Rotavirus Research Articles

The Role of the VP4 Attachment Protein in Rotavirus Host Range Restriction in an In Vivo Suckling Mouse Model.

The basis for rotavirus (RV) host range restriction (HRR) is not fully understood but is likely multigenic. RV genes encoding VP3, VP4, NSP1, NSP2, NSP3, and NSP4 have been associated with HRR in various studies. With the exception of NSP1, little is known about the relative contribution of the other RV genes to HRR. VP4 has been linked to HRR because it functions as the RV cell attachment protein, but its actual role in HRR has not been fully assessed. We generated a collection of recombinant RVs (rRVs) in an isogenic murine-like RV genetic background, harboring either heterologous or homologous VP4 genes from simian, bovine, porcine, human, and murine RV strains, and characterized these rRVs in vitro and in vivo. We found that a murine-like rRV encoding a simian VP4 was shed, spread to uninoculated littermates, and induced diarrhea comparably to rRV harboring a murine VP4. However, rRVs carrying VP4s from both bovine and porcine RVs had reduced diarrhea, but no change in fecal shedding was observed. Both diarrhea and shedding were reduced when VP4 originated from a human RV strain. rRVs harboring VP4s from human or bovine RVs did not transmit to uninoculated littermates. We also generated two rRVs harboring reciprocal chimeric murine or bovine VP4. Both chimeras replicated and caused disease as efficiently as the parental strain with a fully murine VP4. These data suggest that the genetic origin of VP4 partially modulates HRR in the suckling mouse and that both the VP8* and VP5* domains independently contribute to pathogenesis and transmission. IMPORTANCE Human group A rotaviruses (RVs) remain the most important cause of severe acute gastroenteritis among infants and young children worldwide despite the introduction of several safe and effective live attenuated vaccines. The lack of knowledge regarding fundamental aspects of RV biology, such as the genetic basis of host range restriction (HRR), has made it difficult to predictively and efficiently design improved, next-generation live attenuated rotavirus vaccines. Here, we engineered a collection of VP4 monoreassortant RVs to systematically explore the role of VP4 in replication, pathogenicity, and spread, as measures of HRR, in a suckling mouse model. The genetic and mechanistic bases of HRR have substantial clinical relevance given that this restriction forms the basis of attenuation for several replication-competent human RV vaccines. In addition, a better understanding of RV pathogenesis and the determinants of RV spread is likely to enhance our ability to improve antiviral drug and therapy development.

Evaluation of diagnostic methods for the Detection of Bovine Coronavirus and Rotavirus in feces of diarrheic calves

The purpose of the present study was to evaluate the efficacy of Enzyme-Linked Immunosorbent Assay (ELISA), immunochromatographic (ICG), and reverse transcription-polymerase chain reaction (RT-PCR) methods for the detection of rotavirus (RV) and bovine coronavirus (BCV). Feces samples were collected from 90 diarrhoeic calves (male and female) up to one month of age and the immune response against RV and BCV infection was assessed by using ELISA (Ag and Ab), ICG, and RT-PCR. To determine the performance and accuracy of each diagnostic method in comparison to the diagnostic gold standard (RT-PCR) method, different statistical tests including receiver operating characteristic curve (ROC), concordance correlation were used. Results shown the prevalence of RV and BCV and RV+BCV according to RT-PCR were equal to 8.89 (95% CI: 6.64-10.07), 14.44 (95% CI: 11.23-6.90), and 2.22 (95% CI: 0.89-3.72), respectively. The best agreement and the highest sensitivity and specificity were obtained between the RT-PCR and AgELISA (100% and 94.3%), and then AbELISA (100% and 94.3%) was the second-best test and also the ICG test (95% and 94.3%) was less accurate method in comparison to ELISA methods for identifying RV and BCV, but a good correlation and concordance between ICG diagnostic techniques and RT-PCR were observed. To put it in a nutshell, our results suggest that the ICG assay may improve the ability to diagnose calves RV and BCV infections accurately and quickly. Promoting rapid IGG kit with higher accuracy in early diagnosis of the cause of diarrhoea plays an important role in its therapeutic regimes, management protocols, and control procedures, but ELISA is preferred due to more precise results.

