Publications by authors named "Francesca Rovida"

54 Publications

Identification of SARS-CoV-2 RNA in the conjunctival swab of an Italian pediatric patient affected with COVID-19: A case report.

Eur J Ophthalmol 2020 Dec 2:1120672120977822. Epub 2020 Dec 2.

Department of Surgical and Clinical, Diagnostic and Pediatric Sciences, University of Pavia, IRCCS Fondazione Policlinico San Matteo, Pavia, Italy.

Introduction: To report a case of identification of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) RNA in ocular specimen in a pediatric patient affected with Coronavirus disease 2019 (COVID-19) with no signs of ocular involvement.

Case Description: A 11-year old male patient with confirmed COVID-19 infection was hospitalized at the Pediatric Clinic Clinic of the IRCCS Foundation and Hospital San Matteo, Pavia, Italy. Three days after hospital admission, because of the patient complaining very mild ocular symptoms, an ophthalmological evaluation was performed. No signs related to conjunctivitis or keratitis were found but a conjunctival swab was collected as well, based on patient's medical history. The specific SARS-CoV-2 reverse transcription PCR (RT-PCR) was performed, unearthing the presence of viral RNA from the swab. On day 25 from hospitalization, the conjunctival swab was repeated, giving negative result.

Conclusions: This is the first report of the identification of SARS-CoV-2 RNA in ocular specimen in a pediatric patient without signs of ocular involvement. However, despite the transmission through tears is theoretically possible, it is still unclear whether this could be considered as an important route for the spread of SARS-CoV-2.
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http://dx.doi.org/10.1177/1120672120977822DOI Listing
December 2020

The experience of the health care workers of a severely hit SARS-CoV-2 referral Hospital in Italy: incidence, clinical course and modifiable risk factors for COVID-19 infection.

J Public Health (Oxf) 2020 Nov 3. Epub 2020 Nov 3.

Medical Direction, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

Background: During the COVID-19 pandemic, the health care workers (HCWs) at the frontline have been largely exposed to infected patients, running a high risk of being infected by the SARS-CoV-2 virus.Since limiting transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in health care setting is crucial to avoid the community spread of SARS-CoV-2, we want to share our experience as an early hit hospital where standard infection control practices have been conscientiously applied and effective. We believe that our example, as first and hardest hit country, might be a warning and aid not only for those who have been hit later, but also for a second fearful wave of contagion. In addition, we want to offer an insight on modifiable risk factors for HWs-related infection.

Methods: Demographic, lifestyle, work-related and comorbidities data of 1447 HCWs, which underwent a nasopharyngeal swab for SARS-CoV-2, were retrospectively collected. For the 164 HCWs positive for SARS-CoV-2, data about safety in the workplace, symptoms and clinical course of COVID-19 were also collected. Cumulative incidence of SARS-CoV-2 infection was estimated. Risk factors for SARS-CoV-2 infection were assessed using a multivariable Poisson regression.

Results: The cumulative incidence of SARS-CoV-2 infection among the screened HCWs was 11.33% (9.72-13.21). Working in a COVID-19 ward, being a former smoker (versus being a person who never smoked) and BMI was positively associated with SARS-CoV-2 infection, whereas being a current smoker was negatively associated with this variable.

Conclusions: Assuming an equal accessibility and proper use of personal protective equipment of all the HCWs of our Hospital, the great and more prolonged contact with COVID-19 patients remains the crucial risk factor for SARS-CoV-2. Therefore, increased and particular care needs to be focused specifically on the most exposed HCWs groups, which should be safeguarded. Furthermore, in order to limit the risk of asymptomatic spread of SARS-CoV-2 infection, the HCWs mild symptoms of COVID-19 should be considered when evaluating the potential benefits of universal staff testing.
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http://dx.doi.org/10.1093/pubmed/fdaa195DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7665642PMC
November 2020

Residual SARS-CoV-2 RNA in nasal swabs of convalescent COVID-19 patients: Is prolonged quarantine always justified?

Int J Infect Dis 2021 Jan 30;102:299-302. Epub 2020 Oct 30.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy; Department of Clinical, Surgical, Diagnostic and Pediatric Sciences, University of Pavia, Pavia, Italy. Electronic address:

Real-time reverse transcription PCR is currently the most sensitive method to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Defining whether a patient could be contagious or not contagious in the presence of residual SARS-CoV-2 RNA is of extreme importance in the context of public health. In this prospective multicenter study, virus isolation was prospectively attempted in 387 nasal swabs from clinically recovered patients showing low viral load (quantification cycle, Cq, value greater than 30). The median Cq value was 36.8 (range 30.0-39.4). Overall, a cytopathic effect was detected in nine samples, corresponding to a culture positivity rate of 2.3% (9/387). The results of this study help to dissect true virus replication and residual viral RNA detection in recovered patients.
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http://dx.doi.org/10.1016/j.ijid.2020.10.072DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7834568PMC
January 2021

A case of mumps encephalitis imported to Italy from India.

