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Background Ensitrelvir is a novel oral SARS-CoV-2 3C-like protease inhibitor, and under late clinical development stage for COVID-19 disease. In Ph2a and Ph2b studies, ensitrelvir demonstrated rapid decline of viral titer and viral RNA compared with placebo, and tolerability. To investigate treatment-emergent amino acid substitutions (TEAASs), we analyzed viral RNA sequences of nonstructural protein 5 (NSP5), target of ensitrelvir from Ph2a study in Japan. Methods In Ph2a study, 69 patients with confirmed SARS-CoV-2 infection were randomized 1:1:1 to ensitrelvir with the loading dose on Day 1/maintenance dose on Day 2-5 (375/125 mg or 750/250 mg), or placebo. Intent-to-treat (ITT) population was defined as participants confirmed with positivity by qualitative RT-PCR at baseline. TEAASs were defined as novel amino acid substitutions identified after treatment with ensitrelvir. NSP5 sequence analysis was performed with sanger sequencing using nasopharyngeal swab samples at Day 1, 6, 9, 14, and 21 with allowances from ITT population with ensitrelvir. Infectious viral titer was measured by virus-induced cytopathic effects in VeroE6/TMPRSS2 cells. Viral RNA was quantified by RT-PCR. Results By NSP5 sequence analysis, TEAASs were observed in 1 patient from 375/125 mg group and 2 patients from 750/250 mg group. H246Y in 375/125 mg was detected from specimen on Day 8, and A234S and T198I in 750/250 mg were detected from specimens on Day 8 and Day 14, respectively. Structural analysis revealed that these mutations are located outside of the active center of 3C-like protease which is the binding site of ensitrelvir. Viral titer and viral RNA in each specimen, in which TEAASs were observed, were below lower limit of detection and lower limit of quantification, respectively. Furthermore, H246Y, A234S and T198I are rare substitution (< 0.05%) among SARS-CoV-2 variants according to Global Initiative on Sharing Avian Influenza Data. Conclusion In Ph2a study, H246Y, A234S, and T198I in NSP5 were detected as TEAASs. However, results of viral titer and viral RNA, and structural information suggest that these mutations do not have the impact on antiviral efficacy of ensitrelvir. Disclosures All Authors: No reported disclosures.

Purpose: (1) To evaluate the enhancement patterns of an ultrasmall superparamagnetic iron oxide contrast agent (USPIO-CA) compared with those of a gadolinium-based contrast agent (Gd-BCA). (2) To compare the histologic distribution of USPIO-related iron particles (USPIO-IPs) with the USPIO-enhancement area in the early vascular and in the cellular imaging phase (E- and L-phase, respectively) after intravenous CA administration., Methods:We performed USPIO-enhanced MRI of N-ethyl-N-nitrosourea (ENU)-induced endogenous rat glioma, including spin-echo (SE) T1-weighted images (T1WIs) and gradient-recalled-echo (GRE) T2-weighted images (T2WIs), before and at 3-6 h after USPIO-CA administration for E-phase images. For L-phase images, MRI was performed at 16-19 and 62-69 h after administration. Two observers determined the USPIO-enhancement area on E-phase images and Gd-enhancement areas. We compared the USPIO-enhancement size (USPIO-ES) and Gd-ES on SE T1WIs, and the hypo-intense USPIO-ES on GRE T2WIs and Gd-ES using the Wilcoxon signed-rank test. In addition, two raters visually evaluated the correspondence between the histologic distribution of USPIO-IPs and the USPIO-enhancement area on corresponding GRE T2WIs at each phase using a 3-rating scale., Results:Significantly smaller hyper-intense, hypo-intense and combined hyper-/hypo-intense areas were observed on USPIO-enhanced SE T1WIs compared with Gd-enhanced images (all P < 0.001). The hypo-intense USPIO-ES on GRE T2WIs was significantly smaller than the Gd-ES (P = 0.001). The distribution of USPIO-IPs on histopathological specimen and USPIO-enhancement on GRE T2WIs exhibited poor agreement in 5 of 9 tumors with enhancement from rats sacrificed early. The distribution of microglia containing USPIO-IPs corresponded with the pattern of USPIO-enhancement in the 2 tumors with late enhancement., Conclusion:The enhancement pattern and size of USPIO-CA in a rat glioma model were statistically different from those of Gd-BCA. Our histological data suggests that USPIO-enhanced MRI offers vascular bed imaging in E-phase and might depict the intra-tumoral distribution of immune effector cells in L-phase.

