Antimicrobial Susceptibility and Characterization of Extended-Spectrum ß-Lactamase-Producing Escherichia coli Isolated from Stools of Primary Healthcare Patients in Ethiopia.

Antimicrobial resistance of Escherichia coli is a growing problem in both developed and developing countries. This study aimed to investigate the phenotypic antimicrobial resistance of E. coli isolates (n = 260) isolated from the stool specimen of patients attending public health facilities in Addis Ababa and Hossana. This study also aimed to characterize phenotypically confirmed extended-spectrum beta-lactamase (ESBL)-producing E. coli isolates (n = 22) using whole-genome sequencing. Resistance to 18 different antimicrobials was assessed using the disc diffusion method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) guidelines. The highest resistance rate among the E. coli isolates was found for ampicillin (52.7%), followed by trimethoprim-sulfamethoxazole (29.6%). Of all isolates, 50 (19.2%) were multidrug-resistant and 22 (8.5%) were ESBL producers. ESBL genes were detected in 94.7% of the sequenced E. coli isolates, and multiple ß-lactamase genes were detected in 57.9% of the isolates. The predominant ESBL gene identified was blaCTX-M-15 (78.9%). The blaTEM-1B gene was detected in combination with other ESBL genes in 57.9% of the isolates, while only one of the sequenced isolates contained the blaTEM-1B gene alone. The blaCTX-M-3 gene was detected in three isolates. The genes blaCTX-M-15 and blaTEM-1B as well as blaCTX-M-15 and blaTEM-169 were confirmed to coexist in 52.6% and 10.5% of the sequenced E. coli isolates, respectively. In addition, blaOXA-1 was identified together with blaCTX-M-15 and blaTEM-1B in one isolate, and in one isolate, blaTEM-169 together with blaCTX-M-15 and blaTEM-1B was found. The results obtained show that measures need to be taken to reduce the spread of drug resistance and ensure the long-term use of available antimicrobials.

The impact and complete genome characterisation of viruses involved in outbreaks of gastroenteritis in a farrow-to-finish holding.

Viral enteric pathogens continuously burden intensive pig farming, causing gastrointestinal diseases of epidemic and endemic nature. The present study investigated two diarrhoea outbreaks on a large farrow-to-finish holding and subsequent circulation of outbreak-related enteric viruses. These viruses were characterised by whole genome sequencing, and statistical evaluation of the impact on specific production metrics was performed. The results provided evidence that the Porcine epidemic diarrhoea virus-swine enteric coronavirus (PEDV-SeCoV) S gene recombinant strain was responsible for the first outbreak, whilst Rotavirus A (RVA) in a mixed infection with Rotavirus B (RVB) and porcine kobuvirus (PKV) probably caused the second diarrhoea outbreak. Whole genome characterisation revealed a porcine origin of all viruses involved and significant heterogeneity of RVB strain, proposing four novel genotypes and changes in RVB VP1 genotype classification. The statistical evaluation confirmed only a minor disturbance in the number of weaned pigs per sow, with statistical forecasting showing positive trends. A follow-up study corroborated the endemicity of RVA and PKV, in contrast to PEDV-SeCoV. Punctual, comprehensive and timely investigation of diarrhoea outbreaks is a prerequisite for applying adequate pig health and biosecurity management. Calculating such outbreaks' impact on production metrics can potentially shape future decisions on management improvements.

Interspecies transmission of porcine-originated G4P[6] rotavirus A between pigs and humans: a synchronized spatiotemporal approach.

