Publications by authors named "Elisa Franchin"

45 Publications

The first familial cluster of the B.1.1.7 variant of SARS-CoV-2 in the northeast of Italy.

Infection 2021 Apr 10. Epub 2021 Apr 10.

Infectious Diseases Unit, Department of Internal Medicine, Azienda Ospedaliera-Universitaria di Padova, Padua, Italy.

Purpose: We report on the first identified cluster of the B.1.1.7 variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections in the northeast of Italy.

Methods: The cluster was recognized in January 2021 with an epidemiological started from the hospitalization of a 68-year-old man suffering from coronavirus disease 2019 (COVID-19) related pneumonia and we surprisingly found three families involved in the same cluster.

Results: We retrospectively rebuilt the pathway of infection and performed a virological analysis.

Conclusion: This allow us to make clear the very high attack rate and the great infective capacity of this B.1.1.7 variant of SARS-CoV-2.
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http://dx.doi.org/10.1007/s15010-021-01609-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8035884PMC
April 2021

Generation of Combinatorial Lentiviral Vectors Expressing Multiple Anti-Hepatitis C Virus shRNAs and Their Validation on a Novel HCV Replicon Double Reporter Cell Line.

Viruses 2020 09 18;12(9). Epub 2020 Sep 18.

Department of Molecular Medicine, University of Padua, 35121 Padua, Italy.

Despite the introduction of directly acting antivirals (DAAs), for the treatment of hepatitis C virus (HCV) infection, their cost, patient compliance, and viral resistance are still important issues to be considered. Here, we describe the generation of a novel JFH1-based HCV subgenomic replicon double reporter cell line suitable for testing different antiviral drugs and therapeutic interventions. This cells line allowed a rapid and accurate quantification of cell growth/viability and HCV RNA replication, thus discriminating specific from unspecific antiviral effects caused by DAAs or cytotoxic compounds, respectively. By correlating cell number and virus replication, we could confirm the inhibitory effect on the latter of cell over confluency and characterize an array of lentiviral vectors expressing single, double, or triple cassettes containing different combinations of short hairpin (sh)RNAs, targeting both highly conserved viral genome sequences and cellular factors crucial for HCV replication. While all vectors were effective in reducing HCV replication, the ones targeting viral sequences displayed a stronger antiviral effect, without significant cytopathic effects. Such combinatorial platforms as well as the developed double reporter cell line might find application both in setting-up anti-HCV gene therapy approaches and in studies aimed at further dissecting the viral biology/pathogenesis of infection.
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http://dx.doi.org/10.3390/v12091044DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7551853PMC
September 2020

Universal screening of high-risk neonates, parents, and staff at a neonatal intensive care unit during the SARS-CoV-2 pandemic.

Eur J Pediatr 2020 Dec 7;179(12):1949-1955. Epub 2020 Aug 7.

Department of Women's and Children's Health, Neonatal Intensive Care Unit, Padova University Hospital, Padova, Italy.

Since February 21, 2020, SARS-CoV-2 has spread exponentially worldwide. Neonatal patients needing intensive care are considered a vulnerable population. To report the results of a policy based on multi-timepoint surveillance for SARS-CoV-2 of all neonates admitted to the neonatal intensive care unit (NICU), their parents, and all healthcare providers in a part of Italy with a high prevalence of the infection. Observational study conducted from 21 February to 21 April 2020. Intervention consisted of (a) parental triage on arrival at the neonatal ward; (b) universal testing with nasopharyngeal swabs and blood testing for SARS-CoV-2 IgM and IgG antibodies; (c) use of continuous personal protective equipment at the NICU by parents and staff. A total of 6726 triage procedures were performed on 114 parents, and 954 nasopharyngeal swabs were collected from 226 individuals. Five (2.2%) asymptomatic individuals (2 parents and 3 healthcare providers) tested positive on nasopharyngeal swabs and were kept isolated for 14 days. Of 75 admitted newborn, no one tested positive on nasopharyngeal swabs or antibody tests. Three parents presented with fever or flu-like symptoms at triage; they tested negative on swabs.Conclusion: With universal screening of neonates, parents, and staff, there were no cases of SARS-CoV-2 infection among the neonates admitted to a NICU in an area with a high incidence of SARS-CoV-2. Our experience could be usefully compared with other strategies with a view to developing future evidence-based guidelines for managing high-risk neonates in case of new epidemics. What is Known: • The novel coronavirus named SARS-CoV-2 has since spread worldwide at a remarkable rate, with more than 2.5 million confirmed cases. • Pediatric population may be less affected from COVID-19 than adult population but infants and newborn babies seem to be more vulnerable to SARS-CoV-2 infection. What is New: • Using an approach based on triage; testing with nasopharyngeal swabs and serology; and use of personal protective equipment, there were no cases of SARS-CoV-2 infection among neonates in a NICU in a high incidence of SARS-CoV-2 area. • Positive and asymptomatic individuals were identified and isolated early allowing the containment of infection's spread among healthcare providers and parents.
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http://dx.doi.org/10.1007/s00431-020-03765-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7410953PMC
December 2020

Suppression of a SARS-CoV-2 outbreak in the Italian municipality of Vo'.

Nature 2020 08 30;584(7821):425-429. Epub 2020 Jun 30.

School of Medicine, University of Padova, Padua, Italy.