Background Diversity in group A rotavirus (RVA) strains after introduction of RV-vaccines remains an emerging concern worldwide. In this study, we investigated the prevalence and distribution of RVA genotypes in Japanese children with acute gastroenteritis (AGE) from 2015 to 2018. In addition, a comparison of the genotypes in pre-vaccination (2006–2012) and post-vaccination (2012–2018) periods was conducted to understand the impact of these vaccines on genotype distribution. Methods Fecal samples were collected regularly from outpatient clinics in six localities: Hokkaido, Tokyo, Shizuoka, Osaka, Kyoto, and Saga. RVA were screened and genotyped by RT-PCR and sequence-based genotyping. Results During the period 2015–2018, RVA was detected in 307 (19.7%) samples out of 1557 specimens: 29.9% (95% CI: 25.8% to 34.3%), 17.9% (95% CI: 14.7% to 21.5%), and 13% (95% CI: 10.3% to 16.0%) were detected RVA-positive in 2015–2016, 2016–2017 and 2017–2018, respectively. The average detection of RVA in pre-vaccination (2006–2012) and post-vaccination (2012–2018) era remained almost similar (18%-20%). The G2P[4]I2 (52.1%, 95% CI: 43.5%-60.6%) remained the most common genotype in 2015–2016, whereas G8P[8]I2 (55.9%, 95% CI: 45.2%-66.2%) dominated in 2016–2017. In 2017–2018, G9P[8]I2 (42.0%, 95% CI: 30.5%-53.9%) prevailed, followed by G9P[8]I1 (23.0%, 95% CI: 14.0%-34.2%). The detection rate of some common genotypes of pre-vaccination era like G1P[8] and G3P[8] has been reduced after introduction of RV-vaccine, whereas genotypes that were sporadic before the introduction of vaccines like G2P[4], G2P[8], G9P[8] and G8P[8] were emerged/reemerged in post-vaccination period. Conclusions Our study presented the diversity in circulating RVA genotypes in Japan before and after introduction of RV-vaccines. Sudden emergence of DS-1-like (I2) unusual strains in post-vaccination era remains alarming. Continuous monitoring of RVA genotypes is therefore indispensable to refine future vaccine strategy.

Objectives Baloxavir marboxil (formerly S-033188) is a first-in-class, orally available, cap-dependent endonuclease inhibitor licensed in Japan and the USA for the treatment of influenza virus infection. We evaluated the efficacy of delayed oral treatment with baloxavir marboxil in combination with a neuraminidase inhibitor in a mouse model of lethal influenza virus infection. Methods The inhibitory potency of baloxavir acid (the active form of baloxavir marboxil) in combination with neuraminidase inhibitors was tested in vitro. The therapeutic effects of baloxavir marboxil and oseltamivir phosphate, or combinations thereof, were evaluated in mice lethally infected with influenza virus A/PR/8/34; treatments started 96 h post-infection. Results Combinations of baloxavir acid and neuraminidase inhibitor exhibited synergistic potency against viral replication by means of inhibition of cytopathic effects in vitro. In mice, baloxavir marboxil monotherapy (15 or 50 mg/kg twice daily) significantly and dose-dependently reduced virus titre 24 h after administration and completely prevented mortality, whereas oseltamivir phosphate treatments were not as effective. In this model, a suboptimal dose of baloxavir marboxil (0.5 mg/kg twice daily) in combination with oseltamivir phosphate provided additional efficacy compared with monotherapy in terms of virus-induced mortality, elevation of cytokine/chemokine levels and pathological changes in the lung. Conclusions Baloxavir marboxil monotherapy with 96 h-delayed oral dosing achieved drastic reductions in virus titre, inflammatory response and mortality in a mouse model. Combination treatment with baloxavir acid and oseltamivir acid in vitro and baloxavir marboxil and oseltamivir phosphate in mice produced synergistic responses against influenza virus infections, suggesting that treating humans with the combination may be beneficial.

BackgroundDifferences in psychiatric background and dose–response to asenapine in patients with schizophrenia were examined based on efficacy and safety, using data obtained in a double-blind, placebo-controlled trial.MethodsPatients with schizophrenia were classified into three clusters by a cluster analysis based on the Positive and Negative Symptom Scale (PANSS) subscores at baseline, using the data from a 6-week, double-blind, placebo-controlled trial. PANSS Marder factor scores were calculated for each cluster. The efficacy of 10 or 20 mg/day of asenapine on PANSS score was used as the primary endpoint, with the incidence of adverse events evaluated as the secondary endpoint.ResultsA total of 529 asenapine-treated patients were classified into 3 clusters: Cluster-P with the higher scores in positive symptoms, disorganized thoughts, and hostility/excitement, Cluster-N with higher scores in negative symptoms, and Cluster-L with overall lower scores. In Cluster-N and Cluster-L, both 10 and 20 mg/day groups showed significant improvement in PANSS scores, while only the 20 mg/day group showed a significant difference in Cluster-P. Cluster-N and Cluster-L had differences in the incidence of adverse events, but this was not seen in Cluster-P.ConclusionsThe efficacy and safety of asenapine 10 and 20 mg/day differed between the 3 clusters of patients. This suggests that background information regarding baseline psychiatric symptoms may affect the therapeutic response in patients with schizophrenia.

