Norwalk Virus Infection of Volunteers: New Insights Based on Improved Assays

Abstract

Norwalk virus infection is a common cause of gastroenteritis in humans. The clinical features and virologic and immunologic responses following oral administration of Norwalk virus to 50 volunteers were monitored. New ELISAs using recombinant virus particles as the antigen source were used to assess the pattern of virus shedding and the specific immune responses. Forty-one subjects (82%) became infected; 68% were symptomatic and 32% were asymptomatic. The proportion of subjects infected was similar for those with and without preexisting antibody (82% vs. 60%; P > .2). The magnitude of seroconversion was highest in subjects who had vomiting. The peak of viral shedding was between 25 and 72 h, and virus first appeared in stool at 15 h. Specimens collected 7 days after inoculation remained positive. These results show a higher infection rate, more subclinical infections, and longer virus excretion following Norwalk virus inoculation than previously recognized.

Porcine enteric caliciviruses: Genetic and antigenic relatedness to human caliciviruses, diagnosis and epidemiology

“…Stool samples from pigs infected with porcine SaV Cowden strain or porcine NoV QW101 strain have been shown to be positive at dilutions ranging from 1:5 to 1: 125 and 1:5 to 1:12,500, respectively [68]. A similar dilution range (1:10 to 1:10,000) has also been found for human NoVs [80]. A drawback of this strategy is that dilution of weakly positive samples may exceed the detection limit of RT-PCR and therefore become negative.…”

“…The antigen ELISA for detection of NoVs has proven to be sensitive with a detection limit of 0.025 ng of capsid protein [80,89] and 1:10,000 dilutions of viral antigen in the stools of volunteers still being detectable [80]. The sensitivity of ELISA was similar to that of traditional RT-PCR probably due to the large amounts of viral soluble proteins in stools [6,29].…”

“…Hyperimmune antiserum produced to VLPs is often highly specific to the homologous strain or viruses within the same genotype [80,[89][90][91][92][93]. For example, the ELISA using hyperimmune serum against NoV GII-3/Mexico strain did not detect NoV GII-4/Grimsby virus, and vice versa [90].…”

An epidemiological investigation of Norwalk virus infection in South Africa

“…The EIA system of Graham and colleagues [14], as modified by Parker and colleagues [16], was used to detect IgG antibodies to NV. In the assay system horseradish peroxidase (HRP) conjugated rabbit antihuman IgG (Dako immunoglobulins, Denmark) was used with 3,3',5,5'-tetramethylbenzidine dihydrochloride (TMB) (Kirkegaard & Perry, Gaithersburg, MD) as substrate.…”

“…Limited epidemiological studies, carried out with antigens and antisera from human volunteer studies, have shown that in lessdeveloped areas such as Bangladesh, NV is contracted early in life [10], whilst in the USA it was an uncommon childhood infection [11]. The recent cloning and sequencing of the entire NV genome [12], and the consequent production of recombinant NV antigen (rNV) [13], has facilitated the development of sensitive and specific enzyme immunoassays (EIA) for the detection of NV antigen and antibodies [14][15][16]. In addition, the application of the reverse transcriptase polymerase chain reaction and subsequent molecular characterization of these viruses has enabled many SRSVs, including NV, to be classified within the Caliciviridae [17].…”