Molecular evolutionary analysis of novel NSP4 mono-reassortant G1P[8]-E2 rotavirus strains that caused a discontinuous epidemic in Japan in 2015 and 2018.

In the 2010s, several unusual rotavirus strains emerged, causing epidemics worldwide. This study reports a comprehensive molecular epidemiological study of rotaviruses in Japan based on full-genome analysis. From 2014 to 2019, a total of 489 rotavirus-positive stool specimens were identified, and the associated viral genomes were analyzed by next-generation sequencing. The genotype constellations of those strains were classified into nine patterns (G1P[8] (Wa), G1P[8]-E2, G1P[8] (DS-1), G2P[4] (DS-1), G3P[8] (Wa), G3P[8] (DS-1), G8P[8] (DS-1), G9P[8] (Wa), and G9P[8]-E2). The major prevalent genotype differed by year, comprising G8P[8] (DS-1) (37% of that year's isolates) in 2014, G1P[8] (DS-1) (65%) in 2015, G9P[8] (Wa) (72%) in 2016, G3P[8] (DS-1) (66%) in 2017, G1P[8]-E2 (53%) in 2018, and G9P[8] (Wa) (26%) in 2019. The G1P[8]-E2 strains (G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E2-H1) isolated from a total of 42 specimens in discontinuous years (2015 and 2018), which were the newly-emerged NSP4 mono-reassortant strains. Based on the results of the Bayesian evolutionary analyses, G1P[8]-E2 and G9P[8]-E2 were hypothesized to have been generated from distinct independent inter-genogroup reassortment events. The G1 strains detected in this study were classified into multiple clusters, depending on the year of detection. A comparison of the predicted amino acid sequences of the VP7 epitopes revealed that the G1 strains detected in different years encoded VP7 epitopes harboring distinct mutations. These mutations may be responsible for immune escape and annual changes in the prevalent strains.

Whole-genome analysis of human group A rotaviruses in 1980s Japan and evolutionary assessment of global Wa-like strains across half a century.

Historically, the Wa-like strains of human group A rotavirus (RVA) have been major causes of gastroenteritis. However, since the 2010s, the circulation of non-Wa-like strains has been increasingly reported, indicating a shift in the molecular epidemiology of RVA. Although understanding RVA evolution requires the analysis of both current and historical strains, comprehensive pre-1980's sequencing data are scarce globally. We determined the whole-genome sequences of representative strains from six RVA gastroenteritis outbreaks observed at an infant home in Sapporo, Japan, between 1981 and 1989. These outbreaks were mainly caused by G1 or G3 Wa-like strains, resembling strains from the United States in the 1970s-1980s and from Malawi in the 1990s. Phylogenetic analysis of these infant home strains, together with Wa-like strains collected worldwide from the 1970s to 2020, revealed a notable trend: pre-2010 strains diverged into multiple lineages in many genomic segments, whereas post-2010 strains tended to converge into a single lineage. However, Bayesian skyline plot indicated near-constant effective population sizes from the 1970s to 2020, and selection pressure analysis identified positive selection only at amino acid 75 of NSP2. These results suggest that evidence supporting the influence of rotavirus vaccines, introduced globally since 2006, on Wa-like RVA molecular evolution is lacking at present, and phylogenetic analysis may simply reflect natural fluctuations in RVA molecular evolution. Evaluating the long-term impact of RV vaccines on the molecular evolution of RVA requires sustained surveillance.

Genetic analysis of human parechovirus type 5 isolated from children in Sapporo, Japan in the summer of 2018.

Human parechovirus (HPeV) types 1 and 3 are frequently detected in Japan, but HPeV5 is not detected. HPeV5 was isolated for the first time in Japan from seven clinical samples collected from children in Sapporo as part of the National Epidemiological Surveillance of Infectious Diseases from July to August in 2018. Seven HPeV5 strains that were detected in Sapporo (HPeV5 Sa) were analyzed in the VP1 region by direct sequencing using Sanger sequencing methods. Whole genome sequence of these strains was determined by next-generation sequencing. The VP1 region of HPeV5 Sa was closely related to HPeV5 strains detected in Belarus and Germany in 2018, and to those detected in Australia in 2019. The 3D polymerase region of HPeV5 Sa strains showed a high nucleotide identity to HPeV3 strain detected in Australia in 2013. These findings suggest that HPeV5 Sa is a recombinant virus of HPeV5 and HPeV3, and HPeV5 strains that are genetically closely related to each other may have circulated in Europe, Japan, and Australia between 2018 and 2019.

