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Neutralizing antibodies (NAbs) to human cytomegalovirus (HCMV) are associated with the risk of transplacental HCMV infection of the fetus in pregnant women. The IgG-positivity rate to HCMV determined by enzyme immunoassay (EIA) or indirect immunofluorescence assay has decreased from approximately 100% to 70% over the past 30 years in Japan. We tested serum samples from 630 Japanese women aged 20-49 years whose blood samples were obtained between 1980 and 2015. IgG titer was measured using an EIA-based assay. HCMV-NAb titer was measured using a neutralization test assay with an HCMV isolate on human retinal epithelial cells. Longitudinal transitions in HCMV-NAb prevalence were clarified. The prevalence of HCMV-EIA-IgG, and HCMV-NAb at a titer of 16-fold, and HCMV-NAb at a titer of 100-fold, changed from 96.7% to 78.9%, 93.3% to 85.6%, and 35.5% to 41.1%, respectively, between 1980-1990 and 2010-2015. Prevalence of HCMV-NAb at a titer of 16-fold decreased by 7.7%, whereas that at a titer of 100-fold increased by 5.6%. A high titer of HCMV-NAb in pregnant women is expected to reduce the risk of intrauterine HCMV transmission from the mother to the fetus. The association between the risk of congenital HCMV infection and the prevalence of HCMV-NAb remains to be addressed.

Preterm birth is known to be associated with chronic disease risk in adulthood whereby epigenetic memory may play a mechanistic role in disease susceptibility. Gestational age (GA) is the most important prognostic factor for preterm infants, and numerous DNA methylation alterations associated with GA have been revealed by epigenome-wide association studies. However, in human preterm infants, whether the methylation changes relate to transcription in the fetal state and persist after birth remains to be elucidated. Here, we identified 461 transcripts associated with GA (range 23–41 weeks) and 2093 candidate CpG sites for GA-involved epigenetic memory through analysis of methylome (110 cord blood and 47 postnatal blood) and transcriptional data (55 cord blood). Moreover, we discovered the trends of chromatin state, such as polycomb-binding, among these candidate sites. Fifty-four memory candidate sites showed correlation between methylation and transcription, and the representative corresponding gene was UCN, which encodes urocortin.

The cardinal symptom of acute encephalopathy is impairment of consciousness of acute onset during the course of an infectious disease, with duration and severity meeting defined criteria. Acute encephalopathy consists of multiple syndromes such as acute necrotizing encephalopathy, acute encephalopathy with biphasic seizures and late reduced diffusion and clinically mild encephalitis/encephalopathy with reversible splenial lesion. Among these syndromes, there are both similarities and differences. In 2016, the Japanese Society of Child Neurology published 'Guidelines for the Diagnosis and Treatment of Acute Encephalopathy in Childhood', which made recommendations and comments on the general aspects of acute encephalopathy in the first half, and on individual syndromes in the latter half. Since the guidelines were written in Japanese, this review article describes extracts from the recommendations and comments in English, in order to introduce the essence of the guidelines to international clinicians and researchers.

Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) is a severe encephalopathy preceded by viral infections with high fever. AESD is a multifactorial disease, however, few disease susceptibility genes have previously been identified. Here, we conducted a genome-wide association study (GWAS) and assessed functional variants in non-coding regions to study genetic susceptibility in AESD using 254 Japanese children with AESD and 799 adult healthy controls. We also performed a microRNA enrichment analysis using GWAS statistics to search for candidate biomarkers in AESD. The variant with the lowest p-value, rs1850440, was located in the intron of serine/threonine kinase 39 gene (STK39) on chromosome 2q24.3 (p = 2.44 × 10−7, odds ratio = 1.71). The minor allele T of rs1850440 correlated with the stronger expression of STK39 in peripheral blood. This variant possessed enhancer histone modification marks in STK39, the encoded protein of which activates the p38 mitogen-activated protein kinase (MAPK) pathway. In the replication study, the odds ratios of three SNPs, including rs1850440, showed the same direction of association with that in the discovery stage GWAS. One of the candidate microRNAs identified by the microRNA enrichment analysis was associated with inflammatory responses regulated by the MAPK pathway. This study identified STK39 as a novel susceptibility locus of AESD, found microRNAs as potential biomarkers, and implicated immune responses and the MAPK cascade in its pathogenesis.

Objective: This study was carried out to understand the disparities in mortality and survival without major morbidities among very premature and very low birth weight infants between participating Neonatal Intensive Care Units (NICUs) from the Brazilian Network on Neonatal Research (RBPN) and the Neonatal Research Network of Japan (NRNJ). Methods: Secondary data analysis of surveys by the RBPN and NRNJ was performed. The surveys were conducted in 2014 and 2015 and included 187 NICUs. Primary outcome was mortality or survival without any major morbidity. Logistic regression analysis adjustment for confounding factors was used. Results: The study population consisted of 6,406 infants from the NRNJ and 2,319 from the RBPN. Controlling for various confounders, infants from RBPN had 9.06 times higher adjusted odds of mortality (95%CI 7.30–11.29), and lower odds of survival without major morbidities (AOR 0.36; 95%CI 0.32–0.41) compared with those from the NRNJ. Factors associated with higher odds of mortality among Brazilian NICUs included: Air Leak Syndrome (AOR 4.73; 95%CI 1.26–15.27), Necrotizing Enterocolitis (AOR 3.25; 95%CI 1.38–7.26), and Late Onset Sepsis (LOS) (AOR 4.86; 95%CI 2.25–10.97). Conclusions: Very premature and very low birth weight infants from Brazil had significantly higher odds for mortality and lower odds for survival without major morbidities in comparison to those from Japan. Additionally, we identified the factors that increased the odds of in-hospital neonatal death in Brazil, most of which was related to LOS. RESUMO Objetivo: Este estudo foi realizado para compreender as disparidades na mortalidade e sobrevivência sem as principais morbidades entre recém-nascidos muito prematuros e de muito baixo peso entre Unidades de Terapia Intensiva Neonatal (UTINs) participantes da Rede Brasileira de Pesquisas Neonatais (RBPN) e Rede de Pesquisa Neonatal do Japão (NRNJ). Métodos: Foi realizada uma análise dos dados secundários dos bancos de dados da RBPN e da NRNJ. As pesquisas foram realizadas em 2014 e 2015 e incluíram 187 UTINs. O desfecho primário foi mortalidade ou sobrevida sem qualquer morbidade importante. Utilizou-se a análise de regressão logística com ajuste para os fatores de confusão. Resultados: A população do estudo foi composta por 6.406 recém-nascidos do NRNJ e 2.319 do RBPN. Ajustando para diversos fatores de confusão, os prematuros da RBPN tiveram 9,06 vezes maiores chances de mortalidade (IC95% 7,30–11,29) e menores chances de sobrevivência sem morbidades importantes (AOR 0,36; IC95% 0,32–0,41) em comparação com os da NRNJ. Fatores associados a maiores chances de mortalidade entre as UTINs brasileiras incluíram: síndrome de escape de ar (AOR 4,73; IC95% 1,26–15,27), enterocolite necrosante (AOR 3,25; IC95% 1,38–7,26) e sepse de início tardio (AOR 4,86; IC95% 2,25–10,97). Conclusões: Os recém-nascidos muito prematuros e de muito baixo peso do Brasil apresentaram chances significativamente maiores de mortalidade e menores chances de sobrevivência sem as principais morbidades em comparação aos do Japão. Além disso, identificamos os fatores que aumentam as chances da morte neonatal no Brasil, sendo a maioria relacionada à sepse tardia.