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.

Cap-dependent endonuclease (CEN) resides in the PA subunit of the influenza virus and mediates the critical "cap-snatching" step of viral RNA transcription, which is considered to be a promising anti-influenza target. Here, we describe in vitro characterization of a novel CEN inhibitor, baloxavir acid (BXA), the active form of baloxavir marboxil (BXM). BXA inhibits viral RNA transcription via selective inhibition of CEN activity in enzymatic assays, and inhibits viral replication in infected cells without cytotoxicity in cytopathic effect assays. The antiviral activity of BXA is also confirmed in yield reduction assays with seasonal type A and B viruses, including neuraminidase inhibitor-resistant strains. Furthermore, BXA shows broad potency against various subtypes of influenza A viruses (H1N2, H5N1, H5N2, H5N6, H7N9 and H9N2). Additionally, serial passages of the viruses in the presence of BXA result in isolation of PA/I38T variants with reduced BXA susceptibility. Phenotypic and genotypic analyses with reverse genetics demonstrate the mechanism of BXA action via CEN inhibition in infected cells. These results reveal the in vitro characteristics of BXA and support clinical use of BXM to treat influenza.

Background Mobius syndrome is a congenital disorder with facial and abducens palsy. Although a few case series studies have examined comorbid conditions in Mobius syndrome, follow-up and outcome data are sparse. Objectives To examine the clinical characteristics and outcomes of Mobius syndrome. Methods Clinical data were reviewed for 10 patients. Neonatal history, neurological examination, comorbid anomalies, medical home care, outcomes, and neuroimaging were summarized. Results The patients' mean age was 7.3 ± 6.2 years. On neurological examination, absent blink reflex, jaw ankylosis, absent gag reflex, and tongue atrophy were frequently observed. Poland anomaly and clubfoot were present in three and six patients, respectively. Specific therapies required for patients included medical home care (six patients), suction apparatus (six), tube feeding (five), gastrostomy (two), tracheostomy (three), oxygen therapy (three), and home ventilator (two). Punctate calcification in the brainstem was observed in four patients. Pontine and medulla hypoplasia were detected on the basis of anteroposterior diameter in four and seven patients, respectively. Two patients had congenital hydrocephalus with aqueductal stenosis. Global developmental delay occurred in five patients. Three patients died. Conclusion The rate of both the use of home medical devices and death was high in our patients. Mobius syndrome is extremely diverse, not only in clinical manifestation, but also outcome. Early multidisciplinary intervention is important to ensure an optimal outcome. Aqueductal stenosis is an occasional comorbid anomaly resulting from midbrain abnormality.

This is a survey made in thirty nine Schools for Practical Nurses in São Paulo, Brasil, 1978, about library resources. The aspects surveyed were number and type of books and journals, physical area used as library and its organization.

BACKGROUND Group A rotavirus (RVA) vaccines have been introduced in Japan since 2011. To investigate the molecular epidemiological traits of RVA during the transitional period of rotavirus vaccine implementation in Japan, this study was undertaken by following up three-decade long surveillance conducted in the same regions. METHODS RVA were screened and genotyped by RT-PCR from diarrheal samples collected from non-hospitalized patients in six localities (Hokkaido, Tokyo, Shizuoka, Osaka, Kyoto, and Saga) during 2011 - 2014. Selected samples were sequenced to elucidate the evolutionary trend. RESULTS Among 1858 specimens, the detection rate of RVA declined to 4.0% in 2013 - 2014 from 17.9% in 2011 - 2012 and 22.1% in 2012 - 2013. G1P[8] was the most predominant genotype in the first two years accounting for more than half, and G9P[8] showed the highest detection rate as 35.0% in the last year. Interestingly, the proportional rate of G2 strains in the studied period increased from 0% to 25%. VP6 genotyping revealed that DS-1 like reassortant G1P[8] strains were detected all over Japan and their prevalence fluctuated greatly from 35.0% to 89.5%. Sequence analysis of VP6 showed that strains in the current strains were closely related but distinct from the original reference strains, namely Wa and DS-1. CONCLUSIONS The detection rates of RVA, their GP combinations, prevalence of reassortant strains varied greatly after the introduction of rotavirus vaccines in Japan. Continuous monitoring is warranted to refine future vaccine strategy.