Molecular and clinical characterization of the equine-like G3 rotavirus that caused the first outbreak in Japan, 2016.

Since 2013, equine-like G3 rotavirus (eG3) strains have been detected throughout the world, including in Japan, and the strains were found to be dominant in some countries. In 2016, the first eG3 outbreak in Japan occurred in Tomakomai, Hokkaido prefecture, and the strains became dominant in other Hokkaido areas the following year. There were no significant differences in the clinical characteristics of eG3 and non-eG3 rotavirus infections. The eG3 strains detected in Hokkaido across 2 years from 2016 to 2017 had DS-1-like constellations (i.e. G3-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2), and the genes were highly conserved (97.5-100 %). One strain, designated as To16-12 was selected as the representative strain for these strains, and all 11 genes of this strain (To16-12) exhibited the closest identity to one foreign eG3 strain (STM050) seen in Indonesia in 2015 and two eG3 strains (IS1090 and MI1125) in another Japanese prefecture in 2016, suggesting that this strain might be introduced into Japan from Indonesia. Sequence analyses of VP7 genes from animal and human G3 strains found worldwide did not identify any with close identity (>92 %) to eG3 strains, including equine RV Erv105. Analysis of another ten genes indicated that the eG3 strain had low similarity to G2P[4] strains, which are considered traditional DS-1-like strains, but high similarity to DS-1-like G1P[8] strains, which first appeared in Asia in 2012. These data suggest that eG3 strains were recently generated in Asia as mono-reassortant strain between DS-1-like G1P[8] strains and unspecified animal G3 strains. Our results indicate that rotavirus surveillance in the postvaccine era requires whole-genome analyses.

Rotavirus genotype and Vesikari score of outpatients in Japan in the vaccine era.

BACKGROUND: Group A rotaviruses (RVs) are a major cause of severe gastroenteritis among infants and young children. In Japan, RV vaccines were introduced in 2011, leading to a reduction in severe gastroenteritis cases. Studies are required to assess the effectiveness of the vaccines and their effect on the prevalence of RV genotypes. METHODS: Fecal samples were collected from outpatients with RV gastroenteritis in a pediatric clinic in Sapporo, Japan, from 2010 to 2016. GPI genotypes were determined using reverse-transcription polymerase chain reaction. Clinical information and immunization records were obtained from outpatients after 2013. GPI genotypes and clinical features were compared between patients with and without a RV vaccine history. RESULTS: In total, 270 cases were genotyped. G1P[8]I1 (Wa-like G1P[8]) strains were dominant from 2010 to 2012. G1P[8]I2 (DS-1-like G1P[8]) strains appeared in 2012 and dominated in 2013 to 2015. G2P[4]I2 and G9P[8]I1 strains increased every 3 years (G2P[4]I2: 2011 and 2014, G9P[8]I1: 2010, 2013 and 2016). After the 2013 season, 137 cases were collected, 24 of which were vaccinated. Cases requiring drip infusion were fewer in the vaccination group than in the non-vaccination group (16.7% vs 52.2%). No patients required hospitalization in the vaccination group compared with 10.6% in the non-vaccination group. A severe Vesikari score was less common in the vaccination group than in the non-vaccination group (33.3% vs 78.8%). There was no significant difference in the GPI genotype distribution between the two groups. CONCLUSION: Rotaviruses vaccine effectiveness, regardless of GPI genotype, was confirmed in terms of alleviation of disease severity.

Clinical and Molecular Characteristics of Human Rotavirus G8P[8] Outbreak Strain, Japan, 2014.

