OBJECTIVE: To investigate the clinical features and genetic variants associated with Multiple mitochondrial dysfunction syndrome (MMDS) type 3 in two children. METHODS: Two children diagnosed with MMDS type 3 at Zhuhai Maternal and Child Health Care Hospital in January 2021 were selected for this study. A retrospective analysis of their clinical data was carried out. Whole exome sequencing was conducted on the two children and their parents, followed by Sanger sequencing for candidate variants and bioinformatic analysis. Both children received comprehensive rehabilitative therapy and were followed up for 3 years. This study was approved by the Ethics Committee of Zhuhai Maternal and Child Health Hospital (Ethics No. 202380). RESULTS: The two MMDS type 3 children were monozygotic twin girls, aged 9 months, presenting with developmental regression, pyramidal signs, and other clinical manifestations. Cranial MRI revealed widespread abnormal signals and vacuolar changes in the white matter. Whole exome sequencing revealed that both children had harbored compound heterozygous variants of the IBA57 gene, namely c.286T>C (p.Tyr96His) and c.307C>T (p.Gln103Ter). Sanger sequencing confirmed that these variants were inherited from their father and mother, respectively. According to the American College of Medical Genetics and Genomics (ACMG) guidelines, both variants were classified as pathogenic (PM2_Supporting+PM3_Very Strong+PP3_Moderate; PVS1+PM2_Supporting+PM3). After treatment with vitamins, levocarnitine, ATP, coenzyme Q10, and other drugs, both children showed partial recovery of neurodevelopmental regression, with improvement in feeding and sleep. Over the 3-year follow-up, there was slow but progressive improvement in motor, language, and cognitive development. CONCLUSION The compound heterozygous variants c.286T>C (p.Tyr96His) and c.307C>T (p.Gln103Ter) of the IBA57 gene probably underlay the MMDS type 3 in the twin pair. Clinicians should be vigilant about the possibility of MMDS type 3 in children with neurodevelopmental regression and early cranial MRI findings indicating widespread white matter abnormalities with vacuolar changes, as these may be indicative of IBA57 gene variants.
Female ; Humans ; Infant ; Calcium-Binding Proteins/genetics* ; Exome Sequencing ; Genetic Testing/methods* ; Microfilament Proteins/genetics* ; Mitochondrial Diseases/genetics* ; Mutation ; Retrospective Studies ; Carrier Proteins

Objective:To investigate the clinical features and genetic variants associated with Multiple mitochondrial dysfunction syndrome (MMDS) type 3 in two children.Methods:Two children diagnosed with MMDS type 3 at Zhuhai Maternal and Child Health Care Hospital in January 2021 were selected for this study. A retrospective analysis of their clinical data was carried out. Whole exome sequencing was conducted on the two children and their parents, followed by Sanger sequencing for candidate variants and bioinformatic analysis. Both children received comprehensive rehabilitative therapy and were followed up for 3 years. This study was approved by the Ethics Committee of Zhuhai Maternal and Child Health Hospital (Ethics No. 202380).Results:① The two MMDS type 3 children were monozygotic twin girls, aged 9 months, presenting with developmental regression, pyramidal signs, and other clinical manifestations. Cranial MRI revealed widespread abnormal signals and vacuolar changes in the white matter. ② Whole exome sequencing revealed that both children had harbored compound heterozygous variants of the IBA57 gene, namely c. 286T>C (p.Tyr96His) and c. 307C>T (p.Gln103Ter). Sanger sequencing confirmed that these variants were inherited from their father and mother, respectively. ③ According to the American College of Medical Genetics and Genomics (ACMG) guidelines, both variants were classified as pathogenic (PM2_Supporting + PM3_Very Strong + PP3_Moderate; PVS1 + PM2_Supporting + PM3). ④ After treatment with vitamins, levocarnitine, ATP, coenzyme Q10, and other drugs, both children showed partial recovery of neurodevelopmental regression, with improvement in feeding and sleep. Over the 3-year follow-up, there was slow but progressive improvement in motor, language, and cognitive development. Conclusion:The compound heterozygous variants c. 286T>C (p.Tyr96His) and c. 307C>T (p.Gln103Ter) of the IBA57 gene probably underlay the MMDS type 3 in the twin pair. Clinicians should be vigilant about the possibility of MMDS type 3 in children with neurodevelopmental regression and early cranial MRI findings indicating widespread white matter abnormalities with vacuolar changes, as these may be indicative of IBA57 gene variants.