As a leading viral cause of acute gastroenteritis in both humans and pigs, rotavirus A (RVA) poses a potential public health concern. Although zoonotic spillover of porcine RVA strains to humans is sporadic, it has been detected worldwide. The origin of chimeric human-animal strains of RVA is closely linked to the crucial role of mixed genotypes in driving reassortment and homologous recombination, which play a major role in shaping the genetic diversity of RVA. To better understand how genetically intertwined porcine and zoonotic human-derived G4P[6] RVA strains are, the present study employed a spatiotemporal approach to whole-genome characterization of RVA strains collected during three consecutive RVA seasons in Croatia (2018-2021). Notably, sampled children under 2 years of age and weanling piglets with diarrhea were included in the study. In addition to samples tested by real-time RT-PCR, genotyping of VP7 and VP4 gene segments was conducted. The unusual genotype combinations detected in the initial screening, including three human and three porcine G4P[6] strains, were subjected to next-generation sequencing, followed by phylogenetic analysis of all gene segments, and intragenic recombination analysis. Results showed a porcine or porcine-like origin for each of the eleven gene segments in all six RVA strains. The G4P[6] RVA strains detected in children most likely resulted from porcine-to-human interspecies transmission. Furthermore, the genetic diversity of Croatian porcine and porcine-like human G4P[6] strains was propelled by reassortment events between porcine and porcine-like human G4P[6] RVA strains, along with homologous intragenotype and intergenotype recombinations in VP4, NSP1, and NSP3 segments. Described concurrent spatiotemporal approach in investigating autochthonous human and animal RVA strains is essential in drawing relevant conclusions about their phylogeographical relationship. Therefore, continuous surveillance of RVA, following the One Health principles, may provide relevant data for assessing the impact on the protectiveness of currently available vaccines.

Circulation of SARS-CoV-Related Coronaviruses and Alphacoronaviruses in Bats from Croatia.

Bats are natural hosts of various coronaviruses (CoVs), including human CoVs, via an assumed direct zoonotic spillover or intermediate animal host. The present study aimed to investigate the circulation of CoVs in a bat colony in the Mediterranean region of Croatia. Guano and individual droppings from four bat species were sampled and tested with the E-gene sarbecovirus RT-qPCR, the pan-CoV semi-nested RT-PCR targeting the RdRp gene and NGS. Furthermore, bat blood samples were investigated for the presence of sarbecovirus-specific antibodies with the surrogate virus neutralization test (sVNT). The initial testing showed E-gene Sarebeco RT-qPCR reactivity in 26% of guano samples while the bat droppings tested negative. The application of RdRp semi-nested RT-PCR and NGS revealed the circulation of bat alpha- and betaCoVs. Phylogenetic analysis confirmed the clustering of betaCoV sequence with SARS-CoV-related bat sarbecoviruses and alpha-CoV sequences with representatives of the Minunacovirus subgenus. The results of sVNT show that 29% of bat sera originated from all four species that tested positive. Our results are the first evidence of the circulation of SARS-CoV-related coronaviruses in bats from Croatia.

Molecular Typing of Mastadenoviruses in Simultaneously Collected Nasopharyngeal Swabs and Stool Samples from Children Hospitalized for Acute Bronchiolitis, Acute Gastroenteritis, and Febrile Seizures.

This study determines and compares the frequency of human mastadenovirus (HAdV) presence in children with acute bronchiolitis (AB), acute gastroenteritis (AGE), and febrile seizures (FS), ascertains types of HAdVs associated with each individual syndrome and contrasts the findings with a control group of children. The presence of HAdVs was ascertained in simultaneously collected nasopharyngeal (NP) swabs and stool samples amplifying the hexon gene by RT-PCR; these were sequenced to determine the types of HAdVs. HAdVs were grouped into eight different genotypes. Of these, three (F40, F41, and A31) were found solely in stool samples, whereas the others (B3, C1, C2, C5, and C6) were found in both stool samples and NP swabs. The most common genotypes in NP swabs were C2 (found in children with AGE and FS) and C1 (only in children with FS), whereas in stool samples genotypes F41 (in children with AGE) and C2 (in children with AGE and FS) prevailed, and C2 was simultaneously present in both samples. HAdVs were more often detected in stool samples than in NP swabs in patients (with the highest estimated viral load in stool samples in children with AB and AGE) and healthy controls and were more common in NP swabs in children with AGE than in children with AB. In most patients, the characterized genotypes in NP swabs and stool samples were in concordance.

The Efficacy of Probiotics as Antiviral Agents for the Treatment of Rotavirus Gastrointestinal Infections in Children: An Updated Overview of Literature.