During March-July 2014, rotavirus G8P[8] emerged as the predominant cause of rotavirus gastroenteritis among children in Hokkaido Prefecture, Japan. Clinical characteristics were similar for infections caused by G8 and non-G8 strains. Sequence and phylogenetic analyses suggest the strains were generated by multiple reassortment events between DS-1-like P[8] strains and bovine strains from Asia.

[A case of multiple skin metastases from cancer of the descending colon responding to FOLFIRI/cetuximab therapy].

A 64-year-old woman presented to our hospital with subcutaneous tumors in the right thoracic region. After undergoing a thorough medical evaluation, she was diagnosed with multiple skin metastases arising from cancer of the descending colon. Surgical resection of the primary lesion was performed and FOLFIRI (5-fluorouracil, levofolinate calcium, irinotecan) and cetuximab chemotherapy for the metastases was initiated. The patient subsequently entered remission and did not experience any major side effects. This case report details an effective therapy for colon cancer with multiple skin metastases and presents a discussion of the expression profiles of epidermal growth factor receptor in both the primary and metastatic lesions.

Efficacy and safety of pentavalent rotavirus vaccine in Japan: a randomized, double-blind, placebo-controlled, multicenter trial.

Rotavirus is the most common cause of severe gastroenteritis in children under 5 y of age. Estimates of disease burden in Japan suggest that between 26,500 and 78,000 children in this age group need hospitalization each year, resulting in a direct medical cost of 10 to 24 billion Yen. Since being introduced in routine infant immunization schedules in the United States in 2006, the oral live pentavalent rotavirus vaccine RV5 (RotaTeq™) has contributed to dramatic reductions in the incidence of rotavirus gastroenteritis (RVGE) and in health care resource utilization. This was a multicenter, randomized, double-blind, placebo-controlled, parallel-group study to evaluate the efficacy and safety of a 3-dose regimen of RV5 in healthy infants, age 6 to 12 weeks, at 32 sites across Japan. The results indicate that RV5 was significantly efficacious in preventing any severity [74.5% (95% confidence interval [CI]: 39.9%, 90.6%; p<0.001)], moderate-to-severe [80.2% (95% CI: 47.4%, 94.1%)], and severe [100% (95% CI: 55.4%, 100%)] RVGE caused by viruses with serotypes contained in the vaccine. The observed cases of RVGE included rotavirus types G1 (n=19), G3 (n=9), G9 (n=5) and one unspecified G serotype with P1A[8]. No G2 or G4 RVGE cases were observed, and this study was not powered to evaluate efficacy against individual serotypes. RV5 was generally safe and well tolerated in Japanese infants. These results are comparable to those observed in clinical studies conducted in other developed countries. Introduction of the vaccine in Japan may reduce disease burden and associated health care costs.

Efficacy of pectin solution for preventing gastro-esophageal reflux events in patients with percutaneous endoscopic gastrostomy.

The aim of this study was to determine the efficacy of pectin solution, which increases the viscosity of liquid nutrient, for prevention of gastro-esophageal reflux in comparison with half-solid nutrient. The subjects were 10 elderly patients undergoing percutaneous endoscopic gastrostomy feeding. Twenty-four-hour esophageal multichannel intraluminal impedance and pH testing was performed during intake of half-solid nutrient and a combination of pectin solution and liquid nutrient. During 4 h after delivery, there was no significant difference in the total number of gastro-esophageal reflux events between the feeding of the half-solid nutrient and the combination of pectin solution and liquid nutrient (5.7 ± 1.2 vs 5.3 ± 1.0/4 h). Acidic reflux after delivery of the half-solid nutrient was significantly more frequent than that after delivery of the combination of pectin solution and liquid nutrient (80.7% vs 60.4%, p = 0.018). The incidence of gastro-esophageal reflux reaching the upper portion of the esophagus tended to be higher during delivery of the half-solid nutrient than during delivery of the combination of pectin solution and liquid nutrient (47.4% vs 34.0%, p = 0.153). In conclusion, the usage of pectin solution combined with liquid nutrient is effective for preventing acidic gastro-esophageal reflux and gastro-esophageal reflux reaching the upper portion of the esophagus.