Genomic and evolutionary characteristics of G3P[8] group a rotavirus strains in China, 2016 to 2018.

Rotavirus A (RVA) G3P[8] is sporadically detected in China, although G9P[8] predominates. To evaluate their genetic composition at the whole-genome level, 24 G3P[8] RVA strains isolated from children under five years were sequenced and characterized. The 24 strains were genotyped as G3-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1, indicating the Wa-like genotype constellation. A maximum clade credibility (MCC) tree for VP7 indicated that G3 had an estimated mean evolutionary rate of 7.279×10-4 substitutions/site/year; thus, 3-5years would pass from the generation of an ancestor virus to the epidemic spread of that virus throughout China. Considering the ongoing prevalence as well as rapid evolution, it is important to monitor G3P[8] RVA epidemics; continuous nationwide surveillance is essential.

Evaluation of Virucidal Quantitative Carrier Test towards Bovine Viruses for Surface Disinfectants While Simulating Practical Usage on Livestock Farms.

Livestock farming is affected by the occurrence of infectious diseases, but outbreaks can be prevented by effective cleaning and disinfection along with proper farm management. In the present study, bovine coronavirus (BCoV) and bovine rotavirus A (RVA) were inactivated using food additive-grade calcium hydroxide (FdCa(OH)2) solution, quaternary ammonium compound (QAC) and their mixture through suspension tests as the primary screening, and afterward via carrier tests using dropping or dipping techniques as the secondary screenings. Viruses in the aqueous phase can be easily inactivated in the suspension tests, but once attached to the materials, they can become resistant to disinfectants, and require longer times to be inactivated. This highlights the importance of thorough cleaning with detergent before disinfection, and keeping elevated contact durations of proper disinfectants to reduce viral contamination and decrease infectious diseases incidence in farms. It was also reaffirmed that the suspension and carrier tests are necessary to evaluate disinfectants and thus determine their actual use. Particularly, the mixture of QAC and FdCa(OH)2 was found to exhibit synergistic and broad-spectrum effects compared to their use alone, and is now recommended for use on livestock farms.

Establishment of a reverse genetics system for avian rotavirus A strain PO-13

Avian rotavirus A (RVA) is one of major enteric pathogens that cause diarrhoea in young avian individuals. Importantly, some of the avian RVA strains of G18P[17] genotype are naturally transmitted to and cause clinical diseases in mammalian species, indicating their potential risks to animal health. Although molecular information on the pathogenesis by avian RVA strains will be useful for estimating their risks, the absence of a reverse genetics (RG) system for these strains has hindered the elucidation of their pathogenic mechanisms. In this study, we aimed to establish an RG system for the avian G18P[17] prototype strain PO-13, which was isolated from a pigeon in Japan in 1983 and was experimentally shown to be pathogenic in suckling mice. Transfection with plasmids expressing 11 genomic RNA segments of the strain resulted in rescue of the infectious virus with an artificially introduced genetic marker on its genome, indicating that an RG system for the PO-13 strain was successfully established. The rescued recombinant strain rPO-13 had biological properties almost identical to those of its wild-type strain (wtPO-13). Notably, both rPO-13 and wtPO-13 induced diarrhoea in suckling mice with similar efficiencies. It was thus demonstrated that the RG system will be useful for elucidating the pathogenic mechanisms of the PO-13 strain at the molecular level. This is the first report of the establishment of an RG system for an avian RVA strain.

Identification of Pulmonary Infections With Porcine Rotavirus A in Pigs With Respiratory Disease.