Enteric viruses, including the rotavirus, norovirus, and adenoviruses, are the most common cause of acute gastroenteritis. The rotavirus disease is especially prevalent among children, and studies over the past decade have revealed complex interactions between rotaviruses and the gut microbiota. One way to treat and prevent dysbiosis is the use of probiotics as an antiviral agent. This review focuses on the latest scientific evidence on the antiviral properties of probiotics against rotavirus gastroenteric infections in children. A total of 19 studies exhibited a statistically significant antiviral effect of probiotics. The main probiotics that were effective were Saccharomyces cerevisiae var. boulardii, Lacticaseibacillus rhamnosus GG, and various multi-strain probiotics. The underlying mechanism of the probiotics against rotavirus gastroenteric infections in children included immune enhancement and modulation of intestinal microbiota leading to shortening of diarrhoea. However, several clinical studies also found no significant difference in the probiotic group compared to the placebo group even though well-known strains were used, thus showing the importance of correct dosage, duration of treatment, quality of probiotics and the possible influence of other factors, such as the production process of probiotics and the influence of immunisation on the effect of probiotics. Therefore, more robust, well-designed clinical studies addressing all factors are warranted.

The Antimicrobial Effect of Various Single-Strain and Multi-Strain Probiotics, Dietary Supplements or Other Beneficial Microbes against Common Clinical Wound Pathogens.

The skin is the largest organ in the human body and is colonized by a diverse microbiota that works in harmony to protect the skin. However, when skin damage occurs, the skin microbiota is also disrupted, and pathogens can invade the wound and cause infection. Probiotics or other beneficial microbes and their metabolites are one possible alternative treatment for combating skin pathogens via their antimicrobial effectiveness. The objective of our study was to evaluate the antimicrobial effect of seven multi-strain dietary supplements and eleven single-strain microbes that contain probiotics against 15 clinical wound pathogens using the agar spot assay, co-culturing assay, and agar well diffusion assay. We also conducted genera-specific and species-specific molecular methods to detect the DNA in the dietary supplements and single-strain beneficial microbes. We found that the multi-strain dietary supplements exhibited a statistically significant higher antagonistic effect against the challenge wound pathogens than the single-strain microbes and that lactobacilli-containing dietary supplements and single-strain microbes were significantly more efficient than the selected propionibacteria and bacilli. Differences in results between methods were also observed, possibly due to different mechanisms of action. Individual pathogens were susceptible to different dietary supplements or single-strain microbes. Perhaps an individual approach such as a 'probiogram' could be a possibility in the future as a method to find the most efficient targeted probiotic strains, cell-free supernatants, or neutralized cell-free supernatants that have the highest antagonistic effect against individual clinical wound pathogens.

Virus transmission by ultrasonic scaler and its prevention by antiviral agent: An in vitro study.

BACKGROUND: It is well recognized that dental procedures represent a potential way of infection transmission. With the COVID-19 pandemic, the focus of dental procedure associated transmission has rapidly changed from bacteria to viruses. The aim was to develop an experimental setup for testing the spread of viruses by ultrasonic scaler (USS) generated dental spray and evaluate its mitigation by antiviral coolants. METHODS: In a virus transmission tunnel, the dental spray was generated by USS with saline coolant and suspension of Equine Arteritis Virus (EAV) delivered to the USS tip. Virus transmission by settled droplets was evaluated with adherent RK13 cell lines culture monolayer. The suspended droplets were collected by a cyclone aero-sampler. Antiviral activity of 0.25% NaOCl and electrolyzed oxidizing water (EOW) was tested using a suspension test. Antiviral agents' transmission prevention ability was evaluated by using them as a coolant. RESULTS: In the suspension test with 0.25% NaOCl or EOW, the TCID50/mL was below the detection limit after 5 seconds. With saline coolant, the EAV-induced cytopathic effect on RK13 cells was found up to the distance of 45 cm, with the number of infected cells decreasing with distance. By aero-sampler, viral particles were detected in concentration ≤4.2 TCID50/mL. With both antiviral agents used as coolants, no EAV-associated RK-13 cell infection was found. CONCLUSION: We managed to predictably demonstrate EAV spread by droplets because of USS action. More importantly, we managed to mitigate the spread by a simple substitution of the USS coolant with NaOCl or EOW.

