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BackgroundFollowing the low incidence rates of non-SARS-CoV-2 respiratory viruses registered during the strict lockdown enforced in the pandemic, a resurgence of several endemic viruses in Catalonia (Spain) was noted during the early summer of 2021.ObjectivesIn this study, we investigated whether the circulation of non-SARS-CoV-2 respiratory viruses in Catalonia, assessed by Microbiological Reporting System of Catalonia (MRSC) and the Epidemiological Surveillance Network of Catalonia, was affected by the strict lockdown measures, as well as, the implication of the Coronavirus Disease 19 (COVID-19) de-escalation process in the late season outbreaks registered during the 2020-2021 season.Study designA retrospective comparison of epidemic patterns in the respiratory viruses' incidence, using regional public health surveillance data from MRSC, was performed between weeks 26/2016 to week 27/2021. Data were expressed as the weekly total number of test positivity for individual viruses. A segmented negative binomial regression model was conducted, with two parameters included (level and trend) for each segment of the time series (2020 pre-lockdown, 2020 post-lockdown and 2021). Results were reported as a unit changed in the strict lockdown.ResultsA total of 51588 confirmed cases of the different respiratory viruses were included in the analysis, the majority were influenza cases (63.7%). An immediate reduction in the weekly number of cases was observed in 2020 after the COVID-19 outbreak for human adenovirus virus (HAdV) (β2 = -2.606; P ConclusionsOur data not only provides evidence that occurrence of different respiratory virus infections was affected by the strict lockdown taken against SARS-CoV-2 but it also shows a late resurgence of seasonal respiratory viruses' cases during the 2020-2021 season following the relaxation of COVID-19-targeted non-pharmaceutical interventions.

During a waterborne outbreak of norovirus in Spain, we estimated 50% illness doses for a group of exposed (secretor) persons to be 556 (95% CI 319–957) genome copies/day for norovirus GI and 2,934 (95% CI 1,683–5,044) genome copies/day for norovirus GII. Use of a propidium monoazide viability assay reduced these values.

Background: A huge outbreak in the men-having-sex-with-men (MSM) has hit Europe during the years 2016–2018. Outbreak control has been hampered by vaccine shortages in many countries, and to minimize their impact, reduction of antigen doses has been implemented. However, these measures may have consequences on the evolution of hepatitis A virus (HAV), leading to the emergence of antigenic variants. Cases in vaccinated MSM patients have been detected in Barcelona, opening the possibility to study HAV evolution under immune pressure. Methods: We performed deep-sequencing analysis of ten overlapping fragments covering the complete capsid coding region of HAV. A total of 14578255 reads were obtained and used for the analysis of virus evolution in vaccinated versus non-vaccinated patients. We estimated maximum and minimum mutation frequencies, and Shannon entropy in the quasispecies of each patient. Non-synonymous (NSyn) mutations affecting residues exposed in the capsid surface were located, with respect to epitopes, using the recently described crystal structure of HAV, as an indication of its potential role in escaping to the effect of vaccines. Findings: HAV evolution at the quasispecies level, in non-vaccinated and vaccinated patients, revealed higher diversity in epitope-coding regions of the vaccinated group. Although amino acid replacements occurring in and around the epitopes were observed in both groups, their abundance was significantly higher in the quasispecies of vaccinated patients, indicating ongoing processes of fixation. Interpretation: Our data suggest positive selection of antigenic variants in some vaccinated patients, raising concerns for new vaccination polices directed to the MSM group. Keywords: Vaccine escape-mutants, MSM, Quasispecies, Deep-sequencing

Bivalve mollusk contamination by enteric viruses, especially human noroviruses (HuNoV) and hepatitis A virus (HAV), is a problem with health and economic implications. The aim of the study was the evaluation of the effect of heat treatment in clams (Tawera gayi) experimentally contaminated with HuNoV using a PMA-viability RTqPCR assay to minimize measurement of non-infectious viruses, and used HAV as a model to estimate infectivity loss. Spiked clams were immersed in water at 90°C to ensure that internal meat temperature was maintained above 90°C for at least 5 min. The treatment resulted in >3.89 ± 0.24 log10 TCID50/g reduction of infectious HAV, confirming inactivation. For HuNoV, RTqPCR assays showed log10 reductions of 2.96 ± 0.79 and 2.56 ± 0.56, for GI and GII, respectively, and the use of PMA resulted in an additional log10 reduction for GII, providing a better correlation with risk reduction. In the absence of a cell culture system which could be used to determine HuNoV infectivity reduction, a performance criteria based on PMA-RTqPCR log reduction could be used to evaluate food product safety. According to data from this study, heat treatments of clams which cause reductions >3.5 log10 for GII as measured by PMA-RTqPCR assay may be regarded as an acceptable inactivation treatment, and could be set as a performance criterion to test the effectiveness of other time-temperature inactivation processes.