Objectives Acute encephalopathy is an acute brain dysfunction after preceding infection, consisting of multiple syndromes. Some syndromes, such as acute encephalopathy with biphasic seizures and late reduced diffusion (AESD), are severe with poor outcome, whereas others, such as clinically mild encephalitis/encephalopathy with reversible splenial lesion (MERS), are mild with favorable outcome. Previous study reported the association of the thermolabile polymorphism in Carnitine Palmitoyltransferase 2 (CPT2) gene and severe syndromes of acute encephalopathy. To further explore the pathogenetic role of CPT2 in acute encephalopathy, we conducted a case-control association study of a typical thermolabile CPT2 polymorphism, rs2229291, in 416 patients of acute encephalopathy, including both severe and mild syndromes. Methods The case cohort consisted of 416 patients, including AESD, MERS, and other syndromes. The control subjects were 100 healthy Japanese. rs2229291 was genotyped by Sanger sequencing. Genetic distribution was compared between the patients and controls using Cochran-Armitage trend test. Results Minor allele frequency of rs2229291 was significantly higher in AESD (p = 0.044), MERS (p = 0.015) and entire acute encephalopathy (p = 0.044) compared to the controls. The polymorphism showed no significant association with influenza virus, or with outcome. Conclusions This study provided evidence that CPT2 is a susceptibility gene for overall acute encephalopathy, including both severe and mild syndromes, and suggested that impairment of mitochondrial metabolism is common to various syndromes of acute encephalopathy.

Background Epilepsy and autism spectrum disorder (ASD) are the common neurological manifestations of tuberous sclerosis complex (TSC). EXIST-3 study has recently demonstrated that everolimus reduces seizures in patients with TSC and refractory epilepsy. Here we report the efficacy and safety of everolimus for treatment-refractory seizures in Japanese patients of EXIST-3, along with the exploratory analysis evaluating the everolimus effect on comorbid ASD symptoms in these patients. Methods Primary end point was change in seizure frequency from baseline defined as response rate (≥50% reduction) and median percentage reduction in the seizure frequency. Pervasive Developmental Disorders Autism Society Japan Rating Scale (PARS) scores were assessed at baseline and at week-18 for ASD symptoms. Results Overall, 35 Japanese patients were randomized to everolimus low-exposure (LE; n = 10), everolimus high-exposure (HE; n = 14), or placebo (n = 11). The response rate was 30.0% and 28.6% versus 0% with the everolimus LE and HE versus placebo arm, respectively. Similarly, the median percentage reduction in seizure frequency was 6.88% and 38.06% versus −6.67%. Stomatitis was the most frequently reported adverse event (everolimus LE, 100%; HE, 78.6%; placebo, 9.1%). Four of 11 patients with ASD in the everolimus arms and 1 of 8 patients with ASD in the placebo arm showed ≥5 point decrease in PARS scores. Conclusions Adjunctive everolimus treatment improved seizure frequency with a tolerable safety relative to placebo among 35 Japanese patients with TSC-associated refractory seizures, consistent with the results of overall EXIST-3 study involving 366 patients. A favorable trend towards the improvement of ASD symptoms was observed.

Background Human cytomegalovirus (HCMV) causes asymptomatic infections, but also causes congenital infections when women were infected with HCMV during pregnancy, and life-threatening diseases in immunocompromised patients. To better understand the mechanism of the neutralization activity against HCMV, the association of HCMV NT antibody titers was assessed with the antibody titers against each glycoprotein complex (gc) of HCMV. Methods Sera collected from 78 healthy adult volunteers were used. HCMV Merlin strain and HCMV clinical isolate strain 1612 were used in the NT assay with the plaque reduction assay, in which both the MRC-5 fibroblasts cells and the RPE-1 epithelial cells were used. Glycoprotein complex of gB, gH/gL complexes (gH/gL/gO and gH/gL/UL128–131A [PC]) and gM/gN were selected as target glycoproteins. 293FT cells expressed with gB, gM/gN, gH/gL/gO, or PC, were prepared and used for the measurement of the antibody titers against each gc in an indirect immunofluorescence assay (IIFA). The correlation between the IIFA titers to each gc and the HCMV-NT titers was evaluated. Results There were no significant correlations between gB-specific IIFA titers and the HCMV-NT titers in epithelial cells or between gM/gN complex-specific IIFA titers and the HCMV-NT titers. On the other hand, there was a statistically significant positive correlation between the IIFA titers to gH/gL complexes and HCMV-NT titers. Conclusions The data suggest that the gH/gL complexes might be the major target to induce NT activity against HCMV.

Rotavirus vaccines have been available in Japan since 2011. This study conducted to monitor the trend of group A rotavirus (RVA) genotypes 3 years after vaccine introduction. A total of, 539 fecal samples were collected from children with acute gastroenteritis in six regions during July 2014-June 2015. Among them, 178 samples (33.0%) were positive for RVA. The most predominant genotype was G1P[8] (35.9%) followed by G2P[4] (26.4%), G9P[8] (21.3%), G3P[8] (4.5%), and G3P[9] (4.5%). The detection rate of G2P[4] was increased soon after vaccine introduction. Sequence analyses of VP7 and VP4 genes of the representative G2P[4] strains were found to be clustered in sub-lineage IVa of lineage IV. It is noteworthy that one amino acid substitution in the antigenic epitope (Q114P) of VP4 gene was found in representative G2P[4] strains of the current study. However, it is unclear whether the change in antigenic epitope is due to the effect of vaccination or due to natural variation, warranting further continuous monitoring of rotavirus evolution after vaccine introduction.

Background Antiviral-resistant herpes simplex virus type 1 (HSV-1) has been recognized as an emerging clinical problem among patients undergoing hematopoietic stem cell transplantation (HSCT). Methods A prospective observational study was conducted at a hematological center over a 2-year period. Oropharyngeal swab samples were serially collected each week from 1 week before and up to 100 days after HSCT and were tested for virus isolation. The HSV-1 isolates were tested for sensitivity to acyclovir (ACV). The prognosis of patients with ACV-resistant (ACVr) HSV-1 and the genetic background of the ACVr HSV-1 isolates were assessed. Results Herpes simplex virus type 1 was isolated in 39 of 268 (15%) HSCT patients within 100 days after transplantation. Acyclovir-resistant HSV-1 emerged in 11 of these 39 patients (28%). The 100-day death rates of HSCT patients without HSV-1 shedding, those with only ACV-sensitive HSV-1 shedding, and those with ACVr HSV-1 shedding were 31%, 39%, and 64%, respectively. Patients with HSV-1, including ACVr HSV-1, shedding showed a significantly higher mortality rate. Relapsed malignancies were a significant risk factor for the emergence of ACVr HSV-1. Acyclovir resistance was attributable to viral thymidine kinase and DNA polymerase mutations in 6 and 5 patients, respectively. Conclusions Herpes simplex virus type 1, including ACVr HSV-1, shedding was associated with poorer outcome in HSCT patients, even if HSV disease did not always occur. Patients with relapsed malignancies were at especially high risk for the emergence of ACVr HSV-1.

Monkeypox virus (MPXV) causes human monkeypox (human MPX), which is a similar disease to smallpox in humans. A previous study showed that a single vaccination of monkeys with LC16m8, a highly attenuated smallpox vaccine, protected them from MPX from 4-5 weeks post-vaccination. In this study, we evaluated the long-term efficacy of a single vaccination with LC16m8 in a nonhuman primate model of MPXV infection. The monkeys were inoculated with either LC16m8, Lister (parental strain of LC16m8), or a mock-up vaccine, and then challenged with MPXV via a subcutaneous route, at 6 and 12 months after vaccination, which we compared with either Lister or the mock-up vaccination. The LC16m8 monkeys exhibited almost no MPX-associated symptoms, whereas most of the naïve monkeys died. LC16m8 generated the protective memory immune response against MPXV, as suggested by the immediate viremia reduction and the response of the IgG antibody. The results demonstrated that the vaccination of monkeys with a single dose of LC16m8 provided durable protection against MPXV for longer than one year after immunization. The results suggest that the vaccination of humans with LC16m8 could induce long-term protection against MPXV infection.