On 21 February 2020, a resident of the municipality of Vo', a small town near Padua (Italy), died of pneumonia due to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. This was the first coronavirus disease 19 (COVID-19)-related death detected in Italy since the detection of SARS-CoV-2 in the Chinese city of Wuhan, Hubei province. In response, the regional authorities imposed the lockdown of the whole municipality for 14 days. Here we collected information on the demography, clinical presentation, hospitalization, contact network and the presence of SARS-CoV-2 infection in nasopharyngeal swabs for 85.9% and 71.5% of the population of Vo' at two consecutive time points. From the first survey, which was conducted around the time the town lockdown started, we found a prevalence of infection of 2.6% (95% confidence interval (CI): 2.1-3.3%). From the second survey, which was conducted at the end of the lockdown, we found a prevalence of 1.2% (95% CI: 0.8-1.8%). Notably, 42.5% (95% CI: 31.5-54.6%) of the confirmed SARS-CoV-2 infections detected across the two surveys were asymptomatic (that is, did not have symptoms at the time of swab testing and did not develop symptoms afterwards). The mean serial interval was 7.2 days (95% CI: 5.9-9.6). We found no statistically significant difference in the viral load of symptomatic versus asymptomatic infections (P = 0.62 and 0.74 for E and RdRp genes, respectively, exact Wilcoxon-Mann-Whitney test). This study sheds light on the frequency of asymptomatic SARS-CoV-2 infection, their infectivity (as measured by the viral load) and provides insights into its transmission dynamics and the efficacy of the implemented control measures.
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http://dx.doi.org/10.1038/s41586-020-2488-1DOI Listing
August 2020

SARS-CoV-2 RNA identification in nasopharyngeal swabs: issues in pre-analytics.

Clin Chem Lab Med 2020 08 22;58(9):1579-1586. Epub 2020 Jun 22.

Department of Medicine - DIMED, University of Padova, Padova, Italy.

Objectives: The direct identification of SARS-CoV-2 RNA in nasopharyngeal swabs is recommended for diagnosing the novel COVID-19 disease. Pre-analytical determinants, such as sampling procedures, time and temperature storage conditions, might impact on the end result. Our aim was to evaluate the effects of sampling procedures, time and temperature of the primary nasopharyngeal swabs storage on real-time reverse-transcription polymerase chain reaction (rRT-PCR) results.

Methods: Each nasopharyngeal swab obtained from 10 hospitalized patients for COVID-19 was subdivided in 15 aliquots: five were kept at room temperature; five were refrigerated (+4 °C); five were immediately mixed with the extraction buffer and refrigerated at +4 °C. Every day and for 5 days, one aliquot per condition was analyzed (rRT-PCR) for SARS-CoV-2 gene E and RNaseP and threshold cycles (Ct) compared. To evaluate manual sampling, 70 nasopharyngeal swabs were sampled twice by two different operators and analyzed separately one from the other.

Results: A total of 6/10 swabs were SARS-CoV-2 positive. No significant time or storage-dependent variations were observed in SARS-CoV-2 Ct. Re-sampling of swabs with SARS-CoV-2 Ct lower than 33 resulted in highly reproducible results (CV=2.9%), while a high variability was observed when Ct values were higher than 33 (CV=10.3%).

Conclusions: This study demonstrates that time and temperature of nasopharyngeal swabs storage do not significantly impact on results reproducibility. However, swabs sampling is a critical step, and especially in case of low viral load, might be a potential source of diagnostic errors.
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http://dx.doi.org/10.1515/cclm-2020-0749DOI Listing
August 2020

Human West Nile Virus Lineage 2 Infection: Epidemiological, Clinical, and Virological Findings.

Viruses 2020 04 18;12(4). Epub 2020 Apr 18.

Microbiology and Virology Unit, Padova University Hospital, I-35128 Padova, Italy.

West Nile virus (WNV) lineage 2 is expanding and causing large outbreaks in Europe. In this study, we analyzed the epidemiological, clinical, and virological features of WNV lineage 2 infection during the large outbreak that occurred in northern Italy in 2018. The study population included 86 patients with neuroinvasive disease (WNND), 307 with fever (WNF), and 34 blood donors. Phylogenetic analysis of WNV full genome sequences from patients' samples showed that the virus belonged to the widespread central/southern European clade of WNV lineage 2 and was circulating in the area at least since 2014. The incidence of WNND and WNF progressively increased with age and was higher in males than in females. Among WNND patients, the case fatality rate was 22%. About 70% of blood donors reported symptoms during follow-up. Within the first week after symptom onset, WNV RNA was detectable in the blood or urine of 80% of patients, while 20% and 40% of WNND and WNF patients, respectively, were WNV IgM-seronegative. In CSF samples of WNND patients, WNV RNA was typically detectable when WNV IgM antibodies were absent. Blunted or no WNV IgM response and high WNV IgG levels were observed in seven patients with previous flavivirus immunity.
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http://dx.doi.org/10.3390/v12040458DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7232435PMC
April 2020

Measles Virus Infection and Immunity in a Suboptimal Vaccination Coverage Setting.

Vaccines (Basel) 2019 Nov 28;7(4). Epub 2019 Nov 28.

Microbiology and Virology Unit, Padova University Hospital, 35128 Padova, Italy.

