Characterization of intra- and inter-host norovirus P2 genetic variability in linked individuals by amplicon sequencing

Abstract

Noroviruses are the main cause of epidemics of acute gastroenteritis at a global scale. Although chronically infected immunocompromised individuals are regarded as potential reservoirs for the emergence of new viral variants, viral quasispecies distribution and evolution patterns in acute symptomatic and asymptomatic infections has not been extensively studied. Amplicons of 450 nts from the P2 coding capsid domain were studied using next-generation sequencing (454/GS-Junior) platform. Inter-host diversity between symptomatic and asymptomatic acutely infected individuals linked to the same outbreak as well as their viral intra-host diversity over time were characterized. With an average of 2848 reads per sample and a cutoff frequency of 0.1%, minor variant haplotypes were detected in 5 out of 8 specimens. Transmitted variants could not be confirmed in all infected individuals in one outbreak. The observed initial intra-host viral diversity in asymptomatically infected subjects was higher than in symptomatic ones. Viral quasispecies evolution over time within individuals was host-specific, with an average of 2.8 nt changes per day (0.0062 changes per nucleotide per day) in a given symptomatic case. Nucleotide polymorphisms were detected in 28 out of 450 analyzed nucleotide positions, 32.14% of which were synonymous and 67.86% were non-synonymous. Most observed amino acid changes emerged at or near blockade epitopes A, B, D and E. Our results suggest that acutely infected individuals, even in the absence of symptoms, which go underreported and may enhance transmission, may contribute to norovirus genetic variability and evolution.

Porcine circovirus 2 (PCV2) population study in experimentally infected pigs developing PCV2-systemic disease or a subclinical infection

“…On the contrary, the viral genetic variability of pigs developing overt clinical signs, clearly decreased from 2 wpi until the end of the study, although it was comparable to the other groups at the beginning (1 wpi). These data suggest a likely interaction between viral population diversity and disease severity, as reported for other pathogens 19,22,29 .…”

“…Currently, the vast majority of available knowledge has been obtained from a limited number of chronic/ life-lasting infectious diseases caused by RNA viruses, like the Human Immunodeficiency Virus, Hepatitis C Virus, Hepatitis B Virus 26 . A lower amount of data obtained from acute infections (still essentially caused by rapidly evolving RNA viruses) has demonstrated the existence of a noteworthy intra-host variability and suggested its effect on viral evolution and disease features 19,22,27 . However, the relationship between the mutant spectrum and the clinical outcome is often unclear, poorly understood and hardly generalizable.…”

“…However, the relationship between the mutant spectrum and the clinical outcome is often unclear, poorly understood and hardly generalizable. In fact, both positive and negative correlation between quasispecies diversity and disease severity have been described with contradictory results 19,22 .…”

“…Even if a higher sequencing error frequency could be hypothesized, the samples were processed using the same methods and sequenced in the same Ion Torrent run, making this scenario unlikely. Previous studies performed on human pathogens have evidenced the occurrence of remarkable differences in within-host variability between individuals 22,33 . Additionally, fluctuations in the genetic composition were proven to occur in viral population replicating in stable environments, suggesting that this disequilibria may provide a selective advantage to viruses that can be then fully exploited in changing environments 34 , like the ones induced by the host immune system or by replication in different cell or tissues.…”

“…Additionally, PCV2 interaction with the host immune system is particularly complex, since both immune activation and suppression have been proven to enhance viral replication, correlating with the emergence of overt clinical signs 17 , 18 . While an association between intra-host variability and pathogenesis or disease progression has been demonstrated for several human and animal infections 19 – 22 , such issue has never been investigated for PCV2.…”