Fluorescent immunochromatography for rapid and sensitive typing of seasonal influenza viruses.

Akira Sakurai,Katsuyoshi Takayama,Naoki Kajiwara,Yoshihiro Sakoda,Masatoshi Okamatsu,Masako Hashimoto,Futoshi Shibasaki,Jitsuho Yamada,Yukuharu Kobayashi,Naoki Yamamoto,Namiko Nomura,Tsuruki Tamura,Yoshihiko Suda,Hiroshi Kida,Kwok H Chan

doi:10.1371/journal.pone.0116715

Abstract

Lateral flow tests also known as Immunochromatography (IC) is an antigen-detection method conducted on a nitrocellulose membrane that can be completed in less than 20 min. IC has been used as an important rapid test for clinical diagnosis and surveillance of influenza viruses, but the IC sensitivity is relatively low (approximately 60%) and the limit of detection (LOD) is as low as 10³ pfu per reaction. Recently, we reported an improved IC assay using antibodies conjugated with fluorescent beads (fluorescent immunochromatography; FLIC) for subtyping H5 influenza viruses (FLIC-H5). Although the FLIC strip must be scanned using a fluorescent reader, the sensitivity (LOD) is significantly improved over that of conventional IC methods. In addition, the antibodies which are specific against the subtypes of influenza viruses cannot be available for the detection of other subtypes when the major antigenicity will be changed. In this study, we established the use of FLIC to type seasonal influenza A and B viruses (FLIC-AB). This method has improved sensitivity to 100-fold higher than that of conventional IC methods when we used several strains of influenza viruses. In addition, FLIC-AB demonstrated the ability to detect influenza type A and influenza type B viruses from clinical samples with high sensitivity and specificity (Type A: sensitivity 98.7% (74/75), specificity 100% (54/54), Type B: sensitivity 100% (90/90), specificity 98.2% (54/55) in nasal swab samples) in comparison to the results of qRT-PCR. And furthermore, FLIC-AB performs better in the detection of early stage infection (under 13h) than other conventional IC methods. Our results provide new strategies to prevent the early-stage transmission of influenza viruses in humans during both seasonal outbreaks and pandemics.

Highlights

Influenza, commonly known as "the flu", is an infectious disease of birds and mammals caused by negative-strand RNA viruses of the family Orthomyxoviridae, mainly Influenza type A virus (IAV), Influenza B type virus (IBV), and Influenza C type virus [1]
To evaluate the ability of FLIC-AB to detect 7 subtypes of influenza A viruses; A/WSN/1933 for H1N1; A/Hyogo/YS/2011 for H1N1 2009 pdm; A/swine/Missouri/2124514/2006 for H2N3; A/Aichi/2/1968 for H3N2; A/Indiana/08/2011 for H3N2 variant virus (H3N2v), and A/duck/Hokkaido/ Vac-3/2007 for H5N1, A/Anhui/1/2013 for H7N9, and 2 subtypes of influenza B viruses; B/Tokyo/15480/2008 and B/Mass/3/1966 were tested in each single assay by using FLIC-AB or other control ICs; Prorast and Quick (Table 1)
The results showed that FLIC-AB was able to detect 3 IAVs (H1N1, H1N1pdm, H3N2), and one IBV (B/Tokyo/15480/2008) with limit of detection (LOD) of single digits in terms of pfu per reaction within 10 min and 15 min reading time of the device (Table 1)

Summary

Introduction

Commonly known as "the flu", is an infectious disease of birds and mammals caused by negative-strand RNA viruses of the family Orthomyxoviridae, mainly Influenza type A virus (IAV), Influenza B type virus (IBV), and Influenza C type virus [1]. IAVs are classified into 18 hemagglutinin (HA) and 11 neuraminidase (NA) subtypes (http:// www.cdc.gov/flu/about/viruses/types.htm). The influenza viruses with many combinations of subtypes of these 2 surface proteins have been isolated from aquatic birds, poultry, and other avian species. IAVs have the potential to become pandemic because of reassortment between avian and human viruses [2]. In contrast to IAVs, IBVs are only isolated from humans and seals, and consist of only 2 phylogenetic and antigenic lineages in human: the B/Victoria/2/87-like (Victoria) lineage and the B/ Yamagata/16/88-like (Yamagata) lineage [3,4,5]

Methods

Results

Conclusion