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Abstract Food protein amyloid fibrils have superior technological, nutritional, sensorial, and physical properties compared to native monomers, but there is as yet insufficient understanding of their digestive fate and safety for wide consumption. By combining SDS-PAGE, ELISA, fluorescence, AFM, MALDI-MS, CD, microfluidics, and SAXS techniques for the characterization of β-lactoglobulin and lysozyme amyloid fibrils subjected to in-vitro gastrointestinal digestion, here we show that either no noticeable conformational differences exist between amyloid aggregates and their monomer counterparts after the gastrointestinal digestion process (as in β-lactoglobulin), or that amyloid fibrils are digested significantly better than monomers (as in lysozyme). Moreover, in-vitro exposure of human cell lines and in-vivo studies with C. elegans and mouse models, indicate that the digested fibrils present no observable cytotoxicity, physiological abnormalities in health-span, nor accumulation of fibril-induced plaques in brain nor other organs. These extensive in-vitro and in-vivo studies together suggest that the digested food amyloids are at least equally as safe as those obtained from the digestion of corresponding native monomers, pointing to food amyloid fibrils as potential ingredients for human nutrition.

Abstract Background Enterovirus A71 is one of the causative agents of hand, foot, and mouth disease, which is usually a self-limiting disease. Complications of enterovirus infection are also very rare. However, when such complications occur, they can lead to serious neurological diseases or even death. Case presentation In this report, we describe a case of enterovirus A71-associated acute flaccid paralysis in a 13-month-old Caucasian girl that was managed in our hospital. The patient presented with sudden onset of left arm paresis that could not be attributed to any other cause. Establishing a diagnosis was furthermore complicated by negative virological investigations of cerebrospinal fluid and non-pathological radiological findings. A polymerase chain reaction test of the child’s stool sample however tested positive for enterovirus and sequencing results revealed the presence of enterovirus A71. A previous history of febrile gastroenteritis just before the paresis started also supported the suspected diagnosis of enterovirus-associated acute flaccid paralysis. Following this, the child was treated with intravenous immunoglobulin over 5 days and a remarkable improvement was observed in the child’s paresis. Conclusion This case report describes a possible complication of enterovirus A71 infection in a child. It also highlights the prolonged detection of enterovirus in the child’s stool sample as compared with cerebrospinal fluid weeks after the primary infection occurred. Finally, it shows the need for increased clinical and diagnostic awareness especially in the management of sudden and unknown causes of paresis or paralysis in children.

In 2018, an upsurge in echovirus 30 (E30) infections was reported in Europe. We conducted a large-scale epidemiologic and evolutionary study of 1,329 E30 strains collected in 22 countries in Europe during 2016–2018. Most E30 cases affected persons 0–4 years of age (29%) and 25–34 years of age (27%). Sequences were divided into 6 genetic clades (G1–G6). Most (53%) sequences belonged to G1, followed by G6 (23%), G2 (17%), G4 (4%), G3 (0.3%), and G5 (0.2%). Each clade encompassed unique individual recombinant forms; G1 and G4 displayed >2 unique recombinant forms. Rapid turnover of new clades and recombinant forms occurred over time. Clades G1 and G6 dominated in 2018, suggesting the E30 upsurge was caused by emergence of 2 distinct clades circulating in Europe. Investigation into the mechanisms behind the rapid turnover of E30 is crucial for clarifying the epidemiology and evolution of these enterovirus infections.

Abstract Background Aseptic meningitis epidemics may pose various health care challenges. Methods We describe the German enterovirus meningitis epidemics in the university hospital centers of Düsseldorf, Cologne and Berlin between January 1st and December 31st, 2013 in order to scrutinize clinical differences from other aseptic meningitis cases. Results A total of 72 enterovirus (EV-positive) meningitis cases were detected in our multicenter cohort, corresponding to 5.8% of all EV-positive cases which were voluntarily reported within the National Enterovirus surveillance (EVSurv, based on investigation of patients with suspected aseptic meningitis/encephalitis and/or acute flaccid paralysis) by physicians within this period of time. Among these 72 patients, 38 (52.8%) were enterovirus positive and typed as echovirus (18 pediatric and 20 adult cases, median age 18.5 years; echovirus 18 (1), echovirus 2 (1), echovirus 30 (31), echovirus 33 (1), echovirus 9 (4)). At the same time, 45 aseptic meningitis cases in our cohort were excluded to be due to enteroviral infection (EV-negative). Three EV-negative patients were tested positive for varicella zoster virus (VZV) and 1 EV-negative patient for herpes simplex virus 2. Hospitalization was significantly longer in EV-negative cases. Cerebrospinal fluid analysis did not reveal significant differences between the two groups. After discharge, EV-meningitis resulted in significant burden of sick leave in our pediatric cohort as parents had to care for the children at home. Conclusions Voluntary syndromic surveillance, such as provided by the EVSurv in our study may be a valuable tool for epidemiological research. Our analyses suggest that EV-positive meningitis predominantly affects younger patients and may be associated with a rather benign clinical course, compared to EV-negative cases.

