Establishment of the auxin inducible degron system for Babesia duncani: a conditional knockdown tool to study precise protein regulation in Babesia spp.

BACKGROUND: Babesia duncani is a pathogen within the phylum Apicomplexa that causes human babesiosis. It poses a significant threat to public health, as it can be transmitted not only through tick bites but also via blood transfusion. Consequently, an understanding of the gene functions of this pathogen is necessary for the development of drugs and vaccines. However, the absence of conditional gene knockdown tools has hindered the research on this pathogen. The auxin-inducible degron (AID) system is a rapid, reversible conditional knockdown system widely used in gene function studies. Thus, there is an urgent need to establish the AID system in B. duncani to study essential gene functions. METHODS: The endogenous genes of the Skp1-Cullin-F-box (SCF) complex in B. duncani were identified and confirmed through multiple sequence alignment and conserved domain analysis. The expression of the F-box protein TIR1 from Oryza sativa (OsTIR1) was achieved by constructing a transgenic parasite strain using a homologous recombination strategy. Polymerase chain reaction (PCR), western blot, and indirect immunofluorescence assay (IFA) were used to confirm the correct monoclonal parasite strain. The degradation of enhanced green fluorescent protein (eGFP) tagged with an AID degron was detected through western blot and live-cell fluorescence microscopy after treatment of indole-3-acetic acid (IAA). RESULTS: In this study, Skp1, Cul1, and Rbx1 of the SCF complex in B. duncani were identified through sequence alignment and domain analysis. A pure BdTIR1 strain with expression of the OsTIR1 gene was constructed through homologous recombination and confirmed. This strain showed no significant differences from the wild type (WT) in terms of growth rate and proportions of different parasite forms. The eGFP tagged with an AID degron was successfully induced for degradation using 500 µM IAA. Grayscale analysis of western blot indicated a 61.3% reduction in eGFP expression levels, while fluorescence intensity analysis showed a 77.5% decrease in fluorescence intensity. Increasing the IAA concentration to 2 mM accelerated eGFP degradation and enhanced the extent of degradation. CONCLUSIONS: This study demonstrated the functionality of the AID system in regulating protein levels by inducing rapid degradation of eGFP using IAA, providing an important research tool for studying essential gene functions related to invasion, egress, and virulence of B. duncani. Moreover, it also offers a construction strategy for apicomplexan parasites that have not developed an AID system.

Babesia gibsoni Whole-Genome Sequencing, Assembling, Annotation, and Comparative Analysis.

The intracellular protozoan parasite Babesia gibsoni infects canine erythrocytes and causes babesiosis. The hazards to animal health have increased due to the rise of B. gibsoni infections and medication resistance. However, the lack of high-quality full-genome sequencing sets has expanded the obstacles to the development of pathogeneses, drugs, and vaccines. In this study, the whole genome of B. gibsoni was sequenced, assembled, and annotated. The genomic size of B. gibsoni was 7.94 Mbp in total. Four chromosomes with the size of 0.69 Mb, 2.10 Mb, 2.77 Mb, and 2.38 Mb, respectively, 1 apicoplast (28.4 Kb), and 1 mitochondrion (5.9 Kb) were confirmed. KEGG analysis revealed 2,641 putative proteins enriched on 316 pathways, and GO analysis showed 7,571 annotations of the nuclear genome in total. Synteny analysis showed a high correlation between B. gibsoni and B. bovis. A new divergent point of B. gibsoni occurred around 297.7 million years ago, which was earlier than that of B. bovis, B. ovata, and B. bigemina. Orthology analysis revealed 22 and 32 unique genes compared to several Babesia spp. and apicomplexan species. The metabolic pathways of B.gibsoni were characterized, pointing to a minimal size of the genome. A species-specific secretory protein SA1 and 19 homologous genes were identified. Selected specific proteins, including apetala 2 (AP2) factor, invasion-related proteins BgAMA-1 and BgRON2, and rhoptry function proteins BgWH_04g00700 were predicted, visualized, and modeled. Overall, whole-genome sequencing provided molecular-level support for the diagnosis, prevention, clinical treatment, and further research of B. gibsoni. IMPORTANCE The whole genome of B. gibsoni was first sequenced, annotated, and disclosed. The key part of genome composition, four chromosomes, was comparatively analyzed for the first time. A full-scale phylogeny evolution analysis based on the whole-genome-wide data of B. gibsoni was performed, and a new divergent point on the evolutionary path was revealed. In previous reports, molecular studies were often limited by incomplete genomic data, especially in key areas like life cycle regulation, metabolism, and host-pathogen interaction. With the whole-genome sequencing of B. gibsoni, we provide useful genetic data to encourage the exploration of new terrain and make it feasible to resolve the theoretical and practical problems of babesiosis.