Background We previously reported the nationwide, epidemiological data of acute encephalopathy in Japan during 2007–2010. Here we conducted the second national survey of acute encephalopathy during 2014–2017, and compared the results between the two studies to elucidate the trends in the seven years’ interval as well as the influence of changes in pediatric viral infections and guidelines for acute encephalopathy in Japan. Methods The Research Committee on Acute Encephalopathy supported by the Japanese Government sent a questionnaire to 507 hospitals throughout Japan, and collected the responses by mail. Results A total of 1115 cases from 267 hospitals reportedly had acute encephalopathy during April 2014–June 2017. In this study, the age at onset was younger, the ratios of recently established syndromes, such as acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) and clinically mild encephalitis/encephalopathy with a reversible splenial lesion (MERS), were higher, and the ratio of influenza-associated encephalopathy was lower, than in the previous study. The age at onset of influenza-associated encephalopathy was lower, and that of HHV-6/7-associated encephalopathy higher, compared to the first survey. The outcomes of entire acute encephalopathy remained unchanged. Conclusion Some of these changes reflected the recent trends of viral infectious diseases including 2009 influenza pandemic, and others the standardization of the diagnosis of acute encephalopathy in Japan.

Febrile infection-related epilepsy syndrome (FIRES), or acute encephalitis with refractory, repetitive partial seizures (AERRPS), is an epileptic encephalopathy beginning with fever-mediated seizures. The etiology remains unclear. To elucidate the genetic background of FIRES/AERRPS (hereafter FIRES), we recruited 19 Japanese patients, genotyped polymorphisms of the IL1B , IL6 , IL10 , TNFA , IL1RN , SCN1A and SCN2A genes, and compared their frequency between the patients and controls. For IL1RN , the frequency of a variable number of tandem repeat (VNTR) allele, RN2, was significantly higher in the patients than in controls ( p =0.0067), and A allele at rs4251981 in 5′ upstream of IL1RN with borderline significance (p =0.015). Haplotype containing RN2 was associated with an increased risk of FIRES (OR 3.88, 95%CI 1.40–10.8, p =0.0057). For SCN1A , no polymorphisms showed a significant association, whereas a missense mutation, R1575C, was found in two patients. For SCN2A , the minor allele frequency of G allele at rs1864885 was higher in patients with borderline significance ( p =0.011). We demonstrated the association of IL1RN haplotype containing RN2 with FIRES, and showed a possible association of IL1RN rs4251981 G>A and SCN2A rs1864885 A>G, in Japanese patients. These preliminary findings suggest the involvement of multiple genetic factors in FIRES, which needs to be confirmed by future studies in a larger number of FIRES cases.

Objective: To investigate the clinical and epidemiologic features of pediatric acquired demyelinating syndromes (ADS) of the CNS in Japan. Methods: We conducted a nationwide survey and collected clinical data on children with ADS aged 15 years or younger, who visited hospitals between 2005 and 2007. Results: Among 977 hospitals enrolled, 723 (74.0%) responded to our inquiries and reported a total of 439 patients as follows: 244 with acute disseminated encephalomyelitis (ADEM), 117 with multiple sclerosis (MS), 14 with neuromyelitis optica (NMO), and 64 with other ADS. We collected and analyzed detailed data from 204 cases, including those with ADEM (66), MS (58), and NMO (10). We observed the following: (1) the estimated annual incidence rate of pediatric ADEM in Japan was 0.40 per 100,000 children (95% confidence interval [CI], 0.34–0.46), with the lowest prevalence in the north; (2) the estimated prevalence rate of MS was 0.69 per 100,000 children (95% CI, 0.58–0.80), with the lowest prevalence in the south; (3) NMO in Japan was rare, with an estimated prevalence of 0.06 per 100,000 children (95% CI, 0.04–0.08); and (4) the sex ratio and mean age at onset varied by ADS type, and (5) male/female ratios correlated with ages at onset in each ADS group. Conclusions: Our results clarify the characteristic clinical features of pediatric ADS in the Japanese population.

Background Hyperserotonemia in the brain is suspected to be an endophenotype of autism spectrum disorder (ASD). Reducing serotonin levels in the brain through modulation of serotonin transporter function may improve ASD symptoms. Methods We analyzed behavior and gene expression to unveil the causal mechanism of ASD-relevant social deficits using serotonin transporter (Sert) knockout mice. Results Social deficits were observed in both heterozygous knockout mice (HZ) and homozygous knockout mice (KO), but increases in general anxiety were only observed in KO mice. Two weeks of dietary restriction of the serotonin precursor tryptophan ameliorated both brain hyperserotonemia and ASD-relevant social deficits in Sert HZ and KO mice. The expression of rather distinct sets of genes was altered in Sert HZ and KO mice, and a substantial portion of these genes was also affected by tryptophan depletion. Tryptophan depletion in Sert HZ and KO mice was associated with alterations in the expression of genes involved in signal transduction pathways initiated by changes in extracellular serotonin or melatonin, a derivative of serotonin. Only expression of the AU015836 gene was altered in both Sert HZ and KO mice. AU015836 expression and ASD-relevant social deficits normalized after dietary tryptophan restriction. Conclusions These findings reveal a Sert gene dose-dependent effect on brain hyperserotonemia and related behavioral sequelae in ASD and a possible therapeutic target to normalize brain hyperserotonemia and ASD-relevant social deficits. Electronic supplementary material The online version of this article (10.1186/s13229-018-0243-3) contains supplementary material, which is available to authorized users.

Objective Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) is a childhood encephalopathy following severe febrile seizures. The pathogenesis of AESD is considered to be fever-induced seizure susceptibility and excitotoxicity, which may be caused by sodium channel dysfunction in some cases. Here we studied whether mutations in genes encoding sodium channels, SCN1A and SCN2A , predispose children to AESD. Methods We recruited 92 AESD patients in a nationwide survey of acute encephalopathy in Japan from 2008 to 2011. We collected their genomic DNA samples, and sequenced the entire coding region of SCN1A and SCN2A . Results Five out of 92 patients (5.4%) had missense mutations either in SCN1A or SCN2A . After a preceding infection with fever, all the patients showed status epilepticus at the onset. Hemiconvulsion–hemiplegia was recognized in three patients during the acute/subacute phase. One patient had taken theophylline for the treatment of bronchial asthma just before the onset of AESD. Familial history was not remarkable except one patient with a SCN1A mutation (G1647S) whose mother had a similar episode of AESD in her childhood. A different substitution (G1674R) at the same amino acid position, as well as two other SCN1A mutations found in this study, had previously been reported in Dravet syndrome. Another SCN1A mutation (R1575C) had been detected in other types of acute encephahlitis/encephalopathy. One patient had SCN2A mutation, F328V, which had previously been reported in Dravet syndrome. Another SCN2A mutation, I172V, was novel. None of the patients were diagnosed with Dravet syndrome or genetic (generalized) epilepsy with febrile seizure plus in the following-up period. Conclusions Mutations in SCN1A and SCN2A are a predisposing factor of AESD. Altered channel activity caused by these mutations may provoke seizures and excitotoxic brain damage.