Abstract Background Echovirus (E) 30 (E-30) meningitis is characterized by neuroinflammation involving immune cell pleocytosis at the protective barriers of the central nervous system (CNS). In this context, infection of the blood-cerebrospinal fluid barrier (BCSFB), which has been demonstrated to be involved in enteroviral CNS pathogenesis, may affect the tight junction (TJ) and adherens junction (AJ) function and morphology. Methods We used an in vitro human choroid plexus epithelial (HIBCPP) cell model to investigate the effect of three clinical outbreak strains (13-311, 13-759, and 14-397) isolated in Germany in 2013, and compared them to E-30 Bastianni. Conducting transepithelial electrical resistance (TEER), paracellular dextran flux measurement, quantitative real-time polymerase chain reaction (qPCR), western blot, and immunofluorescence analysis, we investigated TJ and AJ function and morphology as well as strain-specific E-30 infection patterns. Additionally, transmission electron and focused ion beam microscopy electron microscopy (FIB-SEM) was used to evaluate the mode of leukocyte transmigration. Genome sequencing and phylogenetic analyses were performed to discriminate potential genetic differences among the outbreak strains. Results We observed a significant strain-dependent decrease in TEER with strains E-30 Bastianni and 13-311, whereas paracellular dextran flux was only affected by E-30 Bastianni. Despite strong similarities among the outbreak strains in replication characteristics and particle distribution, strain 13-311 was the only outbreak isolate revealing comparable disruptive effects on TJ (Zonula Occludens (ZO) 1 and occludin) and AJ (E-cadherin) morphology to E-30 Bastianni. Notwithstanding significant junctional alterations upon E-30 infection, we observed both para- and transcellular leukocyte migration across HIBCPP cells. Complete genome sequencing revealed differences between the strains analyzed, but no explicit correlation with the observed strain-dependent effects on HIBCPP cells was possible. Conclusion The findings revealed distinct E-30 strain-specific effects on barrier integrity and junctional morphology. Despite E-30-induced barrier alterations leukocyte trafficking did not exclusively occur via the paracellular route.

The major aim of the enterovirus surveillance (EVSurv) in Germany is to prove the absence of poliovirus circulation in the framework of the Global Polio Eradication Program (GPEI). Therefore, a free-of-charge enterovirus diagnostic is offered to all hospitals for patients with symptoms compatible with a polio infection. Within the quality proven laboratory network for enterovirus diagnostic (LaNED), stool and cerebrospinal fluid (CSF) samples from patients with suspected aseptic meningitis/encephalitis or acute flaccid paralysis (AFP) are screened for enterovirus (EV), typing is performed in all EV positive sample to exclude poliovirus infections. Since 2006, ≈200 hospitals from all 16 German federal states have participated annually. On average, 2500 samples (70% stool, 28% CSF) were tested every year. Overall, the majority of the patients studied are children

Parents are often uncertain about the vaccination status of their children. In times of vaccine hesitancy, vaccination programs could benefit from active patient participation. The Vaccination App (VAccApp) was developed by the Vienna Vaccine Safety Initiative, enabling parents to learn about the vaccination status of their children, including 25 different routine, special indication and travel vaccines listed in the WHO Immunization Certificate of Vaccination (WHO-ICV). Between 2012 and 2014, the VAccApp was validated in a hospital-based quality management program in Berlin, Germany, in collaboration with the Robert Koch Institute. Parents of 178 children were asked to transfer the immunization data of their children from the WHO-ICV into the VAccApp. The respective WHO-ICV was photocopied for independent, professional data entry (gold standard). Demonstrating the status quo in vaccine information reporting, a Recall Group of 278 parents underwent structured interviews for verbal immunization histories, without the respective WHO-ICV. Only 9% of the Recall Group were able to provide a complete vaccination status; on average 39% of the questions were answered correctly. Using the WHO-ICV with the help of the VAccApp resulted in 62% of parents providing a complete vaccination status; on average 95% of the questions were answered correctly. After using the VAccApp, parents were more likely to remember key aspects of the vaccination history. User-friendly mobile applications empower parents to take a closer look at the vaccination record, thereby taking an active role in providing accurate vaccination histories. Parents may become motivated to ask informed questions and to keep vaccinations up-to-date.