While rotavirus (RV) is primarily known to cause gastroenteritis in many animals, several epidemiological studies have shown concurrent respiratory symptoms with fecal and nasal virus shedding. However, respiratory RV infections have rarely been investigated. By screening clinical samples submitted for diagnostic testing, porcine rotavirus A (RVA) was detected by quantitative reverse transcription PCR (qRT-PCR) in 28 out of 91 (30.8%) lungs obtained from conventionally reared pigs with respiratory signs. Among the positive cases, intensive RVA signals were mainly localized in alveolar macrophages (n = 3) and bronchiolar epithelial cells (n = 1) by RNAscope® in situ hybridization (ISH). The signals of RVA in bronchiolar epithelial cells were verified by ISH with different probes, immunohistochemistry, and transmission electron microscopy. Furthermore, additional cases with RVA ISH-positive signals in alveolar macrophages (n = 9) and bronchial epithelial cells (n = 1) were identified by screening 120 archived formalin-fixed and paraffin-embedded lung samples using tissue microarrays. Overall, our study showed a high frequency of RVA detection in lungs from conventional pigs with respiratory disease. Further research is needed to determine if RVA infection in the respiratory epithelium correlates with nasal shedding of rotavirus and its contribution to respiratory disease.

Binding, recovery, and infectiousness of enveloped and non-enveloped viruses associated with plastic pollution in surface water

Microplastics in wastewater and surface water rapidly become colonised by microbial biofilm. Such ‘plastisphere’ communities are hypothesised to persist longer and be disseminated further in the environment and may act as a vector for human pathogens, particularly as microplastics entering wastewater treatment plants are exposed to high concentrations of pathogenic bacteria. However, the potential for human viral pathogens to become associated with the plastisphere has never before been quantified. Here, we have used rotavirus (RV) SA11 (a non-enveloped enteric virus) and the enveloped bacteriophage Phi6 as model viruses to quantify binding and recovery from biofilm-colonised microplastic pellets in three different water treatments (filtered and non-filtered surface water, and surface water with added nutrients). Viruses associated with biofilm-colonised pellets were more stable compared to those remaining in the water. While infectious particles and genome copies of RV remained stable over the 48 h sampling period, Phi6 stability was highly impacted, with a reduction ranging from 2.18 to 3.94 log10. Virus particles were protected against inactivation factors when associated with the biofilm on microplastic surfaces, and when there was a high concentration of particulate matter in the liquid phase. Although our results suggest that the presence of an envelope may limit virus interaction with the plastisphere, the ability to recover both enveloped and non-enveloped infectious viruses from colonised microplastic pellets highlights an additional potential public health risk of surface waters becoming contaminated with microplastics, and subsequent human exposure to microplastics in the environment.

KNOWLEDGE AND PRACTICE REGARDING ORAL REHYDRATION SALTS (ORS) SOLUTION USE IN DIARRHEAL DISEASE AMONG MOTHERS OF UNDER-FIVE CHILDREN IN RURAL WEST BENGAL: A HOSPITAL BASED CROSS SECTIONAL STUDY

Childhood diarrhea is one of the major public health problems. Diarrhea is the second most common cause of mortality in under-ve children. Use of Oral rehydration salts (ORS) solution is the most cost-effective intervention to prevent diarrhea related mortality and morbidity. Aims And Objectives: To evaluate the knowledge and practice towards ORS use in diarrheal disease among mothers of under-ve children in rural West Bengal. This hospital based cross sectional study was carried out in pediatrics department of BSMCH, Bankura. Data aboutMethodology: demography, knowledge and practice regarding diarrheal management and ORS use were collected with predesigned semi structured questionnaire from 300 mothers of under-ve children who have suffered at least once from diarrhea in last one month. DemographicResults: ndings indicated 54.3% mothers belonged to age of 18-25 years while illiterate and primary level educated mothers constituted 35% and 28% of study sample, respectively. Though 87% mothers heard about ORS but only 68.7% mothers considered ORS as remedy of diarrhea. Knowledge about preparation (42%), initiation (43%), amount (39%), timing of use (35.3%) and storage of ORS (22%) were unsatisfactory. Furthermore, majority had no clue regarding Zinc (9.6%) and Rota virus vaccination (13.3). Age and education of the mother had statistically signicant (P< 0.05) association with knowledge and practice. In our study, mothers had lacuna in their knowledge about proper use of ORS.Conclusion: Successful treatment of diarrhea depends on mother's knowledge and practices. Health Education should be aimed at rectifying those inadequacies to reduce further mortality and morbidity