Saffold virus (SAFV) is a newly discovered human virus in the genus Cardiovirus , family Picornaviridae . The virus was first described from fecal specimens of a child with fever of unknown origin in 2007. A total of 454 fecal specimens were collected from children with diarrhea attended clinics in Japan, 2010–2011, 7 (1.5%) were positive for SAFV. Mixed-infections of SAFV and other enteric viruses (rotavirus, norovirus, and bocavirus) were found in four out of seven cases, while monoinfection by SAFV alone was detected in three cases. In addition to diarrhea, fever and vomiting were observed in three children and mild dehydration in one case. No particular symptoms of cough and rhinorrhea were noted. Analysis of partial VP1 nucleotide sequence of 7 Japanese SAFV strains revealed that 5 SAFV sequences were most closely related with SAFV2 reference strains, but separated into SAFV2-A (3 strains) and SAFV2-B (2 strains). In addition, the other two strains were classified as SAFV3. Our results indicated that SAFVs (SAFV2 and SAFV3) were circulated in children with acute gastroenteritis in Japan during 2010 and 2011 epidemic season.

The molecular epidemiology and characterization of rotaviruses obtained from non-hospitalized children with acute gastroenteritis in five different prefectures (Hokkaido, Saga, Tokyo, Osaka, and Kyoto) from July 2009 to June 2011 was investigated. Among 831 fecal specimens tested, rotavirus was found in 165 specimens (19.9%). The rotavirus detection rate in 2010-2011 (23.3%) was higher than those in 2009-2010 (16.0%). The highest prevalence of rotavirus was found in children aged 12 to 23 months. Rotavirus could be detected throughout the 8 month period with a peak in April. We found that G3P[8] was the most prevalent genotype (54.5%), followed by G1P[8] (29.1%), G9P[8] (9.1%), G3P[4] (3.0%), G2P[4] (2.5%), G1P[4] (1.2%), and G4P[8] (0.6%), respectively. Interestingly, G3 strains emerged as the most predominant genotype and replaced G1 rotavirus which had been reported as the most predominant genotype in the previous studies. Phylogenetic analysis revealed that G3 rotavirus strains were closely related to the "new variant G3" 5091 strain, which emerged in Japan in 2003-2004. A significant increase in the prevalence of rotavirus G3 found in this study indicates that rotavirus G3 strain is the major cause of infection in five geographical areas of Japan and may distribute globally in the near future.

To evaluate the embolic effect and degradability of gelatin microspheres (GMS) using various degrees of cross-linkage and particle sizes in rabbit renal artery embolization. Four types of GMS were used, as follows: 2 types of cross-linkage and 2 types of particle size. Twenty-four rabbits (6 in each group) were used for the renal artery embolization. Renal angiography was performed before and after embolization of right renal artery. Follow-up renal angiography was performed 2 days (n = 2), 5 days (n = 2), and 15 days (n = 2) after embolization in each group, and then kidneys were removed for histopathological evaluation. Vascular areas of the angiography were measured by Image J software, and the reperfusion rate was calculated. In renal specimens, residual GMS were checked and the degree of degradation was classified according to a 4-point scale. The mean amounts of large- and small-particle-size GMS injected were 15.0 and 34.3 mg, respectively. Tissue necrosis was confirmed in each group; however, no difference was observed among groups. Renal reperfusion was observed more with small GMS than with large GMS. Renal reperfusion was also observed more with low cross-linked GMS than with high cross-linked GMS. In histopathological specimens, large GMS were confirmed in lobar artery, and small GMS were confirmed in lobular artery. Low cross-linked GMS completely degraded 15 days after embolization. In contrast, high cross-linked GMS were persistent 15 days after embolization. Degree of cross-linkage and particle size affected degradability and reperfusion.

ATP and its metabolites are important for taste signaling in taste buds, and thus a clearance system for them would play critical roles in maintenance of gustatory function. A previous report revealed that mRNAs for ecto-5'-nucleotidase (NT5E) and prostatic acid phosphatase (PAP) were expressed by taste cells of taste buds, and NT5E-immunoreactivity was detected in taste cells. However, there was no information on PAP-immunoreactivity in taste buds. In this study, we examined the expression profile of PAP in rat taste buds. In the isolated rat taste buds, we detected expression of mRNA for PAP, but NT5E was not detected differing from the case of mouse ones (Dando et al., 2012, J Neuroscience). On immunohistochemical analysis, PAP-immunoreactivity was found predominantly in NTPDase2-positive type I and SNAP25-positive type III taste cells, while there were no apparent signals of it in PLC-β2-positive type II, α-gustducin-positive type II, AADC-positive type III and 5HT-positive type III ones. As for NT5E, we could not detect its immunoreactivity in rat taste buds, and co-localization of it with any taste cell markers, although mouse taste buds expressed NT5E as reported previously. These findings suggest that PAP expressed by type I and one of type III taste cells of rats may contribute to metabolic regulation of the extracellular levels of adenine nucleotides in the taste buds of circumvallate papillae, and the regulating mechanisms for adenine nucleotides in taste buds might be different between rats and mice.