High TXNIP expression accelerates the migration and invasion of the GDM placenta trophoblast.

INTRODUCTION: Our previous study has proofed the glucose sensitive gene-thioredoxin-interacting protein (TXNIP) expression was up in the placenta of the patients with gestational diabetes mellitus (GDM), but the pathological mechanisms underlying abnormal TXNIP expression in the placenta of patients with GDM is completely unclear and additional investigations are required to explain the findings we have observed. In the present study, we simulated the high TXNIP expression via introducing the Tet-On "switch" in vitro, approximate to its expression level in the real world, to explore the following consequence of the abnormal TXNIP. METHODS: The expression and localization of TXNIP in the placenta of GDM patients and the health control was investigated via immunofluorescent staining, western blot and RT-qPCR. Overexpression of TXNIP was achieved through transfecting Tet-on system to the human trophoblastic cell line-HTR-8/Svneo cell. TXNIP knockout was obtained via CRISPR-Cas9 method. The cell phenotype was observed via IncuCyte Imaging System and flow cytometry. The mechanism was explored via western blot and RT-qPCR. RESULTS: The expression level of TXNIP in the GDM placenta was nearly 2-3 times higher than that in the control. The TXNIP located at trophoblastic cells of the placenta. When the expression of TXNIP was upregulated, the migration and invasion of the cells accelerated, but cell apoptosis and proliferation did not changed compared with the control group. Furthermore, the size of the TetTXNIP cells became larger, and the expression level of Vimentin and p-STAT3 increased in the TetTXNIP cells. All the changes mentioned above were opposite in the TXNIP-KO cells. CONCLUSIONS: Abnormal expression of TXNIP might be related to the impairment of the GDM placental function, affecting the migration and invasion of the placental trophoblast cells through STAT3 and Vimentin related pathway; thus, TXNIP might be the potential therapeutic target for repairing the placental dysfunction deficient in GDM patients.

Tandem mass tag (TMT)-based proteomic analysis of Cryptosporidium andersoni oocysts before and after excystation.

BACKGROUND: Cryptosporidium andersoni initiates infection by releasing sporozoites from oocysts through excystation. However, the proteins involved in excystation are unknown. Determining the proteins that participate in the excystation of C. andersoni oocysts will increase our understanding of the excystation process. METHODS: Cryptosporidium andersoni oocysts were collected and purified from the feces of naturally infected adult cows. Tandem mass tags (TMT), coupled with liquid chromatography-tandem mass spectrometry (LC-MS/MS) proteomic analysis, were used to investigate the proteomic expression profiles of C. andersoni oocysts before and after excystation. RESULTS: Proteomic analysis identified a total of 1586 proteins, of which 17 were differentially expressed proteins (DEPs) upon excystation. These included 10 upregulated and seven downregulated proteins. The 17 proteins had multiple biological functions associated with control of gene expression at the level of transcription and biosynthetic and metabolic processes. Quantitative real-time RT-PCR of eight selected genes validated the proteomic data. CONCLUSIONS: This study provides information on the protein composition of C. andersoni oocysts as well as possible excystation factors. The data may be useful in identifying genes for diagnosis, vaccine development, and immunotherapy for Cryptosporidium.

Midterm outcomes of penetrating keratoplasty following allogeneic cultivated limbal epithelial transplantation in patients with bilateral limbal stem cell deficiency.