Impact of maternal and pre-existing antibodies on immunogenicity of inactivated rotavirus vaccines

Rotavirus (RV) is a major pathogen causing severe diarrhea in infants and children aged less than 5 years. Vaccination is an economically feasible and effective strategy to prevent rotavirus infections. However, immune efficacy of live vaccines could be interfered by maternal antibodies and pre-existing antibodies of children. To develop an inactivated rotavirus vaccine (IRV), we had previously isolated a wild-type human rotavirus strain ZTR-68-A (G1P[8]) from the fecal samples of infants having severe diarrhea in a region endemic for the presence of this pathogen. In our present study, we assessed whether the presence of maternal and pre-existing antibodies in newborn BALB/c mice affected the immunogenicity of IRV administered to these animals. Our results indicate that maternal antibodies, generated from either vaccine immunization or rotavirus infection, showed partial influence with the immune responses generated by two doses of IRV vaccination. Increasing the number of immunizations can significantly improve the titer of serum neutralizing antibody and a seroconversion rate of up to 100%. In newborn mice, single-virus infection did not elicit detectable levels of serum neutralizing antibodies. After an IRV vaccination, the immune responses of these mice remained unaffected, with no significant differences in titers compared with those of control-group mice. In summary, choosing a suitable immunization dose and dosing frequency is essential for the immune effectiveness of IRV. The results of this study will provide animal experimental support for the IRV clinical research in future.

Routine Immunization Programs for Children during the COVID-19 Pandemic in Ecuador, 2020-Hidden Effects, Predictable Consequences.

The COVID-19 pandemic has led to a global disruption of several services, including routine immunizations. This effect has been described in several countries, but there are few detailed studies in Latin America and no reports in Ecuador. Therefore, this work aims to quantify the reduction in routine immunizations for infants during the 2020 COVID-19 pandemic in Ecuador. 2018, 2019, and 2020 data were obtained from the Ministry of Health, Ecuador. The number of doses and the extent of immunization coverage was descriptively compared for four vaccines: rotavirus (ROTA), poliovirus (PV), pneumococcal (PCV), and pentavalent (PENTA) vaccines. There was no significant difference in doses applied during the 2018 and 2019 years. However, a significant (p < 0.05) drop of 137,000 delivered doses was observed in 2020 compared to the pre-pandemic years. Reductions in the percentage of coverage were more pronounced for the PENTA vaccine (17.7%), followed by PV (16.4%), ROTA (12%), and PCV vaccines (10.7%). Spatial analysis shows a severe impact on vaccination coverage on provinces from the Coast and Highland regions of the country. The pandemic has significantly impacted the immunization programs for infants across Ecuador. This retrospective analysis shows an urgent need to protect vulnerable zones and populations during public health emergencies.

Frequent Occurrence of Simultaneous Infection with Multiple Rotaviruses in Swiss Pigs.

