Isolation of Batborne Neglected Zoonotic Agent Issyk-Kul Virus, Italy.

We isolated Issyk-Kul virus (ISKV) from a bat sampled from Italy in 2021 and conducted ISKV-specific surveillance in bats collected in Italy during 2017-2023. ISKV circulation among synanthropic and sedentary species of bat, such as Savi's pipistrelle bat (Hypsugo savii) in northern Italy, may have public health implications in this region.

Organic Electrochemical Transistors as Versatile Tool for Real-Time and Automatized Viral Cytopathic Effect Evaluation.

In-vitro viral studies are still fundamental for biomedical research since studying the virus kinetics on cells is crucial for the determination of the biological properties of viruses and for screening the inhibitors of infections. Moreover, testing potential viral contaminants is often mandatory for safety evaluation. Nowadays, viral cytopathic effects are mainly evaluated through end-point assays requiring dye-staining combined with optical evaluation. Recently, optical-based automatized equipment has been marketed, aimed at the real-time screening of cell-layer status and obtaining further insights, which are unavailable with end-point assays. However, these technologies present two huge limitations, namely, high costs and the possibility to study only cytopathic viruses, whose effects lead to plaque formation and layer disruption. Here, we employed poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate) (Pedot:Pss) organic electrochemical transistors (OECTs) for the real-time, electrical monitoring of the infection of cytolytic viruses, i.e., encephalomyocarditis virus (EMCV), and non-cytolytic viruses, i.e., bovine coronavirus (B-CoV), on cells. OECT data on EMCV were validated using a commercially-available optical-based technology, which, however, failed in the B-CoV titration analysis, as expected. The OECTs proved to be reliable, fast, and versatile devices for viral infection monitoring, which could be scaled up at low cost, reducing the operator workload and speeding up in-vitro assays in the biomedical research field.

Assessment of the Costs Related to West Nile Virus Monitoring in Lombardy Region (Italy) between 2014 and 2018.

In Italy, the West Nile Virus surveillance plan considers a multidisciplinary approach to identify the presence of the virus in the environment (entomological, ornithological, and equine surveillance) and to determine the risk of infections through potentially infected donors (blood and organ donors). The costs associated with the surveillance program for the Lombardy Region between 2014 and 2018 were estimated. The costs of the program were compared with a scenario in which the program was not implemented, requiring individual blood donation nucleic acid amplification tests (NAT) to detect the presence of WNV in human samples throughout the seasonal period of vector presence. Considering the five-year period, the application of the environmental/veterinary surveillance program allowed a reduction in costs incurred in the Lombardy Region of 7.7 million EUR. An integrated surveillance system, including birds, mosquito vectors, and dead-end hosts such as horses and humans, can prevent viral transmission to the human population, as well as anticipate the detection of WNV using NAT in blood and organ donors. The surveillance program within a One Health context has given the possibility to both document the expansion of the endemic area of WNV in northern Italy and avoid most of the NAT-related costs.

Hypsugopoxvirus: A Novel Poxvirus Isolated from Hypsugo savii in Italy.

Interest in bat-related viruses has increased considerably during the last decade, leading to the discovery of a rising number of new viruses in several bat species. Poxviridae are a large, diverse family of DNA viruses that can infect a wide range of vertebrates and invertebrates. To date, only a few documented detections of poxviruses have been described in bat populations on three different continents (America, Africa, and Australia). These viruses are phylogenetically dissimilar and have diverse clinical impacts on their hosts. Herein, we report the isolation, nearly complete genome sequencing, and annotation of a novel poxvirus detected from an insectivorous bat (Hypsugo savii) in Northern Italy. The virus is tentatively named Hypsugopoxvirus (HYPV) after the bat species from which it was isolated. The nearly complete genome size is 166,600 nt and it encodes 161 genes. Genome analyses suggest that HYPV belongs to the Chordopoxvirinae subfamily, with the highest nucleotide identity (85%) to Eptesipoxvirus (EPTV) detected from a microbat Eptesicus fuscus in WA, USA, in 2011. To date, HYPV represents the first poxvirus detected in bats in Europe; thus, its viral ecology and disease associations should be investigated further.

Full genome characterization of two novel Alpha-coronavirus species from Italian bats.