We report acute encephalopathy in two cases with severe congenital hydrocephalus. Case 1 was a 23-month-old girl, born at of 36weeks gestation and delivered by cesarean section due to congenital hydrocephalus. Magnetic resonance imaging (MRI) showed prominent ventricular dilation associated with hydrocephalus, Dandy-Walker variant and cortical malformation. The blood test for toxoplasmosis, syphilis, varicella-zoster, rubella, cytomegalovirus, and herpes simplex virus (TORCH) complex and various metabolic tests of blood and urine specimens yielded unremarkable results. She was admitted to our hospital for respiratory failure with fever and her clinical course deteriorated, progressing to hemiconvulsion hemiplegia epilepsy syndrome. Case 2 was a 17-month-old boy, born by spontaneous vertex delivery at 39weeks. Severe, asymmetrical ventricular dilation associated with hydrocephalus, cerebellar and brainstem hypoplasia, and punctuate calcifications of the thalamus, third and fourth ventricles, around the aqueduct, were observed on computed tomography (CT). The blood test for TORCH complex and various metabolic tests of blood and urine specimens yielded unremarkable results. He was admitted to our hospital for status epilepticus with fever and his clinical course progressed to hemorrhagic shock and encephalopathy syndrome. In patients with brain disorders, diagnosis and treatment are likely to be delayed and prognosis may thereby be worsened. When status epileptics or prolonged coma manifests even in patients with severe brain disorders, we must consider encephalopathy in the differential diagnosis.

Of 477 stool specimens, which had been screened for rotavirus, adenovirus, norovirus, sapovirus and astrovirus, collected from infants and children with acute gastroenteritis in pediatric clinics encompassing five localities (Sapporo, Tokyo, Maizuru, Osaka, and Saga) in Japan from July 2007 to June 2008, 247 negative samples (51.7%) were subjected to screening for human parechovirus. Human parechovirus (HPeV) was detected by RT-PCR using a primer pair to amplify 5′UTR region of its genome and was genotyped by sequencing of the VP1 gene. HPeV was detected in 20 of 247 specimens tested, and the detection rate was found to be 8.1%. Seventeen of the 20 strains that tested positive for HPeV were sequenced successfully the VP1 gene. The majority of the HPeV strains (n = 15) could be identified as HPeV1, and the remaining 2 strains could be typed as HPeV3. By phylogenetic and identical matrix analyses of HPeV VP1 sequences, HPeV1 should be divided into two lineages, and all of the Japanese studied HPeV1 strains belong to the lineage 2 accordingly. This is the first report of the circulation of HPeV, especially HPeV1 in Japan. J. Med. Virol. 83:331–336, 2011. © 2010 Wiley-Liss, Inc.

SUMMARY The buttonhole technique of access of needle insertion into a single selected site in the arteriovenous fistula has proved to be a reliable alternative to older methods due to its overall low complication rates. Although the use of blunt needles improves the technique, the success rate of cannulation with these needles is difficult to predict. We analysed the short-term outcome of 16 patients receiving in-centre haemodialysis and compared clinical relevant parameters between patients with and without buttonhole technique failure. Our dialysis unit treats about 180 patients and is located in a tertiary hospital in Sao Paulo, Brazil. The variables as discussed in the paper were the same for both groups. The incidence of technique failure was 43.7%. Patients enrolled later in the study had a better buttonhole failure-free survival rates than patients enrolled at the beginning (p < 0.05). Patients’ clinical characteristics did not predict the success rate of buttonhole tunnel tracks cannulation with blunt needles. This paper also reports on our successes and failures in buttonhole technique and gives some reasons and reflections for both.

The DnaK/Hsp70 family is a molecular chaperone that binds non-native states of other proteins, and affects various physiological processes in bacterial, plant and animal cells. In this study, a dnaK gene from a halotolerant cyanobacterium, Aphanothece halophytica, was introduced into Eustoma grandiflorum. The ApdnaK transformed plants could grow at a similar rate with the control plants expressing the vector alone. Although exposure to low temperature is required for the bolting of control Eustoma, it was not required for the ApdnaK transformant. Under normal growth conditions, the glutathione content in the ApdnaK transformants was higher than that of control plants. In the transformant, the pistil is not evident and consequently, no seed formation. The results suggest the involvement of molecular chaperone for the bolting of Eustoma.