Despite efforts to improve surveillance and vaccination coverage, measles virus (MeV) continues to cause outbreaks also in high-income countries. As the reference laboratory of the Veneto Region, Italy, we analyzed changes in population immunity, described measles outbreaks, investigated MeV genetic diversity, and evaluated cross-protection of measles vaccination against MeV epidemic strains. Like most European areas, the Veneto Region has suboptimal measles vaccination coverage and is facing a growing public mistrust of vaccination. A progressive decline of measles vaccine uptake was observed during the last decade in the Veneto Region, leading to immunity gaps in children and young adults. Measles outbreaks were caused by the same MeV genotype B3, D4, and D8 strains that were circulating in other European countries. Eleven cases of measles were observed in immunized subjects. These cases were not associated with particular MeV genotypes nor with mutations in epitopes recognized by neutralizing antibodies. Accordingly, sera from fully vaccinated subjects cross-neutralized epidemic MeV strains, including the genotypes B3, D4, and D8, with the same high efficiency demonstrated against the vaccine strain. In fully vaccinated subjects, high MeV IgG antibody titers persisted up to 30 years following vaccination. These results support the use of the current measles-containing vaccines and strategies to strengthen vaccination.
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http://dx.doi.org/10.3390/vaccines7040199DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6963570PMC
November 2019

Clinical and virological findings in patients with Usutu virus infection, northern Italy, 2018.

Euro Surveill 2019 Nov;24(47)

Department of Molecular Medicine, University of Padova, Padova, Italy.

BackgroundUsutu virus (USUV) is a mosquito-borne flavivirus, which shares its transmission cycle with the phylogenetically related West Nile virus (WNV). USUV circulates in several European countries and its activity has increased over the last 5 years.AimTo describe human cases of USUV infection identified by surveillance for WNV and USUV infection in the Veneto Region of northern Italy in 2018.MethodsFrom 1 June to 30 November 2018, all cases of suspected autochthonous arbovirus infection and blood donors who had a reactive WNV nucleic acid test were investigated for both WNV and USUV infection by in-house molecular methods. Anti-WNV and anti-USUV IgM and IgG antibodies were detected by ELISA and in-house immunofluorescence assay, respectively; positive serum samples were further tested by WNV and USUV neutralisation assays run in parallel.ResultsEight cases of USUV infection (one with neuroinvasive disease, six with fever and one viraemic blood donor who developed arthralgia and myalgia) and 427 cases of WNV infection were identified. A remarkable finding of this study was the persistence of USUV RNA in the blood and urine of three patients during follow-up. USUV genome sequences from two patients shared over 99% nt identity with USUV sequences detected in mosquito pools from the same area and clustered within lineage Europe 2.ConclusionsClinical presentation and laboratory findings in patients with USUV infection were similar to those found in patients with WNV infection. Cross-reactivity of serology and molecular tests challenged the differential diagnosis.
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http://dx.doi.org/10.2807/1560-7917.ES.2019.24.47.1900180DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6885746PMC
November 2019

West Nile virus infection in individuals with pre-existing Usutu virus immunity, northern Italy, 2018.

Euro Surveill 2019 May;24(21)

Microbiology and Virology Unit, Padova University Hospital, Padova, Italy.

In 2018, there was a large West Nile virus (WNV) outbreak in northern Italy. We observed five atypical cases of WNV infection that were characterised by the presence of WNV RNA and WNV IgG at the time of diagnosis, but no IgM response during follow-up. Neutralisation assays demonstrated pre-existing Usutu virus immunity in all patients. Besides challenging diagnosis, the immunological crosstalk between the two viruses warrants further investigation on possible cross-protection or infection enhancement effects.
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http://dx.doi.org/10.2807/1560-7917.ES.2019.24.21.1900261DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6540647PMC
May 2019

Antiviral treatment and virological monitoring of oseltamivir-resistant influenza virus A(H1N1)pdm09 in a patient with chronic B lymphocytic leukemia.

J Infect Chemother 2019 Jul 20;25(7):543-546. Epub 2019 Apr 20.

Department of Molecular Medicine, University of Padova, Padova, Italy; Microbiology and Virology Unit, Azienda Ospedaliera di Padova, Padova, Italy. Electronic address:

We report the virological monitoring and the antiviral therapy adopted for the treatment of a patient affected by chronic B lymphocytic leukemia, who experienced a severe pneumonia with long-term shedding of influenza virus A(H1N1)pdm09, characterized by an early development of oseltamivir resistance.
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http://dx.doi.org/10.1016/j.jiac.2018.11.008DOI Listing
July 2019

Cytomegalovirus, Epstein-Barr virus and human herpesvirus 8 salivary shedding in HIV positive men who have sex with men with controlled and uncontrolled plasma HIV viremia: a 24-month longitudinal study.

BMC Infect Dis 2018 Dec 19;18(1):683. Epub 2018 Dec 19.

Department of Molecular Medicine, University of Padova, Via Gabelli 63, 35100, Padova, Italy.

Background: This longitudinal study described Cytomegalovirus (CMV) DNA, Epstein-Barr (EBV) DNA and human herpesvirus 8 (HHV-8) DNA asymptomatic salivary shedding in HIV-positive men who have sex with men (MSM). We aimed to 1-analyze frequency and persistence of herpesvirus shedding, 2-correlate herpesvirus positivity and HIV viroimmunological parameters and 3-assess the association between HIV-RNA suppression and herpesvirus replication.