In 2014, sepsis-like illness affected 9 full-term newborns in 1 hospital in Austria. Although results of initial microbiological testing were negative, electron microscopy identified picornavirus. Archived serum samples and feces obtained after discharge were positive by PCR for human parechovirus 3. This infection should be included in differential diagnoses of sepsis-like illness in newborns.

Infectious and inflammatory diseases of the central nervous system are difficult to identify early. Case definitions for aseptic meningitis, encephalitis, myelitis, and acute disseminated encephalomyelitis (ADEM) are available, but rarely put to use. The VACC-Tool (Vienna Vaccine Safety Initiative Automated Case Classification-Tool) is a mobile application enabling immediate case ascertainment based on consensus criteria at the point-of-care. The VACC-Tool was validated in a quality management program in collaboration with the Robert-Koch-Institute. Results were compared to ICD-10 coding and retrospective analysis of electronic health records using the same case criteria. Of 68,921 patients attending the emergency room in 10/2010–06/2013, 11,575 were hospitalized, with 521 eligible patients (mean age: 7.6 years) entering the quality management program. Using the VACC-Tool at the point-of-care, 180/521 cases were classified successfully and 194/521 ruled out with certainty. Of the 180 confirmed cases, 116 had been missed by ICD-10 coding, 38 misclassified. By retrospective application of the same case criteria, 33 cases were missed. Encephalitis and ADEM cases were most likely missed or misclassified. The VACC-Tool enables physicians to ask the right questions at the right time, thereby classifying cases consistently and accurately, facilitating translational research. Future applications will alert physicians when additional diagnostic procedures are required.

In 2015, several provinces in Lao People's Democratic Republic (Lao PDR) experienced a vaccine-derived poliovirus outbreak. This survey was conducted (i) to evaluate the vaccination coverage in different settings and cohorts using the seroprevalence of anti-poliovirus (PV) antibodies as a surrogate measure, and (ii) to explore the usefulness of an ELISA in a country with limited resources and a specific epidemiological setting. IgG antibodies were assessed by ELISA in Lao children (n = 1216) and adults (n = 1228), including blood donors and health care workers. Protective antibody titers against the 3 vaccine serotypes were determined by microneutralization (VNT) in a subset of participants. More than 92% of the children had anti-poliovirus antibodies, regardless of nutritional status or access to health care, highlighting the success of the vaccination outreach activities in the country. In contrast, anti-poliovirus seroprevalence reached only 81.7% in blood donors and 71.9% in health care workers. Participants born before the introduction of poliovirus vaccination in Lao PDR were considerably less likely to be seropositive. These findings align with the epidemiology of the outbreak. Neutralizing antibodies against at least one of the 3 poliovirus serotypes were detected in all children (99/99) and 93/99 had antibodies against all serotypes. Similarly, all health care workers had neutralizing antibodies against at least one and 92/99 against all serotypes. The comparison of both assays shows an acceptable underestimation of vaccine coverage in children by ELISA, but a low sensitivity of the ELISA in the adults. We show that the ELISA is a reasonable alternative to the VNT in particular in vaccinated children, that an improved version should be serotype specific, and that negativity thresholds should be revisited for optimal sensitivity and specificity. Thus, polio-free countries with an uncertain vaccination coverage and limited laboratory capacity, that are at risk of vaccine-derived poliovirus outbreaks or of re-importation of wild poliovirus may benefit from an improved ELISA for cohort studies to evaluate their immunization program in children.

We used physician sentinel surveillance to identify 25 (7.7%) mild to severe infections with enterovirus D68 (EV-D68) in children and adults among 325 outpatients with acute respiratory infections in Germany during August–October 2014. Results suggested low-level circulation of enterovirus D68 in Germany. Viruses were characterized by sequencing viral protein (VP) 1 and VP4/VP2 genomic regions.