J Med Virol 2020 Jul 7. Epub 2020 Jul 7.

Italian National Reference Laboratory for Measles and Rubella, Istituto Superiore di Sanità, Rome, Italy.

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http://dx.doi.org/10.1002/jmv.26263DOI Listing
July 2020

Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infection in Children and Adolescents: A Systematic Review.

JAMA Pediatr 2020 Sep;174(9):882-889

Pediatric Clinic, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

Importance: The current rapid worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection justifies the global effort to identify effective preventive strategies and optimal medical management. While data are available for adult patients with coronavirus disease 2019 (COVID-19), limited reports have analyzed pediatric patients infected with SARS-CoV-2.

Objective: To evaluate currently reported pediatric cases of SARS-CoV-2 infection.

Evidence Review: An extensive search strategy was designed to retrieve all articles published from December 1, 2019, to March 3, 2020, by combining the terms coronavirus and coronavirus infection in several electronic databases (PubMed, Cochrane Library, and CINAHL), and following the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines. Retrospective cross-sectional and case-control studies, case series and case reports, bulletins, and national reports about the pediatric SARS-CoV-2 infection were included. The risk of bias for eligible observational studies was assessed according to the Strengthening the Reporting of Observational Studies in Epidemiology reporting guideline.

Findings: A total of 815 articles were identified. Eighteen studies with 1065 participants (444 patients were younger than 10 years, and 553 were aged 10 to 19 years) with confirmed SARS-CoV-2 infection were included in the final analysis. All articles reflected research performed in China, except for 1 clinical case in Singapore. Children at any age were mostly reported to have mild respiratory symptoms, namely fever, dry cough, and fatigue, or were asymptomatic. Bronchial thickening and ground-glass opacities were the main radiologic features, and these findings were also reported in asymptomatic patients. Among the included articles, there was only 1 case of severe COVID-19 infection, which occurred in a 13-month-old infant. No deaths were reported in children aged 0 to 9 years. Available data about therapies were limited.

Conclusions And Relevance: To our knowledge, this is the first systematic review that assesses and summarizes clinical features and management of children with SARS-CoV-2 infection. The rapid spread of COVID-19 across the globe and the lack of European and US data on pediatric patients require further epidemiologic and clinical studies to identify possible preventive and therapeutic strategies.
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http://dx.doi.org/10.1001/jamapediatrics.2020.1467DOI Listing
September 2020

West Nile or Usutu Virus? A Three-Year Follow-Up of Humoral and Cellular Response in a Group of Asymptomatic Blood Donors.

Viruses 2020 01 29;12(2). Epub 2020 Jan 29.

Molecular Virology Unit, Microbiology and Virology Department, IRCCS Policlinico San Matteo, Via Taramelli 5, 27100 Pavia, Italy.

West Nile virus (WNV) and Usutu virus (USUV) are two related arboviruses (genus , family ), with birds as a reservoir and mosquitoes as transmitting vectors. In recent years, WNV epidemiology changed in many European countries with increased frequency of outbreaks posing the issue of virus transmission risks by blood transfusion. USUV emerged for the first time in birds of the Tuscany region (Italy) in 1996 and in 2001 in Austria. While WNV is responsible for both mild and neuroinvasive diseases, USUV infection is usually asymptomatic and neuroinvasive symptoms are rare. Since WNV and USUV co-circulate, the surveillance of WNV allows also the detection of USUV. Due to the great similarity in amino-acid sequence of major surface proteins of the two viruses, a high cross-reactivity can lead to misinterpretation of serological results. Here, we report the results obtained from 54 asymptomatic blood donors during a three-year follow-up showing an unexpected high positivity (46.3%) for USUV. The major obstacle encountered in the differential diagnosis between these two viruses was the high cross-reactivity found in neutralizing antibodies (NT Abs) and, in some cases, a long follow-up was mandatory for a correct diagnosis. Moreover, two new ELISpot assays were developed for a more rapid and specific differential diagnosis, especially in those cases in which NT Abs were not determinant. Using a combination of Enzyme-linked immunospot (ELISpot), molecular, and serological tests, we could identify 25 true positive WNV and 25 true positive USUV blood donors. Our data highlight the importance of raising awareness for increasing USUV infections in endemic countries involved in blood transfusion and organ donation.
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http://dx.doi.org/10.3390/v12020157DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7077259PMC
January 2020

Zika Virus Infection in Pregnancy: Advanced Diagnostic Approaches in Dengue-Naive and Dengue-Experienced Pregnant Women and Possible Implication for Cross-Reactivity and Cross-Protection.