A novel reassortant mammalian orthoreovirus with a divergent S1 genome segment identified in a traveler with diarrhea.

Mammalian orthoreoviruses with reassortant genomes have recently been detected in various mammals and humans with respiratory, central nervous system, and gastrointestinal symptoms. This study describes the detection of the novel reassortant mammalian orthoreovirus SI-MRV07 in a traveler with gastroenteritis that returned from southeast Asia. The virus was initially detected with electron microscopy in stool, followed by propagation in the epithelial-like monkey kidney Marc145 cell line. Whole-genome sequencing revealed the reassortant nature of the genome segments, whereby the S1 genome segment was the most variable according to known sequences deposited in GenBank. Based on the nucleotide sequence of the S1 genome segment, the isolate clusters to serotype 2, close to the reference strain Jones T2J. The patient's serum showed the highest virus neutralization capacity toward SI-MRV07 and T2J isolates. This study provides additional insight into emerging mammalian orthoreoviruses with reassortant genomes and possible zoonotic potential, which should be carefully monitored in the future.

Identification of novel reassortant mammalian orthoreoviruses from bats in Slovenia.

BACKGROUND: Recently, mammalian orthoreoviruses (MRVs) were detected for the first time in European bats, and the closely related strain SI-MRV01 was isolated from a child with severe diarrhoea in Slovenia. Genetically similar strains have also been reported from other mammals, which reveals their wide host distribution. The aim of this study was to retrospectively investigate the occurrence and genetic diversity of MRVs in bats in Slovenia, from samples obtained throughout the country in 2008 to 2010, and in 2012 and to investigate the occurrence of the novel SI-MRV01 MRV variant in Slovenian bats. RESULTS: The detection of MRVs in bat guano was based on broad-range RT-PCR and specific bat MRV real-time RT-PCR. Subsequently, MRV isolates were obtained from cell culture propagation, with detailed molecular characterisation through whole-genome sequencing. Overall, bat MRVs were detected in 1.9% to 3.8% of bats in 2008, 2009 and 2012. However, in 2010 the prevalence was 33.0%, which defined an outbreak of the single SI-MRV01 strain. Here, we report on the identification of five MRV isolates of different serotypes that are designated as SI-MRV02, SI-MRV03, SI-MRV04, SI-MRV05 and SI-MRV06. There is high genetic variability between these characterised isolates, with evident genome reassortment seen across their genome segments. CONCLUSIONS: In conclusion, we have confirmed the presence of the SI-MRV01 strain in a Slovenian bat population. Moreover, according to genetic characterisation of S1 genome segment, all three MRV serotypes were present in the bat population. In this study, five independent MRV isolates were obtained and detailed whole genome analysis revealed high diversity between them. This study generates new information about the epidemiology and molecular characteristics of emerging bat MRV variants, and provides important molecular data for further studies of their pathogenesis and evolution.

Study of the In Vitro Antagonistic Activity of Various Single-Strain and Multi-Strain Probiotics against Escherichia coli.

Escherichia coli is an important commensal of our gut, however, many pathogenic strains exist, causing various severe infections in the gut or beyond. Due to several antibiotic resistance patterns of E. coli, research of alternative treatments or adjuvant therapy is important. One of these is the use of probiotics as antagonistic agents against E. coli. Most published studies investigate only one strain of E. coli and single-strain probiotics. The objectives of this study were to evaluate the antagonistic activity of selected single-strain and multi-strain probiotic supplements against selected clinical E. coli pathotypes using the in vitro agar spot test and the co-culturing method. Molecular methods were used to determine the presence of the genus lactobacilli and bifidobacteria as well as certain selected strains in the probiotic supplements. The agar-spot test showed that the multi-strain probiotics were more effective than the single-strain probiotics. On the other hand, the co-culturing method showed the opposite result, indicating that results are importantly influenced by the chosen method. The most effective single-strain probiotics against E. coli strains were Bifidobacterium animalis subsp. lactis BB-12 and Lactobacillus reuteri DSM 17938. The most effective multi-strain probiotics contained lactobacilli, bifidobacteria and enterococci strains, thus proving that most effective probiotics against E. coli strains are the lactic acid bacteria and bifidobacteria. The overall results from both in vitro tests reveal that all selected probiotics exhibited an antagonistic activity against all E. coli strains. From a public health perspective probiotics have thus proved to be successful in inhibiting the growth of E. coli and could therefore be used as adjuvant therapy or alternative therapy in E. coli infections.