In dicotyledonous plants, nicotianamine synthase (NAS) is thought to play a role in the intercellular transport of iron (Fe). Fe is an essential metal for nitrogen-fixing root nodules of legumes, prompting us to characterize the role of the NAS gene in detail. We previously compared gene-expression profiles in ineffective nodules formed on a Lotus japonicus Fix(-) mutant, sen1, with those in wild-type-effective nodules, and showed that expression of an expressed sequence tag (EST) clone encoding an NAS (EC 2.5.1.43) homologue was repressed in the ineffective nodules. In the present study, two EST clones encoding NAS homologues were found in the EST database. We named them LjNAS1 and LjNAS2. Both were detected as single-copy genes in the L. japonicus genome, and conferred NAS activities in transformed Saccharomyces cerevisiae. LjNAS2 was expressed only in nodules, but LjNAS1 was expressed mainly in leaves, stems, and cotyledons. The level of LjNAS2 transcripts was highest in the nodules 24 days after inoculation with Mesorhizobium loti, and was localized in vascular bundles within the nodules. Expression of LjNAS2 was suppressed in ineffective nodules formed on Fix(-) mutants other than sen1. By contrast, nitrogenase activities of nodules were not influenced in LjNAS2-suppressed plants. We discuss the role of LjNAS2 from the aspect of Fe translocation in nodules.

Astrocytes, one of the predominant types of glial cells, function as both supportive and metabolic cells for the brain. Under cerebral ischemia/reperfusion-induced oxidative conditions, astrocytes accumulate and activate in the ischemic region. DJ-1 has recently been shown to be a sensor of oxidative stress in living cells. However, the function of astrocytic DJ-1 is still unknown. In the present study, to clarify the effect of astrocytic DJ-1 protein under massive oxidative insult, we used a focal ischemic rat model that had been subjected to middle cerebral artery occlusion (MCAO) and reperfusion. We then investigated changes in the distribution of DJ-1 in astrocytes, DJ-1 release from cultured astrocytes, and the effects of recombinant DJ-1 protein on hydrogen peroxide (H2O2)-induced death in normal and DJ-1-knockdown SH-SY5Y cells and on in vitro scavenging of hydroxyl radicals (•OH) by electron spin resonance spectrometry. At 24 h after 2-h MCAO and reperfusion, an infarct lesion was markedly observed using magnetic resonance imaging and 2,3,5-triphenyltetrazolium chloride staining. In addition, reactive astrocytes enhanced DJ-1 expression in the penumbral zone of the ischemic core and that DJ-1 protein was extracellularly released from astrocytes by H2O2 in in vitro primary cultures. Although DJ-1-knockdown SH-SY5Y cells were markedly vulnerable to oxidative stress, treatment with glutathione S-transferase-tagged recombinant human DJ-1 protein (GST-DJ-1) significantly inhibited H2O2-induced cell death. In addition, GST-DJ-1 protein directly scavenged•OH. These results suggest that oxidative stress induces the release of astrocytic DJ-1 protein, which may contribute to astrocyte-mediated neuroprotection.

Norovirus is one of the most common causes of nonbacterial gastroenteritis in humans. A total of 603 fecal specimens collected from sporadic pediatric cases of acute gastroenteritis in Japan from 2004 to 2005 were tested for the presence of norovirus by RT-PCR. It was found that 51 (8.5%) specimens were positive for norovirus. The norovirus genotypes detected in this study were GII/1, GII/2, GII/3, GII/4, GII/6, and GII/7. Of these, GII/3 was the most predominant (52.9%), followed by GII/4 (37.2%) and others. It was noticed that four distinct types of recombinant noroviruses were co-circulating and the variant norovirus GIIb suddenly emerged to be the leading strain in Japan for the first time. A novel norovirus nomenclature was proposed, in which worldwide noroviruses were classified into seven distinct genogroups (I-VII). Norovirus GI and GII consisted of 16 genotypes with 32 subgenotypes and 23 genotypes with 34 subgenotypes, respectively. Of note, human and porcine noroviruses had a close genetic relationship within GII. Interestingly, multiple short amino acid motifs located at N terminus, S domain, P1 domain, P2 domain, and C terminus of capsid gene correctly defined the phylogenetic norovirus genogroups, genotypes, and subgenotypes. Another interesting feature of the study was the identification of eight hitherto unreported recombinant noroviruses. It was noteworthy that three different types (intergenogroup, intergenotype, and intersubgenotype) of recombination in noroviruses were also found. This is the first report to demonstrate the existence of intergenogroup and intersubgenotype recombinations in noroviruses and highlights a possible route of zoonoses in humans because porcine, bovine and murine noroviruses belong to genogroups II, III, and V, respectively.