Microorganisms 2019 Dec 28;8(1). Epub 2019 Dec 28.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100 Pavia, Italy.

Zika virus (ZIKV) infection has been linked to congenital defects in fetuses and infants, as exemplified by the microcephaly epidemic in Brazil. Given the overlapping presence of Dengue virus (DENV) in the majority of ZIKV epidemic regions, advanced diagnostic approaches need to be evaluated to establish the role of pre-existing DENV immunity in ZIKV infection. From 2015 to 2017, five pregnant women with suspected ZIKV infection were investigated in Pavia, Italy. Among the five pregnant women, three were DENV-ZIKV immunologically cross-reactive, and two were DENV-naïve. Advanced diagnosis included the following: (i) NS1 blockade-of-binding (BOB) ELISA assay for ZIKV specific antibodies and (ii) ELISpot assay for the quantification of effector memory T cells for DENV and ZIKV. These novel assays allowed to distinguish between related flavivirus infections. The three DENV-experienced mothers did not transmit ZIKV to the fetus, while the two DENV-naive mothers transmitted ZIKV to the fetus. Pre-existing immunity in DENV experienced mothers might play a role in cross-protection
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http://dx.doi.org/10.3390/microorganisms8010056DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7023361PMC
December 2019

Rotavirus molecular epidemiology in hospitalized patients, Northern Italy, 2015-2018.

New Microbiol 2020 Jan 9;43(1):1-5. Epub 2019 Dec 9.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100, Pavia, Italy.

About 15,000 hospitalizations due to group A Rotavirus gastroenteritis (RVA) are recorded each year in Italy. In the present study, we report the seasonal distribution and molecular characterization of RVA in pediatric and adult hospitalized patients in the period September 2015-April 2018 in Pavia province, Lombardy Region. During the study period, stool samples of 1450 patients with acute gastroenteritis were analyzed and 122 were RVA positive, the majority belonging to pediatric patients (94.0%) while only a minority of patients (6.0%) were adults. G3P[8], G1P[8], G9P[8] and G2P[4] were the most detected RVA strains, with a prevalence of 82.4%. However, a variety of RVA strains circulated in Northern Italy in hospitalized patients over a period of three years, emphasizing distinct patterns of distribution in different age groups and between years.
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January 2020

Evolutionary Dynamics of the Lineage 2 West Nile Virus That Caused the Largest European Epidemic: Italy 2011-2018.

Viruses 2019 09 3;11(9). Epub 2019 Sep 3.

Department of Biomedical and Clinical Sciences "L.Sacco", University of Milan, 20157 Milano, Italy.

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.
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http://dx.doi.org/10.3390/v11090814DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6784286PMC
September 2019

Molecular Characterization of Influenza Strains in Patients Admitted to Intensive Care Units during the 2017-2018 Season.

Int J Mol Sci 2019 May 30;20(11). Epub 2019 May 30.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100 Pavia, Italy.

This study aimed at assessing the frequency and the distribution of influenza virus types/subtypes in 172 laboratory-confirmed influenza-positive patients admitted to intensive care units (ICUs) during the 2017-2018 season in the Lombardy region (Northern Italy), and to investigate the presence of molecular pathogenicity markers. A total of 102/172 (59.3%) patients had influenza A infections (83 A/H1N1pdm09, 2 H3N2 and 17 were untyped), while the remaining 70/172 (40.7%) patients had influenza B infections. The 222G/N mutation in the hemagglutinin gene was identified in 33.3% (3/9) of A/H1N1pdm09 strains detected in the lower respiratory tract (LRT) samples and was also associated with more severe infections, whereas no peculiar mutations were observed for influenza B strains. A single-point evolution was observed in site 222 of A/H1N1pdm09 viruses, which might advantage viral evolution by favouring virus binding and replication in the lungs. Data from 17 paired upper respiratory tract (URT) and LRT samples showed that viral load in LRT samples was mostly higher than that detected in URT samples. Of note, influenza viruses were undetectable in 35% of paired URT samples. In conclusion, LRT samples appear to provide more accurate clinical information than URT samples, thus ensuring correct diagnosis and appropriate treatment of patients with severe respiratory infections requiring ICU admission.
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http://dx.doi.org/10.3390/ijms20112664DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6600310PMC
May 2019

Time-scaled phylogeography of complete Zika virus genomes using discrete and continuous space diffusion models.

