ADGRL1 variants: From developmental and epileptic encephalopathy to genetic epilepsy with febrile seizures plus.

AIM: To expand the phenotypic spectrum of ADGRL1 and explore the correlation between epilepsy and the ADGRL1 genotype. METHOD: We performed whole-exome sequencing in a cohort of 115 families (including 195 males and 150 females) with familial febrile seizure or epilepsy with unexplained aetiology. The damaging effects of variants was predicted using protein modelling and multiple in silico tools. All reported patients with ADGRL1 pathogenic variants were analysed. RESULTS: One new ADGRL1 variant (p.Pro753Leu) was identified in one family with genetic epilepsy with febrile seizures. Further analysis of 12 ADGRL1 variants in 16 patients revealed that six patients had epilepsy. Epilepsy types ranged from early-onset epileptic encephalopathy to genetic epilepsy with febrile seizures plus (GEFS+). All four variants associated with epilepsy were located in the non-helix or sheet region of ADGRL1. Three of the four epilepsy-associated variants were missense variants. Thus, all three variants located in the G-protein-coupled receptor autoproteolytic-inducing domain exhibited epilepsy. INTERPRETATION: We found one new missense variant of ADGRL1 in one family with GEFS+. ADGRL1 may be a potential candidate or susceptibility gene for epilepsy. ADGRL1-associated epilepsy ranged from benign GEFS+ to severe epileptic encephalopathy; the genotypes and variant locations may help explain the phenotypic heterogeneity of patients with the ADGRL1 variant.

Role of TRAK1 variants in epilepsy: genotype-phenotype analysis in a pediatric case of epilepsy with developmental disorder.

Purpose: The TRAK1 gene is mapped to chromosome 3p22.1 and encodes trafficking protein kinesin binding 1. The aim of this study was to investigate the genotype-phenotype of TRAK1-associated epilepsy. Methods: Trio-based whole-exome sequencing was performed on a cohort of 98 patients with epilepsy of unknown etiologies. Protein modeling and the VarCards database were used to predict the damaging effects of the variants. Detailed neurological phenotypes of all patients with epilepsy having TRAK1 variants were analyzed to assess the genotype-phenotype correlations. Results: A novel TRAK1 compound heterozygous variant comprising variant c.835C > T, p.Arg279Cys and variant c.2560A > C, p.Lys854Gln was identified in one pediatric patient. Protein modeling and VarCards database analyses revealed that the variants were damaging. The patient received a diagnosis of early infantile epileptic spasms with a developmental disorder; he became seizure-free through valproate and adrenocorticotropic hormone treatment. Further results for six variants in 12 patients with epilepsy indicated that biallelic TRAK1 variants (including homozygous or compound heterozygous variants) were associated with epilepsy with developmental disorders. Among these patients, eight (67%) had epileptic spasms and seven (58%) were intractable to anti-seizure medicines. Moreover, eight patients experienced refractory status epilepticus, of which seven (88%) died in early life. To our knowledge, this is the first reported case of epilepsy caused by TRAK1 compound heterozygous variants. Conclusion: Biallelic TRAK1 variants can cause epilepsy and developmental disorders. In these patients, seizures progress to status epilepticus, suggesting a high risk for poor outcomes and the requirement of early treatment.

The increase of Nrf2 m6A modification induced by FTO downregulation promotes hippocampal neuron injury and aggravates the progression of epilepsy in a rat model.