AIM: To evaluate the midterm outcomes of penetrating keratoplasty (PK) following allogeneic cultivated limbal epithelial transplantation (CLET) for bilateral total limbal stem cell deficiency (LSCD). METHODS: Ten patients (10 eyes) with bilateral LSCD were enrolled in this prospective noncomparative case series study. Each participant underwent PK approximately 6mo after a CLET. Topical tacrolimus, topical and systemic steroids, and oral ciclosporin were administered postoperatively. Best-corrected visual acuity (BCVA), intraocular pressure (IOP), ocular surface grading scores (OSS), corneal graft epithelial rehabilitation, persistent epithelial defect (PED), immunological rejection, and graft survival rate were assessed. RESULTS: The time interval between PK and allogeneic CLET was 6.90±1.29 (6-10)mo. BCVA improved from 2.46±0.32 logMAR preoperatively to 0.77±0.55 logMAR post-PK (P<0.001). Kaplan-Meier analysis of mean graft survival revealed graft survival rates of 100% at 12 and 24mo and 80.0% at 36mo. PEDs appeared in 5 eyes at different periods post-PK, and graft rejection occurred in 4 eyes. The total OSS decreased from 12.4±4.4 before allogeneic CLET to 1.4±1.51 after PK. CONCLUSION: A sequential therapy design of PK following allogeneic CLET can maintain a stable ocular surface with improved BCVA despite the relatively high graft rejection rate.

Enhanced Gating Performance of Single-Molecule Conductance by Heterocyclic Molecules.

Enhancing the gating performance of single-molecule conductance is significant for realizing molecular transistors. Herein, we report a new strategy to improve the electrochemical gating efficiency of single-molecule conductance with fused molecular structures consisting of heterocyclic rings of furan, thiophene, or selenophene. One order magnitude of gating ratio is achieved within a potential window of 1.2 V for the selenophene-based molecule, which is significantly greater than that of other heterocyclic and benzene ring molecules. This is caused by the different electronic structures of heterocyclic molecules and transmission coefficients T(E), and preliminary resonance tunneling is achieved through the highest occupied molecular orbital at high potential. The current work experimentally shows that electrochemical gating performance can be significantly modulated by the alignment of the conducting orbital of the heterocyclic molecule relative to the metal Fermi energy.

OncoPDSS: an evidence-based clinical decision support system for oncology pharmacotherapy at the individual level.

BACKGROUND: Precision oncology pharmacotherapy relies on precise patient-specific alterations that impact drug responses. Due to rapid advances in clinical tumor sequencing, an urgent need exists for a clinical support tool that automatically interprets sequencing results based on a structured knowledge base of alteration events associated with clinical implications. RESULTS: Here, we introduced the Oncology Pharmacotherapy Decision Support System (OncoPDSS), a web server that systematically annotates the effects of alterations on drug responses. The platform integrates actionable evidence from several well-known resources, distills drug indications from anti-cancer drug labels, and extracts cancer clinical trial data from the ClinicalTrials.gov database. A therapy-centric classification strategy was used to identify potentially effective and non-effective pharmacotherapies from user-uploaded alterations of multi-omics based on integrative evidence. For each potentially effective therapy, clinical trials with faculty information were listed to help patients and their health care providers find the most suitable one. CONCLUSIONS: OncoPDSS can serve as both an integrative knowledge base on cancer precision medicine, as well as a clinical decision support system for cancer researchers and clinical oncologists. It receives multi-omics alterations as input and interprets them into pharmacotherapy-centered information, thus helping clinicians to make clinical pharmacotherapy decisions. The OncoPDSS web server is freely accessible at https://oncopdss.capitalbiobigdata.com .

Experimental Study on the Effect of a Weifufang on Human Gastric Adenocarcinoma Cell Line BGC-823 Xenografts and PTEN Gene Expression in Nude Mice.