Infect Genet Evol 2019 09 9;73:33-43. Epub 2019 Apr 9.

Department of Biomedical and Clinical Sciences "L. Sacco", University of Milan, Milano, Italy; CRC-Coordinated Research Center "EpiSoMI", University of Milan, Milano, Italy. Electronic address:

Zika virus (ZIKV), a vector-borne infectious agent that has recently been associated with neurological diseases and congenital microcephaly, was first reported in the Western hemisphere in early 2015. A number of authors have reconstructed its epidemiological history using advanced phylogenetic approaches, and the majority of Zika phylogeography studies have used discrete diffusion models. Continuous space diffusion models make it possible to infer the possible origin of the virus in real space by reconstructing its ancestral location on the basis of geographical coordinates deduced from the latitude and longitude of the sampling locations. We analysed all the ZIKV complete genome isolates whose sampling times and localities were available in public databases at the time the study began, using a Bayesian approach for discrete and continuous phylogeographic reconstruction. The discrete phylogeographic analysis suggested that ZIKV emerged to become endemic/epidemic in the first decade of the 1900s in the Ugandan rainforests, and then reached Western Africa and Asia between the 1930s and 1950s. After a long period of about 40 years, it spread to the Pacific islands and reached Brazil from French Polynesia. Continuous phylogeography of the American epidemic showed that the virus entered in north-eastern Brazil in late 2012 and started to spread in early 2013 from two high probability regions: one corresponding to the entire north-east Brazil and the second surrounding the city of Rio de Janeiro, in a mainly northwesterly direction to Central America, the north-western countries of south America and the Caribbean islands. Our data suggest its cryptic circulation in both French Polynesia and Brazil, thus raising questions about the mechanisms underlying its undetected persistence in the absence of a known animal reservoir, and underline the importance of continuous diffusion models in making more reliable phylogeographic reconstructions of emerging viruses.
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http://dx.doi.org/10.1016/j.meegid.2019.04.006DOI Listing
September 2019

Expanding Usutu virus circulation in Italy: detection in the Lazio region, central Italy, 2017 to 2018.

Euro Surveill 2019 Jan;24(3)

Laboratory of Virology, National Institute for Infectious Diseases 'Lazzaro Spallanzani' IRCCS, Rome, Italy.

Blood donation screening for West Nile virus (WNV) was mandatory in the Lazio region in 2017 and 2018 (June-November) according to the national surveillance plan. In these years, all five donations reactive in WNV nucleic acid amplification tests harboured instead Usutu virus (USUV). Clade 'Europe 2' was identified in four blood donations and a 2018 mosquito pool. The cocirculation of WNV and USUV in Lazio warrants increased laboratory support and awareness of possible virus misidentification.
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http://dx.doi.org/10.2807/1560-7917.ES.2019.24.3.1800649DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6344840PMC
January 2019

West-Nile virus encephalitis in an immunocompetent pediatric patient: successful recovery.

Ital J Pediatr 2018 Nov 20;44(1):140. Epub 2018 Nov 20.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100, Pavia, Italy.

Background: West Nile virus (WNV) is a mosquito-borne RNA virus belonging to the Flaviviridae family. Symptomatic infection happens in only about 20% of the cases, while WNV neuroinvasive disease (WNND) is rare and accounts for less than 1%. There is insufficient information about natural history and clinical course in children, because underdiagnosis is common, and reports are scarce. On the other hand, Europe has seen a dramatic increase of WNV infections in the last decades, and the Po valley itself, in Northern Italy, has become an endemic region since 2013.

Case Presentation: We hereby report a case of West-Nile virus neuroinvasive disease in a 12-year-old boy. This is one of the very few cases diagnosed in the Italian pediatric population. The clinical presentation was compatible with acute encephalitis. Diagnosis was made by detection of specific IgM in both serum and cerebrospinal fluid. He finally was discharged with complete recovery, and no neurologic sequelae after a 12-months follow up period.

Conclusions: Given its non-specific clinical presentation, the diffusion of WNV constitutes a crucial and emerging concern. Even though rare, neuroinvasive WNV infection should always be suspected in pediatric patients, living or traveling in endemic areas, presenting with meningitis, encephalitis or acute flaccid paralysis during the WNV transmission season.
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http://dx.doi.org/10.1186/s13052-018-0574-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6245891PMC
November 2018

Cellular DNA quantification in respiratory samples for the normalization of viral load: a real need?