Intrahost Norovirus Evolution in Chronic Infection Over 5 Years of Shedding in a Kidney Transplant Recipient.

Noroviruses are the leading cause of acute gastroenteritis, and they can affect humans of all age groups. In immunocompromised patients, norovirus infections can develop into chronic diarrhea or show prolonged asymptomatic virus shedding. Chronic norovirus infections are frequently reported for solid organ transplant recipients, with rapid intrahost norovirus evolution seen. In this report, we describe a case of chronic norovirus infection in an immunocompromised patient who was followed up for over 5 years. The purpose of the study was to specify the norovirus evolution in a chronically infected immunocompromised host and identify possible selection sites in norovirus capsid protein. During the follow-up period, 25 sequential stool samples were collected and nine of them were selected to generate amplicons covering viral RNA-dependent RNA polymerase (RdRp) and viral capsid protein (VP1) genes. Amplicons were sequenced using next-generation sequencing. Single nucleotide polymorphisms were defined, which demonstrated a nearly 3-fold greater mutation rate in the VP1 genome region compared to the RdRp genome region (7.9 vs. 2.8 variable sites/100 nucleotides, respectively). This indicates that mutations in the virus genome were not accumulated randomly, but are rather the result of mutant selection during the infection cycle. Using ShoRAH software we were able to reconstruct haplotypes occurring in each of the nine selected samples. The deduced amino-acid haplotype sequences were aligned and the positions were analyzed for selective pressure using the Datamonkey program. Only 12 out of 25 positive selection sites were within the commonly described epitopes A, B, C, and D of the VP1 protein. New positive selection sites were determined that have not been described before and might reflect adaptation of the norovirus toward optimal histo-blood-group antigen binding, or modification of the norovirus antigenic properties. These data provide new insights into norovirus evolutionary dynamics and indicate new putative epitope "hot-spots" of modified and optimized norovirus-host interactions.

Whole genome sequence and a phylogenetic analysis of the G8P[14] group A rotavirus strain from roe deer.

BACKGROUND: Group A rotaviruses (RVA) are associated with acute gastroenteritis in children and in young domestic and wild animals. A RVA strain was detected from a roe deer for the first time during a survey of game animals in Slovenia in 2014. A further RVA strain (SLO/D110-15) was detected from a roe deer during 2015. The aim of this study was to provide a full genetic profile of the detected RVA strain from roe deer and to obtain additional information about zoonotic transmitted strains and potential reassortments between human rotavirus strains and zoonotic transmitted rotavirus strains. The next generation sequencing (NGS) analysis on Ion Torrent was performed and the whole genome sequence has been determined together with a phylogenetic analysis. RESULTS: The whole genome sequence of SLO/D110-15 was obtained by NGS analyses on an IonTorrent platform. According to the genetic profile, the strain SLO/D110-15 clusters with the DS-1-like group and expresses the G8-P[14]-I2-R2-C2-M2-A3-N2-T6-E2-H3 genome constellation. Phylogenetic analysis shows that this roe deer G8P[14] strain is most closely related to RVA strains found in sheep, cattle and humans. A human RVA strain with the same genotype profile was detected in 2009 in Slovenia. CONCLUSIONS: The G8P[14] genotype has been found, for the first time, in deer, a newly described host from the order Artiodactyla for this RVA genotype. The finding of a rotavirus with the same genome segment constellation in humans indicates the possible zoonotic potential of this virus strain.

Detection and Whole-Genome Analysis of a Zoonotic G8P[14] Rotavirus Strain Isolated from a Child with Diarrhea.

