Evaluation of the BioFire Gastrointestinal Panel to Detect Diarrheal Pathogens in Pediatric Patients

Infectious diarrhea is a global pediatric health concern; therefore, rapid and accurate detection of enteropathogens is vital. We evaluated the BioFire® FilmArray® Gastrointestinal (GI) Panel with that of comparator laboratory tests. Stool samples of pediatric patients with diarrhea were prospectively collected and tested. As a comparator method for bacteria, culture, conventional PCR for diarrheagenic E. coli, and Allplex GI-Bacteria(I) Assay were tested. For discrepancy analysis, BD MAX Enteric Bacterial Panel was used. As a comparator method for virus, BD MAX Enteric Virus Panel and immunochromatography was used and Allplex GI-Virus Assay was used for discrepancy analysis. The “true positive” was defined as culture-positive and/or positive results from more than two molecular tests. Of the 184 stool samples tested, 93 (50.5%) were true positive for 128 pathogens, and 31 (16.9%) were positive for multiple pathogens. The BioFire GI Panel detected 123 pathogens in 90 of samples. The BioFire GI Panel demonstrated a sensitivity of 100% for 12 targets and a specificity of >95% for 16 targets. The overall positive rate and multiple pathogen rate among patients in the group without underlying diseases were significantly higher than those in the group with hematologic disease (57.0% vs. 28.6% (p = 0.001) and 20.4% vs. 4.8% (p = 0.02), respectively). The BioFire GI Panel provides comprehensive results within 2 h and may be useful for the rapid identification of enteropathogens.

Molecular Detection of Enteric Viruses in Under-Five Children with Diarrhea in Debre Tabore, Northwest Ethiopia

Viral gastroenteritis belongs to the major public health problems of infant and children worldwide. The largest proportion of morbidity and mortality occurs in Sub-Saharan Africa. This preliminary study aimed to assess the burden and genetic diversity of enteric viruses among children with diarrhea in Debre Tabore. A cross-sectional study was undertaken from December 2015 to April 2016. A total of thirty-eight children, who presented with diarrhea at Debre Tabore health centers were included. Fecal samples were collected and screened for enteric viruses by RT-PCR. Data were analyzed by using SPSS statistical software. Descriptive statistical summary techniques were used to display the study findings. Out of the thirty-eight children screened, 52.6% were positive for at least one enteric virus. Six (30.0%) of the children had mixed enteric virus infections. Human adenovirus (HAdV) 7 (18.4%) was predominant followed by noroviruses 5 (13.2%), enterovirus 5 (13.2%), rotavirus 4 (10.5%), human astrovirus (HAstV) 2 (5.3%), and human parechovirus (HPeV) 1(2.6%). Overall nineteen different types of enteric virus genotypes were identified. Diverse adenovirus within species A (HAdV-12,-31), B (HAdV-3), C (HAdV-2), and F (HAdV-4) were detected. Norovirus II (GII.4 and GII.6) and norovirus I (GI.2, GI.3, and GI.5) genotypes were found. Sapovirus genotypes within genogroup II (GII.1, GII.5, and GII.6) were identified. Wild-type rotavirus G9[P8] genotype was detected in one of the rotavirus positive samples. Non-polio enteroviruses within species A (coxsackie A virus (CAV) 5, CAV6, and CAV14) and C (enterovirus (EV-C) 99) were also identified. HPeV-6 genotype was identified in one of the samples. In two of the fecal samples, classic HAstV-2 was detected. Phylogenetic analysis of these enteric viruses revealed that they have close phylogenetic relatedness with previous genotypes reported from Ethiopia. Diverse enteric viruses were detected in fecal samples from under-five children with diarrhea. The detection of heterogeneous enteric viruses in this small data set highlights the need for extended multicenter studies to describe the burden and genetic diversity.