Background Theophylline has recently been suspected as a risk factor of acute encephalopathy with biphasic seizures and late reduced diffusion (AESD), although there has been no systematic study on the relationship between acute encephalopathy in children taking theophylline (AET) and AESD. Methods We recruited 16 Japanese patients (11 male and 5 female, median age of 2 years and 7 months) with AET from 2008 to 2013. We evaluated their clinical features, such as the duration of first seizure, biphasic clinical course and cranial CT/MRI imaging and compared them with those of AESD. We analyzed the polymorphisms or mutations of genes which are associated with AESD. Results Clinically, 12 patients had neurological and/or radiological features of AESD. Only one patient died, whereas all 15 surviving patients were left with motor and/or intellectual deficits. Genetically, 14 patients had at least one of the following polymorphisms or mutations associated with AESD: thermolabile variation of the carnitine palmitoyltransferase 2 ( CPT2 ) gene, polymorphism causing high expression of the adenosine receptor A2A ( ADORA2A ) gene, and heterozygous missense mutation of the voltage gated sodium channel 1A ( SCN1A ) and 2A ( SCN2A ) gene. Conclusions Our results demonstrate that AET overlaps with AESD, and that AET is a multifactorial disorder sharing a genetic background with AESD.

The symptoms of attention-deficit/hyperactivity disorder (ADHD) are characterized by inattention and hyperactivity-impulsivity. It is a common childhood neurodevelopmental disorder that often persists into adulthood. Improvements in ADHD symptoms using psychostimulants have been recognized as a paradoxical calming effect. The psychostimulant methylphenidate (MPH) is currently used as the first-line medication for the management of ADHD. Recent studies have drawn attention to altered dopamine-mediated neurotransmission in ADHD, particularly reuptake by the dopamine transporter (DAT). This hypothesis is supported by the observation that DAT knockout mice exhibit marked hyperactivity that is responsive to acute MPH treatment. However, other behaviors relevant to ADHD have not been fully clarified. In the present study, we observed learning impairment in shuttle-box avoidance behavior together with hyperactivity in a novel environment in DAT knockout mice. Methylphenidate normalized these behaviors and enhanced escape activity in the tail suspension test. Interestingly, the effective dose of MPH increased extracellular dopamine in the prefrontal cortex but not striatum, suggesting an important role for changes in prefrontal dopamine in ADHD. Research that uses rodent models such as DAT knockout mice may be useful for elucidating the pathophysiology of ADHD.

Two live attenuated oral rotavirus vaccines, Rotarix and RotaTeq, have been introduced as voluntary vaccination in Japan since 2011 and 2012, respectively. Effectiveness of the vaccines has been confirmed, whereas concerns such as shedding of the vaccine strains and gastroenteritis cases caused by vaccine strains are not well assessed. We aimed to identify the vaccine strains in children with acute gastroenteritis (AGE) to investigate the prevalence of AGE caused by vaccination or horizontal transmission of vaccine strains. A total of 1,824 stool samples were collected from children with AGE at six outpatient clinics in 2012-2015. Among all, 372 group A rotavirus (RVA) positive samples were screened for vaccine components by real-time RT-PCR which were designed to differentiate vaccine strains from rotavirus wild-type strains with high specificity. For samples possessing both vaccine and wild-type strains, analyses by next-generation sequencing (NGS) were conducted to characterize viruses existed in the intestine. As a result, Rotarix-derived strains were identified in 6 of 372 (1.6%) RVA positive samples whereas no RotaTeq strain was detected. Among six samples, four possessed Rotarix-derived strains while two possessed both Rotarix-derived strains and wild-type strains. In addition, other pathogens such as norovirus, enterovirus and E.coli were detected in four samples. The contribution of these vaccine strains to each patient's symptoms was unclear as all of the cases were vaccinated 2-14 days before sample collection. Proportion of average coverage for each segmented gene by NGS strongly suggested the concurrent infection of the vaccine-derived strain and the wild-type strain rather than reassortment of these two strains in one sample. This is the first study to report the prevalence of vaccine-derived strains in patients with RVA AGE in Japan as 1.6% without evidence of horizontal transmission. The results emphasized the importance of continuous monitoring on vaccine strains and their clinical impacts on children.

Aichivirus C is the third species in the genus Kobuvirus, family Picornaviridae, and the virus is circulating in pigs worldwide. Aichivirus A in humans and Aichivirus B in cows have been shown to associate with diarrheal diseases, however, the pathogenesis of Aichivirus C has not been demonstrated clearly. In this study, the full genome nucleotide sequence of the Thai strain, CMP06/2007/THA collected from stool sample of a diarrheal piglet was analyzed and identified as a variant type with a 90-nt deletion in the 2B-coding region. In addition, molecular characterization of nucleotide sequences of the 2B-coding region of Aichivirus C strains from six diarrheal and six healthy piglets in Thailand, and four strains from healthy pigs in Japan revealed that all of the strains in this study were variant types. These findings indicate that variant strains of Aichivirus C are circulating in Asian countries such as China, Thailand and Japan, and deletion of tandem repeat of 2B-region is unlikely to associate with the pathogenesis of the virus.

We studied the altered molecular species of lipids in brain and liver tissues, and fibroblasts from patients with Zellweger syndrome (ZS). ZS cerebellum samples contained a higher amount of sphingomyelin with shorter chain fatty acids compared to that in normal controls. The amount of phosphatidylethanolamine (PE) was less than half of that in controls, with the absence of the PE-type of plasmalogen. Gangliosides were accumulated in the brains and fibroblasts of ZS patients. To investigate whether or not impaired beta-oxidation of very long chain fatty acids and/or plasmalogen synthesis affects glycolipids metabolism, RNAi of peroxisomal acylCo-A oxidase (ACOX1) and glyceronephosphate O-acyltransferase (GNPAT) was performed using cultured neural cells. In neuronal F3-Ngn1 cells, ACOX1 and GNPAT silencing up-regulated ceramide galactosyltransferase (UGT8) mRNA expression, and down-regulated UDP-glucose ceramide glucosyltransferase (UGCG). These results suggest that both impaired beta-oxidation of very long chain fatty acids and plasmalogen synthesis affect glycolipid metabolism in neuronal cells.

Objective: Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) is a childhood encephalopathy following severe febrile seizures, leaving neurologic sequelae in many patients. However, its pathogenesis remains unclear. In this study, we clarified that genetic variation in the adenosine A2A receptor ( ADORA2A ), whose activation is involved in excitotoxicity, may be a predisposing factor of AESD. Methods: We analyzed 4 ADORA2A single nucleotide polymorphisms in 85 patients with AESD. The mRNA expression in brain samples, mRNA and protein expression in lymphoblasts, as well as the production of cyclic adenosine monophosphate (cAMP) by lymphoblasts in response to adenosine were compared among ADORA2A diplotypes. Results: Four single nucleotide polymorphisms were completely linked, which resulted in 2 haplotypes, A and B. Haplotype A (C at rs2298383, T at rs5751876, deletion at rs35320474, and C at rs4822492) frequency in patients was significantly higher than in controls ( p = 0.005). Homozygous haplotype A (AA diplotype) had a higher risk of developing AESD (odds ratio 2.32, 95% confidence interval 1.32–4.08; p = 0.003) via a recessive model. mRNA expression was significantly higher in AA than AB and BB diplotypes, both in the brain ( p = 0.003 and 0.002, respectively) and lymphoblasts ( p = 0.035 and 0.003, respectively). In lymphoblasts, ADORA2A protein expression ( p = 0.024), as well as cellular cAMP production ( p = 0.0006), was significantly higher in AA than BB diplotype. Conclusions: AA diplotype of ADORA2A is associated with AESD and may alter the intracellular adenosine/cAMP cascade, thereby promoting seizures and excitotoxic brain damage in patients.