Methods: Herpesvirus DNA was tested with an in-house real-time PCR in 2 salivary samples obtained at T0 and T1 (24 months after T0). HIV-RNA was evaluated in the 24 months prior to T0 and in the 24 months prior to T1; MSM were classified as successfully suppressed patients (SSPs), viremic patients (VPs) and partially suppressed patients (PSPs). EBV DNA load was classified as low viral load (EBV-LVL, value ≤10,000 copies/ml) and as high viral load (EBV-HVL,> 10,000 copies/ml). Mann-Whitney U test tested the difference of the median between groups of patients. Chi-squared test and Fisher's exact test compared categorical variables according to the frequencies. Kruskal-Wallis test compared continuous data distributions between levels of categorical variables.

Results: Ninety-two patients (median CD4+ count 575 cells/mm3, median nadir 330 CD4+ cells/mm3) were included: 40 SSPs,33 VPs and 19 PSPs. The more frequently single virus detected was EBV, both at T0 and at T1 (in 67.5 and 70% of SSPs, in 84.8 and 81.8% of VPs and in 68.4 and 73.7% of SPSs) and the most frequently multiple positivity detected was EBV + HHV-8. At T1, the percentage of CMV positivity was higher in VPs than in SSPs (36.4% vs 5%, p < 0.001), the combined shedding of HHV-8, CMV and EBV was present only in VPs (15.1%, p = 0.01 respect to SSPs) and no VPs confirmed the absence of shedding found at T0 (vs 17.5% of SSPs, p = 0.01). EBV-HVL was more frequent in VPs than in SSPs: 78.6% at T0 (p = 0.03) and 88.9% at T1 (p = 0.01).

Conclusions: The relationship between uncontrolled plasma HIV viremia and CMV, EBV, and HHV-8 shedding is multifaceted, as demonstrated by the focused association with EBV DNA load and not with its frequency and by the persistent combined detection of two oncogenic viruses as EBV and HHV-8 regardless of HIV virological control.
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http://dx.doi.org/10.1186/s12879-018-3591-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6300014PMC
December 2018

The complete genome sequence analysis of West Nile virus strains isolated in Slovakia (central Europe).

Arch Virol 2019 Jan 4;164(1):273-277. Epub 2018 Oct 4.

Department of Microbiology and Immunology, University of Veterinary Medicine and Pharmacy in Košice, Košice, Slovakia.

The present study reports the first complete genome sequence analysis of West Nile virus (WNV) strains isolated from brain samples from raptors. The results prove the circulation of closely related WNV lineage II strains in central Europe and genetic analysis revealed seven amino acid substitutions in structural (PrM, E and E) and in non-structural (NS1, NS5, NS5 and NS5) proteins. Observed amino acid substitutions Phe and Ser were common only within the lineage VII Koutango strain isolated from Rhipicephalus guilhoni tick in Senegal. Further research could reveal whether these substitutions influence the biological properties of WNV, including virulence and neuroinvasiveness.
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http://dx.doi.org/10.1007/s00705-018-4056-7DOI Listing
January 2019

Prevalence, molecular epidemiology and intra-hospital acquisition of Klebsiella pneumoniae strains producing carbapenemases in an Italian teaching hospital from January 2015 to September 2016.

Int J Infect Dis 2017 Jun 12;59:103-109. Epub 2017 Apr 12.

Department of Molecular Medicine, University of Padova, Via Gabelli 63, 35100 Padova, Italy. Electronic address:

Objectives: We described Klebsiella pneumoniae producing carbapenemase (CPKP) spread from 01/01/2015 to 13/09/16 in a tertiary level hospital.

Methods: The first positive surveillance rectal swab (SRS) or clinical sample (CS) collected in the medical department (MD), surgical department (SD) and intensive care department (ICD) were included in the study. A validated in-house Real-Time PCR method was used to detect carbapenemases; multilocus sequence typing (MLST) was used for further characterization of the strains.

Results: 21535 patients were included: 213 CPKP strains from surveillance rectal swab (SRS) and 98 from clinical samples (CS) were collected. The percentage of CPKP detected in SRS with respect to CS increased in the medical MD from 2015 to 2016 (p=0.01) and in ICD from 2012 to 2015 (p=0.0001), while it decreased in SD from 2014 to 2016 (p=0.003); 68.5% of the positive SRS had a previous negative SRS; CPKP was more frequently identified in CS than in SRS in MD. Twelve strains harboured more than one carbapenemase gene. Many other species harbouring a carbapenemase gene were collected.

Conclusions: MDs need more inclusive surveillance criteria. The late detection of positive SRS underlined the risk of colonization during hospitalization.
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http://dx.doi.org/10.1016/j.ijid.2017.04.007DOI Listing
June 2017

Infection dynamics in a traveller with persistent shedding of Zika virus RNA in semen for six months after returning from Haiti to Italy, January 2016.

Euro Surveill 2016 Aug;21(32)

Department of Molecular Medicine, University of Padova, Padova, Italy.

We describe the dynamics of Zika virus (ZIKV) infection in a man in his early 40s who developed fever and rash after returning from Haiti to Italy, in January 2016. Follow-up laboratory testing demonstrated detectable ZIKV RNA in plasma up to day 9 after symptom onset and in urine and saliva up to days 15 and 47, respectively. Notably, persistent shedding of ZIKV RNA was demonstrated in semen, still detectable at 181 days after onset.
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http://dx.doi.org/10.2807/1560-7917.ES.2016.21.32.30316DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4998504PMC
August 2016

Virological testing of cerebrospinal fluid in children aged less than 14 years with a suspected central nervous system infection: A retrospective study on 304 consecutive children from January 2012 to May 2015.

Eur J Paediatr Neurol 2016 Jul 16;20(4):588-96. Epub 2016 Apr 16.