Background Unintended exposure to antitumor agents from an oral medicine may place healthcare workers and patients taking medicine at risk. In this study, the exposure to blister pack by CP (cyclophosphamide) and appropriate preventive procedures were examined. Findings CP detected inside the blister pack of the tested seven lots by LC-MS/MS ranged from 8.2 to 199.6 ng. Raman imaging clearly showed that CP ingredient was completely covered by the tablet coating layer and had not leached out of the tablet. In addition, the amount of CP detected inside the vials was suppressed under the lower detection limit until day 28, and only 6.0 ng was detected only at day 56. Conclusions Various amounts of CP were contaminated to not only the inside of the blister pack but also the outside. This contamination may be caused not only by the manufacturing environment but also by the CP oral tablets themselves through volatilization of CP. Refrigerated storage of CP oral tablets may protect healthcare workers and patients from contact with CP. Electronic supplementary material The online version of this article (doi:10.1186/s40780-015-0020-9) contains supplementary material, which is available to authorized users.

A total of 402 fecal specimens collected during July 2003-June 2004 from infants and children with acute gastroenteritis, encompassing five localities (Maizuru, Tokyo, Sapporo, Saga, and Osaka) of Japan, were tested for the presence of norovirus by RT-PCR. It was found that 58 (14.4%) fecal specimens were positive for norovirus. Norovirus infection was detected throughout the year with the highest prevalence in December. Norovirus GII was the most predominant genogroup (98.3%; 57 of 58). The genotypes detected in this study were GI/4, GII/2, GII/3, GII/4, and GII/6. Of these, NoV GII/3 (known as the Arg320 virus cluster) was the most predominant genotype (43.9%), followed by NoV GII/4 (the Lordsdale virus cluster; 35.1%) and others. Two norovirus strains clustered with a "new variant designated GIIb" and a "new variant of GII/4" were found circulating in Japan for the first time. It was interesting to note that NoV GIIb and NoV GII/3 appeared to be the recombinant strains and the recombination site was demonstrated at the overlap of ORF1 and ORF2. The majority (96%) of the dominant norovirus strains were identified as the recombination of GII/3 capsid and GII/12 polymerase. The recombination in the NoV GIIb capsid gene at the breakpoint located at P1 domain was also identified. Obviously, NoV GIIb isolate in Japan had double recombination. This is the first report demonstrating the existence of different "new variants" co-circulating in Japanese infants and children with acute gastroenteritis.

A tumor within the right globe in a 9-year-old Shetland sheepdog was examined pathologically. The tumor was composed of spindle or oval cells arranged in interwoven bundles with intervening collagenous or mucinous matrices. Immunohistochemically, the tumor bound antibody directed to S-100 protein and vimentin, but not to desmin, actin smooth muscle, or neurofilament. Electron microscopy revealed that the tumor cells had poorly developed cytoplasmic processes, desmosomes between closely contiguous cells, and a discontinuous basement membrane-like material. Based on these findings, the tumor was diagnosed as a peripheral nerve sheath tumor (PNST) histologically. To the authors' knowledge, this is the first reported case of intraocular PNST in dogs.

Transgenic tobacco plants expressing the ascorbate oxidase (AAO) gene in sense and antisense orientations, and an Arabidopsis mutant in which the T-DNA was inserted into a putative AAO gene, were used to examine the potential roles of AAO for salt-stress tolerance in plants. AAO activities in the transgenic tobacco plants expressing the gene in sense and antisense orientations were, respectively, about 16-fold and 0.2-fold of those in the wild type. Under normal growth conditions, no significant differences in phenotypes were observed, except for a delay in flowering time in the antisense plants. However, at high salinity, the percentage germination, photosynthetic activity, and seed yields were higher in antisense plants, with progressively lower levels in the wild type and the sense plants. The redox state of apoplastic ascorbate in sense plants was very low even under normal growth conditions. Upon salt stress, the redox state of symplastic and apoplastic ascorbate decreased among the three types of plants, but was lowest in the sense plants. The hydrogen peroxide contents in the symplastic and apoplastic spaces were higher in sense plants, progressively lower in the wild type, followed by the antisense plants. The Arabidopsis T-DNA inserted mutant exhibited very low ascorbate oxidase activity, and its phenotype was similar to that of antisense tobacco plants. These results suggest that the suppressed expression of apoplastic AAO under salt-stress conditions leads to a relatively low level of hydrogen peroxide accumulation and a high redox state of symplastic and apoplastic ascorbate which, in turn, permits a higher seed yield.

Two experiments were designed to investigate shifting attention from an adult’s face to an adult’s hand by 3- and 4-month-olds. In Experiment 1, 24 infants were presented with five types of hand gestures by their mothers and a stranger. Experiment 2 was given to 22 infants with the same procedures, except for the addition of a head inclining while pointing to objects. The results were: (1) after encountering an averted head repeatedly, the infants shifted their attention from the adult’s face to the moving hand and objects; they oriented to what the adult was attending to. (2) The moving head improved the rate of infants turning their heads to the same direction as the adult. The conclusion was averted head and eyes play a major role in infants’ orienting to an adult’s hand. A hand was a shared visual target during the adult’s object performance, indicating that infants’ orientation to the adults’ hand is a precursor stage of joint visual attention.