Objective:To investigate the clinical features and genetic variants associated with Multiple mitochondrial dysfunction syndrome (MMDS) type 3 in two children.Methods:Two children diagnosed with MMDS type 3 at Zhuhai Maternal and Child Health Care Hospital in January 2021 were selected for this study. A retrospective analysis of their clinical data was carried out. Whole exome sequencing was conducted on the two children and their parents, followed by Sanger sequencing for candidate variants and bioinformatic analysis. Both children received comprehensive rehabilitative therapy and were followed up for 3 years. This study was approved by the Ethics Committee of Zhuhai Maternal and Child Health Hospital (Ethics No. 202380).Results:① The two MMDS type 3 children were monozygotic twin girls, aged 9 months, presenting with developmental regression, pyramidal signs, and other clinical manifestations. Cranial MRI revealed widespread abnormal signals and vacuolar changes in the white matter. ② Whole exome sequencing revealed that both children had harbored compound heterozygous variants of the IBA57 gene, namely c. 286T>C (p.Tyr96His) and c. 307C>T (p.Gln103Ter). Sanger sequencing confirmed that these variants were inherited from their father and mother, respectively. ③ According to the American College of Medical Genetics and Genomics (ACMG) guidelines, both variants were classified as pathogenic (PM2_Supporting + PM3_Very Strong + PP3_Moderate; PVS1 + PM2_Supporting + PM3). ④ After treatment with vitamins, levocarnitine, ATP, coenzyme Q10, and other drugs, both children showed partial recovery of neurodevelopmental regression, with improvement in feeding and sleep. Over the 3-year follow-up, there was slow but progressive improvement in motor, language, and cognitive development. Conclusion:The compound heterozygous variants c. 286T>C (p.Tyr96His) and c. 307C>T (p.Gln103Ter) of the IBA57 gene probably underlay the MMDS type 3 in the twin pair. Clinicians should be vigilant about the possibility of MMDS type 3 in children with neurodevelopmental regression and early cranial MRI findings indicating widespread white matter abnormalities with vacuolar changes, as these may be indicative of IBA57 gene variants.

The E protein of dengue virus type Ⅱ was presented on ferritin of Helicobacter pylori to construct a novel dengue nanoparticle vaccine candidate,and the immunological indexes of the vac-cine were evaluated,aiming to provide new ideas for the development of dengue vaccine.The re-combinant plasmid of E-Ferritin was optimized and synthesized,and then transfected into HEK-293F cells.The recombinant protein was expressed,identified,purified and analyzed.Mice were im-munized with E-Ferritin nanoparticle vaccine by intramuscular injection on the hind limbs on the day 0,14 and 28.ELISA,neutralization test,flow cytometry and lymphocyte proliferation test were used to detect the levels of specific antibodies,neutralizing antibodies,CD3+,CD4+and CD8+T lymphocytes in spleen cells and the proliferation of spleen lymphocytes after specific stimulation.The target protein with a size of about 69 kDa was expressed in the cells with a single band.The purified protein concentration was 0.407 g/L,and the purity was 82.32％.The results from transmission electron microscopy showed that E-Ferritin protein could be recombined into a particle structure with a particle size of about 50 nm.The results of mouse immune experiments showed that E-Ferritin protein had good immunogenicity.The average specific antibody titer of E-ferritin protein in serum was 1∶92 160 after immunization 42 d.The main subclass of antibody was IgGl.The results of flow cytometry showed that E-Ferritin as an immunogen could induce higher levels of CD4+and CD8+T lymphocyte immune response.In lymphocyte proliferation test,the level of specific stimulation in the vaccine group was significantly higher than that in the non-specific stimulation group.In conclusion,the dengue virus envelope protein ferritin nanoparticle vaccine constructed in this study has good immunogenicity,which can provide reference for the de-velopment of new dengue vaccine candidates.

The SYBR Green Ⅰ real-time fluorescence quantitative PCR detection method was used to determine the prevalence of Japanese encephalitis virus(JEV)in mosquito vectors and cattle se-rum samples in Xinjiang.The E gene fragment of the JEV strain was amplified by PCR,cloned into a pEASY-Blunt vector,produced as a recombinant plasmid,and its sensitivity,specificity and re-producibility were verified.Between 2021 and 2022,serum samples were taken in the regions of Hami,Altay,Ili,Aksu,and Kashi in order to monitor the prevalence of JEV in livestock in Xin-jiang.The positive rate was discovered and evaluated using the established detection method.The established detection method showed a good linear relationship,and the detection interval was 4.03X102-4.03×109 copies/pL.The correlation coefficient was 0.995,the slope was-3.431,and the extreme value of the lower limit of sensitivity was 4.03 × 102 copies/pL.This method has no specific amplification for Zika virus(ZIKV)and Dengue virus(DENV).The intra group coefficient of variation of reproducibility was 0.53％-1.27％,and the inter group coefficient of variation was 0.48％-1.43％.Using this method to detect serum samples from livestock in Xinjiang from 2021 to 2022,the total positive rate was 3.28％,with positive detection rates in horses,cows,and sheep being 2.35％,6.77％,and 3.74％respectively,the virus was identified as Type Ⅰ JEV.A SYBR Green Ⅰ real-time PCR method for the detection of genotype 1 JEV was established.JEV was de-tected in the serum of horses,cattle and sheep in Xinjiang,with a total positive rate of 3.11％.

