Introduction
Norovirus（NV）is one of the major causative agents of nonbacterial gastroenteritis. Other vi- ruses causing gastroenteritis include rotavirus
（RV）,astrovirus（AstV）,and adenovirus（Ad）.
NV can be genetically divided into two groups, genogroup I（GI）and genogroup II（GII）. Each genogroup can be further divided into nine GI genotypes and ten GII genotypes at present1）. Electron microscopy（EM）and reverse transcri- ption-PCR（RT-PCR）are used to detect NV, how- ever these methods require skillful techniques and are time consuming . Immunochromatogra- phy（IC）is a rapid, easy, and economical assay for NV screening. RV-IC and Ad-IC have been suc- cessfully developed, whereas an IC method for NV has not yet been reported2）. In this study, we developed an IC for NV（NV-IC）and evaluated it
using patient samples.
Materials and Methods
A total of 55 stool samples were collected from patients with gastroenteritis in five geographi- cally different pediatric clinics in Japan（Sapporo, Tokyo, Maizuru, Osaka, Saga）between July 2000 and June 2001. These samples were negative for RV, Ad, AstV, and bacteria. RT-PCR with com- mon primers3）4）was used to screen all samples for NV. A recombinant capsid protein from the NV 1207 strain（Lordsdale［LD］virus type, GII）was expressed in a baculovirus expression system5）6）. The polyclonal antibody for the NV-IC was ob- tained from a rabbit immunized with the recom- binant capsid protein （ recovered from SDS- PAGE）. The NV-IC strip contained a judgment line coated with the polyclonal antibody and a control line coated with anti-rabbit IgG antibody.
The stool suspension and a reagent（polyclonal antibody coated with latex ） were mixed and dropped onto the NV-IC strip. The line was ex- amined visually for a positive or negative result.
Evaluation of a Newly Developed Immunochromatographic Method for Detection of Norovirus
Michio OKAME
1）, Hainen YAN
1）, Shiho AKIHARA
1）, Shoko OKITSU
1）, Hideki TANI
2）, Yoshiharu MATSUURA
2）& Hiroshi USHIJIMA
1）1）Department of Developmental Medical Sciences, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan
2）Research Center for Emerging Infectious Diseases, Research Institute for Microbial Diseases, Osaka University, Osaka, Japan
（Received：March 17, 2003）
（Accepted：May 26, 2003）
〔J.J.A. Inf. D. 77：637〜639, 2003〕
Correspondence to：Hiroshi USHIJIMA
Dept. of Developmental Medical Sciences Graduate School of Medicine The Univ. of Tokyo
Hongo 7―3―1 Bunkyo-ku Tokyo 113―8654 Japan
Key words： norovirus, immunochromatography, RT-PCR
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平成15年 8 月20日
Table 1 Results of detection by RT-PCR and IC
RT-PCR
TOTAL
−
＋ Others LD virus type
26
2
0 24
IC ＋
29 20
5
4
−
55 22
5 28
TOTAL
Phylogenetic analysis of the capsid N!S domain was evaluated by NJ the method1）.
Results
The results are shown in Table 1. Thirty three of the 55 samples were positive for NV by RT- PCR and 26 were positive by NV-IC. Two sam- ples were positive for NV by NV-IC, but negative by RT-PCR. The sensitivity of NV-IC was 72.7％
（24!33）and specificity was 90.9％（20!22）.The concordance ratio with NV-IC and RT-PCR was 80.0％（44!55）. Phylogenetic analysis divided the NV sequences into four distinct genotypes. The majority of the RT-PCR positive sequences, 28 of the 33（84.8％），were grouped into the LD cluster, whereas 24 of the 26 NV-IC positive samples were grouped into the LD cluster.
Discussion
Compared with the result of RT-PCR, the NV- IC method had a high sensitivity and specificity.
This result shows that NV-IC is useful for NV screening . The fact that NV-IC detected 24
（85.7％）of the LD-cluster samples indicated that this NV-IC is specific for LD-like strains. NV GI and GII are antigenically distinct7）8）, but the anti- genic diversity within a genogroup remains un- clear . Other recombinant capsid proteins from different genotypes are needed to detect other strains using the NV-IC method . This was the first attempt to develop an IC assay for NV , therefore the NV-IC is still under development .
However, our results showed that NV-IC has the potential to become a new screening method for NV in the near future.
Acknowledgements
We are grateful to Mr . Nomura H , and Mr . Mori K. for their help in making the antibody and the immunochromatography.
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イムノクロマト法によるノロウイルス迅速診断法の開発と評価
1）東京大学大学院医学系研究科発達医科学教室，2）大阪大学微生物研究所エマージング感染症研究センター
大亀 路生1） 顔 海念1） 秋原 志穂1） 沖津 祥子1）
谷 英樹2） 松浦 善治2） 牛島 廣治1）
Immunochromatography for Norovirus 639
平成15年 8 月20日