Department of Molecular Medicine, University of Padova, Via Gabelli 63, 35100 Padova, Italy; Microbiology and Virology Unit, Padova University Hospital, Indirizzo: Via Giustiniani, 2, 35128 Padova, Italy.

Objective: The study aimed to describe the prevalence of HSV DNA, VZV DNA, Enterovirus RNA, Parechovirus RNA, CMV DNA, EBV DNA, adenovirus DNA, HHV-6 DNA, HHV-7 DNA, HHV-8 DNA and Parvovirus B19DNA in children aged less 14 years with a suspected viral infection of the central nervous system in a clinical practice setting.

Methods: Between January 2012 and May 2015, cerebrospinal fluids from 304 children were tested with an in-house real-time PCR method.

Results: A positive PCR was detected in 64 subjects (21%): the mean number of tests performed in patients who showed a viral infection was 7.5, significantly higher (p = 0.001) with respect to that reported in negative samples (6.4). Enterovirus is the leading virus detected: 12 out of the 37 positive children reported were newborns (85.7% of all the newborns with a positive result). The second most frequently identified virus was HHV-7 (5 positive PCR out of 105 samples tested, 4.8%, if we excluded a child with a concomitant S. pneumoniae isolated), a prevalence significantly higher with respect to VZV (p = 0.02) and to CMV (p = 0.04). HHV-6 was the third most commonly identified aetiology (4.2%). All children were immunocompetent.

Significance: Only a minority of children had a specific viral aetiology identified: the rate of HHV-7 positivity suggests a routine testing of these viruses within the diagnostic algorithm in immunocompetent paediatric patients. This approach could help to define the clinical role of this herpesvirus.
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http://dx.doi.org/10.1016/j.ejpn.2016.04.002DOI Listing
July 2016

Isolation of infectious Zika virus from saliva and prolonged viral RNA shedding in a traveller returning from the Dominican Republic to Italy, January 2016.

Euro Surveill 2016 ;21(10):30159

Department of Molecular Medicine, University of Padova, Padova, Italy.

We report the isolation of infectious Zika virus (ZIKV) in cell culture from the saliva of a patient who developed a febrile illness after returning from the Dominican Republic to Italy, in January 2016. The patient had prolonged shedding of viral RNA in saliva and urine, at higher load than in blood, for up to 29 days after symptom onset. Sequencing of ZIKV genome showed relatedness with strains from Latin America.
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http://dx.doi.org/10.2807/1560-7917.ES.2016.21.10.30159DOI Listing
September 2016

Characterization of Intra-Type Variants of Oncogenic Human Papillomaviruses by Next-Generation Deep Sequencing of the E6/E7 Region.

Viruses 2016 Mar 14;8(3):79. Epub 2016 Mar 14.

Department of Molecular Medicine, University of Padova, via A. Gabelli 63, 35121 Padova, Italy.

Different human papillomavirus (HPV) types are characterized by differences in tissue tropism and ability to promote cell proliferation and transformation. In addition, clinical and experimental studies have shown that some genetic variants/lineages of high-risk HPV (HR-HPV) types are characterized by increased oncogenic activity and probability to induce cancer. In this study, we designed and validated a new method based on multiplex PCR-deep sequencing of the E6/E7 region of HR-HPV types to characterize HPV intra-type variants in clinical specimens. Validation experiments demonstrated that this method allowed reliable identification of the different lineages of oncogenic HPV types. Advantages of this method over other published methods were represented by its ability to detect variants of all HR-HPV types in a single reaction, to detect variants of HR-HPV types in clinical specimens with multiple infections, and, being based on sequencing of the full E6/E7 region, to detect amino acid changes in these oncogenes potentially associated with increased transforming activity.
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http://dx.doi.org/10.3390/v8030079DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4810269PMC
March 2016

Full Genome Sequence-Based Comparative Study of Wild-Type and Vaccine Strains of Infectious Laryngotracheitis Virus from Italy.

PLoS One 2016 18;11(2):e0149529. Epub 2016 Feb 18.

Department of Molecular Medicine, University of Padua (DMM), Padua, Italy.

Infectious laryngotracheitis (ILT) is an acute and highly contagious respiratory disease of chickens caused by an alphaherpesvirus, infectious laryngotracheitis virus (ILTV). Recently, full genome sequences of wild-type and vaccine strains have been determined worldwide, but none was from Europe. The aim of this study was to determine and analyse the complete genome sequences of five ILTV strains. Sequences were also compared to reveal the similarity of strains across time and to discriminate between wild-type and vaccine strains. Genomes of three ILTV field isolates from outbreaks occurred in Italy in 1980, 2007 and 2011, and two commercial chicken embryo origin (CEO) vaccines were sequenced using the 454 Life Sciences technology. The comparison with the Serva genome showed that 35 open reading frames (ORFs) differed across the five genomes. Overall, 54 single nucleotide polymorphisms (SNPs) and 27 amino acid differences in 19 ORFs and two insertions in the UL52 and ORFC genes were identified. Similarity among the field strains and between the field and the vaccine strains ranged from 99.96% to 99.99%. Phylogenetic analysis revealed a close relationship among them, as well. This study generated data on genomic variation among Italian ILTV strains revealing that, even though the genetic variability of the genome is well conserved across time and between wild-type and vaccine strains, some mutations may help in differentiating among them and may be involved in ILTV virulence/attenuation. The results of this study can contribute to the understanding of the molecular bases of ILTV pathogenicity and provide genetic markers to differentiate between wild-type and vaccine strains.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0149529PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4758665PMC
July 2016

Viral infections of the central nervous system in elderly patients: a retrospective study.