Wide Diversity of Recombinant Noroviruses Circulating in Spain, 2016 to 2020

Noroviruses are the leading cause of sporadic cases and outbreaks of viral gastroenteritis. For more than 20 years most norovirus infections have been caused by the pandemic genotype GII.4, yet recent studies have reported the emergence of recombinant strains in many countries. In the present study, 4,950 stool samples collected between January 2016 and April 2020 in Valencia (Spain) from patients with acute gastroenteritis were analyzed to investigate the etiological agent. Norovirus was the most frequently detected enteric virus with a positive rate of 9.5% (471/4,950). Among 224 norovirus strains characterized, 175 belonged to genogroup GII and 49 to genogroup GI. Using dual genotyping based on sequencing the ORF1/ORF2 junction region we detected 25 different capsid-polymerase type associations. The most common GII capsid genotype was GII.4 Sydney 2012, followed by GII.2, GII.3, GII.6 and GII.17. A high prevalence of recombinant strains (90.4%) was observed among GII infections between 2018 and 2020. GII.4 Sydney[P16] was the predominant genotype from 2019 to 2020. In addition, GII.P16 polymerase was found harboring within six different capsid genes. A new subcluster of GII.4 Sydney associated with the P31 polymerase was identified by phylogenetic analysis. GI.4 and GI.3 were the predominant genotypes in genogroup GI, in which recombinant strains were also found, such as GI.3[P10], GI.3[P13] and GI.5[P4]. Interestingly, the GI.3[P10] strain could represent a new capsid genotype. This study shows the extensive diversity of recombinant noroviruses circulating in Spain and highlights the role of recombination events in the spread of noroviruses.

A Review of the Emerging White Chick Hatchery Disease

White chick hatchery disease is an emerging disease of broiler chicks with which the virus, chicken astrovirus, has been associated. Adult birds typically show no obvious clinical signs of infection, although some broiler breeder flocks have experienced slight egg drops. Substantial decreases in hatching are experienced over a two-week period, with an increase in mid-to-late embryo deaths, chicks too weak to hatch and pale, runted chicks with high mortality. Chicken astrovirus is an enteric virus, and strains are typically transmitted horizontally within flocks via the faecal–oral route; however, dead-in-shell embryos and weak, pale hatchlings indicate vertical transmission of the strains associated with white chick hatchery disease. Hatch levels are typically restored after two weeks when seroconversion of the hens to chicken astrovirus has occurred. Currently, there are no commercial vaccines available for the virus; therefore, the only means of protection is by good levels of biosecurity. This review aims to outline the current understanding regarding white chick hatchery disease in broiler chick flocks suffering from severe early mortality and increased embryo death in countries worldwide.

Single-cell sequencing of rotavirus-infected intestinal epithelium reveals cell-type specific epithelial repair and tuft cell infection

Intestinal epithelial damage is associated with most digestive diseases and results in detrimental effects on nutrient absorption and production of hormones and antimicrobial defense molecules. Thus, understanding epithelial repair and regeneration following damage is essential in developing therapeutics that assist in rapid healing and restoration of normal intestinal function. Here we used a well-characterized enteric virus (rotavirus) that damages the epithelium at the villus tip but does not directly damage the intestinal stem cell, to explore the regenerative transcriptional response of the intestinal epithelium at the single-cell level. We found that there are specific Lgr5+ cell subsets that exhibit increased cycling frequency associated with significant expansion of the epithelial crypt. This was accompanied by an increase in the number of immature enterocytes. Unexpectedly, we found rotavirus infects tuft cells. Transcriptional profiling indicates tuft cells respond to viral infection through interferon-related pathways. Together these data provide insights as to how the intestinal epithelium responds to insults by providing evidence of stimulation of a repair program driven by stem cells with involvement of tuft cells that results in the production of immature enterocytes that repair the damaged epithelium.

