Rotavirus Genome Segment 7 (NSP3) Is a Determinant of Extraintestinal Spread in the Neonatal Mouse

Abstract

We used the neonatal mouse model of rotavirus infection to study extraintestinal spread following oral inoculation. Five-day-old pups were inoculated with either SA11-Cl3, SA11-Cl4, SA11-4F, RRV, or B223. By using virus detection in the liver as a proxy determination for extraintestinal spread, rotavirus strains capable of extraintestinal spread at high frequency (rhesus rotavirus [RRV]) and very low frequency (SA11-Cl4) were identified. Both strains productively infected the gastrointestinal tract. Oral inoculation of mice with RRV/ SA11-Cl4 reassortants and determination of virus titers in the gut and liver revealed that the extraintestinal spread phenotype segregated with RRV genome segment 7 to a high level of significance (P ‫؍‬ 10 ؊3 ). RRV segment 7 also segregated with the growth of virus in the gut (P ‫؍‬ 10 ؊5 ). Although infection of the gut was clearly required for tropism to the liver, there was no correlation between virus titers in the gut and detection of virus in the liver. Five days after intraperitoneal administration to bypass the gut barrier to virus spread, RRV and SA11-Cl4 both were recovered in the liver. However, only RRV was found in the liver following subcutaneous inoculation, suggesting that this peripheral site presented a similar barrier to virus spread as the gut. Sequence analysis of segment 7 from parental RRV and SA11-Cl4 and selected reassortants showed that (i) amino acid differences were distributed throughout the coding sequences and not concentrated in any particular functional motif and (ii) parental sequence was preserved in reassortants. These data support the hypothesis that NSP3, coded for by genome segment 7, plays a significant role in viral growth in the gut and spread to peripheral sites. The mechanism of NSP3-mediated tropism is under investigation.

Effect of Rotavirus Strain on the Murine Model of Biliary Atresia

“…Single-gene reassortants (i.e., all but one gene derived from one parent) allow for the determination of the function of that gene. Using such techniques, the extraintestinal spread of rotavirus has been linked to the NSP3 gene derived from RRV (23). The results of the current studies provide the framework for further manipulation.…”

“…Inoculation with the mouse strain EDIM and the human strain Wa caused no clinical signs of hepatobiliary injury, and consistent with the lack of injury, we found no detectable infectious virus within the liver. Previous studies have shown that the extraintestinal spread of rotavirus is strain dependent (22,23,32). In those studies, the presence of virus within the liver was used as a marker of spread (22,23).…”

“…Previous studies have shown that the extraintestinal spread of rotavirus is strain dependent (22,23,32). In those studies, the presence of virus within the liver was used as a marker of spread (22,23). The results of the current study expand on these observations showing that not only is the extraintestinal spread of rotavirus strain dependent but the specific targets of inoculation within the liver vary, which in turn, impacts the subsequent development of clinical manifestations of disease and the survival of the injected mouse.…”

A Lymphatic Mechanism of Rotavirus Extraintestinal Spread in the Neonatal Mouse

“…While the extraintestinal spread of rotavirus in the neonatal mouse model is well documented, the determinants and mechanisms of the spread of the virus are not as clear. It is known that extraintestinal spread occurs in a rotavirus straindependent manner (16,29). We previously demonstrated that among SA11-clone 4 (SA11-Cl4) and Rhesus rotavirus (RRV) reassortants, spread of the virus to the liver was significantly associated with RRV-derived genome segment 7, which encodes NSP3 (16).…”

“…Animal studies support these observations. Rotavirus and/or rotavirus antigen was recovered from the blood, liver, spleen, kidneys, lungs, and mesenteric lymph nodes (MLN) of orally inoculated mice (1,2,4,10,16,27), and mouse models for rotavirusinduced hepatitis (29) and biliary atresia (25) were previously described.…”