Int J Infect Dis 2016 Mar 25;44:8-10. Epub 2016 Jan 25.

Department of Molecular Medicine, University of Padova, Via Gabelli 63, 35100 Padova, Italy; Microbiology and Virology Unit, Padova Hospital, Via Giustiniani 2, Padova, Italy.

Objectives: Very few data exist on viral meningitis and encephalitis in elderly patients (>65 years old).

Methods: This study investigated the detection of herpes simplex virus (HSV), varicella zoster virus (VZV), human herpes virus 6 (HHV-6), HHV-7, HHV-8, cytomegalovirus (CMV), Epstein-Barr virus (EBV), enterovirus (EV), human adenovirus (HAdV), human parechoviruses (HPeVs), and tick-borne encephalitis virus (TBEV) through real-time PCR (RT-PCR) in patients >65 years old who had cerebrospinal fluid (CSF) tested for a suspected central nervous system infection.

Results: A total of 2868 RT-PCRs were performed on 502 CSF samples. Overall, 65 positive RT-PCRs were found: 23 for HSV (35.4% of positives), 15 for EV (23.1% of positives), 14 for EBV (21.5% of positives), 12 for VZV (18.5% of positives), and one for CMV (1.5% of positives). A positive RT-PCR in CSF was detected in 24 (17.4%) patients aged ≥ 80 years and in 35 (9.6%) patients aged 65-79 years (p=0.02). VZV was more frequently detected in the oldest subjects (5.9% vs. 1.6%, p=0.03).

Conclusions: HSV was the most common viral aetiology identified in the study, with VZV infection being recognized more frequently in those patients aged ≥ 80 years.
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http://dx.doi.org/10.1016/j.ijid.2016.01.012DOI Listing
March 2016

Epstein-Barr and cytomegalovirus DNA salivary shedding correlate with long-term plasma HIV RNA detection in HIV-infected men who have sex with men.

J Med Virol 2016 Jul 15;88(7):1211-21. Epub 2015 Dec 15.

Department of Molecular Medicine, University of Padova, Padova, Italy.

The aim of the study was to evaluate cytomegalovirus (CMV) and Epstein-Barr virus (EBV) DNA salivary shedding in HIV-positive men who have sex with men (MSM) and to determine whether viro-immunological parameters and long-term (24 months) plasma HIV RNA (pHIV) detection may predict herpesviruses replication. A total of 193 HIV-positive MSM were consecutively recruited (mean CD4+ cell count 607 cells/mm(3) and mean nadir value 333 cells/mm(3) ); pHIV was analyzed for 24 months prior to saliva sampling: patients were categorized as successfully suppressed (SS) and not suppressed (NS). The EBV viral load was categorized as high viral load (HVL), intermediate (IVL), or low (LVL), CMV DNA as positive or negative. NS patients experienced both herpesviruses detectability more frequently respect to SS patients (P = 0.034); conversely, no salivary shedding was more frequent in SS patients (P = 0.014). HVL EBV was more frequent in NS patients than in SS subjects (P = 0.038 for isolated EBV detection and P = 0.001 when CMV shedding was associated). NS subjects with HVL EBV had a median pHIV of 43,820 copies/ml, significantly higher respect to IVL and LVL patients (P = 0.027 and P = 0.0005, respectively). CMV shedding was mostly associated to EBV shedding. NS patients showed a significantly higher frequency of saliva HVL EBV detection compared to SS patients; moreover, NS patients with HVL EBV had a higher pHIV respect to those with IVL and LVL shedding. Our results suggest that a successful pHIV suppression could reduce the burden of salivary EBV replication and likely the risk of herpesviruses-related cancers.
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http://dx.doi.org/10.1002/jmv.24441DOI Listing
July 2016

Prevalence of Klebsiella pneumoniae strains producing carbapenemases and increase of resistance to colistin in an Italian teaching hospital from January 2012 To December 2014.

BMC Infect Dis 2015 Jun 27;15:244. Epub 2015 Jun 27.

Department of Molecular Medicine, University of Padova, Via Gabelli 63, 35100, Padova, Italy.

Background: The aim of this study was to characterize the spread of carbapenemase-producing Klebsiella pneumoniae (CPKP) in a tertiary level hospital using ongoing active surveillance with rectal swab cultures. Furthermore, this study analyzed the presence of CPKP in the clinical samples (CS) of a single patient as well as the evolution of Colistin-sensitive strains (CoS) to Colistin-resistant strains (CoR).

Methods: This study was performed from January 1, 2012 to December 31, 2014. In 2012, a survey was conducted in the Intensive Care Department. In autumn 2013, active monitoring was extended to the Surgery Department, and since mid-2014, the surveillance has included the Medical Department as well. Only the first isolated strain from each patient was included. Antimicrobial susceptibility testing was performed on CPKP isolates: Klebsiella pneumoniae carbapenemase, oxacillinase-48, Verona integron-encoded metallo-β-lactamase and New Delhi metallo-β-lactamase were detected using a validated in-house PCR method, and multilocus sequence typing (MLST) was used to investigate the clonal transmission of strains.