Background: This study aims at investigating the effect of the Weifufang, an effective prescription for the treatment of gastric cancer developed by the Traditional Chinese Medicine (TCM)/Combination of TCM and Western Medicine Department of the Hunan Cancer Hospital, on gastric cancer xenografts in nude mice and its effect on the PTEN gene; it also aims at exploring the possible tumor suppression mechanism. Methods: Nude mice with xenografts were treated with different concentrations of the Weifufang for 2 weeks, and changes in tumor volume were observed. The histopathology of the tumor was detected by hematoxylin and eosin staining; PTEN gene expression in tumor tissues was detected by immunohistochemistry (IHC) and western blot. Results: After 2 weeks of treatment, tumor inhibition rates in the 5-flourouracil (5-FU) group, and in the Weifufang low-, middle-, and high-dose groups were 30.67%, 19%, 49.52%, and 29.36%, respectively. The IOD of the PTEN gene was detected by IHC. The values in the water group, the 5-FU group, and the Weifufang low-, middle-, and high-dose groups were 0.013 ± 0.004, 0.085 ± 0.062, 0.041 ± 0.024, 0.128 ± 0.032, and 0.061 ± 0.052, respectively. Except for the 5-FU group, the differences between the gastric compound middle dose-group and the other groups were statistically significant (p < 0.05). Results of PTEN expression detection by western blot: The expression levels in the water group, 5-FU group, and the Weifufang low-, middle-, and high-dose groups were 0.2240 ± 0.0172, 0.4200 ± 0.0228, 0.2760 ± 0.0163, 0.3840 ± 0.0133, and 0.3040 ± 0.0211, respectively. Except for the 5-FU group, differences between the Weifufang middle-dose group and the other groups were statistically significant (p < 0.05). Conclusion: The Weifufang may inhibit the growth of gastric cancer xenografts by upregulating PTEN gene expression. The middle-dose group had the best effect.

[Characteristics of humidity and temperature variations and CO2 exchange of mobile dunes at different space-time scales in Horqin sandy land, China].

Terrestrial carbon cycle plays a key role in driving climate change and ecosystem carbon balance. Understanding the variations of humidity and temperature and CO2 exchanges are meaningful to reveal the law and mechanism of regional carbon cycles in deserts. We examined the near surface humidity, temperature variations, and CO2 exchanges by eddy covariance and Bowen ratio systems in a typical mobile dune of Horqin sandy land. We analyzed the relationships between water-heat and CO2 exchanges of 0 to 10 m vertical height at daily and seasonal scales were analyzed. The results showed that the vertical variations of near surface temperature ranged from 0.4 ℃ to 2 ℃ and decreased with the increases of height from April to September, but with an opposite pattern in other months. The seasonal variation of air relative humidity was greater than 40%. During the growing season of 2018, the averaged daily net ecosystem carbon exchange (NEE) was -0.02 mg·m-2·s-1. The annual averaged daily NEE was 0.003 mg·m-2·s-1, indicating that the mobile dunes were carbon sources at the whole year scale. The vertical differences of temperature and humidity well fitted the NEE. The inflexion points of the fitting curve were at 10% humidity and 0.5 ℃ temperature, respectively. At the scalem of the year, the NEE fitting result of temperature was better than that of humidity, with the inflexion points at 17 ℃ and 65% humidity, respectively. In the growing season, the near surface vertical temperature difference was negative, which would inhibit CO2 absorption of mobile dunes. The circumstances of high humidity would promote the absorption of atmospheric CO2. Across different time and vertical height, the variations of humidity and temperature were closely related to CO2 exchanges, which affected carbon sink and source of mobile dunes. Carbon budget was more sensitive to temperature than humidity.

[Deep water leakage from semi-mobile dunes in semi-arid regions and its response to rainfall patterns]. / åŠå¹²æ—±åœ°åŒºåŠæµåŠ¨æ²™ä¸˜æ°´åˆ†æ·±å±‚æ¸—æ¼é‡åŠå ¶å¯¹é™é›¨æ ¼å±€çš„å“åº”.