J Clin Virol 2018 10 27;107:6-10. Epub 2018 Jul 27.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy; Department of Clinical-Surgical, Diagnostic and Pediatric Sciences, University of Pavia, Pavia, Italy. Electronic address:

Background: Respiratory tract infections have an enormous social economic impact, with high incidence of hospitalization and high costs. Adequate specimen collection is the first crucial step for the correct diagnosis of viral respiratory infections.

Objectives: The present retrospective study aimed: i) to verify the cell yield obtained from sampling the nasal respiratory tract using mid-turbinate flocked swabs; ii) to evaluate the normalization of viral load, based on cell number; and iii) to compare the kinetics of viral infection obtained with normalized vs non-normalized viral load.

Study Design: The number of cells were quantified by real-time PCR in residual extract of nasal swabs tested for respiratory viruses detection and stored at -80 °C in a universal transport medium (UTM™).

Results: A total of 513 virus-positive and 226 virus-negative samples were analyzed. Overall, a median of 4.42 log β2-microgolubin DNA copy number/ml of UTM (range 1.17-7.26) was detected. A significantly higher number of cells was observed in virus-positive as compared to virus-negative samples (4.75 vs 3.76; p < 0.001). Viral loads expressed as log RNA copies/ml of UTM and log RNA copies/median number of cells were compared in virus-positive samples and a strict correlation (r = 0.89, p < 0.001) and agreement (R2 = 0.82) were observed. In addition, infection kinetics were compared using the two methods with a follow-up series of eight episodes of viral infection and the mean difference was -0.57 log (range -1.99 to 0.40).

Conclusions: The normalization of viral load using cellular load compliments the validation of real-time PCR results in the diagnosis of respiratory viruses but is not strictly needed.
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http://dx.doi.org/10.1016/j.jcv.2018.07.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7173160PMC
October 2018

Reply to Fontaine.

Clin Infect Dis 2018 09;67(7):1144-1145

Molecular Virology Unit, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

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http://dx.doi.org/10.1093/cid/ciy262DOI Listing
September 2018

Virus and Antibody Dynamics in Travelers With Acute Zika Virus Infection.

Clin Infect Dis 2018 04;66(8):1173-1180

Molecular Virology Unit, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

Background: To improve our understanding of the natural history of Zika virus (ZIKV) infection in humans, we described the dynamics of ZIKV RNA shedding in different body fluids and antibody responses in patients with acute infection.

Methods: Twenty-nine adults with travel-associated infection and 1 case of sexual transmission were enrolled and followed up with weekly ZIKV RNA testing in blood, urine, saliva, and semen samples and antibody testing.

Results: ZIKV RNA was detected in plasma, urine, and saliva of 57%, 93.1%, and 69.2% of participants, with estimated median times to clearance of 11.5 days (interquartile range [IQR] 6-24 days), 24 days (IQR, 17-34), and 14 days (IQR, 8-31), respectively. In 2 pregnant women, ZIKV RNA persisted in blood until delivery of apparently healthy infants. ZIKV RNA was detected in semen of 5 of 10 tested men; median time to clearance was 25 days (IQR 14-29), and the longest time of shedding in semen was 370 days. In flavivirus-naive patients, the median times to detection of ZIKV nonstructural protein 1 (NS1)-specific immunoglobulin M (IgM) and immunoglobulin G (IgG) antibodies were estimated as 8 days (IQR, 5-15 days) and 17 days (IQR, 12-26 days), respectively. ZIKV NS1 IgM antibodies were undetectable in patients with previous dengue.

Conclusions: Prolonged viremia and ZIKV RNA shedding in urine, saliva, and semen occur frequently in patients with acute ZIKV infection. At the time of diagnosis, about half of patients are ZIKV IgM negative. ZIKV NS1 IgM antibodies remain undetectable in patients with previous dengue. Estimates of the times to viral clearance and seroconversion are useful to optimize diagnostic algorithms.
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http://dx.doi.org/10.1093/cid/cix967DOI Listing
April 2018

An autochthonous sexually transmitted Zika virus infection in Italy 2016.

New Microbiol 2018 Jan 7;41(1):80-82. Epub 2017 Nov 7.

Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

We describe two cases of Zika virus infection involving an Italian patient returning from the Dominican Republic and his wife, who remained in Italy and had not travelled to Zika virus endemic areas in the previous months. The infection was transmitted through unprotected sexual intercourse after the man's return to Italy.
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January 2018

Molecular epidemiology of influenza B virus among hospitalized pediatric patients in Northern Italy during the 2015-16 season.

PLoS One 2017 19;12(10):e0185893. Epub 2017 Oct 19.

Pediatric Highly Intensive Care Unit, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico and University of Milan, Milano, Italy.