Norovirus (NoV) is recognized as one of the most common causative agents of diarrhea disease in young children. A total of 187 fecal specimens collected from non-hospitalized children with acute gastroenteritis in Shizuoka, Japan during July 2008 to June 2009 were investigated for the presence of diarrhea viruses by a multiplex RT-PCR. Diarrhea viruses were overall detected in 158 of 187 (84.5%). Of the viruses detected, NoV was the most prevalent (55.6%). Most of the NoV sequences belonged to GII.4 (53.8%). NoV GII.6 emerged as the second most common strain (40.4%). The full-length capsid sequences of five representative Shizuoka GII.6 strains were compared with all 12 GII.6 strains available in GenBank database between 1990 and 2009. At least three distinct GII.6 subclusters (a–c) appeared in different parts of the world. Shizuoka GII.6 strains formed their own subcluster c, distinct from other complete GII.6 reference sequences. The Shizuoka strains had significant amino acid divergence, particularly in the P2 domain up to 10.9–17.5% and contained eight unique mutations in the P domains, compared with subcluster a and b viruses. The homology model showed that the eight mutations were predicted to be located at the surface-exposed P1 and P2 domains. The data suggest the emergence of a new NoV GII.6 variant in Shizuoka, with a high level of genetic variation. J. Med. Virol. 84: 1089–1096, 2012. © 2012 Wiley Periodicals, Inc.

A patient with a large deletion of the distal part of the long arm of chromosome 13 showed severe psychomotor retardation, a characteristic face, nystagmus, retinopathy, cystic kidney disease, and brain malformation with molar tooth sign and cerebellar vermis hypoplasia, a phenotype typical of Arima syndrome. This patient also had bilateral retinoblastoma. Fluorescent in situ hybridization and single-nucleotide-polymorphism genotyping microarray demonstrated an interstitial deletion of 54 Mbp, ranging from 13q14.13 to 13q32.3 and involving the RB1 gene. This patient is the first case of Arima syndrome, or a Joubert syndrome-related disorder, that showed linkage to chromosome 13q.

Breast milk contains immunological factors, such as IgA antibody, which help to prevent infectious diseases. A total of 197 paired samples of colostrum and breast milk was collected from postpartum mothers in Gunma City, Japan, and examined for anti-rubella IgA antibody by enzyme-linked immunosorbent assay (ELISA) and Western blotting (WB). The anti-rubella virus IgA ranged from 0.5 to 78.5 U/ml with a mean of 6.05 U/ml and a median of 3.6 U/ml in colostrum, and from 0.5 to 32.7 U/ml with a mean of 2.74 U/ml and a median of 2 U/ml in milk. The differences between the means of titers of total IgA and anti-rubella virus IgA in colostrum and in milk were significant statistically. The levels of anti-rubella virus IgA in both colostrum and breast milk correlated positively with the anti-rubella virus hemagglutination inhibition (HI) titers in the sera of mother, indicating that the levels of these different classes of antibodies correlated. Based on WB, anti-rubella virus IgA in both colostrum and breast milk reacted with the rubella viral protein E1 and C, but not with the E2 protein. J. Med. Virol. 82:1475–1479, 2010. © 2010 Wiley-Liss, Inc.

Mutations in the Cyclin-dependent kinase-like 5 (CDKL5) gene cause severe neurodevelopmental disorders. Recently we have generated Cdkl5 KO mice by targeting exon 2 on the C57BL/6N background, and demonstrated postsynaptic overaccumulation of GluN2B-containing N-methyl-D-aspartate (NMDA) receptors in the hippocampus. In the current study, we subjected the Cdkl5 KO mice to a battery of comprehensive behavioral tests, aiming to reveal the effects of loss of CDKL5 in a whole perspective of motor, emotional, social, and cognition/memory functions, and to identify its undetermined roles. The neurological screen, rotarod, hot plate, prepulse inhibition, light/dark transition, open field, elevated plus maze, Porsolt forced swim, tail suspension, one-chamber and three-chamber social interaction, 24-h home cage monitoring, contextual and cued fear conditioning, Barnes maze, and T-maze tests were applied on adult Cdkl5 -/Y and +/Y mice. Cdkl5 -/Y mice showed a mild alteration in the gait. Analyses of emotional behaviors revealed significantly enhanced anxiety-like behaviors of Cdkl5 -/Y mice. Depressive-like behaviors and social interaction of Cdkl5 -/Y mice were uniquely altered. The contextual and cued fear conditioning of Cdkl5 -/Y mice were comparable to control mice; however, Cdkl5 -/Y mice showed a significantly increased freezing time and a significantly decreased distance traveled during the pretone period in the altered context. Both acquisition and long-term retention of spatial reference memory were significantly impaired. The morphometric analysis of hippocampal CA1 pyramidal neurons revealed impaired dendritic arborization and immature spine development in Cdkl5 -/Y mice. These results indicate that CDKL5 plays significant roles in regulating emotional behaviors especially on anxiety- and fear-related responses, and in both acquisition and long-term retention of spatial reference memory, which suggests that focus and special attention should be paid to the specific mechanisms of these deficits in the CDKL5 deficiency disorder.

The aim of this study was to clarify the difference between influenza and non-influenza cases in clinical symptoms, laboratory and neuroimaging findings, and outcome in children with ANE. We retrospectively studied 22 children with ANE. Eleven of them had virological proof of influenza infection and the other 11 were judged as non-influenza infection. There was no significant difference between influenza and non-influenza cases in sex, antipyretics use and neurological symptoms. Although laboratory data were not different between the two groups, brainstem lesions were relatively more frequent in influenza cases than in non-influenza cases. Outcome was not different between the two groups. The results of our study suggest that the pathogenesis of acute necrotizing encephalopathy will not be dependent on infectious agents.

Objective. To examine the relation between outcome and treatment with steroids and gammaglobulin in children with acute necrotizing encephalopathy. Methods. We retrospectively evaluated the clinical course and outcome of 34 children with acute necrotizing encephalopathy. They were divided into two groups; 17 patients with brainstem lesion and 17 patients without brainstem lesion. Early steroid use was defined as when steroids were administered within 24 h after the onset. The outcome was judged as good when a patient had no or mild cognitive impairment and poor when a patient had more severe sequelae, or died. Results. Among patients without brainstem lesions, the outcome was good in 7 of 12 with early steroid, whereas it was poor in all 5 patients without early steroid. There was no significant difference in sex, age, and laboratory data between patients with and without early steroid. The outcome was not correlated with gammaglobulin treatment. As to patients without brainstem lesions, the outcome was not correlated with early steroid or gammaglobulin treatment. Conclusions. Steroid within 24 h after the onset was related to better outcome of children with acute necrotizing encephalopathy without brainstem lesions. Early steroid treatment will be an important option of the treatment for acute necrotizing encephalopathy.

To elucidate the neuropathological mechanism of Zellweger syndrome (ZS), we studied changes in the molecular species of glycerophospholipids in the cerebral tissue by thin-layer chromatography (TLC) and fast atom bombardment mass spectrometry (FABMS). First, we estimated the amount of plasmalogens by TLC. Plasmalogen-type phosphatidyl ethanolamine (PE) accounted for 30% of the total PE in the control brain, but was absent in the ZS brain. Plasmalogen-type phosphatidyl choline (PC) was undetectable in both control and ZS brains. Next, we analyzed plasmalogen-type PE by FABMS. Oleic (18:1), arachidonic (20:4) and docosapentanoic (22:5) acids were present in the control gray matter, but not in the ZS gray matter. In compensation for the defect of plasmalogen, the level of diacyl PE with polyunsaturated fatty acids, 20:4, 22:4, 22:5 and 22:6, was higher in the ZS brain than that in the control brain. These results indicate an alteration in the molecular species of PE, which may cause abnormal neural membrane fluidity and excessive vulnerability to oxygen stress.

To determine G and P genotypes, we performed nested PCR on 307 rotavirus specimens collected in Dhaka, Bangladesh, during 2004–2005. G2 (43.3%) was detected at the highest frequency, followed by G4 (19.5%), G9 (13.7%), G1 (12.7%), and G3 (2.6%). P[8] was the most predominant genotype (53.2%), followed by P[4] (42.9%).