As the main source of soil moisture supply in desertified areas, rainfall has a profound impact on soil moisture changes and plays an important role in deep soil moisture replenishment. Based on the Hydrus-1D model with optimized parameters, we analyzed the dynamic change process of the leakage in the 200 cm deep layer of the semi-mobile dunes in Horqin Sandy Land and its response to the rainfall patterns. The results showed that the averaged leakage replenishment of semi-mobile dunes was 254.31 mm from April to October each year during 2016 to 2019, accoun-ting for 61.8% of the rainfall in the same period. Deep leakage mainly occurred from June to August, accounting for 72.8% of the total. The leakage rate was distributed between 0.03-2.70 mm·h-1, with the maximum leakage rate occurring under heavy rainfall and frequent rainfall events. The deep soil water supplied by rainfall infiltration was affected by the amount of rainfall, rainfall intensity, duration of precipitation and soil moisture content in the earlier period. Precipitation events with long duration and small rainfall intensity were more conducive to deep water lea-kage, with a significant positive correlation between the leakage and rainfall (R2=0.85). 16-18 mm rainfall was the threshold for the leakage of 200 cm soil depth. The high-frequency rainfall event usually reached peak after 17-38 hours, with the entire leakage process being more than 164 hours. Accurate estimation of deep leakage has theoretical and practical significance for water resource assessment and ecological construction in desertified areas.

Genotyping and Zoonotic Potential of Enterocytozoon bieneusi in Pigs in Xinjiang, China.

Enterocytozoon bieneusi is an obligate intracellular fungus, infecting various invertebrate and vertebrate hosts, it is common in humans and causes diarrhea in the immunocompromised. In the present study, 801 fecal specimens were collected from pigs on seven large-scale pig farms in Xinjiang, China. Nested polymerase chain reaction (PCR) amplification of the internal transcribed spacer (ITS) gene showed that the overall E. bieneusi infection rate was 48.6% (389/801). The E. bieneusi infection rates differed significantly among the collection sites (20.0-73.0%) (χ2 = 75.720, df = 6, p < 0.01). Post-weaned pigs had the highest infection rate (77.2%, 217/281), followed by fattening pigs (67.4%, 87/129) and pre-weaned suckling pigs (35.5%, 60/169). Adult pigs had the lowest infection rate (11.3%, 25/222). The E. bieneusi infection rates also differed significantly among age groups (χ2 = 246.015, df = 3, p < 0.01). Fifteen genotypes were identified, including 13 known genotypes (CHC, CS-1, CS-4, CS-7, CS-9, D, EbpA, EbpC, EbpD, H, PigEb4, PigEBITS5, and WildBoar8) and two novel genotypes (XJP-II and XJP-III). Among them, six genotypes (CS-4, D, EbpA, EbpC, H, and PigEBITS5) have been reported in humans. Phylogenetic analysis showed that all the genotypes belonged to Group 1 of E. bieneusi. These findings suggest that pigs may play an important role in transmitting E. bieneusi infections to humans.

Mechanism of Placenta Damage in Gestational Diabetes Mellitus by Investigating TXNIP of Patient Samples and Gene Functional Research in Cell Line.

INTRODUCTION: Gestational diabetes mellitus (GDM) is a gestational complication that affects maternal and child health. The placenta provides the fetus with the necessary nutrition and oxygen and takes away the metabolic waste. Patients with GDM are diagnosed and treated merely on the basis of the blood glucose level; this approach does nothing to help evaluate the status of the placenta, which is worth noting in GDM. The purpose of this research was to clarify the relation between thioredoxin-interacting protein (TXNIP) and reactive oxygen species (ROS) in the placenta of patients with GDM, which has thus far remained unclear. METHODS: The expression of TXNIP in the placentas of 10 patients with GDM and 10 healthy puerperae (control group) was investigated via immunofluorescence. The relation among TXNIP, ROS, and the function of mitochondria was explored in HTR-8/SVneo cells stimulated by high glucose (HG). RESULTS: The results showed the expression of TXNIP in the placentas of patients with GDM was higher than that in the control group, and the expression of TXNIP in HTR-8/SVneo cells treated with HG was higher than that in the control group, causing the accumulation of ROS and changes of mitochondria, promoting apoptosis and inhibition of migration. CONCLUSIONS: High expression of TXNIP caused by HG mediates the increasing ROS and the mitochondria dysfunction in GDM; this impairs the function of the placenta and is the basis for the prediction of perinatal outcome.