1331. Seasonality of Common Human Coronaviruses in the United States, 2014-2021

Background The four common human coronavirus (HCoV) types, including two alpha (NL63 and 229E) and two beta (HKU1 and OC43) coronaviruses, generally cause mild, upper respiratory illness. Common HCoV seroprevalence increases rapidly during the first five years of life and remains high throughout adulthood. HCoVs are known to have seasonal patterns, with variation in predominant types each year, but more defined measures of seasonality are needed. Methods We describe laboratory detection, percent positivity, and seasonality of the four common HCoVs during July 2014 to May 2021 in the United States reported to the National Respiratory and Enteric Virus Surveillance System (NREVSS). We also describe age, sex, and co-detection with other respiratory viruses for a subset of specimens available through the Public Health Laboratory Interoperability Project (PHLIP). We used a method previously validated for respiratory syncytial virus, characterized by a centered 5-week moving average and normalization to peak, to define seasonal inflections, including season onset, peak, and offset. Results Any HCoV type was detected in 96,336 (3.4%) of 2,487,736 specimens. Predominant common HCoV types fluctuated by surveillance year (Figure 1) and were generally consistent across geographic regions. In a subset of 4,576 specimens with a common HCoV detection, those with type 229E had a higher median age compared to other HCoV types (30.8 versus 24.8 years, p< 0.001), but there were no differences by sex. Influenza was the most commonly co-detected virus. In the last six complete HCoV seasons, onsets ranged from October to November, peaks from January to February, and offsets from April to June; >95% of all HCoV detections occurred within these ranges. The 2020-2021 common HCoV season onset, dominated by types NL63 and OC43, was delayed by approximately two months compared to prior seasons. Figure 1. The top panel represents total specimens tested and the bottom panel shows percent positivity of the four common human coronavirus (HCoV) types by week starting July 5, 2014 through May 8, 2021. Data are from the National Respiratory and Enteric Virus Surveillance System (NREVSS). Conclusion Common HCoVs demonstrate relatively consistent seasonal patterns. The delayed onset of the 2020-2021 season may be attributable to mitigation measures implemented across the US including masking, improved hand hygiene, and social distancing. Better defining HCoV seasonality can inform clinical preparedness and testing practices and may provide insights into the behavior of emerging coronaviruses. Disclosures All Authors: No reported disclosures

Specific bacterial cell wall components influence the stability of Coxsackievirus B3

Enteric viruses infect the mammalian gastrointestinal tract and lead to significant morbidity and mortality worldwide. Data indicate that enteric viruses can utilize intestinal bacteria to promote viral replication and pathogenesis. However, the precise interactions between enteric viruses and bacteria are unknown. Here we examined the interaction between bacteria and Coxsackievirus B3, an enteric virus from the picornavirus family. We found that bacteria enhance the infectivity of Coxsackievirus B3 (CVB3) in vitro . Notably, specific bacteria are required as Gram-negative Salmonella enterica , but not Escherichia coli , enhanced CVB3 infectivity and stability. Investigating the cell wall components of both S. enterica and E. coli revealed that structures in the O-antigen or core of lipopolysaccharide, a major component of the Gram-negative bacterial cell wall, were required for S. enterica to enhance CVB3. To determine if these requirements were necessary for similar enteric viruses, we investigated if S. enterica and E. coli enhanced infectivity of poliovirus, another enteric virus in the picornavirus family. We found that while E. coli did not enhance the infectivity of CVB3, E. coli enhanced poliovirus infectivity. Overall, these data indicate that distinct bacteria enhance CVB3 infectivity and stability, and specific enteric viruses may have differing requirements for their interactions with specific bacterial species. Importance Previous data indicate that several enteric viruses utilize bacteria to promote intestinal infection and viral stability. Here we show that specific bacteria and bacterial cell wall components are required to enhance infectivity and stability of Coxsackievirus B3 in vitro . These requirements are likely enteric virus-specific as the bacteria for CVB3 differs from poliovirus, a closely related virus. Therefore, these data indicate that specific bacteria and their cell wall components dictate the interaction with various enteric viruses in distinct mechanisms.

DEVELOPMENT OF A UPL PROBE-BASED REAL-TIME PCR ASSAY OF PORCINE DELTACORONAVIRUS IN TAIWAN

Porcine deltacoronavirus (PDCoV) causes clinical symptoms characterized by severe diarrhea and vomiting in neonatal piglets and pregnant sows, which is similar to those resulted from transmissible gastroenteritis coronavirus (TGEV) and porcine epidemic diarrhea virus (PEDV). Since PEDV was considered as the dominant enteric virus all round the world, PDCoV has been unwittingly overlooked due to its indistinguishable clinical signs with other porcine coronaviruses and relatively low death rates in the pig farm. Specimens which have been previously performed for the detection of PEDV in Animal Disease and Diagnostic Center, National Pingtung University of Science and Technology (NPUST) from January 5, 2015 to January 11, 2016 were examined by a novel universal probe library (UPL) probe-based real-time polymerase chain reaction (PCR). A total of 527 clinical specimens from pigs with diarrhea suspected were examined for PDCoV. Positive rates of PDCoV in small intestine and rectal swab were 4.3% (13/305) and 1.8% (4/222), respectively. Collectively, as to the total specimens, the detection rate is 3.2% (17/527). Our results provide development of a UPL probe-based real-time PCR assay and retrospective investigation of potentially circulating PDCoVs in the field in the whole 2015 and early 2016.