The group A rotavirus strain SI-2987/09 was detected in a child with severe diarrhea. The whole-genome analysis revealed the G8-P[14]-I2-R2-C2-M2-A3-N2-T6-E2-H3 genome constellation, which reflects the zoonotic transmission of the strain most probably from the ungulate group. This was also confirmed by the high nucleotide identity to those animal rotavirus strains.

Global Spread of Norovirus GII.17 Kawasaki 308, 2014-2016.

Analysis of complete capsid sequences of the emerging norovirus GII.17 Kawasaki 308 from 13 countries demonstrated that they originated from a single haplotype since the initial emergence in China in late 2014. Global spread of a sublineage SL2 was identified. A new sublineage SL3 emerged in China in 2016.

Respiratory and Enteric Virus Detection in Children.

The majority of children with febrile seizures have viral infections and viruses were detected in 22% to 63% of children in published studies. Using molecular methods, viruses were also detected in asymptomatic persons. A prospective study was conducted to detect respiratory and enteric viruses in 192 children with febrile seizures and compare the detection rates to those found in 156 healthy age-matched controls. A respiratory or enteric virus was detected in 72.9% of children with febrile seizures and in 51.4% of healthy controls. The viruses most strongly associated with febrile seizures were influenza, respiratory syncytial virus, parainfluenza, human coronavirus, and rotavirus. Compared to healthy controls, the age-adjusted odds ratios for nasopharynx virus positivity in febrile seizure patients were 79.4, 2.8, 7.2, and 4.9 for influenza virus, parainfluenza virus, respiratory syncytial virus, and human coronavirus, respectively, and 22.0 for rotavirus in stool. The detected virus did not influence clinical features of febrile seizure.

The first detection and whole genome characterization of the G6P[15] group A rotavirus strain from roe deer.

Although rotaviruses have been detected in a variety of host species, there are only limited records of their occurrence in deer, where their role is unknown. In this study, group A rotavirus was identified in roe deer during a study of enteric viruses in game animals. 102 samples of intestinal content were collected from roe deer (56), wild boars (29), chamois (10), red deer (6) and mouflon (1), but only one sample from roe deer was positive. Following whole genome sequence analysis, the rotavirus strain D38/14 was characterized by next generation sequencing. The genotype constellation, comprising 11 genome segments, was G6-P[15]-I2-R2-C2-M2-A3-N2-T6-E2-H3. Phylogenetic analysis of the VP7 genome segment showed that the D38/14 rotavirus strain is closely related to the various G6 zoonotic rotavirus strains of bovine-like origin frequently detected in humans. In the VP4 segment, this strain showed high variation compared to that in the P[15] strain found in sheep and in a goat. This finding suggests that rotaviruses from deer are similar to those in other DS-1 rotavirus groups and could constitute a source of zoonotically transmitted rotaviruses. The epidemiological status of group A rotaviruses in deer should be further investigated.

Narrowing of the Diagnostic Gap of Acute Gastroenteritis in Children 0-6 Years of Age Using a Combination of Classical and Molecular Techniques, Delivers Challenges in Syndromic Approach Diagnostics.

BACKGROUND: Twenty-five percent to 50% of acute gastroenteritis (AGE) cases remain etiologically undiagnosed. Our main aim was to determine the most appropriate list of enteric pathogens to be included in the daily diagnostics scheme of AGE, ensuring the lowest possible diagnostic gap. METHODS: Two hundred ninety seven children ≤6 years of age, admitted to hospital in Slovenia, October 2011 to October 2012, with AGE, and 88 ≤6 years old healthy children were included in the study. A broad spectrum of enteric pathogens was targeted with molecular methods, including 8 viruses, 6 bacteria and 2 parasites. RESULTS: At least one enteric pathogen was detected in 91.2% of cases with AGE and 27.3% of controls. Viruses were the most prevalent (82.5% and 15.9%), followed by bacteria (27.3% and 10.2%) and parasites (3.0% and 1.1%) in cases and controls, respectively. A high proportion (41.8%) of mixed infections was observed in the cases. For cases with undetermined etiology (8.8%), stool samples were analyzed with next generation sequencing, and a potential viral pathogen was detected in 17 additional samples (5.8%). CONCLUSIONS: Our study suggests that tests for rotaviruses, noroviruses genogroup II, adenoviruses 40/41, astroviruses, Campylobacter spp. and Salmonella sp. should be included in the initial diagnostic algorithm, which revealed the etiology in 83.5% of children tested. The use of molecular methods in diagnostics of gastroenteritis is preferable because of their high sensitivity, specificity, fast performance and the possibility of establishing the concentration of the target. The latter may be valuable for assessing the clinical significance of the detected enteric, particularly viral pathogens.