Human enteroviruses and human parechoviruses are associated with a broad range of diseases and even severe and fatal conditions. For human cosaviruses, the etiological role is yet unknown. Little is known about the circulation of non-polio enteroviruses, human parechoviruses, and human cosaviruses in Nigeria. A total of 113 stool samples were collected from healthy individuals in Osun State between February 2016 and May 2017. RT-PCR assays targeting the 5′ non-coding region (5′ -NCR) were used to screen for human enteroviruses, human parechoviruses, and human cosaviruses. For human enteroviruses, species-specific RT-PCR assays targeting the VP1 regions were used for molecular typing. Inoculation was carried out on RD-A, CaCo-2, HEp-2C, and L20B cell lines to compare molecular and virological assays. Ten samples tested positive for enterovirus RNA with 11 strains detected, including CV-A13 (n = 3), E-18 (n = 2), CV-A20 (n = 1), CV-A24 (n = 1), EV-C99 (n = 1), and EV-C116 (n = 2). Three samples tested positive for human parechovirus RNA, and full genome sequencing on two samples allowed assignment to a new Parechovirus A type (HPeV-19). Thirty-three samples tested positive for cosavirus with assignment to species Cosavirus D and Cosavirus A based on the 5′-NCR region. Screening of stool samples collected from healthy individuals in Nigeria in 2016 and 2017 revealed a high diversity of circulating human enteroviruses, human parechoviruses, and human cosaviruses. Molecular assays for genotyping showed substantial benefits compared with those of cell-culture assays.

Background Human enteroviruses (EVs) and parechoviruses (HPeVs) belong to the family Picornaviridae. Although most EV and HPeV infections remain asymptomatic, both pathogens can cause a wide spectrum of clinical manifestations ranging from respiratory or gastrointestinal symptoms to myocarditis, neonatal sepsis, and infections of the central nervous system. Objectives Aim of the present study was to investigate the spectrum of EVs and HPeVs in apparently healthy adults and children living in the South of Côte d’Ivoire. Study design The study included 105 stool samples obtained from healthy individuals aged 0–53 years between June 2013 and December 2014 in the Sud-Como region of Côte d’Ivoire. After collection and shipment to Germany, the samples were analyzed by real-time PCR for the presence of EVs and HPeVs RNA. Molecular typing and virus isolation of all samples were performed.’’é Results Out of 105 samples, 24 (22.8%) were EV positive and six (5.2%) were HPeV positive. Twenty-one EV positive samples could be characterized with serotypes belonging to EV group A–C, while three could not be further specified. Interestingly, several rarely described serotypes were identified, e.g., EV-C99, EV-B93, EV-C116, and EV-A119. Typing of HPeV positive samples resulted in HPeV-1 and -5 detections, while one isolate could not be assigned to the known HPeV types. Conclusions This study showed a large variety of EV strains in healthy people in the South of Côte d’Ivoire and provided the first available data about HPeV infections in a sub-Saharan African country. [ABSTRACT FROM AUTHOR]

Background: Human enteroviruses (EVs) and parechoviruses (HPeVs) belong to the family Picornaviridae. Although most EV and HPeV infections remain asymptomatic, both pathogens can cause a wide spectrum of clinical manifestations ranging from respiratory or gastrointestinal symptoms to myocarditis, neonatal sepsis, and infections of the central nervous system., Objectives: Aim of the present study was to investigate the spectrum of EVs and HPeVs in apparently healthy adults and children living in the South of Côte d'Ivoire., Study Design: The study included 105 stool samples obtained from healthy individuals aged 0-53 years between June 2013 and December 2014 in the Sud-Como region of Côte d'Ivoire. After collection and shipment to Germany, the samples were analyzed by real-time PCR for the presence of EVs and HPeVs RNA. Molecular typing and virus isolation of all samples were performed.''é, Results: Out of 105 samples, 24 (22.8%) were EV positive and six (5.2%) were HPeV positive. Twenty-one EV positive samples could be characterized with serotypes belonging to EV group A-C, while three could not be further specified. Interestingly, several rarely described serotypes were identified, e.g., EV-C99, EV-B93, EV-C116, and EV-A119. Typing of HPeV positive samples resulted in HPeV-1 and -5 detections, while one isolate could not be assigned to the known HPeV types., Conclusions: This study showed a large variety of EV strains in healthy people in the South of Côte d'Ivoire and provided the first available data about HPeV infections in a sub-Saharan African country., (Copyright © 2015 Elsevier B.V. All rights reserved.)