Full sequence analysis of the original Sapporo virus.

In this study, the full-length genome sequence of the prototype of sapovirus, namely Sapporo virus (SV82), was identified. Sapporo virus RNA was extracted from a fecal sample, amplified by RT-PCR and the PCR products sequenced directly and analyzed. Sequence analysis showed that Sapporo virus consists of 7433 nucleotides and has three open reading frames. The Sapporo strain shows 91.7% nucleotide sequence identity to the Manchester virus. Phylogenic analysis has also revealed the closeness of Sapporo virus to other sapovirus/genogroup I strains. Basic information on the evolutionary history of sapovirus analysis is provided here.

Effects of mosapride on esophageal functions and gastroesophageal reflux.

BACKGROUND AND AIM: A substantial number of patients with gastroesophageal reflux disease show symptomatic resistance to high-dose proton pump inhibitors. In those cases, prokinetics are possible candidates for treatment. The aim of the present study was to determine whether mosapride, a prokinetic agent, stimulates esophageal functions, and prevents acidic and non-acidic gastroesophageal reflux. METHODS: Normal volunteers (nine and 13 for two experiments, respectively) were enrolled. Salivary secretion, esophageal peristaltic contractions, and resting lower esophageal sphincter pressure with and without mosapride administration were recorded using a cross-over protocol. Post-prandial acidic and non-acidic reflux levels were also recorded. RESULTS: Mosapride at a standard dose of 15 mg/day did not stimulate salivary secretion or any esophageal motor functions. It also failed to prevent acidic and non-acidic post-prandial gastroesophageal reflux. CONCLUSIONS: Mosapride at 15 mg/day, a standard dose in Japan, did not change the esophageal motility and salivary secretion in healthy volunteers. Future study on a larger number of individuals with higher dose of mosapride is worthwhile.

Effects of anti-hypertensive drugs on esophageal body contraction.

AIM: To clarify the effects of anti-hypertensive drugs on esophageal contraction and determine their possible relationship with gastro-esophageal reflux disease. METHODS: Thirteen healthy male volunteers were enrolled. Esophageal body peristaltic contractions and lower esophageal sphincter (LES) pressure were measured using high resolution manometry. All subjects were randomly examined on four separate occasions following administrations of nifedipine, losartan, and atenolol, as well as without any drug administration. RESULTS: Peristaltic contractions by the esophageal body were separated into three segments by two troughs. The peak peristaltic pressures in the mid and lower segments of the esophageal body under atenolol administration were significantly higher than those without medication in a supine position. On the other hand, peristaltic pressures under nifedipine administration were lower than those observed without drug administration. Losartan did not change esophageal body peristalsis. Atenolol elevated LES pressure and slowed peristaltic wave transition, while the effects of nifedipine were the opposite. CONCLUSION: Among the anti-hypertensive drugs tested, atenolol enhanced esophageal motor activity, which was in contrast to nifedipine.

Comparative Study of 5-Day and 10-Day Cefditoren Pivoxil Treatments for Recurrent Group A beta-Hemolytic Streptococcus pharyngitis in Children.

Efficacy of short-course therapy with cephalosporins for treatment of group A beta-hemolytic streptococcus (GABHS) pharyngitis is still controversial. Subjects were 226 children with a history of at least one episode of GABHS pharyngitis. Recurrence within the follow-up period (3 weeks after initiation of therapy) occurred in 7 of the 77 children in the 5-day treatment group and in 1 of the 149 children in the 10-day treatment group; the incidence of recurrence being significantly higher in the 5-day treatment group. Bacteriologic treatment failure (GABHS isolation without overt pharyngitis) at follow-up culture was observed in 7 of the 77 children in the 5-day treatment group and 17 of the 149 children in the 10-day treatment group. There was no statistical difference between the two groups. A 5-day course of oral cephalosporins is not always recommended for treatment of GABHS pharyngitis in children who have repeated episodes of pharyngitis.

Detection of enteric viruses in rectal swabs from children with acute gastroenteritis attending the pediatric outpatient clinics in Sapporo, Japan.