Ellis-van Creveld syndrome (EvC) is a ciliopathy with cardiac anomalies, disproportionate short stature, polydactyly, dystrophic nails and oral defects. To obtain further insight into the genetics of EvC, we screened EVC/EVC2 mutations in eight Vietnamese EvC patients. All the patients had a congenital heart defect with atypical oral and/or skeletal abnormalities. One had compound heterozygous EVC2 mutations: a novel mutation c.769G T-p.E177X in exon 6 inherited from father and another previously reported c.2476C T-p.R826X mutation in exon 14 inherited from mother. The EVC2 mRNA expression level was significantly lower in the patient and her parents compared to controls. Another case had a novel heterozygous EVC mutation (c.1717C G-p.S572X) in exon 12, inherited from his father. Of note, the mother without any EVC mutation on Sanger sequencing showed a lower expression level of EVC mRNA compared with controls. SNP array analysis revealed that the patient and mother had a heterozygous 16.4 kb deletion in EVC. This patient also had a heterozygous novel variant in exon 9 of EFCAB7 (c.1171 T C-p.Y391H), inherited from his father. The atypical cardiac phenotype of this patient and the father suggested that EFCAB7 may modify the phenotype by interacting with EVC. In conclusion, we detected two novel nonsense mutations and a partial deletion of EVC/EVC2 in two Vietnamese families with EvC. Moreover, we found in one family a missense mutation of EFCAB7, a possible modifier gene in EvC and its related disorders.

SUMMARYMolecular epidemiology and clinical impact of human rhinovirus (HRV) are not well documented in tropical regions. This study compared the clinical characteristics of HRV to other common viral infections and investigated the molecular epidemiology of HRV in hospitalized children with acute respiratory infections (ARIs) in Vietnam. From April 2010 to May 2011, 1082 nasopharyngeal swabs were screened for respiratory viruses by PCR. VP4/VP2 sequences of HRV were further characterized. HRV was the most commonly detected virus (30%), in which 70% were diagnosed as either pneumonia or bronchiolitis. Children with single HRV infections presented with significantly higher rate of hypoxia than those infected with respiratory syncytial virus or parainfluenza virus (PIV)-3 (12·4% vs. 3·8% and 0%, respectively, P < 0·05), higher rate of chest retraction than PIV-1 (57·3% vs. 34·5%, P = 0·028), higher rate of wheezing than influenza A (63·2% vs. 42·3%, P = 0·038). HRV-C did not differ to HRV-A clinically. The genetic diversity and changes of types over time were observed and may explain the year-round circulation of HRV. One novel HRV-A type was discovered which circulated locally for several years. In conclusion, HRV showed high genetic diversity and was associated with significant morbidity and severe ARIs in hospitalized children.

Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) is the most common subtype of infectious pediatric encephalopathy in Japan. It is sometimes difficult to make an early diagnosis of AESD; excitotoxicity is postulated to be the pathogenesis based on elevated glutamine (Gln) and glutamate (Glu) complex (Glx = Glu + Gln) observed on MR spectroscopy. It is uncertain whether Gln or Glu contributes to the elevated Glx, or whether MR spectroscopy is useful for an early diagnosis. Five Japanese patients with AESD (three boys and two girls, 1 year of age) were enrolled in this study. MR spectroscopy was acquired from the frontal white matter (repetition time (TR) of 5000 ms, echo time (TE) of 30 ms) with a 1.5- or 3.0-T scanner. MR spectroscopy was performed four times for two patients, three times for one patient, and two times for two patients. Quantification of Glu and Gln was performed using LCModel. Glu was elevated in three of four studies on days 1–4 and became normal or low afterward. Gln was normal in three studies on days 1–2, elevated in all seven studies on days 4–12, and became normal or low afterward. These findings suggest that MR spectroscopy may be useful for an early diagnosis. Acute Glu elevation changes to subacute Gln elevation, suggesting that a disrupted Glu-Gln cycle may play an important role.

Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) at onset manifests an early seizure (ES) usually lasting more than 30 min. Following ES, some patients exhibit almost clear consciousness with no neurological symptoms, and no MRI abnormality for a few days, which may lead to an initial misdiagnosis of prolonged febrile seizures (PFS). To allow an early diagnosis of AESD, we retrospectively analyzed clinical manifestations, laboratory data, and radiologic and EEG findings in patients with AESD (n=62) having ES of over 30 min, and ones with PFS (n=54), using logistic regression analyses. Multivariate logistic regression analysis revealed that an age below 1.5 years and a Glasgow Coma Scale score of 14 or less than 14 (Japan Coma Scale score of 1 or higher) were high risk factors of developing AESD. We proposed an AESD prediction score system consisting of consciousness level, age, duration of convulsions, enforcement of mechanical intubation, and aspartate aminotransferase, blood glucose and creatinine levels (full score: 9), the mean scores in AESD and PFS being 5.9 and 1.8, which were significantly different (p

Acute gastroenteritis continues to be a major public health problem worldwide. A wide variety of viruses associated with diarrhea disease is being reported continually. This study investigated the epidemiological situation of viruses that cause diarrhea in Japanese pediatric patients. This study enrolled a total of 2,381 fecal specimens collected between 2009 and 2013 from Japanese children with acute gastroenteritis. There is currently a 70.4% prevalence of viruses causing diarrhea among these Japanese pediatric outpatients. Norovirus was detected in 39.3% of the patients, whereas the prevalence of rotavirus, human parechovirus, enterovirus, adenovirus, sapovirus, astrovirus, and Aichi virus was 20.1, 6.6, 6.1, 5.6, 4.8, 2.3, and 0.1%, respectively. Co-infections were observed at the prevalence rates of 13.4 and 0.5% for double infections and triple infections, respectively. Mixed viral infections were found commonly in Japanese outpatients, and the norovirus seemed to play a major role in co-infections. Viral diarrhea cases were detected mostly in children younger than 3 years of age. The norovirus and rotavirus can be detected throughout the year, with a peak during the cold and dry seasons, whereas other common viruses are found during no specific season. Surveillance data revealed that a wide variety of viruses has caused diarrhea to circulate currently in Japanese pediatric outpatients, with very high detection rates; and norovirus and rotavirus are the most important pathogens. The data obtained from this study are valuable for compiling the overall picture of several viruses that causes diarrhea and associates with acute gastroenteritis in the Japanese pediatric population.

SummaryWe describe herein a patient with Panayiotopoulos syndrome (PS) showing an atypical course. The patient initially had seizures typical of this syndrome from 3 to 5 years of age. EEG showed right occipital high-amplitude sharp and slow-wave complexes followed by brief generalized discharges of slow waves. Sequential EEGs obtained from 5 to 11 years of age showed both multifocal discharges and generalized spike and wave complexes. With these changes in EEG findings, the patient experienced various types of seizures. The seizures were frequent and showed oculocephalic deviation followed by absence, atonic seizures, generalized tonic clonic convulsions and clonic seizures of the eyelids, which were observed between 7 and 10 years of age. Antiepileptic drugs were only partially effective for these seizures. Ictal EEG recorded at 8 years of age revealed high-voltage slow waves from the bilateral frontal and occipital regions prior to diffuse high-amplitude spike–wave bursts. At 9 years of age, magnetoencephalography (MEG) revealed the calculated dipoles of the preceding bifrontal spike–wave discharges to be in the frontal areas, while those of the following generalized spike–wave bursts were in the bilateral mid-temporal areas. In PS, reportedly, dipoles of multifocal epileptic discharges are usually located in the occipital and Rolandic areas. The unique clinical evolution in our case may be associated with the unusual frontal localization of dipoles detected by MEG.