Results: A total of 15,104 patients were included in the study, and 496 consecutive non-replicated strains of CPKP were collected: 149 strains were collected in 2012 (39 [26.2 %] from surveillance rectal swabs [SRS]), 133 strains were collected in 2013 (70 [52.6 %] from SRS) and 214 strains were collected in 2014 (164 [76.6 %] from SRS). We observed a significant increase in the percentage of positive SRS cases in 2014 relative to 2013 and 2012 (p = 0.0001 and p = 0.0172, respectively) and in the proportion of CPKP first isolated by SRS relative to those identified by CS (p < 0.0001). Among all available samples, the number of CoR isolated from SRS was higher in 2013 and 2014 compared with 2012 (p = 0.0019 and p = 0.008, respectively). ST-258 and ST-512 were more prevalent in the tested specimens, and a new single locus variant (SLV) of ST-512 (ST-745) was isolated.

Conclusions: The results of this 3-year study of 15,104 patients highlight the clinical relevance of antimicrobial resistance as well as the drug-selection pressure of colistin therapy. The active surveillance in the three different departments increased the level of CPKP cases isolated by SRS.
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http://dx.doi.org/10.1186/s12879-015-0996-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4482166PMC
June 2015

Isolation of West Nile virus from urine samples of patients with acute infection.

J Clin Microbiol 2014 Sep 20;52(9):3411-3. Epub 2014 Jun 20.

Department of Molecular Medicine, University of Padua, Padua, Italy Microbiology and Virology Unit, Padua University Hospital, Padua, Italy.

This study demonstrated that West Nile virus (WNV) excreted in the urine of patients with acute infection can be isolated in cell cultures. In addition, the protocols for WNV isolation from urine samples were standardized, and factors that may affect the efficiency of WNV isolation were identified.
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http://dx.doi.org/10.1128/JCM.01328-14DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4313171PMC
September 2014

Four year longitudinal study of Mycobacterium tuberculosis complex isolates in a region of North-Eastern Italy.

Infect Genet Evol 2014 Aug 10;26:58-64. Epub 2014 May 10.

Department of Molecular Medicine, University of Padova, Padova, Italy; U.O. Microbiologia e Virologia, Azienda Ospedaliera di Padova, Padova, Italy.

Recent reports have suggested a change of Mycobacterium tuberculosis complex genetic diversity in Western Europe due to an increasing proportion of imported cases of tuberculosis (TB). This study analyzed a total of 705 M. tuberculosis strains isolated from 2006 to 2009 in Veneto, a North-Eastern Italian region, to see the impact of foreign-born cases vs. Italian patients on prevailing TB epidemiology. Strains were genotyped using spoligotyping followed by comparison with international genotyping database SITVIT2. Six spoligotyping clusters with suspected phylogeographical specificity for imported cases, were typed by 15-loci MIRUs for a finer characterization. Overall, 410 (58.16%) strains were isolated from foreign-born patients, while 295 (41.84%) were isolated from Italian patients. Older patients (>70 years, i.e., 46.4% of cases) predominated among Italians while younger age groups prevailed among foreign-born patients. Our results suggest that despite a high proportion of reactivation of latent TB infection in elderly Italian-born patients, active TB transmission between foreign-born and Italian patients may be ongoing, and argue in favor of an increased TB surveillance among immigrants to combat TB epidemic in Italy.
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http://dx.doi.org/10.1016/j.meegid.2014.05.004DOI Listing
August 2014

Large human outbreak of West Nile virus infection in north-eastern Italy in 2012.

Viruses 2013 Nov 22;5(11):2825-39. Epub 2013 Nov 22.

Department of Molecular Medicine, University of Padova, Padova I-35121, Italy.

Human cases of West Nile virus (WNV) disease have been reported in Italy since 2008. So far, most cases have been identified in north-eastern Italy, where, in 2012, the largest outbreak of WNV infection ever recorded in Italy occurred. Most cases of the 2012 outbreak were identified in the Veneto region, where a special surveillance plan for West Nile fever was in place. In this outbreak, 25 cases of West Nile neuroinvasive disease and 17 cases of fever were confirmed. In addition, 14 WNV RNA-positive blood donors were identified by screening of blood and organ donations and two cases of asymptomatic infection were diagnosed by active surveillance of subjects at risk of WNV exposure. Two cases of death due to WNND were reported. Molecular testing demonstrated the presence of WNV lineage 1 in all WNV RNA-positive patients and, in 15 cases, infection by the novel Livenza strain was ascertained. Surveillance in other Italian regions notified one case of neuroinvasive disease in the south of Italy and two cases in Sardinia. Integrated surveillance for WNV infection remains a public health priority in Italy and vector control activities have been strengthened in areas of WNV circulation.
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http://dx.doi.org/10.3390/v5112825DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3856417PMC
November 2013

Genomic comparative analysis and gene function prediction in infectious diseases: application to the investigation of a meningitis outbreak.

BMC Infect Dis 2013 Nov 19;13:554. Epub 2013 Nov 19.

Department of Molecular Medicine, University of Padova, Padova, Italy.

Background: Next generation sequencing (NGS) is being increasingly used for the detection and characterization of pathogens during outbreaks. This technology allows rapid sequencing of pathogen full genomes, useful not only for accurate genotyping and molecular epidemiology, but also for identification of drug resistance and virulence traits.

Methods: In this study, an approach based on whole genome sequencing by NGS, comparative genomics, and gene function prediction was set up and retrospectively applied for the investigation of two N. meningitidis serogroup C isolates collected from a cluster of meningococcal disease, characterized by a high fatality rate.