Rotavirus (RV) infections are the most important viral cause of diarrhea in piglets in Switzerland and are thought to cause substantial economic losses to the pig industry. However, no data are available on the occurrence and dynamics of the main porcine RV species, namely RVA, RVB, and RVC, and the diversity of the circulating strains. We therefore tested fecal samples from a cross-sectional (n = 95) and a longitudinal (n = 48) study for RVA, RVB, and RVC by real-time RT-PCR and compared the results of the cross-sectional study to postmortem findings. In addition, eight samples were fully genotyped by using next-generation sequencing. In the cross-sectional study, triple RV infections significantly correlated with diarrhea and wasting and were most frequent in the weaned age group. In the longitudinal study, the shedding of RV peaked one week after weaning and decreased thereafter. Here, mainly double infections were seen, and only a few animals showed diarrhea. The full-genome sequencing revealed a genotype pattern similar to other European countries and, importantly, co-infection by up to four RVA strains. Our results imply that the weaning of piglets may trigger not only RV shedding but facilitate co-infection of multiple RV species and strains in the same host.

A Galactooligosaccharide Product Decreases the Rotavirus Infection in Suckling Rats.

The leading cause of gastroenteritis among young children worldwide is the Group A rotaviruses (RV), which produce a wide range of symptoms, from a limited diarrhea to severe dehydration and even death. After an RV infection, immunity is not complete and less severe re-infections usually occur. These infections could be ameliorated by nutritional interventions with bioactive compounds, such as prebiotics. The aim of this research was to study the impact of a particular galactooligosaccharide (B-GOS) on the RV symptomatology and immune response during two consecutive infections. Lewis neonatal rats were inoculated with SA11 (first RV infection) on day 6 of life and with EDIM (second RV infection) on day 17 of life. B-GOS group was administered by oral gavage with a daily dose of B-GOS between days three to nine of life. Clinical and immunological variables were assessed during both infective processes. In the first infection, after the prebiotic intervention with B-GOS, a lower incidence, duration, and overall severity of the diarrhea (p < 0.05) was observed. In addition, it improved another severity indicator, the fecal weight output, during the diarrhea period (p < 0.05). The second RV infection failed in provoking diarrhea in the groups studied. The immune response during first infection with SA11 was not affected by B-GOS administration and had no impact on second infection, but the prebiotic intervention significantly increased IFN-γ and TNF-α intestinal production after the second infection (p < 0.05). In summary, B-GOS supplementation is able to reduce the incidence and severity of the RV-associated diarrhea and to influence the immune response against RV infections.

In Vitro Inactivation of Respiratory Viruses and Rotavirus by the Oral Probiotic Strain Weissella cibaria CMS1

Weissella cibaria CMS1 (oraCMS1) has been commercially used in Korea as an oral care probiotic for several years. Human respiratory syncytial virus (RSV) and the influenza A virus (H1N1) are representative viruses that cause infantile lower respiratory tract infections. Rotavirus A (RVA) is the most common cause of diarrhea in infants and young children. Here, we aimed to evaluate the efficacy of the cell-free supernatant (CFS) of oraCMS1 in inactivating RSV, H1N1, and RVA in suspension as per ASTM (American Society for Testing and Materials) E1052-20. The mixture of oraCMS1 and these viruses was evaluated at contact times of 1, 2, and 4 h. Virucidal activity was measured using a 50% tissue culture infective dose assay (log10TCID50) after infecting the host cells with the viruses. The CFS of oraCMS1 inactivated RSV by up to 99.0% after 1 h and 99.9% after 2 and 4 h, and H1N1 and RVA were inactivated by up to 99.9% and 99.0% at 2 h, respectively. Although these in vitro results cannot be directly interpreted as implying clinical efficacy, our findings suggest that oraCMS1 provides a protective barrier against RSV, H1N1, and RVA, and therefore, it can help decrease the risk of respiratory tract and intestinal infections.