Acute necrotizing encephalopathy of childhood is a novel type of parainfectious encephalopathy with a racial and geographic predilection, rarely reported from other than East Asian areas. The objective was to describe the clinical, imaging, and other laboratory findings of non-Asian patients with acute necrotizing encephalopathy. Data were collected from three patients diagnosed in Athens over a 4-year period plus 16 cases reported from other European and North American countries. One of the Greek children died, and the other two had a normal outcome. A neuropathologic examination in the fatal case showed edematous necrosis without inflammatory, reactive, or proliferative changes. Data from Greek and other non-Asian patients support the homogeneity of the disease worldwide. (J Child Neurol 2006;21:872—879; DOI 10.2310/7010.2006.00210).

Acute infantile encephalopathy predominantly affecting the frontal lobes (AIEF) is proposed as a novel form of acute encephalopathy in infancy. To establish the diagnostic criteria for AIEF, we reviewed the clinical data of 10 patients who were seen by us and diagnosed as having AIEF, and those of 7 patients in the literature compatible with the diagnosis of AIEF. The mean age of onset was 1 year and 7 months. Boys and girls were equally affected. There is always an association with hyperpyrexia due to viral illness. Manifestations at the onset were convulsive status epilepticus and prolonged coma followed by signs of frontal lobe dysfunction such as a lack of spontaneity and regression of verbal functions. Imaging studies demonstrated edematous changes of the bilateral frontal lobes, which showed increased cerebral perfusion initially but attenuated perfusion several weeks later. The recovery of intellectual deficit was generally slower than that of motor disability. Based on these findings, we propose tentative diagnostic criteria of AIEF.

To test the hypothesis that the disruption of fukutin protein produces the brain pathology through hypoglycosylation of alpha-dystroglycan (alpha-DG), we immunostained Fukuyama congenital muscular dystrophy (FCMD) brains with an antibody that recognizes the polysaccharide epitope of alpha-DG. Immunoreactivity of the glia-limitans along the cortical surface, as well as that of the glial endfeet around vessel walls, was preserved in the FCMD cerebrum. However, fragmentation of the immunostained glia-limitans was noted in association with parenchymal protrusion and gyral fusion. In the FCMD cerebellum, this fragmentation of alpha-DG labeling was limited to the area of micropolygyria, and immunostaining at the glia-limitans and vessel walls was comparable to that of the control brains, in structurally normal areas. In the hippocampus, neurons of the dentate gyrus and corpus ammonis were immunopositive for alpha-DG in control subjects, but this staining was markedly decreased in FCMD brains. In contrast, immunolabeling of blood vessels and the glia-limitans was preserved in this region. Fukutin antisera clearly labeled hippocampal neurons in control brains, while this labeling was decreased in FCMD brains. Thus, hypoglycosylation of alpha-DG was evident in neurons, but not in the glial cell population of FCMD brains. This suggests that the mechanism of alpha-DG glycosylation may differ between neurons and glial cells, and that a fukutin gene defect may result in functional disruption through hypoglycosylation of both neuronal and glial alpha-DG.

Human uterine leiomyomata are the most common tumors in women of reproductive age. The pathogenesis of leiomyomata remains unknown. An animal model of Eker rats with deleted tuberous sclerosis complex gene 2 (tuberin) shows increased incidence of leiomyomata. The role of tuberin in human leiomyomata is unknown. In this study, we designed a tissue microarray with tissue cores of leiomyomata and the matched myometrium from 60 hysterectomy specimens. We examined the expression of tuberin and tuberous sclerosis complex gene 1 product hamartin, proteins of the insulin-signaling pathway, steroid receptors and some of their cofactors, and human mobility group gene A2 by immunohistochemistry. We found that nearly half of the cases displayed either reduction or loss of tuberin in leiomyomata compared with matched normal myometrium. No change of hamartin was noted. Furthermore, a significant reduction of glucocorticoid receptor was found in leiomyomata with reduced tuberin. The proteins insulin like growth factor 1, insulin-like growth factor receptor beta, AKT kinase, and phosphatidylinositol 3-kinase were upregulated. Nearly half of leiomyomata show upregulation of human mobility group gene A2, along with the steroid receptor cofactors. Our findings suggest that there are two broad groups of uterine leiomyomata. One group is associated with an alteration of tuberin and glucocorticoid receptor. The other group is associated with upregulation of human mobility group gene A2 and steroid receptor cofactors.

This female patient was the second-born child to nonconsanguinous, healthy parents. The first child, her elder brother, had respiratory distress and metabolic acidosis at 3 days of age, and died at 5 days. After an uneventful gestation for 39 weeks, this patient was born by normal delivery. Immediately after birth, she weighed 2608 g and looked well. Twenty-one hours later, however, she became dyspneic and underwent mechanical ventilation until 8 days of age. Blood examination revealed severe lactic acidosis (pH 6.97, PaCO 2 23 mmHg, base excess 26 mEq/ l, lactate 135 mg/dL), which was treated with sodium bicarbonate and thiamine. Despite improvement of her general condition, metabolic acidosis persisted. At the age of 1 month, lactate and pyruvate levels were 36.8 and 1.80 mg/dL in the blood (normal values, 7.0–25.0 and 0.6– 1.35 mg/dL), and 51.1 and 2.77 mg/dL in the cerebrospinal fluid (normal values, 11.1–16.3 and 0.75–1.29 mg/dL), respectively. An analysis of mitochondrial DNA (mtDNA) of the peripheral blood cells disclosed a T-to-G mutation at nucleotide position 8993. When she was discharged at 2 months of age, she was unresponsive to environmental stimuli, and had spastic paraplegia. She began to follow objects with her eyes at 2 months, to show social smile at 3 months, to control her head at 5 months, to roll over at 7 months, and to sit unsupported at 7 months. Cranial MRI at 4 months of age showed no abnormal signals in the brain. After 8 months of age, she showed no further development. She smiled less frequently than previously. Dysphagia and stridor became more prominent. Ubidecarenone and levocarnitine were prescribed. MRI at 10 months of age showed T 1 and T 2 prolongation of the bilateral putamina, caudate nuclei, periaqueductal gray matter, and substantia nigra (Fig. 1). Spastic tetraparesis worsened. At 10.5 months, sudden appearance of nystagmus was followed by difficulty in sucking milk, loss of smile and other responses, and irregular respiration. When she was admitted again, she showed deep coma, no spontaneous movements, and minimal response to painful stimuli. The pupils were meiotic and unresponsive to light. There were marked muscle hypotonia and positive pathological reflexes. Blood examination showed lactic acidosis (pH 7.44, PaCO 2 21 mmHg, base excess 8 mEq/l, lactate 72.3 mg/dL, pyruvate 2.05 mg/dL). Despite mechanical ventilation, and administration of sodium dichloroacetate and cytochrome C, her condition deteriorated further with intercurrent pneumonia, appearance of mydriasis, and loss of spontaneous respiration. Cranial computed tomography showed hypodensity of the entire brain. Electroencephalogram and auditory evoked response showed a total loss of electrical activity. Two weeks later, she died at the age of 11 months.

During influenza epidemics in Japan, the number of children with acute encephalopathies and encephalitis has recently increased. Although the pathophysiologies remain unclear, there is usually brain edema with evidence of damage to the blood-brain-barrier (BBB). We investigated the glial reaction and apoptosis in brains of eight such cases comprising two of acute necrotizing encephalopathy and six of influenza encephalopathy, and compared the results with those in five control brains. Apoptosis, evidenced by chromatin condensation and fragmentation in hematoxylin sections, in situ end labeling of fragmented DNA (TUNEL) and DNA laddering, was observed in neurons and glial cells in four brains with influenza encephalopathy. In the TUNEL-positive brains, the increase in microglia was greater than in the TUNEL-negative brains. Immunoreactivity for active-caspase 3, demonstrated by immunohistochemistry, and the overexpression of a caspase-cleaved fragment of poly(ADP-ribose) polymerase, demonstrated by Western blotting, indicated that activation of caspase 3 is involved in the apoptotic pathway in the brains of influenza encephalopathy cases. Apoptosis or specific pathological processes that cause apoptosis may give rise to aggravated encephalopathy.