Results: According to conventional molecular typing methods, all the isolates had the same typing results and were classified as outbreak isolates within the same N. meningitidis sequence type ST-11, while full genome sequencing demonstrated subtle genetic differences between the isolates. Looking for these specific regions by means of 9 PCR and cycle sequencing assays in other 7 isolates allowed distinguishing outbreak cases from unrelated cases. Comparative genomics and gene function prediction analyses between outbreak isolates and a set of reference N. meningitidis genomes led to the identification of differences in gene content that could be relevant for pathogenesis. Most genetic changes occurred in the capsule locus and were consistent with recombination and horizontal acquisition of a set of genes involved in capsule biosynthesis.

Conclusions: This study showed the added value given by whole genome sequencing by NGS over conventional sequence-based typing methods in the investigation of an outbreak. Routine application of this technology in clinical microbiology will significantly improve methods for molecular epidemiology and surveillance of infectious disease and provide a bulk of data useful to improve our understanding of pathogens biology.
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http://dx.doi.org/10.1186/1471-2334-13-554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4225559PMC
November 2013

The complex epidemiological scenario of West Nile virus in Italy.

Int J Environ Res Public Health 2013 Sep 30;10(10):4669-89. Epub 2013 Sep 30.

Department of Molecular Medicine, University of Padova, Via Gabelli, 63, 35121 Padova, Italy.

Entomological, veterinary, and human surveillance systems for West Nile virus (WNV) infection have been implemented in Italy since the first detection of the virus in 1998. These surveillance activities documented a progressive increase of WNV activity and spread in different regions and the emergence of new WNV lineages and strains. Italy is a paradigmatic example of the complex epidemiology of WNV in Europe, where sporadic cases of WNV infection, clusters, and small outbreaks have been reported in several regions. In addition, different strains of both WNV lineage 1 and lineage 2 have been identified, even co-circulating in the same area.
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http://dx.doi.org/10.3390/ijerph10104669DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3823324PMC
September 2013

Genome sequencing of West Nile Virus from human cases in Greece, 2012.

Viruses 2013 Sep 24;5(9):2311-9. Epub 2013 Sep 24.

Department of Molecular Medicine, University of Padova, 35122 Padova, Italy.

A West Nile Virus (WNV) lineage 2 strain, named Nea Santa-Greece-2010, has been demonstrated to be responsible for the large outbreaks of neuroinvasive disease (WNND) that have been occurring in Greece since 2010, based on sequence similarities of viral isolates identified between 2010-2012. However, knowledge on the evolution of this strain is scarce because only partial WNV genome sequences are available from Greece. The aim of this study was to get the complete genome sequence of WNV from patients with infection. To this aim, plasma and urine samples collected during the 2012 Greek outbreak were retrospectively investigated. Full WNV genome sequence was obtained from a patient with WNND. The genome had 99.7% sequence identity to Nea Santa, higher than to other related WNV lineage 2 strains, and five amino acid changes apparently not relevant for viral pathogenicity or fitness. In addition, infection by WNV lineage 2 was confirmed in additional nine patients with WNND; in three of them the infection with WNV Nea Santa was demonstrated by sequencing. In conclusion, this study characterized for the first time a WNV full genome from a patient with WNND from Greece, demonstrated the persistence of the Nea Santa strain, and suggested that the virus might have locally evolved.
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http://dx.doi.org/10.3390/v5092311DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3798904PMC
September 2013

Excretion of West Nile virus in urine during acute infection.

J Infect Dis 2013 Oct 2;208(7):1086-92. Epub 2013 Jul 2.

Department of Molecular Medicine, University of Padova, Padova, Italy.

Detection of West Nile virus (WNV) RNA in urine has been anecdotally described and proposed for the diagnosis of WNV infection. This study reports the routine use of real-time reverse-transcription polymerase chain reaction for the detection of WNV RNA in urine to support diagnosis of WNV infection during the large outbreak that occurred in northeastern Italy in 2012. Fourteen of 32 patients (43.8%) with symptomatic WNV infection, defined as neuroinvasive disease and fever, had detectable WNV RNA in urine at the time of diagnosis, at a higher rate and load and for a longer time than detection of WNV RNA in blood. Detection of WNV RNA in urine was less frequent (2 of 14 patients [14.2%]) in blood donors in whom WNV infection was identified by WNV nucleic acid amplification testing. Infectious virus was isolated from the urine of a patient with neuroinvasive disease and a high WNV RNA load in urine.
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http://dx.doi.org/10.1093/infdis/jit290DOI Listing
October 2013

Next-generation sequencing technologies in diagnostic virology.

J Clin Virol 2013 Oct 21;58(2):346-50. Epub 2013 Mar 21.

Department of Molecular Medicine, University of Padova, Via A. Gabelli 63, I-35121 Padova, Italy. Electronic address:

The data deluge produced by next-generation sequencing (NGS) technologies is an appealing feature for clinical virologists that are involved in the diagnosis of emerging viral infections, molecular epidemiology of viral pathogens, drug-resistance testing, and also like to do some basic and clinical research. Indeed, NGS platforms are being implemented in many clinical and research laboratories, as the costs of these platforms are progressively decreasing. We provide here some suggestions for virologists who are planning to implement a NGS platform in their clinical laboratory and an overview on the potential applications of these technologies in diagnostic virology.
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http://dx.doi.org/10.1016/j.jcv.2013.03.003DOI Listing
October 2013