Coronaviruses (CoVs) have been detected worldwide in several bat species, which are considered the main reservoir. The attention to the high diversity of CoVs hosted by bats has increased during the last decade due to the high number of human infections caused by two zoonotic Beta-CoVs, SARS-CoV and MERS-CoV, that cause several respiratory diseases. Among coronaviruses, two Alpha-CoV strains (HuCoV-229E and HuCoV-NL63) cause mild respiratory disease that can change to severe disease in children, elderly and individuals affected by illnesses. Phylogenetic analysis conducted on bat Alpha-CoV strains revealed their evolutive correlation to human strains, suggesting their origin in bats. The genome of CoVs is characterized by a high frequency of mutations and recombination events, increasing their ability to switch hosts and their zoonotic potential. In this study, three strains of Alpha-CoV genera detected in Italian bats (Pipistrellus kuhlii) were fully sequenced by Next Generation Sequencing (NGS) and characterized. The complete genome analysis showed the correlation of the Italians strains with a Chinese strain detected in 2013 and, based on CoV molecular species demarcation, two new Alpha-CoV species were established. The analysis of a fragment of the RNA-dependent RNA polymerase (RdRp) showed the correlation of the Italian strains with CoVs that was only detected in the bat Pipistrellus genera (Pipistrellus kuhlii and Pipistrellus Pipistrellus) in European countries.

GI-16 lineage (624/I or Q1), there and back again: The history of one of the major threats for poultry farming of our era.

The genetic variability of Infectious bronchitis virus (IBV) is one of the main challenges for its control, hindering not only the development of effective vaccination strategies but also its classification and, consequently, epidemiology understanding. The 624/I and Q1 genotypes, now recognized to be part of the GI-16 lineage, represent an excellent example of the practical consequences of IBV molecular epidemiology limited knowledge. In fact, being their common origin unrecognized for a long time, independent epidemiological pictures were drawn for the two genotypes. To fix this misinterpretation, the present study reconstructs the history, population dynamics and spreading patterns of GI-16 lineage as a whole using a phylodynamic approach. A collection of worldwide available hypervariable region 1 and 2 (HVR12) and 3 (HVR3) sequences of the S1 protein was analysed together with 258 HVR3 sequences obtained from samples collected in Italy (the country where this genotype was initially identified) since 1963. The results demonstrate that after its emergence at the beginning of the XX century, GI-16 was able to persist until present days in Italy. Approximately in the late 1980s, it migrated to Asia, which became the main nucleus for further spreading to Middle East, Europe and especially South America, likely through multiple introduction events. A remarkable among-country diffusion was also demonstrated in Asia and South America. Interestingly, although most of the recent Italian GI-16 strains originated from ancestral viruses detected in the same country, a couple were closely related to Chinese ones, supporting a backward viral flow from China to Italy. Besides to the specific case-study results, this work highlights the misconceptions that originate from the lack of a unified nomenclature and poor molecular epidemiology data generation and sharing. This shortcoming appears particularly relevant since the described scenario could likely be shared by many other IBV genotypes and pathogens in general.

Surveillance of Mosquitoes and Selected Arthropod-Borne Viruses in the Context of Milan EXPO 2015.

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.

Detection and Characterization of a Novel Reassortant Mammalian Orthoreovirus in Bats in Europe.

A renewed interest in mammalian orthoreoviruses (MRVs) has emerged since new viruses related to bat MRV type 3, detected in Europe, were identified in humans and pigs with gastroenteritis. This study reports the isolation and characterization of a novel reassortant MRV from the lesser horseshoe bat (Rhinolophus hipposideros). The isolate, here designated BatMRV1-IT2011, was first identified by electron microscopy and confirmed using PCR and virus-neutralization tests. The full genome sequence was obtained by next-generation sequencing. Molecular and antigenic characterizations revealed that BatMRV1-IT2011 belonged to serotype 1, which had not previously been identified in bats. Phylogenetic and recombination detection program analyses suggested that BatMRV1-IT2011 was a reassortant strain containing an S1 genome segment similar to those of MRV T1/bovine/Maryland/Clone23/59 and C/bovine/ Indiana/MRV00304/2014, while other segments were more similar to MRVs of different hosts, origins and serotypes. The presence of neutralizing antibodies against MRVs has also been investigated in animals (dogs, pigs, bovines and horses). Preliminary results suggested that MRVs are widespread in animals and that infections containing multiple serotypes, including MRVs of serotype 1 with an S1 gene similar to BatMRV1-IT2011, are common. This paper extends the current knowledge of MRVs and stresses the importance to continue and improve MRV surveillance in bats and other mammals through the development and standardization of specific diagnostic tools.