Background: The influenza B viruses belong to two lineages distinguished by their genetic and antigenic characteristics, which are referred to as the Yamagata and Victoria lineages, designated after their original isolates, B/Yamagata/16/88 and B/Victoria/2/87. The primary aim of this study was to evaluate the molecular characteristics of influenza B viruses circulating in a region of Northern Italy, Lombardia, during the influenza season of 2015-2016.

Methods: Influenza B virus was detected using a respiratory virus panel of assays and an influenza B-specific real-time polymerase chain reaction. The complete influenza B hemagglutinin (HA) gene was amplified and sequenced directly from clinical specimens. Phylogenetic analysis was performed using nucleotide sequences.

Results: A total of 71 hospitalized pediatric patients were influenza B positive. Phylogenetic analysis showed that the great majority of influenza B strains (66/71, 93.0%) belonged to the Victoria-lineage and were antigenically like vaccine strain (B/Brisbane/60/2008) included only in the quadrivalent vaccine. In the detected influenza B strains, a series of amino acid changes were observed in the antigenic regions: I117V, V124A, N129D, V146I, N197D, T199A, and A202T. However, only 2 amino acid changes were observed in the HA regions involved in receptor binding or in antibody recognition.

Conclusions: All the influenza B strains identified in this study belonged to the influenza B Victoria lineage not included in the trivalent vaccine commonly used by the general population during the 2015-2016 influenza season in Italy. This indicates that protection against influenza B infection in the vaccinated population was in general very poor during the 2015-2016 influenza season.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0185893PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5648122PMC
November 2017

Usutu Virus Antibodies in Blood Donors and Healthy Forestry Workers in the Lombardy Region, Northern Italy.

Vector Borne Zoonotic Dis 2017 09 31;17(9):658-661. Epub 2017 Jul 31.

1 S.C. Microbiologia e Virologia, Fondazione IRCCS Policlinico San Matteo , Pavia, Italy .

Usutu virus (USUV), a member of the genus Flavivirus, is known to circulate at low prevalence in Northern Italy, and has been reported to cause overt infection. USUV was first reported in Europe in 2001, but a retrospective study showed that it has been present in Italy at least since 1996. Seroprevalence data for USUV antibodies in sera are being collected in different European countries, showing circulation at low prevalence in human populations. Interestingly, two consecutive studies in Northern Italy indicate a possible increase in the presence of the virus, from 0% to 0.23% seroprevalence in blood donors. In this study, antibodies against USUV were measured in 3 consecutive blood samples collected from October 2014 to December 2015 from 33 forestry workers in the Po river valley, while samples from 200 blood donors from the same geographical area were tested in parallel. Neutralizing and IgG antibodies were found in six forestry workers (18.1%) and in two blood donors (1%). Our results indicate that USUV circulation in the examined area, part of a highly populated region in Northern Italy, is higher than expected. Healthy subjects exhibit a higher prevalence than what was found in a previous report in an adjoining region (0.23%), while the population at risk shows a much higher prevalence value (18.1%).
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http://dx.doi.org/10.1089/vbz.2017.2126DOI Listing
September 2017

Antibody-based assay discriminates Zika virus infection from other flaviviruses.

Proc Natl Acad Sci U S A 2017 08 17;114(31):8384-8389. Epub 2017 Jul 17.

Humabs BioMed SA, 6500 Bellinzona, Switzerland;

Zika virus (ZIKV) is a mosquito-borne flavivirus that emerged recently as a global health threat, causing a pandemic in the Americas. ZIKV infection mostly causes mild disease, but is linked to devastating congenital birth defects and Guillain-Barré syndrome in adults. The high level of cross-reactivity among flaviviruses and their cocirculation has complicated serological approaches to differentially detect ZIKV and dengue virus (DENV) infections, accentuating the urgent need for a specific and sensitive serological test. We previously generated a ZIKV nonstructural protein 1 (NS1)-specific human monoclonal antibody, which we used to develop an NS1-based competition ELISA. Well-characterized samples from RT-PCR-confirmed patients with Zika and individuals exposed to other flavivirus infections or vaccination were used in a comprehensive analysis to determine the sensitivity and specificity of the NS1 blockade-of-binding (BOB) assay, which was established in laboratories in five countries (Nicaragua, Brazil, Italy, United Kingdom, and Switzerland). Of 158 sera/plasma from RT-PCR-confirmed ZIKV infections, 145 (91.8%) yielded greater than 50% inhibition. Of 171 patients with primary or secondary DENV infections, 152 (88.9%) scored negative. When the control group was extended to patients infected by other flaviviruses, other viruses, or healthy donors ( = 540), the specificity was 95.9%. We also analyzed longitudinal samples from DENV-immune and DENV-naive ZIKV infections and found inhibition was achieved within 10 d postonset of illness and maintained over time. Thus, the Zika NS1 BOB assay is sensitive, specific, robust, simple, low-cost, and accessible, and can detect recent and past ZIKV infections for surveillance, seroprevalence studies, and intervention trials.
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http://dx.doi.org/10.1073/pnas.1704984114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5547631PMC
August 2017

Reconstructing the recent West Nile virus lineage 2 epidemic in Europe and Italy using discrete and continuous phylogeography.