Norovirus infections are a leading cause of acute gastroenteritis outbreaks worldwide and across all age groups, with two main genogroups (GI and GII) infecting humans. The aim of our study was to investigate the occurrence of norovirus in saliva samples from individuals involved in outbreaks of acute gastroenteritis in closed and semiclosed institutions, and its relationship with the virus strain, virus shedding in stool, the occurrence of symptoms, age, and the secretor status of the individual. Epidemiological and clinical information was gathered from norovirus outbreaks occurring in Catalonia, Spain during 2017–2018, and stool and saliva samples were collected from affected and exposed resident individuals and workers. A total of 347 saliva specimens from 25 outbreaks were analyzed. Further, 84% of individuals also provided a paired stool sample. For GII infections, norovirus was detected in 17.9% of saliva samples from symptomatic cases and 5.2% of asymptomatic individuals. Positivity in saliva occurred in both secretors and nonsecretors. None of the individuals infected by norovirus GI was positive for the virus in saliva. Saliva positivity did not correlate with any of the studied symptoms but did correlate with age ≥ 65 years old. Individuals who were positive in saliva showed higher levels of virus shedding in stool. Mean viral load in positive saliva was 3.16 ± 1.08 log10 genome copies/mL, and the predominance of encapsidated genomes was confirmed by propidium monoazide (PMA)xx-viability RTqPCR assay. The detection of norovirus in saliva raises the possibility of oral-to-oral norovirus transmission during the symptomatic phase and, although to a lesser extent, even in cases of asymptomatic infections.

[This corrects the article DOI: 10.1371/journal.pone.0201850.].

Noroviruses are the main cause of epidemics of acute gastroenteritis at a global scale. Although chronically infected immunocompromised individuals are regarded as potential reservoirs for the emergence of new viral variants, viral quasispecies distribution and evolution patterns in acute symptomatic and asymptomatic infections has not been extensively studied. Amplicons of 450 nts from the P2 coding capsid domain were studied using next-generation sequencing (454/GS-Junior) platform. Inter-host diversity between symptomatic and asymptomatic acutely infected individuals linked to the same outbreak as well as their viral intra-host diversity over time were characterized. With an average of 2848 reads per sample and a cutoff frequency of 0.1%, minor variant haplotypes were detected in 5 out of 8 specimens. Transmitted variants could not be confirmed in all infected individuals in one outbreak. The observed initial intra-host viral diversity in asymptomatically infected subjects was higher than in symptomatic ones. Viral quasispecies evolution over time within individuals was host-specific, with an average of 2.8 nt changes per day (0.0062 changes per nucleotide per day) in a given symptomatic case. Nucleotide polymorphisms were detected in 28 out of 450 analyzed nucleotide positions, 32.14% of which were synonymous and 67.86% were non-synonymous. Most observed amino acid changes emerged at or near blockade epitopes A, B, D and E. Our results suggest that acutely infected individuals, even in the absence of symptoms, which go underreported and may enhance transmission, may contribute to norovirus genetic variability and evolution.

A remarkable percentage of acute gastroenteritis cases remain etiologically undiagnosed. The aim of the study was to determine the prevalence of common and emerging enteric viruses, such as novel human astroviruses, among undiagnosed samples from children with acute gastroenteritis. Epidemiological studies for novel human astroviruses are still scarce. Stool samples collected over two consecutive winter seasons (2016−2017) from children with gastroenteritis in Spain, which were negative for bacteria, rotavirus, and adenovirus by routine diagnostics were screened by real-time RT-PCR assays for the presence of classical and novel astrovirus, rotavirus, norovirus GI and GII, sapovirus, and adenovirus. Overall, 220/384 stool samples (57.3%) were positive for at least one virus. Co-infections were identified in 21% of cases. Among a total of 315 viruses identified, adenovirus was the most prevalent (n = 103), followed by rotavirus (n = 51), sapovirus (n = 50), classical astrovirus (n = 43), novel astroviruses (n = 42), and norovirus (n = 26). Novel astroviruses were present in 13.3% of virus-positive cases. Most novel astroviruses were found in children p = 0.01) and were found in co-infection (66%). Only classical astroviruses demonstrated significant differences in the Cq values during mono-infections compared to co-infections. In conclusion, common enteric viruses may be frequently found in children with undiagnosed gastroenteritis, indicating the need to implement more sensitive diagnostic methods. Novel astroviruses circulate in the community and could be the cause of gastroenteritis among young children.