Epilepsy is a common chronic neurological disorder characterized by widespread neuronal death. The purpose of this study was to investigate the role of nuclear factor erythroid 2-related factor 2 (Nrf2) m6A methylation in epilepsy. To create epileptic models, the rats were given Lithium chloride and pilocarpine, and isolated primary rat hippocampal neurons were cultured in an Mg2+ -free medium. The frequency of seizures was recorded in the epilepsy group of rats. The functional tests included TUNEL, MTT, and flow cytometry. Mechanistically, RNA degradation assay, RNA immunoprecipitation, and methylated RNA immunoprecipitation were performed. In epileptic models, Nrf2 and fat mass and obesity-associated (FTO) levels were downregulated, whereas YT521-B homology (YTH) domain family protein 2 (YTHDF2) was upregulated. Additionally, in epileptic models, there was a rise in the m6A methylation level of Nrf2 mRNA. Overexpressing FTO increased cell viability and reduced apoptosis, but Nrf2 interference reversed these effects. Meanwhile, FTO overexpression decreased the m6A methylation of Nrf2 mRNA. Moreover, YTHDF2 bound to Nrf2 mRNA and decreased its stability. Furthermore, FTO overexpression reduced seizure frequency in rats and inhibited hippocampal neuron apoptosis via lowering the m6A methylation level of Nrf2 mRNA. Overexpressing FTO reduced m6A methylation of Nrf2 mRNA, increased cell viability, suppressed apoptosis, and slowed the progression of epileptic diseases, which is linked to YTHDF2 binding to m6A-modified Nrf2 and promoting its degradation, as well as downregulating Nrf2 expression in hippocampal neurons.

First report of brown rot caused by Monilia mumecola on Chinese plum (Prunus salicina) in Fujian, China.

Brown rot caused by Monilinia spp. (anamorph Monilia) is a common disease in stone fruits worldwide, but the species in different hosts or regions may vary. Monilia mumecola is a recently identified species, only reported in some regions. Although the pathogen has been found on plum in Yunnan, Hubei, and Zhejiang provinces, and Chongqing municipality in China (Yin et al. 2015), it has not been reported in southeast coast of China. In May 2022, brown rot with grey spores on fruit was observed in a plum orchard with 15% disease incidence in Sanming City, Fujian Province, located in southeast coast of China. Four single-spored isolates were obtained from germinating conidia on the water agar for further investigation. The colonies on potato dextrose agar (PDA) were initially white, gradually turned gray to brown, with lobbed margins and rare sporulation. Average mycelial growth rate ranged from 0.74 to 1.08 cm/day at 25 oC. Conidia were lemon-shaped or subglobose, hyaline, with an average size of 17.64 to 19.35×11.14 to 14.44 µm (n=30). Each isolate produced one to three or four germ tubes. Such characteristics are similar to M. mumecola (Yin et al. 2015). To confirm the identity of the isolates, genomic DNA was extracted and species-specific primers of Hu et al. (2011) were used to amplify the 712 bp sequence. In addition, the ITS region, partial genes of glyceraldehyde-3- phosphate dehydrogenase (GAPDH) and beta-tubulin (TUB2) were also amplified using primers sets ITS1/ITS4 (Glass and Donaldson 1995), Mon-G3pdhF/Mon-G3pdhR and Mon-TubF1/ Mon-TubR1, respectively (Hu et al. 2011). Sequences obtained for those three regions were 478, 762 and 1527 bp, respectively. Each region of all four isolates was identical, so one sequence for each region was submitted to GenBank with accession numbers OQ207672, OQ225251 and OQ225252, respectively, which had 100% identity with M. mumecola HQ908786 (ITS), HQ908784 (GAPDH), and HQ908775 (TUB2) using BLAST analysis in NCBI database, respectively. Pathogenicity was conducted with mycelium plugs from the edge of 7-day-old colony on three mature 'Angeleno' plums fruit (Prunus salicina) creating nine inoculations with three wounds per fruit, and each wound was 5 mm in diameter and 2 mm in depth. The same amount fruit and wounds inoculated with PDA plugs without fungi were used as a control. Brown rot symptoms were observed on all inoculated plums 4 days post-inoculation under room temperature with 100% humidity, whereas control plums remained symptomless. Fungal colonies re-isolated from the lesions showed the same morphological features as the original isolate , thus fulfilling Koch's postulates. To our knowledge, this is the first report of M. mumecola on plum in Fujian Province of China. The findings in this studies have important management implications for local plum growers because more than one Monilia species have been reported, where only M. fructicola was present in this region.

Prevalence of H6Y mutation in ß-tubulin causing thiophanate-methyl resistant in Monilinia fructicola from Fujian, China.