Association of Anti-Rotavirus IgA Seroconversion with Growth, Environmental Enteric Dysfunction and Enteropathogens in Rural Pakistani Infants

BackgroundThe underperformance of oral vaccines in children of low- and middle-income countries is partly attributable to underlying environmental enteric dysfunction (EED). MethodologyWe conducted a longitudinal, community-based study to evaluate the association of oral rotavirus vaccine (Rotarix®) seroconversion with growth anthropometrics, EED biomarkers and intestinal enteropathogens in Pakistani infants. Children were enrolled between three to six months of their age based on their nutritional status. We measured serum anti-rotavirus immunoglobulin A (IgA) at enrollment and nine months of age with EED biomarkers and intestinal enteropathogens. ResultsA total of 391 infants received two doses of rotavirus (RV) vaccine. 331/391 provided paired blood samples. Of these 331 children, 45% seroconverted at 9 months of age, 35% did not seroconvert and 20% were seropositive at baseline. Non-seroconverted children were more likely to be stunted, wasted and underweight at enrollment. In univariate analysis, insulin-like growth factor (IGF) concentration at 6 months were higher in seroconverters, median (25th, 75th percentile): 26.3 (16.5, 43.5) ng/ml vs. 22.5 (13.6, 36.3) ng/ml for non-seroconverters, p-value = 0.024. At nine months, fecal myeloperoxidase (MPO) concentrations were significantly lower in seroconverters, 3050(1250, 7587) ng/ml vs. 4623.3 (2189, 11650) ng/ml in non-seroconverted children, p-value = 0.017. In multivariable logistic regression analysis, alpha-1 acid glycoprotein (AGP) and IGF-1 concentrations were positively associated with seroconversion at six months. The presence of sapovirus and rotavirus in fecal samples at the time of rotavirus administration, was associated with non-seroconversion and seroconversion, respectively. ConclusionWe detected high baseline RV seropositivity and impaired RV vaccine immunogenicity in this high-risk group of children. Healthy growth, serum IGF-1 and AGP, and fecal shedding of rotavirus were positively associated with RV IgA seroconversion following immunization, whereas the presence of sapovirus was more common in non-seroconverters.Trial registration: Clinical Trials ID: NCT03588013.

Rotaviruses A and C in dairy cattle in the state of Rio de Janeiro, Brazil.

Stool samples were collected from calves from nine family-based small dairy farms in the state of Rio de Janeiro, for detection and characterization of rotavirus (RV) species A, B, and C (RVA, RVB, and RVC, respectively) by reverse transcription polymerase chain reaction. Twenty-six samples (27.7%) were positive for at least one of the species: 22 (23.4%) samples were positive only for RVA, 3 (3.2%) were positive for RVC, and one sample (1.1%) had co-infection of RVA and RVC. RVB was not detected. Seven (21.9%; n = 32) animals with diarrhea and 19 (30.1% n = 62) asymptomatic animals were positive, with no significant difference in positivity (p = 0.3677). RV was detected in all properties studied, at rates between 14.3 and 80%, demonstrating the widespread circulation of RV in four of the seven geographic regions of the state of Rio de Janeiro. Infection was more prevalent among animals ≤ 6months of age. Sequence analysis of a portion of the RVA VP6-encoding gene identified the I2 genotype. RVC was also detected; to our knowledge, this is the first description of this agent in cattle in Brazil. The data presented here should add knowledge regarding the importance and prevalence of RV in our national territory, and may facilitate the planning and implementation of control and prevention measures for bovine rotavirus infections in Brazil.

Determination and Genotype Distribution of Rotavirus Gastroenteritis in Pediatric Patients Admitted to a Tertiary Care Hospital

