Sensitive Genotyping of Foodborne-Associated Human Noroviruses and Hepatitis A Virus Using an Array-Based Platform

Abstract

Human noroviruses (NoV) are the leading cause of human gastroenteritis in populations of all ages and are linked to most of the foodborne outbreaks worldwide. Hepatitis A virus (HAV) is another important foodborne enteric virus and is considered the most common agent causing acute liver disease worldwide. In the present study, a focused, low-density DNA microarray was developed and validated for the simultaneous identification of foodborne-associated genotypes of NoV and HAV. By employing a novel algorithm, capture probes were designed to target variable genomic regions commonly used for typing these foodborne viruses. Validation results showed that probe signals, specific for the tested NoV or HAV genotypes, were on average 200-times or 38-times higher than those detected for non-targeted genotypes, respectively. To improve the analytical sensitivity of this method, a 12-mer oligonucleotide spacer sequence was added to the capture probes and resulted in a detection threshold of less than 10 cRNA transcripts. These findings have indicated that this array-based typing sensor has the accuracy and sensitivity for identifying NoV and HAV genotypic profiles predominantly linked to food poisoning. The implementation of this typing sensor would thus provide highly relevant and valuable information for use in surveillance and outbreak attribution.

Molecular epidemiology of hepatitis A outbreaks in two districts in Indonesia in 2018: Same subtype, but different strains

“…Rapid tests have been widely used as screening tools in developing countries (5,23). Viral genotypic profiles are required to identify foodborne outbreaks, implement preventative measures and recognize transmission routes (3,(24)(25)(26). The VP1 region of HAV is an area that contains variable amino acids, which is why this region was used as one of the areas for molecular detection in the present study (19,27).…”

“…As subgenotype IA is prevalent, genotyping/subgenotyping alone can rarely be utilized to identify the source of an HAV outbreak or the chain of transmission (1,11). The HAV isolates that have been identified thus far display a high degree of genetic conservation, and modest genetic heterogeneity exists in several genomic regions, with the exception of the 5' untranslated region, which has demonstrated high levels of conservation, supporting the use of RT-PCR for the sensitive detection of HAV RNA (1,3,11,32,33).…”

“…Hepatitis A virus (HAV) infection, which is a major cause of acute hepatitis, poses an important public health problem worldwide (1)(2)(3). The virus can spread through the faecal-oral route, e.g.…”

A Survey of Analytical Techniques for Noroviruses

“…This was achieved through the use of RT-PCR amplicons in a single base extension reaction with labeled genotype specific probes and hybridization to a GenFlex TM Tag Array. In addition, Quinones et al [188] developed a microarray based platform for sensitive genotyping of 12 (GI/GII) HuNoV genotypes and 2 HAV genotypes from clinical samples, with a detection threshold of < 10 transcript copies. Won et al [189] also developed an oligonucleotide-based microarray for the detection of foodborne viruses (including NoV, HAV, human rotavirus and human astrovirus), with the selected probes having a detection limit of 100 ng DNA for each virus.…”

Advances in Diagnostic Approaches for Viral Etiologies of Diarrhea: From the Lab to the Field

“…The high rate of EPEC and EAEC detection in the stools of patients but also in controls has previously been reported by others (Jensen et al, 2014; Gomes et al, 2016). Recently, genotyping of Norovirus and Hepatitis A viruses was conducted using DNA microarray platform and it could detect less than 10 cRNA transcripts (Quiñones et al, 2017).…”