The Role of Human Coronaviruses in Children Hospitalized for Acute Bronchiolitis, Acute Gastroenteritis, and Febrile Seizures: A 2-Year Prospective Study.

UNLABELLED: Human coronaviruses (HCoVs) are associated with a variety of clinical presentations in children, but their role in disease remains uncertain. The objective of our prospective study was to investigate HCoVs associations with various clinical presentations in hospitalized children up to 6 years of age. Children hospitalized with acute bronchiolitis (AB), acute gastroenteritis (AGE), or febrile seizures (FS), and children admitted for elective surgical procedures (healthy controls) were included in the study. In patients with AB, AGE, and FS, a nasopharyngeal (NP) swab and blood sample were obtained upon admission and the follow-up visit 14 days later, whereas in children with AGE a stool sample was also acquired upon admission; in healthy controls a NP swab and stool sample were taken upon admission. Amplification of polymerase 1b gene was used to detect HCoVs in the specimens. HCoVs-positive specimens were also examined for the presence of several other viruses. HCoVs were most often detected in children with FS (19/192, 9.9%, 95% CI: 6-15%), followed by children with AGE (19/218, 8.7%, 95% CI: 5.3-13.3%) and AB (20/308, 6.5%, 95% CI: 4.0-9.8%). The presence of other viruses was a common finding, most frequent in the group of children with AB (19/20, 95%, 95% CI: 75.1-99.8%), followed by FS (10/19, 52.6%, 95% CI: 28.9-75.6%) and AGE (7/19, 36.8%, 95% CI: 16.3-61.6%). In healthy control children HCoVs were detected in 3/156 (1.9%, 95% CI: 0.4-5.5%) NP swabs and 1/150 (0.7%, 95% CI: 0.02-3.3%) stool samples. It seems that an etiological role of HCoVs is most likely in children with FS, considering that they had a higher proportion of positive HCoVs results than patients with AB and those with AGE, and had the highest viral load; however, the co-detection of other viruses was 52.6%. TRIAL REGISTRATION: ClinicalTrials.gov NCT00987519.

Detection and Characterization of a Novel Reassortant Mammalian Orthoreovirus in Bats in Europe.

A renewed interest in mammalian orthoreoviruses (MRVs) has emerged since new viruses related to bat MRV type 3, detected in Europe, were identified in humans and pigs with gastroenteritis. This study reports the isolation and characterization of a novel reassortant MRV from the lesser horseshoe bat (Rhinolophus hipposideros). The isolate, here designated BatMRV1-IT2011, was first identified by electron microscopy and confirmed using PCR and virus-neutralization tests. The full genome sequence was obtained by next-generation sequencing. Molecular and antigenic characterizations revealed that BatMRV1-IT2011 belonged to serotype 1, which had not previously been identified in bats. Phylogenetic and recombination detection program analyses suggested that BatMRV1-IT2011 was a reassortant strain containing an S1 genome segment similar to those of MRV T1/bovine/Maryland/Clone23/59 and C/bovine/ Indiana/MRV00304/2014, while other segments were more similar to MRVs of different hosts, origins and serotypes. The presence of neutralizing antibodies against MRVs has also been investigated in animals (dogs, pigs, bovines and horses). Preliminary results suggested that MRVs are widespread in animals and that infections containing multiple serotypes, including MRVs of serotype 1 with an S1 gene similar to BatMRV1-IT2011, are common. This paper extends the current knowledge of MRVs and stresses the importance to continue and improve MRV surveillance in bats and other mammals through the development and standardization of specific diagnostic tools.