A unique taste-modifying activity that converts the sense of sourness to the sense of sweetness occurs in the fruit of the plant Curculigo latifolia, intrinsic to West Malaysia. The active component, known as curculin, is a protein consisting of two identical subunits. We have found a new taste-modifying protein, named neoculin, of the same origin. Both chemical analysis and cDNA cloning characterized neoculin as a heterodimeric protein consisting of an acidic, glycosylated subunit of 113 amino acid residues and a basic subunit that is the monomeric curculin itself.

Background S-033447, an active form of orally available prodrug S-033188, is a novel small molecule inhibitor of cap-dependent endonuclease that is essential for influenza virus transcription and replication. In this study, we evaluated the inhibitory effect of S-033188 in combination with neuraminidase inhibitors on the replication of influenza A/H1N1 virus in cultured cells. Methods The inhibitory effects of S-033447 in combination with NA inhibitors on the cytopathic effect of A/PR/8/34 strain in Madin–Darby canine kidney cells cultured for 2 days were tested and EC50 were determined. The combination index (CI), which were obtained when S-033188 and NA inhibitor were added at the closest ratio of each EC50 value, were used for the evaluation of these combinational effects (Table 1). CI values were calculated by the Chou and Talalay method, in which combinational effect were determined according to the criteria as follows: synergistic if CI ≤ 0.8, additive if 0.8 < CI < 1.2, and antagonistic if CI ≥ 1.2. CI = (DA/A + B)/DA + (DB/A + B)/DB + (DA/A + B × DB/A + B)/(DA × DB) DA: the EC50 of S-033447 DB: the EC50 of NA inhibitor DA/A + B: the concentration of S-033447 giving 50% inhibition in combination with NA inhibitor at the closest ratio of each EC50 value DB/A + B: the concentration of NA inhibitor giving 50% inhibition in combination with S-033447 at the closest ratio of each EC50 value Results All CI values were lower than 0.8, under the condition that both S-033447 and NA inhibitor (oseltamivir acid, zanamivir hydrate, laninamivir, or peramivir trihydrate) were added at the closest ratio of each EC50 value (Table 1). Conclusion S-033447 in combination with oseltamivir acid, zanamivir hydrate, laninamivir, or peramivir trihydrate synergistically inhibited the replication of influenza A/H1N1 virus in MDCK cells. Disclosures All authors: No reported disclosures.

portanto perfeitamen-te adequado. Esse termo tambem ja foi empregado pela comunidade medica por -tuguesa. Sugerimos ao JBN, e a comuni-dade nefrologica brasileira em geral, a normatizacao deste termo em trabalhos futuros.O segundo comentario refere-se aos aspectos medicos. Em nossa experien-cia, recentemente publicada (Galante

Purpose To evaluate the efficacy and safety of fosphenytoin (fPHT) for the treatment of seizures in children with acute encephalopathy. Methods Using responses from physicians on the Annual Zao Conference on Pediatric Neurology mailing list we chose patients who met the following criteria: clinical diagnosis of acute encephalopathy and use of intravenous fPHT for the treatment of seizures. We divided the patients into two groups: acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) and other encephalopathies. The efficacy of fPHT was considered effective when a cessation of seizures was achieved. Results Data of 38 children were obtained (median age, 27 months). Eighteen children were categorized into the AESD group and 20 into the other encephalopathies group. fPHT was administered in 48 clinical events. The median loading dose of fPHT was 22.5 mg/kg and was effective in 34 of 48 (71%) events. The rate of events in which fPHT was effective did not differ according to the presence or absence of prior antiepileptic treatment, subtype of acute encephalopathy, or the type of seizures. One patient experienced apnea and oral dyskinesia as adverse effects of fPHT, whereas arrhythmia, hypotension, obvious reduction of consciousness, local irritation, phlebitis and purple grove syndrome were not observed in any patient. Conclusion fPHT is effective and well tolerated among children with acute encephalopathy.

"Two different methods were used to clarify whether total measured vitamin C (ascorbic acid plus dehydro-ascorbic acid) content of fresh, frozen and cooked vegetables from commercial sources depends on the method used. Total vitamin C content in frozen and cooked vegetables measured by HPLC was only 47.5% of that measured by the colorimetric method. Such an overestimation of vitamin C of prepared vegetables by the colorimetric method was due to the color development from other compounds such as 2, 3-diketogulonic acid, seasonings, sugar and starch. Total vitamin C content in cooked vegetables from commercial sources measured by HPLC should be one third to one fifth of that in fresh vegetables."