Evaluation of the 50% Infectious Dose of Human Norovirus Cin-2 in Gnotobiotic Pigs: A Comparison of Classical and Contemporary Methods for Endpoint Estimation

Human noroviruses (HuNoVs) are the leading causative agents of epidemic and sporadic acute gastroenteritis that affect people of all ages worldwide. However, very few dose–response studies have been carried out to determine the median infectious dose of HuNoVs. In this study, we evaluated the median infectious dose (ID50) and diarrhea dose (DD50) of the GII.4/2003 variant of HuNoV (Cin-2) in the gnotobiotic pig model of HuNoV infection and disease. Using various mathematical approaches (Reed–Muench, Dragstedt–Behrens, Spearman–Karber, logistic regression, and exponential and approximate beta-Poisson dose–response models), we estimated the ID50 and DD50 to be between 2400–3400 RNA copies, and 21,000–38,000 RNA copies, respectively. Contemporary dose–response models offer greater flexibility and accuracy in estimating ID50. In contrast to classical methods of endpoint estimation, dose–response modelling allows seamless analyses of data that may include inconsistent dilution factors between doses or numbers of subjects per dose group, or small numbers of subjects. Although this investigation is consistent with state-of-the-art ID50 determinations and offers an advancement in clinical data analysis, it is important to underscore that such analyses remain confounded by pathogen aggregation. Regardless, challenging virus strain ID50 determination is crucial for identifying the true infectiousness of HuNoVs and for the accurate evaluation of protective efficacies in pre-clinical studies of therapeutics, vaccines and other prophylactics using this reliable animal model.

1101. Comparison of Clinical Characteristics and Demographics of GII.4 vs. Other GII Noroviruses Associated With Sporadic Acute Gastroenteritis in Children in Nashville, TN, 2012–2015

Background Norovirus is a leading cause of acute gastroenteritis (AGE) in all age groups. Although at least 28 different genotypes infecting humans have been reported, most outbreaks over the last 15 years have been caused by genogroup II (GII) viruses, of which GII.4 viruses have caused more than 50%. Since clinical differences between different genotypes are poorly understood, we sought to compare clinical characteristics in children infected with GII.4 and non-GII.4 viruses. Methods Children between 15 days and 17 years who presented with AGE defined as diarrhea (≥3 loose stools in a 24 hour period) or vomiting (≥1 episodes in a 24 hour period) within 10 days duration were recruited in outpatient, emergency, and inpatient settings in Nashville, TN, during 2012–2015. Stool specimens were tested by RT-qPCR for GI and GII norovirus. Norovirus-positive specimens were genotyped by sequencing of a partial region of the capsid gene. In this study, we excluded children infected with GI, mixed GI/GII and non-typeable GII viruses. Results Of 3,705 AGE subjects enrolled, 2,892 (78%) specimens were collected, 637 (22%) tested norovirus-positive (567 [89%] GII, 62 [10%] GI, and 8 [1%] mixed GI/GII). Of the 567 GII viruses, 461 (81%) were able to be genotyped and of those 238/461 (51.6%) were typed as GII.4 and 223/461 (48.3%) were typed as other GII genotypes (non-GII.4, primarily GII.3 [65/ 461, 14.1%], GII.6 [48/461, 10.4%] and GII.7 [36/461, 7.8%]). Over three AGE seasons, GII.4 represented 64/117 (54%), 79/178 (44%), and 71/166 (57%), of the GII infections, respectively. Compared with non-GII.4 subjects, GII.4 subjects were more likely to be younger (15.5 vs. 21.3 months, P < 0.01), and less likely to attend daycare (23% vs. 39%, P < 0.01). GII.4 subjects also were more likely to present with diarrhea (75% vs. 57%, P < 0.01) and had higher median modified Vesikari score (7 vs. 6, P < 0.01). Conclusion Children infected with GII.4 viruses were younger, less likely to attend child care, more likely to present with diarrhea, and had a more severe illness compared with those with non-GII.4 infections. These data provide important information on the genotype distribution of norovirus in children with AGE in Tennessee and highlight GII.4 as the most prevalent strain. Disclosures N. Halasa, sanofi pasteur: Investigator, Research support. GSK: Consultant, Consulting fee. Moderna: Consultant, Consulting fee.

NOROVIRUS GENOTYPES THAT CAUSED CASES OF ACUTE GASTROENTERITIS IN THE KHABAROVSK REGION

Noroviruses are the leading etiologic cause of outbreaks and sporadic cases of acute gastroenteritis worldwide. The objective of research was to study the genotypes of noroviruses, that caused outbreaks and sporadic incidence of norovirus infection in the Khabarovsk region in 2015-2018. The analysis of outbreaks due to norovirus infection in the Khabarovsk Region in 2015-2018 was performed. The molecular genetic study of samples from 60 patients from three norovirus outbreaks in the Khabarovsk Region and from 164 children with sporadic acute gastroenteritis in Khabarovsk region was performed. Genotype of noroviruses was determined by sequencing method, phylogenetic analysis of the obtained nucleotide sequences was carried out. The norovirus genotypes GII.17, GII.4 Sydney_2012 and GII.6 had caused the outbreaks of norovirus infection in the Khabarovsk region in 2015-2018. Sporadic cases of acute gastroenteritis in children in Khabarovsk in 2016 were due to GII.4 Sydney_2012, GII.3 and GII.6 norovirus genotypes. Detection of the GII.4 Sydney_2012 strain in both outbreaks and sporadic norovirus infection cases in the Khabarovsk region in 2016 evidenced of active circulation of this variant type during this period. The genotype GII.6 had been identified in Khabarovsk from 2016 to 2018.

Molecular Diagnostic Methods for Detection and Characterization of Human Noroviruses

Human noroviruses are a group of viral agents that afflict people of all age groups. The viruses are now recognized as the most common causative agent of nonbacterial acute gastroenteritis and foodborne viral illness worldwide. However, they have been considered to play insignificant roles in the disease burden of acute gastroenteritis for the past decades until the recent advent of new and more sensitive molecular diagnostic methods. The availability and application of the molecular diagnostic methods have led to enhanced detection of noroviruses in clinical, food and environmental samples, significantly increasing the recognition of noroviruses as an etiologic agent of epidemic and sporadic acute gastroenteritis. This article aims to summarize recent efforts made for the development of molecular methods for the detection and characterization of human noroviruses.