PLoS One 2017 5;12(7):e0179679. Epub 2017 Jul 5.

Department of Biomedical and Clinical Sciences "L.Sacco", University of Milan, Milano, Italy.

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.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0179679PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5497961PMC
September 2017

Frequency of respiratory virus infections and next-generation analysis of influenza A/H1N1pdm09 dynamics in the lower respiratory tract of patients admitted to the ICU.

PLoS One 2017 7;12(6):e0178926. Epub 2017 Jun 7.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

Recent molecular diagnostic methods have significantly improved the diagnosis of viral pneumonia in intensive care units (ICUs). It has been observed that 222G/N changes in the HA gene of H1N1pdm09 are associated with increased lower respiratory tract (LRT) replication and worse clinical outcome. In the present study, the frequency of respiratory viruses was assessed in respiratory samples from 88 patients admitted to 16 ICUs during the 2014-2015 winter-spring season in Lombardy. Sixty-nine out of 88 (78.4%) patients were positive for a respiratory viral infection at admission. Of these, 57/69 (82.6%) were positive for influenza A (41 A/H1N1pdm09 and 15 A/H3N2), 8/69 (11.6%) for HRV, 2/69 (2.9%) for RSV and 2/69 (2.9%) for influenza B. Phylogenetic analysis of influenza A/H1N1pdm09 strains from 28/41 ICU-patients and 21 patients with mild respiratory syndrome not requiring hospitalization, showed the clear predominance of subgroup 6B strains. The median influenza A load in LRT samples of ICU patients was higher than that observed in the upper respiratory tract (URT) (p<0.05). Overall, a greater number of H1N1pdm09 virus variants were observed using next generation sequencing on partial HA sequences (codons 180-286) in clinical samples from the LRT as compared to URT. In addition, 222G/N/A mutations were observed in 30% of LRT samples from ICU patients. Finally, intra-host evolution analysis showed the presence of different dynamics of viral population in LRT of patients hospitalized in ICU with a severe influenza infection.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0178926PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5462403PMC
September 2017

Frequency of respiratory viruses among patients admitted to 26 Intensive Care Units in seven consecutive winter-spring seasons (2009-2016) in Northern Italy.

J Clin Virol 2017 07 8;92:48-51. Epub 2017 May 8.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy; Department of Clinical, Surgical, Diagnostic and Pediatric Sciences, University of Pavia, Pavia, Italy. Electronic address:

Background: The role of respiratory viruses in the etiology of community-acquired pneumonia (CAP) is still debated. The advent of molecular assays has improved the identification of viruses in patients with CAP and according to published studies, viruses account for 11-55% of adult CAP cases.

Objectives And Study Design: In the present study, the frequency of respiratory viruses was evaluated in respiratory samples collected from 414 patients with CAP admitted to 26 ICUs in the Lombardy Region (10 million inhabitants) during seven winter-spring seasons (2009-2016).

Results: In 226 (54.6%) patients one or more respiratory viruses were identified, while 188 (45.4%) patients were negative. A single virus infection was observed in 214/226 (94.7%) patients; while, in 12/226 (5.3%) at least two respiratory viruses were detected. Influenza A was the most common virus in 140/226 patients (61.9%) followed by rhinoviruses (33/226, 14.6%), respiratory syncytial virus (13/226, 5.8%), influenza B virus (9/226, 4.0%), human coronaviruses (9/226, 4.0%), cytomegalovirus (9/226, 4.0%) and human metapneumovirus (1/226, 0.4%).

Conclusions: Viral infections are present in a consistent proportion of patients admitted to the ICU for CAP. Influenza A and rhinovirus accounted for three-quarters of all CAP in ICU patients. The use of lower respiratory instead of upper respiratory samples might be useful in the diagnosis of viral CAP.
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http://dx.doi.org/10.1016/j.jcv.2017.05.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7172430PMC
July 2017

Zika virus isolation from semen.

New Microbiol 2017 Jul 17;40(3):197-198. Epub 2017 May 17.

Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

Zika virus (ZIKV) can be sexually transmitted and replicative particles were first detected in a semen sample from a patient during the 2013-14 French Polynesia outbreak. Here we describe the virus isolation from semen of a patient returning to Italy from Brazil.
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July 2017

Swine influenza A (H1N1) virus (SIV) infection requiring extracorporeal life support in an immunocompetent adult patient with indirect exposure to pigs, Italy, October 2016.

Euro Surveill 2017 Feb;22(5)

Dipartimento di Scienze Clinico-Chirurgiche, Diagnostiche e Pediatriche, Università degli Studi di Pavia, Pavia, Italy.

We describe a case of severe swine influenza A(H1N1) virus infection in an immunocompetent middle-aged man in October 2016 in Italy who had only indirect exposure to pigs. The patient developed a severe acute distress respiratory syndrome which was successfully supported by extracorporeal membrane oxygenation and treated with antiviral therapy. The sole risk factor for influenza was a body mass index > 30 kg/m. After a month of hospitalisation, the patient was discharged in good health.
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http://dx.doi.org/10.2807/1560-7917.ES.2017.22.5.30456DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5388119PMC
February 2017

Miscarriage following dengue virus 3 infection in the first six weeks of pregnancy of a dengue virus-naive traveller returning from Bali to Italy, April 2016.

Euro Surveill 2016 Aug;21(31)

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy.

We report miscarriage following dengue virus (DENV)-3 infection in a pregnant woman returning from Bali to Italy in April 2016. On her arrival, the woman had fever, rash, asthenia and headache. DENV RNA was detected in plasma and urine samples collected the following day. Six days after symptom onset, she had a miscarriage. DENV RNA was detected in fetal material, but in utero fetal infection cannot be demonstrated due to possible contamination by maternal blood.
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http://dx.doi.org/10.2807/1560-7917.ES.2016.21.31.30308DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4998508PMC
August 2016

Evaluation of Xpert® Norovirus Assay performance in comparison with real-time RT-PCR in hospitalized adult patients with acute gastroenteritis.

Diagn Microbiol Infect Dis 2016 Aug 7;85(4):426-7. Epub 2016 May 7.

Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo, 27100, Pavia, Italy; Department of Clinical, Surgical, Diagnostic and Pediatric Sciences, University of Pavia, 27100, Pavia, Italy. Electronic address:

Xpert® Norovirus Assay (Cepheid, Sunnyvale, CA) was compared with a laboratory-developed real-time RT-PCR assay for the detection of Norovirus GI and GII in hospitalized patients with acute gastroenteritis. The two assays showed a high level of concordance but Xpert® Norovirus Assay allowed faster detection of Norovirus and a simpler sample handling.
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http://dx.doi.org/10.1016/j.diagmicrobio.2016.05.002DOI Listing
August 2016

The novel influenza A virus protein PA-X and its naturally deleted variant show different enzymatic properties in comparison to the viral endonuclease PA.

Nucleic Acids Res 2015 Oct 17;43(19):9405-17. Epub 2015 Sep 17.

Institute of Molecular Genetics IGM-CNR, via Abbiategrasso 207, 27100 Pavia, Italy

The PA protein of Influenza A virus (IAV) encoded by segment 3 acts as a specialized RNA endonuclease in the transcription of the viral genome. The same genomic segment encodes for a second shorter protein, termed PA-X, with the first 191 N-terminal aminoacids (aa) identical to PA, but with a completely different C-ter domain of 61 aa, due to a ribosomal frameshifting. In addition, it has been shown that several IAV isolates encode for a naturally truncated PA-X variant, PAXΔC20, missing the last 20 aa. The biochemical properties of PA-X and PAXΔC20 have been poorly investigated so far. Here, we have carried out an enzymatic characterization of PA-X and its naturally deleted form, in comparison with PA from the human IAV strain A/WSN/33 (H1N1). Our results showed, to the best of our knowledge for the first time, that PA-X possesses an endonucleolytic activity. Both PA and PA-X preferentially cut single stranded RNA regions, but with some differences. In addition, we showed that PAXΔC20 has severely reduced nuclease activity. These results point to a previously undetected role of the last C-ter 20 aa for the catalytic activity of PA-X and support distinct roles for these proteins in the viral life cycle.
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http://dx.doi.org/10.1093/nar/gkv926DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4627086PMC
October 2015

West Nile virus outbreak in the Lombardy region, northern Italy, summer 2013.

Vector Borne Zoonotic Dis 2015 Apr;15(4):278-83

1 Molecular Virology Unit, Microbiology and Virology Department, Fondazione IRCCS Policlinico San Matteo , Pavia, Italy .

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).
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http://dx.doi.org/10.1089/vbz.2014.1711DOI Listing
April 2015