Acute infectious gastroenteritis is a prevalent disease worldwide, and most of the cases are caused by viral pathogens. Many different viruses, including Rotavirus (RV), Norovirus, Adenovirus, and Astroviruses, are responsible for most acute viral gastroenteritis cases. According to the Centers for Disease Control and Prevention (CDC), viral gastroenteritis infections cause more than 200 000 child mortalities each year worldwide. One of the best strategies to reduce the global burden of RV gastroenteritis is the development and administration of effective vaccines. However, since there are differences in the coverage of the vaccines, the choice of appropriate vaccine for localized genotypes based on regions is important. The aim of this study was to detect the RV infections in our region and to perform genotyping using real time polymerase chain reaction (Rt-PCR) and high resolution melting (HRM) analysis. A total of 341 stool samples collected from pediatric patients were tested. Lateral flow immunochromatographic assay principle based assay was used for antigen detection. RT-PCR and HRM were applied for genotype analysis. Similar to the data from our country and Eastern Mediterranean region, RV positivity in stool samples was 23.1%. The majority of the patients (51%) were aged 0-2 years and the vast majority of the patients, with a rate of 77%, were between the ages of 0-5. Most of the cases were detected in the winter months, especially in February. The distribution of 40 samples, whose G genotype could be detected, was as follows: G2, 21 (52.5%); G1, 11 (27.5%); G9, 5 (12.5%); G3, 2 (5%); G4, 1 (2.5%). The distribution of 53 samples, whose P genotype could be detected, was as follows: P4, 44 (83.0%); P9, 8 (15.1%); P10, 1 (1.9%). Among those whose genotype could be detected, the most prevalent genotypes were G2 with 52.5% and P4 with 83%. When the distribution of 25 samples was evaluated, in which RV G and P genotypes were detected simultaneously, G1P [4] 11 (44%), G2P[9] 5 (20%), G9P[4] 5 (20%), G2P[4] 2 (8%), G3P[10] 1 (4%), and G4P[4] 1 (4%) genotypes were determined, respectively. The most commonly observed genotype was G1P[4]. In the HRM analysis, it was observed that the melting curve peaks were at different temperatures in nine of the G2 genotype samples and 16 of the P4 and P9 genotype samples. Thus, genotyping with HRM analysis could not be fully finalized, especially for G2 and P. Of the Rota Teq® and RotarixTM vaccines administered on demand in our country, Rota Teq® is considered the vaccine that has the widest coverage for the genotypes observed in our country and region. ROTASIIL® vaccine, which covers all the genotypes in our region (G1, G2, G3, G4, G9) is not available in our country. The emergence of the strains with the potential to increase the current burden of RV disease should be continuously monitored, as different results are obtained by region and year, even within the same country. Thus, the emergence of vaccine-resistant strains can be followed up, especially in countries with higher viral diversity.

Clinical characteristics, health care resource utilization and direct medical costs of Rotavirus hospitalizations in Spain (2013–2018)

ABSTRACT Rotavirus (RV) is the most common cause of severe gastroenteritis (GE) in infants and young children worldwide and is associated with a significant clinical and economic burden. The objective of this study was to analyze the characteristics, healthcare resource utilization and the direct medical costs related to RVGE hospitalizations in Spain. An observational, multicenter, cross-sectional study was conducted from June 2013 to May 2018 at the pediatric departments of 12 hospitals from different Spanish regions. Children under 5 years of age admitted to the hospital with a confirmed diagnosis of RVGE were selected. Data on clinical characteristics, healthcare resource use and costs were collected from patient records and hospital databases. Most children hospitalized for RVGE did not have any previous medical condition or chronic disease. Forty-seven percent had previously visited the Emergency Room (ER), 27% had visited a primary care pediatrician, and 15% had received pharmacological treatment prior to hospital admission due to an RVGE episode. The average length of a hospital stay for RVGE was 5.6 days, and the mean medical costs of RVGE hospitalizations per episode ranged from 3,940€ to 4,100€. The highest direct medical cost was due to the hospital stay. This study showed a high burden of health resource utilization and costs related to the management of cases of RVGE requiring hospitalization. RV vaccination with high coverage rates should be considered to minimize the clinical and economic impacts of this disease on the health-care system.