Brown rot disease broke out in stone fruit orchards of Fujian, China in 2019, despite pre-harvest application of methyl benzimidazole carbamate (MBC). To determine the reason, a total of 44 Monilinia fructicola strains were collected from nectarine, plum and peach fruits in this study, among which 79.5% strains were resistant to thiophanate-methyl, indicated by discriminatory dose of 5 µg/mL. The resistance of these strains was confirmed by treating detached peach fruit with label rates of formulated thiophanate-methyl which only completely inhibit infection of the sensitive strains, but not the resistant strains. Further analysis of the mechanism of MBC resistance revealed that all resistant strains carry a H6Y mutation in ß-tubulin protein Tub2, which was only reported previously in the M. fructicola strains from California, USA, and do not display obvious fitness penalties, as no significant defects in mycelial growth rate, sporulation, conidia germination, aggressiveness on detached peach fruit and temperature sensitivity was detected. In addition, we found that diethofencarb, the agent for managing MBC-resistance strains, was unable to inhibit growth of the H6Y strains. Taken together, our study, for the first time, identified a mutation form of H6Y in the ß-tubulin protein of M. fructicola in China, rendering the strains wide resistance to thiophanate-methyl. This mechanism of M. fructicola gaining resistance to MBC fungicides needs to be fully considered, when designing management strategies to control brown rot disease in stone fruit orchards.

Damaging novel mutations in PIGN cause developmental epileptic-dyskinetic encephalopathy: a case report.

BACKGROUND: Mutations in PIGN, resulting in a glycosylphosphatidylinositol (GPI) anchor deficiency, typically leads to multiple congenital anomalies-hypotonia-seizures syndrome. However, the link between PIGN and epilepsy or paroxysmal non-kinesigenic dyskinesia (PNKD) is not well-described. This study reported a patient with PIGN mutation leading to developmental and epileptic encephalopathy and PNKD, to expand upon the genotype-phenotype correlation of PIGN. CASE PRESENTATION: During the first 10 days of life, a girl exhibited paroxysmal staring episodes with durations that ranged from several minutes to hours. These episodes occurred 2-5 times daily and always occurred during wakefulness. Ictal electroencephalography revealed no abnormalities, and PNKD was diagnosed. The patient also exhibited severely delayed psychomotor development and generalized seizures at the age of 4 months. Results of brain magnetic resonance imaging and metabolic screenings were normal, but trio-based whole-exome sequencing identified two novel compound heterozygous PIGN mutations (NM_176787; c.163C > T [p.R55 > X] and c.283C > T [p.R95W]). Flow cytometry analysis of the patient's granulocytes revealed dramatically reduced expression of GPI-anchored proteins. This indicated that the mutations compromised GPI functions. The patient got seizure-free for 1 year, and her dyskinesia episodes reduced significantly (1-2 times/month) after treatment with levetiracetam (600 mg/day) and clonazepam (1.5 mg/day). No progress was observed with respect to psychomotor development; however, no craniofacial dysmorphic features, cleft lip/palate, brachytelephalangy with nail hypoplasia, and internal malformations have been observed until now (6 years of age). CONCLUSION: This is the first study to document developmental and epileptic encephalopathy with PNKD in a human with PIGN mutations. This report expanded our understanding of the genotype-phenotype correlation of PIGN, and PIGN may be considered a potentially relevant gene when investigating cases of epilepsy or PNKD.

[Fever for 2 months and disturbance of consciousness for 1 week in a preschool-aged girl].

