We have developed an assay for the detection of Norwalk-like viruses (NLVs) based on reverse transcription-PCR (RT-PCR) that is highly sensitive to a broad range of NLVs. We isolated virus from 71 NLV-positive stool specimens from 37 outbreaks of nonbacterial acute gastroenteritis and sequenced the open reading frame 1 (ORF1)-ORF2 junction region, the most conserved region of the NLV genome. The data were subjected to multiple-sequence alignment analysis and similarity plot analysis. We used the most conserved sequences that react with diverse NLVs to design primers and TaqMan probes for the respective genogroups of NLV, GI and GII, for use in a real-time quantitative RT-PCR assay. Our method detected NLV in 99% (80 of 81) of the stool specimens that were positive by electron microscopy, a better detection rate than with the two available RT-PCR methods. Furthermore, our new method also detected NLV in 20 of 28 stool specimens from the same NLV-related outbreaks that were negative for virus by electron microscopy. Our new assay is free from carryover DNA contamination and detects low copy numbers of NLV RNA. It can be used as a routine assay for diagnosis as well as for elucidation of the epidemiology of NLV infections.

Broadly Reactive and Highly Sensitive Assay for Norwalk-Like Viruses Based on Real-Time Quantitative Reverse Transcription-PCR

Norovirus classification and proposed strain nomenclature

https://link.springer.com/content/pdf/10.1007%2Fs10156-009-0669-9.pdf

Loop-mediated isothermal amplification (LAMP): a rapid, accurate, and cost-effective diagnostic method for infectious diseases

Norwalk-like caliciviruses (Noroviruses) cause over 90% of nonbacterial epidemic gastroenteritis worldwide, but the pathogenesis of norovirus infection is poorly understood because these viruses do not grow in cultured cells and there is no small animal model. Here, we report a previously unknown murine norovirus. Analysis of Murine Norovirus 1 infection revealed that signal transducer and activator of transcription 1-dependent innate immunity, but not T and B cell-dependent adaptive immunity, is essential for norovirus resistance. The identification of host molecules essential for murine norovirus resistance may provide targets for prevention or control of an important human disease.

https://science.sciencemag.org/content/sci/299/5612/1575.full.pdf

STAT1-Dependent Innate Immunity to a Norwalk-Like Virus

Norwalk-like viruses

To the Editor: Coxsackievirus A6 (CVA6) belongs to human enterovirus species A of the genus Enterovirus. According to a Japanese Infectious Agents Surveillance Report, this virus is one of the major causes of herpangina, an acute febrile disease characterized by vesicles, ulcers, and redness around the uvula, which occurs mainly in young children and infants (1).

In June 2011, a sudden increase in cases of hand, foot, and mouth disease (HFMD) at pediatric sentinel sites (≈3,000 pediatric hospitals and clinics) was reported to the National Epidemiologic Surveillance of Infectious Diseases System in Japan. Compared with past numbers of cases over 30 years of surveillance, the number of cases of HFMD per sentinel site peaked in week 28 (July) of 2011 (10.97 cases per sentinel), particularly in western Japan (2). According to the Infectious Agents Surveillance Report (as of September, 18, 2011), CVA6 was detected in 709 HFMD cases and 156 herpangina cases throughout Japan (1).

Clinical samples (throat swab specimens and feces) obtained from sentinel sites in Shimane, Hyogo, Hiroshima, and Shizuoka, Japan, were screened for enteroviruses by using an enterovirus-specific reverse transcription PCR and sequence analysis of the partial viral protein (VP)4/VP2 or VP1 region (3). Among 93 clinical samples from 108 HFMD case-patients, we identified 74 case-patients as CVA6 positive by sequence analysis.

On the basis of sequence analysis of the entire VP1 region (GenBank accession nos. {"type":"entrez-nucleotide-range","attrs":{"text":"AB649286-AB649291","start_term":"AB649286","end_term":"AB649291","start_term_id":"342166534","end_term_id":"342166544"}}AB649286-AB649291), the consensus sequence had 82.3%–82.5% nt identity (94.8%–95.4% aa identity) with the prototype CVA6 Gdula strain (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"AY421764","term_id":"40068436"}}AY421764). CVA6 was not isolated from clinical samples in a cell culture system. Therefore, most CVA6 strains were identified by molecular detection directly from clinical samples and sequence analysis. Some CVA6 strains were grown and isolated in suckling mice; these strains were antigenically identified as CVA6 by a neutralization test with specific antiserum against CVA6 (4).

In Japan, HFMD and herpangina are classified as category V infectious diseases. On the basis of clinical diagnosis, suspected infections were reported by pediatric sentinel sites on a weekly basis to the Infectious Disease Surveillance Center of the National Institute of Infectious Diseases (Tokyo, Japan). Typical clinical signs and symptoms of HFMD cases caused by CVA6 were fever, mild vesicles in oral mucosa, and skin blisters on hands, arms, feet, legs, buttocks, and nail matrixes (Figure). Some patients with HFMD had onychomadesis (periodic shedding of the nails) 1–2 months after onset of HFMD. Most cases of HFMD were self-limited. However, additional follow-up may be necessary for patients with onychomadesis who are treated at dermatology clinics.



Figure

Typical clinical manifestations of hand, foot, and mouth disease associated with coxsackievirus CVA6 in Shizuoka, Japan, June–July, 2011. A) Hand and arm of a 2.5-year-old boy; B) foot and C) buttocks of a 6-year-old boy; D) nail matrix of a 20-month ...