Prevalence and Population Genetics Analysis of Enterocytozoon bieneusi in Dairy Cattle in China.

Enterocytozoon bieneusi, an obligate intracellular pathogen, can infect various hosts. In this study, 3527 dairy cattle fecal specimens were collected from different geographic locations in China (including 673 from Shandong province, 1,440 from Guangdong province and 1,414 from Gansu province) and examined for the presence of E. bieneusi using polymerase chain reactions targeting the ribosomal internal transcribed spacer (ITS). The dominant genotypes identified were further subtyped by multilocus sequence typing. The overall prevalence of E. bieneusi was 14.2% (501/3527), with a significant difference in prevalence among the different geographical locations (P < 0.001). Our logistic regression analysis showed that all four variables (farming model, location, age, and clinical manifestations) had strong effects on the risk of contracting E. bieneusi. Sequence analysis revealed 11 genotypes: eight known genotypes (J, I, BEB4, BEB10, D, EbpC, CM19, and CM21) and three novel genotypes (named here as CGC1, CGC2, and CGC3). Genotypes J and I, the commonest, were found on all farms across the three provinces. Our linkage disequilibrium analysis showed a clonal population structure in the E. bieneusi dairy cattle population but the ITS genotypes had different population structures. Phylogenetic and haplotype network analysis showed the absence of geographical segregation in the E. bieneusi dairy cattle populations. Instead, they revealed the presence of host adaptation to the E. bieneusi populations in various animals. Our findings augment the current understanding of E. bieneusi transmission dynamics.

The novel GLP-1/GIP dual receptor agonist DA3-CH is neuroprotective in the pilocarpine-induced epileptogenesis rat model.

AIMS: Glia-mediated neuro-inflammation and oxidative stress-induced neuronal apoptosis can contribute to epileptogenesis. We have demonstrated previously that mimetics of glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP) and dual-GLP-1/GIP receptor agonists protect the brain from inflammation, oxidative stress, apoptosis and neuronal loss in animal models of central nervous system diseases. METHODS: This study investigated for the first time whether the novel dual GLP-1/GIP receptor agonist DA3-CH has neuroprotective effects in the pilocarpine-induced status epilepticus (SE) rat model and the studies the underlying mechanisms. DA3-CH was administered once daily at 10 nmol/kg ip. following SE induction. The effect of DA3-CH was evaluated by immunohistochemistry and western blot at 12 h, 1 d, 3 d, 7 d after kindling. RESULTS: Our findings show that DA3-CH reduced the chronic inflammation response (astrogliosis and microgliosis), and the associated release of the pro-inflammatory cytokines interleukin-1ß (IL-ß) and tumor necrosis factor-α (TNF-α) in the hippocampal CA1 area. Furthermore, DA3-CH reduced the expression of the mitochondrial pro-apoptotic protein Bax, while increasing the expression of the anti-apoptotic protein Bcl-2. Neuronal numbers in the CA1 area were much reduced by pilocarpine treatment, and DA3-CH protected neurons from neurotoxicity. CONCLUSION: These results demonstrated that DA3-CH could mitigate pilocarpine-induced neuro-inflammation, mitochondrial apoptosis and neuronal loss. The findings encourage the development of dual agonists as novel therapeutic interventions for epilepsy.

Post-treatment with the GLP-1 analogue liraglutide alleviate chronic inflammation and mitochondrial stress induced by Status epilepticus.

Glucagon-like peptide-1(GLP-1) is a growth factor that has neuroprotective and anti-inflammatory properties. The protease resistant GLP-1 analogue liraglutide has been shown to be neuroprotective in previous studies in animal models of Alzheimer's disease or Parkinson's disease. Status epilepticus (SE) is a complex disorder, involving many underlying pathological processes, including excitotoxic and chronic inflammatory events. The present pilot study aims to investigate whether liraglutide alleviates the chronic inflammation response and mitochondrial stress induced by SE in the lithium-pilocarpine animal model. We found that treatment with 25nmol/kg. i.p. once-daily after the induction of SE for 7 days reduced chronic inflammation as shown by reduced numbers of activated microglia and astrocytes, and reduced levels of TNF-α and IL-1ß in the hippocampus. The mitochondrial stress marker BAX was reduced and the survival factor Bcl-2 was enhanced by liraglutide. Blood glucose levels were not affected by liraglutide. We show for the first time that liraglutide can reduce the chronic inflammation and mitochondrial stress induced by SE, and the results suggest that GLP-1 receptor agonists such as liraglutide have restorative and protective effects in the brain after SE and could serve as a potential treatment.