Rapid-onset obesity with hypothalamic dysfunction, hypoventilation, and autonomic dysregulation syndrome (ROHHADS) is a rare multi-system disease, and delayed diagnosis and treatment may lead to catastrophic cardiopulmonary complications. As far as we know, no patient with ROHHADS has been reported in China, and this article reports a child with ROHHADS to improve the awareness of this disease among clinicians. A girl, aged 3 years, had the clinical manifestations of rapid weight gain, fever, disturbance of consciousness, and convulsion. The physical examination showed a body weight of 20 kg, somnolence, irregular breathing, and stiff neck. She had increased blood levels of prolactin and follicle-stimulating hormone and hyponatremia. The lumbar puncture showed an increased intracranial pressure. The brain MRI and magnetic resonance venography showed symmetrical lesions in the periventricular region and venous thrombosis in the right transverse sinus and the superior sagittal sinus. The sleep monitoring showed hypopnea. The girl was finally diagnosed with ROHHADS and intracranial venous thrombosis. She recovered after symptomatic treatment including decreasing intracranial pressure, anticoagulation, and respiratory support. The possibility of ROHHADS should be considered for patients with unexplained obesity, fever, and hypoventilation, with or without central nervous system symptoms. Early diagnosis and standardized follow-up can improve the prognosis of children with ROHHADS.

Vaccine-Associated Neurological Adverse Events: A Case Report and Literature Review.

Vaccination is an effective strategy to reduce the burden of preventable illness. However, many clinical reports revealed that various vaccinations may associate with neurological disorders, mainly including autoimmune disease, febrile seizure, and vaccine-associated paralytic poliomyelitis (VAPP). Although more and more reports revealed that part of the above post-vaccine neurological disorders is not directly related to vaccination, it may be merely a coincidence. However, these reports may increase the hesitancy on vaccination for the public population and influence the coverage of vaccination. In this report, we described a child with acute flaccid paralysis possibly caused by a poliovirus vaccine. To provide feasible ways to realize or reduce the risk of neurological adverse events caused by vaccines, we further provide a mini-review of the literature of vaccination associated with neurological adverse events. This revealed that oral poliomyelitis vaccine use exclusively and type 2 serotype poliomyelitis vaccine virus were the risk factors for VAPP. The combination vaccine was associated with an increased risk of ADEM and FS following immunization when compared with the administration of vaccines separately. Even though cases have been reported that vaccination may be a trigger of anti-NMDARe and GBS, there is no direct evidence to prove that vaccination increased the risk of GBS and anti-NMDARe.

Japanese Encephalitis Virus-Induced Anti-N-Methyl-D-Aspartate Receptor Encephalitis: A Case Report and Review of Literature.

Anti-N-methyl-D-aspartate receptor encephalitis (anti-NMDARe) was originally described as a paraneoplastic disease with more than 50% cases involving a tumor. However, tumor incidence in anti-NMDARe in children is much lower. Herpes simplex virus-induced anti-NMDARe has been well-described; however, findings on Japanese encephalitis virus (JEV)-induced anti-NMDARe are scarce. Here, we describe a 7-year-old boy who presented with fever and headache that progressed to seizures and disturbance of consciousness. Brain magnetic resonance imaging (MRI) revealed abnormalities in the bilateral globus pallidus. The diagnosis of JE was made based on a positive JE antibody test results in serum and cerebrospinal fluid. Antiviral and symptomatic therapies led to rapid recovery. Four weeks after the onset of JE, the patient presented with emotional and behavioral disturbances, sleep difficulty, and extrapyramidal symptoms. MRI showed symmetrical lesions in the bilateral thalami and basal ganglia which were expanded than those on the original scan. Antibodies against NMDAR were detected and immunotherapy led to significant recovery. This case and our literature review suggest that JEV may be a clinically important cause of anti-NMDARe in children. Patients with JE-induced anti-NMDARe present with symptoms similar to those of patients with primary anti-NMDARe. Most patients with JE-induced anti-NMDARe showed a good response to first-line immunotherapies.

Invasive Laryngeal Squamous Cell Carcinoma in a Boy.

Laryngeal squamous cell carcinoma (SCC) is rare in children. Usually, laryngeal SCC in children has a poor prognosis. A 9-year-old boy is reported who was diagnosed as having poorly differentiated laryngeal squamous cell carcinoma with neck metastasis. This report aims to highlight the importance of a comprehensive knowledge of differential diagnosis, putting great attention to the onset of symptoms, early application of flexible laryngoscopy, and intensive studies on similar cases.