As in other countries in the Asia–Pacific region, major causes of HFMD in Japan were CVA16 and enterovirus 71. In 2010, enterovirus 71 was identified as a major cause of HFMD (2). In contrast, CVA6 was consistently associated with herpangina, as were CVA2, CVA4, CVA5, and CVA10, but CVA6 was occasionally detected in HFMD case-patients. CVA6 was the major cause of herpangina in 2007, but an increase in the detection rate of CVA6 in HFMD case-patients was reported in Japan in 2009 (2).

HFMD outbreaks caused by CVA6 were reported in Singapore, Finland, and Taiwan in 2007–2009 (5–8). Recent HFMD outbreaks in Finland and Spain were associated with cases of onychomadesis 1–2 months after onset of HFMD (6,8,9). In Japan, cases of onychomadesis after onset of HFMD were reported in 2009 (10). Therefore, changes in clinical outcomes of CVA6-associated diseases should be investigated.

Although most HFMD cases caused by CVA6 in Japan were mild, CVA6 was also detected in other clinical samples, including cerebrospinal fluid from a patient with acute encephalitis in Hiroshima, which reaffirmed possible additional clinical manifestations during an HFMD outbreak caused by CVA6. Careful surveillance of disease and infectious agent activities are crucial in monitoring CVA6-associated HFMD, onychomadesis, and neurologic diseases. Nucleotide identity between CVA6 strains in Finland (2008) (7) and Japan (2011) was ≈95% in the partial VP1 region. More detailed genetic, phenotypic, and epidemiologic analyses of CVA6 are needed to determine the role of CVA6 in HFMD outbreaks with or without onychomadesis.

Hand, foot, and mouth disease caused by coxsackievirus A6, Japan, 2011.

Surveillance results from pediatric cases and outbreaks of viral gastroenteritis in Japan are presented. In winter, both small round structured virus (SRSV, or Norwalk-like viruses) and rotavirus were detected from infants with gastroenteritis; however, in recent years, the prevailing time of SRSV infection has preceded that of rotavirus infection. Most nonbacterial gastroenteritis outbreaks were related to SRSV infection, and >60% of the outbreaks were caused by contaminated food. In small-sized outbreaks, raw oysters were the primary source of transmission. In large-sized outbreaks, school lunches and catered meals that were served at schools, banquet halls, and hospitals were most often implicated in the transmission of foodborne gastroenteritis.

Surveillance of viral gastroenteritis in Japan : pediatric cases and outbreak incidents

Aim: To understand the situation of influenza epidemics and neurologic complications during the influenza season in Japan.



Methods: The influenza surveillance system in Japan was reviewed and the data on influenza/influenza-like illness and neurologic complications, such as acute encephalitis/encephalopathy, were evaluated.



Results: From the data of the National Epidemiological Surveillance of Infectious Diseases (NESID), an increase in clinically diagnosed encephalitis/encephalopathy was clearly noted recently. Most cases reported were acute encephalitis/encephalitis. The number of cases of Reye’s syndrome reported was stable and low in number, since the time of the recommendation of not using aspirin for pediatric febrile disease, particularly for influenza-like illness, in Japan. These acute encephalitis/encephalopathy cases were not correlated clearly with clinically diagnosed influenza patients before the 1997/98 influenza season from the surveillance data, although several case reports have shown a relationship.



Conclusions: It is clarified from the NESID that neurologic complications such as acute encephalitis/ encephalopathy, apart from Reye’s syndrome, have occurred associated with influenza virus infection, especially among young children. Further investigation should be necessary to find the pathogenesis of this serious complication of influenza virus infection and whether this is unique in Japan, due to unclarified factor/factors, or whether it has not been recognized in other countries.

Influenza surveillance system of Japan and acute encephalitis and encephalopathy in the influenza season.

Nationwide rubella epidemic - Japan, 2013

The program of the National Epidemiological Surveillance of Infectious Diseases under the auspices of the Ministry of Health and Welfare started in 1981 apprehended in 1995 emergence of adenovirus type 7 in Japan. We analyzed the reported data of type 7 comparing with those of type 3, both belonging to the same subgenus B, and the following results were obtained: After 1981, the main serotypes in the reports of adenovirus isolation/detection were types 3, 2, and 1 in this order. The reports of isolation of adenovirus type 7 used to be extremely few, however, suddenly increased in 1995. In 1997, reports of isolation of adenovirus type 3 decreased and those of type 7 acquired the third place after those of types 2 and 3. Type 7 infection occur almost every month, but most frequently during May-September. The ages of cases from which type 7 was isolated were 0-4 years accounting for 55%, 5-9 years 35%, teens 6.3% and adults 4.0%, being similar proportions to those yielding type 3. Clinical diagnoses of cases yielding adenovirus type 7 were pharyngo-conjunctival fever (PCF) and influenza-like illness, these two accounting for half. The symptoms were severe, being characterized by higher maximum body temperature during the feverish period and severe pneumonia. Encephalitis and arthro-muscular pain were seen in only type 7-infected cases, although such cases were few.

Kazuyo Yamashita

Papers

Hand, foot, and mouth disease caused by coxsackievirus A6, Japan, 2011.

Surveillance of viral gastroenteritis in Japan : pediatric cases and outbreak incidents

Influenza surveillance system of Japan and acute encephalitis and encephalopathy in the influenza season.

Nationwide rubella epidemic - Japan, 2013

Trend of adenovirus type 7 infection, an emerging disease in japan