Cerebral cortical lesions of tuberous sclerosis (TSC) and focal cortical dysplasia (FCD) show disturbances in laminar architecture and cellular differentiation. We immunohistochemically studied the expression of doublecortin, a fetal neuronal protein that regulates neuronal migration, in the surgical specimens of five TSC and eight FCD patients. In both TSC and FCD, bizarre giant cells showed a variable degree of doublecortin immunoreactivity. Both cytomegalic neurons and balloon cells were positive. The staining tended to be more intense in TSC than in FCD, although there were exceptional cases in both groups. Doublecortin immunoreactivity of normal-sized neural cells was restricted to a small number of astrocytes, and comparable to that in control patients. The persistent expression of doublecortin by giant cells in the postnatal cerebrum is additional evidence of abnormal differentiation, which may be relevant to the pathogenesis of cortical disarray in TSC and FCD.

We describe an 8-year-old girl with the mildest form of acute necrotizing encephalopathy, associated with pandemic influenza A. She manifested a convulsion engendering deterioration of consciousness, although cranial computed tomography and magnetic resonance imaging within 4 hours after the convulsion revealed no abnormalities. Cranial magnetic resonance imaging 20 hours after the convulsion revealed lesions of the thalamus bilaterally, brainstem tegmentum, internal capsule, and white matter. She was diagnosed with acute necrotizing encephalopathy. Typically, the prognosis of acute necrotizing encephalopathy with a brainstem lesion is poor. Nevertheless, she recovered almost completely, after early intervention with pulsed methylprednisolone and high-dose γ-globulin therapy. She manifested a thermolabile phenotype of carnitine palmitoyltransferase II variants such as cystine-isoleucine-methionine phenotype type 9 (FVM-CIM; Phe352Cys-Val388Ile-Met647Met alleles), resulting in a predisposition to encephalopathy during influenza infection. This case is the first, to the best of our knowledge, of pandemic influenza A-associated acute necrotizing encephalopathy with a good outcome despite severe magnetic resonance imaging findings.

We retrospectively collected three patients with clinically mild encephalitis with a reversible splenial lesion (MERS) after mumps vaccination, and reviewed five patients, including two patients previously reported. The five patients (all males, aged 1 to 9) presented with fever, vomiting, or headache as the initial symptoms (day 0), suggesting meningitis, at 13 to 21 days after mumps vaccination. Consciousness disturbance, delirious behavior, seizures, or dysarthria was observed on days 1 to 3, which had completely resolved before day 11. Hyponatremia was observed in all patients. A cerebrospinal fluid study showed pleocytosis, and confirmed the vaccine strain genome. MRI revealed reduced diffusion in the splenium of the corpus callosum on days 2 to 4, which had completely disappeared on the follow-up studies performed on days 7-15. EEG showed high voltage slow wave in three patients, which later normalized. These findings led to a diagnosis of MERS after mumps vaccination. MERS after mumps vaccination may be more common than previously considered. MERS is suspected when a male patient after mumps vaccination presents with neurological symptoms with hyponatremia, following symptoms of aseptic meningitis, and MRI would be performed to examine the splenium of the corpus callosum.

We detected Campylobacter spp. in 5% (20/380) of diarrheal stool samples collected at an outpatient clinic in Kyoto using a commercial loop-mediated isothermal amplification (LAMP) kit with a fluorescent detection reagent after DNA extraction. The sensitivity and specificity were 100% in comparison with those of semi-nested PCR for the differentiation of Campylobacter jejuni and Campylobacter coli. Fourteen of the 20 samples were already determined as C. jejuni by the culture method. All 20 samples were also positive for C. jejuni by the PCR method. Among the 58 cultured samples, the sensitivity of the culture method against the LAMP method was 93.3% (14/15) and the specificity was 100% (43/43). The detection rate of Campylobacter spp. from the heated supernatants by the LAMP method was lower than that from the supernatant after DNA extraction. In total, 25% (5/20) of the Campylobacter-positive samples by the LAMP method were co-infected with norovirus (3/20), rotavirus (1/20), and human parechovirus (1/20), although no other bacterial co-infection was identified by the culture method. C. jejuni was mostly detected in children aged >5 years throughout the year. Based on these results, we concluded that care should be taken while diagnosing Campylobacter infection in children. Our newly modified LAMP method is a rapid, easy, and useful method for this diagnosis.

Objective: To elucidate the clinical and radiologic features and analyze factors associated with neurologic outcomes of encephalopathy secondary to Shiga toxin–producing Escherichia coli (STEC) O111. Methods: We reviewed medical records and neuroimaging in 22 patients with neurologic symptoms among 86 with STEC O111 infection. Results: Twenty-one (6 males and 15 females, 10 children and 11 adults) of the 22 patients were diagnosed with encephalopathy. All patients with encephalopathy also presented with hemolytic-uremic syndrome. Five patients died, from day 1 to 6 months (days 1–5 in 4 patients), due to progressive encephalopathy with severe cerebral edema observed in neuroimaging (4 patients). Fifteen of the 16 surviving patients clinically recovered completely. Statistical analysis revealed differences between patients with poor (n = 6) and good (n = 15) outcomes in the interval from hemolytic-uremic syndrome presentation to encephalopathy, creatinine levels, and the methylprednisolone administration ratio. Conclusion: We note a high incidence of encephalopathy in the Toyama STEC O111 outbreak. All fatal cases resulted from progressive encephalopathy. Methylprednisolone pulse therapy represents a possible therapeutic choice. Classification of evidence: This study provides Class III evidence that methylprednisolone pulse therapy increases the probability of a good outcome for patients with encephalopathy associated with STEC O111.

We studied the expression of Down's syndrome cell adhesion molecule (DSCAM) in Down's syndrome (DS) and control brains, using antisera against peptide fragments of DSCAM. On Western blots of human, mouse and rat brain homogenates, the antisera recognized a product at approximately 200 kDa. In the brain of a 2-year-old patient with DS, Western blotting revealed an overexpression of DSCAM compared to an age-matched control. Immunohistochemistry demonstrated DSCAM in the cerebral and cerebellar white matter of both control and DS subjects, in accordance with the temporal and spatial sequence of myelination. In DS brains, immunoreactivity for DSCAM, compared to that for controls, was enhanced in the Purkinje cells at all ages, and in the cortical neurons during adulthood. In demented DS patients, DSCAM immunoreactivity was observed in the core and periphery of senile plaques. The pattern of DSCAM expression suggests that it may play a role as an adhesion molecule regulating myelination. The overexpression of DSCAM may also play a role in the mental retardation and the precocious dementia of DS patients, although the mechanism of neuronal dysfunction is undetermined.

Telencephalin (TLN), a telencephalon-specific glycoprotein, is exclusively expressed in neurons of the mammalian telencephalon. In the normally developing human brain, TLN immunoreactivity appeared and increased from 35 gestational weeks (GW) in the temporal cortex, and reached adult level at 5 months of postnatal age, being strong in the molecular layer, and weak in the external and internal granular layers. TLN expression corresponded with the development of neuronal dendrites and synapses. In brains with holoprosencephaly TLN immunoreactivity was already strong from as early as 28 GW. Staining was weak in the molecular layer, but strong in the external sparse and middle cellular layers in most cases. Notably, TLN was abundant in the glomerular structures in the internal pyramidal and multiform layers of fetal brains with alobar holoprosencephaly, which disappeared with increasing age. These results indicate premature and ectopic development of the dendrites and synaptic network in holoprosencephaly.