Near-Complete Genome Sequence of a Human Norovirus GII.1[Pg] Strain Associated with Acute Gastroenteritis, Determined Using Long-Read Sequencing

Yang, Zhihui; Mammel, Mark K.; Wales, Samantha Q.

doi:10.1128/mra.00401-21

Cited by 2 publications

(2 citation statements)

References 10 publications

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“…Limited quantities of HuNoV fecal samples belonging to GII.1 [ 22 ], GII.4 [ 21 ] and GII.6 [ 23 ] genotypes that were known to replicate in HIE were received courtesy of Dr. Samantha Wales (FDA/CFSAN). Fecal suspensions (10%) were prepared in 1X PBS and these were processed by either serial centrifugation [ 17 ] or serial filtration [ 13 ].…”

Section: Methodsmentioning

confidence: 99%

Replication of Human Norovirus in Human Intestinal Enteroids Is Affected by Fecal Sample Processing

Narwankar,

Esseili

2024

Viruses

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Human intestinal enteroids (HIEs) culture is an emerging model for assessing the infectivity of human noroviruses (HuNoVs). The model is based on detecting an increase in HuNoV RNA post-infection of HIEs. HuNoV fecal samples used for HIE infection are traditionally processed by serial filtration. Recently, processing HuNoV fecal samples by serial centrifugation was shown to retain vesicles containing HuNoV. The objective of this study was to investigate whether serially centrifuged fecal samples, RNA extraction kit (QIAamp versus MagMaX) and HIE age (newer versus older) affect HuNoV RNA fold increase in HIE. HuNoV GII.1, GII.4 and GII.6 fecal samples were prepared by serial centrifugation and filtration and the viral RNA in HIE was quantified at 1 and 72 h post-infection (hpi) following RNA extraction and RT-qPCR. The serially filtered GII.1, GII.4 and GII.6 showed successful replication in HIE, resulting in mean log increases of 2.2, 2 and 1.2, respectively, at 72 vs. 1 hpi. In contrast, only serially centrifuged GII.1 showed consistently successful replication. However, using newer HIE passages and the MagMAX kit resulted in mean log fold increases for serially centrifuged GII.1, GII.4 and GII.6 (1.6, 2.3 and 1.8 log, respectively) that were similar to serially filtered samples. Therefore, HuNoV fecal sample processing and HIE age can affect virus replication in the HIE model.

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Section: Methodsmentioning

confidence: 99%

Replication of Human Norovirus in Human Intestinal Enteroids Is Affected by Fecal Sample Processing

Narwankar,

Esseili

2024

Viruses

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“…We had one paired set of GII.6[P7] stool and vomit, in which the NoV from stool [26] replicated, but not that from the vomit. We also had a second paired set (GII.1) that was acquired from an adult and child within the same household [30]. In this set, the virus from the child's sample did not replicate despite having a very high titer (average of 7.20 × 10 5 genome copies added per well at a 1:200 dilution of the 10% filtrate).…”

Section: Screening Of Nov Fecal Filtrate Replication With J2 Hiesmentioning

confidence: 99%

Use of Human Intestinal Enteroids for Recovery of Infectious Human Norovirus from Berries and Lettuce

Wales,

Kulka,

Keinard

et al. 2023

Foods

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Norovirus (NoV) is the leading cause of viral foodborne gastroenteritis globally. Currently, the gold standard for detecting NoV in clinical, food, and environmental samples is via molecular-based methods, primarily RT-PCR. Nevertheless, there is a great need for confirmatory assays that can determine the infectivity of viral particles recovered from contaminated matrices. The use of the human intestinal enteroids system (HIEs) has allowed for the expansion of norovirus replication, although it still suffers from limitations of strain preferences and the requirement of high titer stocks for infection. In this study, we wanted to explore the feasibility of using the HIEs to support the replication of NoV that had been recovered from representative food matrices that have been associated with foodborne illness. We first confirmed that HIEs can support the replication of several strains of NoV as measured by RT-qPCR. We subsequently chose two of those strains that reproducibly replicated, GII.4 and GII.6, to evaluate in a TCID50 assay and for future experiments. Infectious NoV could be recovered and quantified in the HIEs from lettuce, frozen raspberries, or frozen strawberries seeded with high titers of either of these strains. While many experimental challenges still remain to be overcome, the results of this study represent an important step toward the detection of infectious norovirus from representative produce items.

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Near-Complete Genome Sequence of a Human Norovirus GII.1[Pg] Strain Associated with Acute Gastroenteritis, Determined Using Long-Read Sequencing

Cited by 2 publications

References 10 publications

Replication of Human Norovirus in Human Intestinal Enteroids Is Affected by Fecal Sample Processing

Replication of Human Norovirus in Human Intestinal Enteroids Is Affected by Fecal Sample Processing

Use of Human Intestinal Enteroids for Recovery of Infectious Human Norovirus from Berries and Lettuce

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