BACKGROUND: Gastroenteritis is a world-wide disorder. Numerous studies to identify causative viral agents have been reported for hospitalized patients but there are only a few for outpatients with mild symptoms who are usually managed in the outpatient clinics. OBJECTIVES: Our aim was to clarify the epidemiological and clinical characteristics of acute gastroenteritis in children who visited the outpatient clinics with various complaints suggestive of gastroenteritis. STUDY DESIGN: From December 2003 to December 2005, 877 rectal swabs were collected from patients attending outpatient clinics in Sapporo, Japan. Viral genomes of major five enteric viruses (rotavirus, norovirus, adenovirus, astrovirus and sapovirus) and bocavirus were investigated by RT-PCR or PCR. RESULTS: At least one viral agent was found in 326 (37.2%) cases of the 877 studied. Rotaviruses were the most prevalent and were detected in 143 (16.3%) followed by norovirus in 116 (13.2%), adenovirus in 42 (4.8%), astrovirus in 40 (4.6%) and sapovirus in 15 (1.7%) cases. Bocavirus was detected in only 4 (0.5%) cases. Frequent diarrhea and frequent vomiting were prominent in rotavirus and norovirus infection, respectively. CONCLUSIONS: The prevalence of each enteric virus in outpatients resembled that previously estimated in hospitalized patients, although the detection rate of rotavirus was slightly low. The contribution of bocavirus appears to be small.

Half-solidification of nutrient does not decrease gastro-esophageal reflux events in patients fed via percutaneous endoscopic gastrostomy.

BACKGROUND & AIMS: Percutaneous endoscopic gastrostomy (PEG) tube feeding is widely used for patients with swallowing dysfunction, and aspiration pneumonitis induced by reflux of gastric contents is one of the most frequent and life-threatening events in such patients. We performed this study to clarify the effect of half-solidification of nutrient on gastro-esophageal reflux (GER) of acidic and non-acidic gastric contents in patients with PEG. METHODS: The subjects were 14 elderly patients undergoing PEG feeding (female: 11, mean age 83.1 y). Twenty-four hour esophageal multichannel intraluminal impedance (MII) and pH (MII-pH) monitoring were performed during and after feeding with half-solid and liquid nutrient, respectively. The numbers of GER events during 4h after the start of feeding were compared. RESULTS: There was no significant difference in the total number of GER events between half-solid nutrient feeding and liquid nutrient feeding (5.6+/-1.5 vs. 6.6+/-1.5). The number of acid reflux events after half-solid nutrient feeding tended to be higher than that after liquid nutrient feeding (1.9+/-1.1 vs. 1.4+/-0.8). CONCLUSION: Half-solidification of nutrition does not appear to be effective for preventing GER caused by liquid PEG feeding.

Esophageal epithelial surface in patients with gastroesophageal reflux disease: an electron microscopic study.

AIM: To investigate the intercellular spaces between the most superficially located esophageal epithelial cells in patients with gastroesophageal reflux disease (GERD). METHODS: Eighteen patients with erosive esophagitis, 10 patients with non-erosive reflux disease (NERD), and 18 normal asymptomatic volunteers were enrolled. Biopsy specimens were obtained from the lower esophageal mucosa without ulcer or erosion. Scanning electron microscopy was employed to investigate the tightness of the superficial cellular attachment. RESULTS: The intercellular space between the most superficially located epithelial cells in patients with erosive esophagitis or NERD was not different from that in asymptomatic healthy individuals. CONCLUSION: Widened luminal intercellular spaces of esophageal superficial epithelium are not responsible for the induction of reflux symptoms in patients with GERD.

Efficacy of antibiotic prophylaxis for intrafamilial transmission of group A beta-hemolytic streptococci.

