Use of Human Intestinal Enteroids to Evaluate Persistence of Infectious Human Norovirus in Seawater

Abstract

H uman noroviruses are the major cause of viral gastroenteritis worldwide (1) and the most common cause of foodborne or waterborne outbreaks in Europe (2). Noroviruses spread through fecal-oral transmission, mainly person to person, but also spread through environmental contamination (1). Food and drinks can be contaminated by infected food handlers and, during production, by human sewage spillover (3). When grown in contaminated seawater, filterfeeding shellfish bioaccumulate human noroviruses in their tissues (2,4). Shellfish, especially those eaten raw, are among the main foods involved in foodborne epidemics (2,5).Noroviruses are diverse, positive-stranded RNA viruses, classified into >10 genogroups (G) and many genotypes; most noroviruses that infect humans belong to genogroups GI and GII (6). Since 1995, the epidemiology of human noroviruses has been dominated by the GII.4 genotype (1). Of note, GII.4 appears to be predominantly transmitted person to person, whereas other genotypes, such as GII.6, GII.

Persistence of Human Norovirus (GII) in Surface Water: Decay Rate Constants and Inactivation Mechanisms

“…This variability has also been demonstrated for infectious HuNoV (GII.4) persistence in seawater, where HuNoV seeded into filter-sterilized seawater and was detectable via the HIE assay for between 7 and 14 days in one sample and for at least 35 days in another from the same location (no k values were reported). 24 These findings suggest that k values may vary greatly within environmental sample matrices; the cause is not clear but could be due to the transient presence of variable constituents in the environmental matrix, which could be biotic (e.g., enzymatic) or abiotic.…”

“…At the time of those reviews, there were only two studies , with available first-order decay rate constants for HuNoV (quantified via RT-qPCR). The only other data available for caliciviruses used murine norovirus (quantified via RT-qPCR and/or culture methods). − Since then, additional studies have been published describing inactivation (i.e., loss of infectious virus) of murine norovirus using cell culture (RAW-264.7) and HuNoV using cell culture (human intestinal enteroid (HIE) cells). , Given the importance of norovirus globally as an etiology of acute gastrointestinal illness, there is an urgent need to better understand its inactivation in water.…”

“…20−22 Since then, additional studies have been published describing inactivation (i.e., loss of infectious virus) of murine norovirus using cell culture (RAW-264.7) 23 and HuNoV using cell culture (human intestinal enteroid (HIE) cells). 24,25 Given the importance of norovirus globally as an etiology of acute gastrointestinal illness, there is an urgent need to better understand its inactivation in water.…”

Development of an amplicon-based high throughput sequencing method for genotypic characterisation of norovirus in oysters

“…As the target regions for RT-qPCR of norovirus are < 100 bp, it is not surprising to observe less than a perfect agreement between the HTS reads for 340/344 bp amplicons from a semi-nested PCR and gc/g as per RT-qPCR amplicons. Therefore, it is challenging to define the probability that norovirus viral RNA detected by RT-qPCR in shellfish is an intact and/or infectious virus (67, 68).…”

“…While this study builds on previous work (5, 8, 67) and enhances the capacity for surveillance and outbreak response on a national level, there are limitations. Much of the work presented was performed with spiked samples, which are not necessarily representative of naturally contaminated shellfish due to the quality and concentration of the RNA.…”