Atrial septal defect in a patient with congenital disorder of glycosylation type 1a: a case report.

BACKGROUND: Atrial septal defect often become more severe when encountered in genetic syndromes. Congenital disorder of glycosylation type 1a is an inherited metabolic disorder associated with mutations in PMM2 gene and can affect almost all organs. Cardiac abnormalities vary greatly in congenital disorder of glycosylation type 1a and congenital heart defects have already been reported, but there is little knowledge about the effect of this inherited disorder on an existing congenital heart defect. Herein we report for the first time on a baby with congenital disorder of glycosylation type 1a with atrial septal defect and make a comparison of changes in atrial septal defect by follow-ups to the age of 3. CASE PRESENTATION: Our patient was an 8-month-old Han Chinese boy. At the initial visit, he presented with recurrent lower respiratory infection, heart murmur, psychomotor retardation, inverted nipples, and cerebellar atrophy. Echocardiography revealed a 8 mm secundum atrial septal defect with left-to-right shunt (Qp/Qs ratio 1.6). Enzyme testing of phosphomannomutase 2 demonstrated decreased levels of phosphomannomutase 2 activities in fibroblasts. Whole exon sequencing showed he was heterozygous for a frameshift mutation (p.I153X) and a missense mutation (p.I132T) in PMM2 gene. The diagnosis of congenital disorder of glycosylation type 1a with atrial septal defect was issued. Now, he is 3-years old at the time of this writing, with the development of congenital disorder of glycosylation type 1a (cerebellar atrophy become more severe and the symptom of nystagmus emerged), the size of atrial septal defect increased to 10 mm and the Qp/Qs ratio increased to 1.9, which suggested exacerbation of the atrial septal defect. Congenital heart defect-associated gene sequencing is then performed and shows there are no pathogenic mutations, which suggested intrinsic cardiac factors are not the cause of exacerbation of the atrial septal defect in our patient and it is reasonable to assume congenital disorder of glycosylation type 1a can worsen the situation of the existing atrial septal defect. CONCLUSIONS: This report highlights the view that congenital disorders of glycosylation type 1a should be excluded when faced with congenital heart defect with cerebellar atrophy or neurodevelopmental delay, especially when the situation of congenital heart defect becomes more and more severe.

[MECP2 duplication syndrome: a clinical analysis of three cases and literature review].

MECP2 duplication syndrome (MDS) is a rare pediatric disease and mainly manifests as delayed motor development, language loss or delay, recurrent infection, severe intellectual disability, epilepsy, autistic symptoms, and early infantile hypotonia. In this article, the three children with this disease were all boys. Cases 1 and 2 had delayed motor development, and language loss or delay as initial manifestations, and case 3 had recurrent infection as initial manifestation. Physical examination showed hypotonia and negative pathological signs in each case. Case 1 had tonic-clonic seizures and electroencephalography showed focal seizures, for which he was given oxcarbazepine, levetiracetam, and clonazepam as the antiepileptic treatment to control seizures. Case 3 experienced one absence seizure and three head-nodding seizures with normal electroencephalographic findings during these seizures, and therefore, he was not given antiepileptic treatment. In each case, recurrent infection was improved with the increase in age, but there were no significant improvements in language or intelligence. Array-based comparative genomic hybridization (aCGH) showed MECP2 duplication in X chromosome in each case, and so they were diagnosed with MDS. MDS should be considered for children with delayed development complicated by recurrent infection and epileptic seizures, and early aCGH helps with the diagnosis of this disease.