BACKGROUND: The role of chemoprophylaxis for household contacts of patients with acute streptococcal disease is uncertain. METHODS: The subjects were 1440 sibling contacts of 1181 index patients with group A beta-hemolytic streptococcal (GABHS) pharyngitis. Instances of subsequent GABHS pharyngitis in sibling contacts who received chemoprophylaxis and in a control group without prophylaxis were compared. RESULTS: Of the 948 siblings in the prophylaxis group, 507 were treated with cephalosporins and 441 were treated with penicillins for 3 to 5 days. Subsequent GABHS pharyngitis occurred within 30 days in 28 (3.0%) of the 948 siblings in the prophylaxis group and in 26 (5.3%) of the 492 siblings in the control group. Among siblings in the prophylaxis group, subsequent GABHS pharyngitis occurred in 9 (1.8%) of the 507 siblings in the cephalosporin prophylaxis group and in 19 (4.3%) of the 441 siblings in the penicillin prophylaxis group. When these data were each compared with that in the control group (5.3%), a significant statistical difference was seen in the cephalosporin prophylaxis group (P = 0.003) but not in the penicillin prophylaxis group (P = 0.542). Only 5-day cephalosporin prophylaxis showed significant reduction in the rate of subsequent GABHS pharyngitis compared with that in the control group (P = 0.002). CONCLUSIONS: In view of the low incidence of subsequent GABHS pharyngitis in the nonprophylaxis group, the usual self-limited nature of GABHS pharyngitis, the cost of prophylaxis and the risk for selecting resistant flora, routine chemoprophylaxis against GABHS pharyngitis for sibling contacts is not recommended.

Virological, serological, and clinical features of an outbreak of acute gastroenteritis due to recombinant genogroup II norovirus in an infant home.

Norovirus (NV) is an important cause of acute nonbacterial gastroenteritis worldwide. Recently, several sporadic cases due to naturally occurring recombinant NVs have been reported. In January 2000, there was an outbreak of gastroenteritis in an infant home in Sapporo, Japan. Of 34 residents of the home that were less than 2 years old, 23 developed gastrointestinal symptoms and NV infection was confirmed by conventional reverse transcription-PCR to detect the RNA polymerase region of genogroup II NV. In this virus, the RNA polymerase region shared 86% nucleotide identity with Hawaii virus but only 77% with Mexico virus; however, its capsid region shared only 70% identity with Hawaii virus but 90% with Mexico virus. On the other hand, both regions shared a higher 96% nucleotide identity with Arg320 virus, which was found in Mendoza, Argentina, in 1995 and considered to be a recombinant of Hawaii and Mexico viruses. The findings indicate that the virus involved in the outbreak was similar and may have evolved from the Arg320 virus. Clinically the cases were more severe than those of previously reported sporadic or outbreak cases of NV infection.

[Multi-center analysis of calcinosis in children with juvenile dermatomyositis].

OBJECTIVES: To reveal the frequency and the clinical characteristics of dystrophic calcification that occurs in children with juvenile dermatomyositis, multi-center analysis was constructed. METHOD: Fifty children with JDM were enrolled, and 14 of them (28.0%) were complicated with calcinosis. Clinical symptoms and laboratory tests at onset, initial therapy and disease course were compared in children with and without calcinosis. RESULTS: The mean age of the onset of calcinosis was 4.78 +/- 3.33 years, and it was younger than those of children without calcinosis (8.66 +/- 3.85 years) (P = 0.0017). No differences of clinical manifestation except Gower's sign were observed. The frequency of positive anti-nuclear antibody was 7.1% in children with calcinosis and 52.9% without calcinosis (P = 0.0112). The initial therapy of methylprednisolon pulses gave no effects on prognosis of calcium deposition. The calcinosis appeared in 1.56 +/- 1.91 year after the onset of the disease. The various types of calcium deposition including large tumorous clumps, subcutaneous plaques or nodules, sheet-type calcification were deserved. They appeared over knee joints (64.3%), elbow joint (64.3%), and hip processes (50.0%). Calcinosis affecting the subcutaneous tissues frequently resulted in painful superficial ulceration of the overlying skin (42.9%), local infection (50.0%), and limitation of joint movement (14.3%). Although aluminum phosphate was effective in 2 children among 7, no other effective treatment was recommended. In 5 cases, surgical removal of tumorous clumps was operated. Thus, juvenile dermatomyositis is frequently complicated with calcinosis. This type of calcinosis was found to be unlikely to resolve completely, and resulted in severe disability in children.