Protective effects of the postbiotic deriving from cow\u2019s milk fermentation with L. paracasei CBA L74 against Rotavirus infection in human enterocytes

Rotavirus (RV) is the leading cause of acute gastroenteritis-associated mortality in early childhood. Emerging clinical evidence suggest the efficacy of the postbiotic approach based on cow’s milk fermentation with the probiotic Lacticaseibacillus paracasei CBAL74 (FM-CBAL74) in preventing pediatric acute gastroenteritis, but the mechanisms of action are still poorly characterized. We evaluated the protective action of FM-CBAL74 in an in vitro model of RV infection in human enterocytes. The number of infected cells together with the relevant aspects of RV infection were assessed: epithelial barrier damage (tight-junction proteins and transepithelial electrical resistance evaluation), and inflammation (reactive oxygen species, pro-inflammatory cytokines IL-6, IL-8 and TNF-α, and mitogen-activated protein kinase pathway activation). Pre-incubation with FM-CBA L74 resulted in an inhibition of epithelial barrier damage and inflammation mediated by mitogen-activated protein kinase pathway activation induced by RV infection. Modulating several protective mechanisms, the postbiotic FM-CBAL74 exerted a preventive action against RV infection. This approach could be a disrupting nutritional strategy against one of the most common killers for the pediatric age.

Assessing the Epidemiology of Rotavirus A, B, C and H in Diarrheic Pigs of Different Ages in Northern Italy.

Rotaviruses are classified in 10 groups (A to J), where rotavirus A (RVA) is the major cause of diarrhea in humans and animals. With some exceptions, there is scarce information on the epidemiology of non-A rotaviruses in human and animal hosts. Currently, five species (A, B, C, E and H) have been identified in pigs. In the present study we investigated the prevalence of RVA, RVB, RVC and RVH among diarrheic pigs of different ages, in different seasons and in the presence of co-infections. Two molecular assays were developed for the detection of porcine RVA, RVB, RVC and RVH and were used to screen 962 stool specimens from suckling, weaning and fattening pigs with acute enteritis. Overall, rotaviruses were detected in a high percentage of samples (78%), with RVA being predominant (53%), followed by RVC (45%), RVB (43%) and RVH (14%). RVA was more common in the suckling (58%) and weaning cohorts (64%), while RVB, RVC and RVH were also frequently detected in fattening pigs. Only RVA and RVB infections followed a seasonal trend and exhibited age-related differences. Rotavirus infections were frequently present in combination with other pathogens. The present study depicts a portrait of rich rotavirus diversity in porcine herds, identifying seasonal and age-related patterns of circulation of the different rotavirus species in the surveyed areas.

Development of a Multiplex RT-PCR Assay for Simultaneous Detection of Four Potential Zoonotic Swine RNA Viruses.

Swine viruses like porcine sapovirus (SaV), porcine encephalomyocarditis virus (EMCV), porcine rotavirus A (RVA) and porcine astroviruses (AstV) are potentially zoonotic viruses or suspected of potential zoonosis. These viruses have been detected in pigs with or without clinical signs and often occur as coinfections. Despite the potential public health risks, no assay for detecting them all at once has been developed. Hence, in this study, a multiplex RT-PCR (mRT-PCR) assay was developed for the simultaneous detection of SaV, EMCV, RVA and AstV from swine fecal samples. The PCR parameters were optimized using specific primers for each target virus. The assay’s sensitivity, specificity, reproducibility, and application to field samples have been evaluated. Using a pool of plasmids containing the respective viral target fragments as a template, the developed mRT-PCR successfully detected 2.5 × 103 copies of each target virus. The assay’s specificity was tested using six other swine viruses as a template and did not show any cross-reactivity. A total of 280 field samples were tested with the developed mRT-PCR assay. Positive rates for SaV, EMCV, RVA, and AstV were found to be 24.6% (69/280), 5% (14/280), 4.3% (12/280), and 17.5% (49/280), respectively. Compared to performing separate assays for each virus, this mRT-PCR assay is a simple, rapid, and cost-effective method for detecting mixed or single infections of SaV, EMCV, RVA, and AstV.