labelled (4 muCi/kg body wt) and unlabelled (20 mg) UDCA, UDCA appeared in serum almost entirely in conjugated form (taurine conjugated); UDCA was present in bile mostly as taurine conjugated; the more relevant metabolite is 3 alpha,6 alpha, 7 beta-trihydroxycholanoic acid which represents 10% of the total bile acid pool. UDCA increased bile flow and selectively decreased biliary cholesterol secretion, while phospholipid secretion was unaffected. Faecal UDCA excretion was 15-20% while the urinary extraction was 1.5% during 24 h. The data show that UDCA, when administered in high dose, is promptly secreted into bile almost entirely metabolized to tauroursodeoxycholic acid, where it (1) desaturates the cholesterol in bile, (2) exerts choleretic properties.

Report of 27 cases and comments on their epidemiological, clinical, immunological features and on the question of their treatment and prophylaxis.

Staphyococcal septicemia is rare (0.19 p. 100 of admittances). The origin of this kind of infection could be located in all cases out of the sanitary units, and in 12 out of the 14 cases, patients under 30 years old were concerned. The main disorders were in pulmonary tract, kidneys and heart. A fatal issue occured 6 times due in most cases to a delayed admittance. A good study of sensitivity to antibiotics and a treatment associating several antibiotics are necessary.