Bluetongue virus(BTV)is classified as a category Ⅱ animal epidemic disease in China,infecting multiple species and posing significant threats to the ruminant breeding industry.There are 29 serotypes of BTV,with BTV16 being one of the major serotypes currently prevalent in Chi-na.Bluetongue virus infection mainly manifests as a latent infection,making the establishment of ELISA assays crucial for epidemiological detection.In this study;the expression of the BTV16 VP2 protein was achieved using a prokaryotic expression system,and polyclonal antibodies were pre-pared using BALB/c mice.An indirect ELISA assay using VP2 protein as the encapsulated antigen was established and optimized.Clinical samples from the Guangxi Zhuang Autonomous Region were tested and analyzed for compliance with commercial kits.The results showed that the BTV16 VP2 protein was successfully expressed and purified,and the prepared polyclonal antibody exhibi-ted good immunogenicity.The ELISA assay had good specificity,with no cross-reactivity against ruminant diseases such as AKAV,FMDV and GETV.The critical values for negativity and positivity were determined to be 0.314,and the coefficients of variation(Cv)between batches and within batches were both less than 5％,indicating good reproducibility.The ELISA assay revealed a positive rate of 92.4％for 79 samples from the Guangxi Zhuang Autonomous Region,with a compliance rate of 98.7％when compared to the commercialized kit.In conclusion,this study suc-cessfully established an indirect ELISA method for BTV16,facilitating the detection of bovine clin-ical samples.

Bluetongue virus(BTV)is classified as a category Ⅱ animal epidemic disease in China,infecting multiple species and posing significant threats to the ruminant breeding industry.There are 29 serotypes of BTV,with BTV16 being one of the major serotypes currently prevalent in Chi-na.Bluetongue virus infection mainly manifests as a latent infection,making the establishment of ELISA assays crucial for epidemiological detection.In this study;the expression of the BTV16 VP2 protein was achieved using a prokaryotic expression system,and polyclonal antibodies were pre-pared using BALB/c mice.An indirect ELISA assay using VP2 protein as the encapsulated antigen was established and optimized.Clinical samples from the Guangxi Zhuang Autonomous Region were tested and analyzed for compliance with commercial kits.The results showed that the BTV16 VP2 protein was successfully expressed and purified,and the prepared polyclonal antibody exhibi-ted good immunogenicity.The ELISA assay had good specificity,with no cross-reactivity against ruminant diseases such as AKAV,FMDV and GETV.The critical values for negativity and positivity were determined to be 0.314,and the coefficients of variation(Cv)between batches and within batches were both less than 5％,indicating good reproducibility.The ELISA assay revealed a positive rate of 92.4％for 79 samples from the Guangxi Zhuang Autonomous Region,with a compliance rate of 98.7％when compared to the commercialized kit.In conclusion,this study suc-cessfully established an indirect ELISA method for BTV16,facilitating the detection of bovine clin-ical samples.

Target identification of bioactive compounds is important for understanding their mechanisms of action and provides critical insights into their therapeutic utility. While it remains a challenge, unbiased chemoproteomics strategy using clickable photoaffinity probes is a useful and validated approach for target identification. One major limitation of this approach is the efficient synthesis of appropriately substituted clickable photoaffinity probes. Herein, we describe an efficient and consistent method to prepare such probes. We further employed this method to prepare a highly stereo-congested probe based on naturally occurring triterpenoid betulinic acid. With this photoaffinity probe, we identified tropomyosin as a novel target for betulinic acid that can account for the unique biological phenotype on cellular cytoskeleton induced by betulinic acid.

Aggregatibacter aphrophilus is a member of the normal flora of the human oral cavity and pharynx, a Gram-negative fastidous bacteria, belonging to agglomerates, which is a normal mixed oropharyngeal flora in humans, most commonly colonized on the surface of oral mucosa. This bacterial infection is rare in clinical practice, and it is difficult to culture and identify the bacteria, which is easy to be missed. A patient with intracranial infection was admitted to Huaihe Hospital, who showed fever and headache as the main clinical manifestations, and Aggregatibacter aphrophilus was detected by the metagenomic next-generation sequencing of cerebrospinal fluid. The patient′s symptoms were significantly improved after anti-infection treatment.

OBJECTIVE: To discuss the advantages and technical limitations of various molecular genetic techniques in the diagnosis of two infants featuring all-round developmental retardation. METHODS: The two patients were initially screened by using chromosomal microarray analysis (CMA). For patient 1, his parents were also subjected to CMA analysis, and the data was analyzed by using ChAS and UPD-tool software. For patient 2, methylation-specific PCR (MS-PCR) was carried out. RESULTS: Patient 1 was diagnosed with maternal uniparental disomy (UPD) type Prader-Willi syndrome (PWS) by CMA and UPD-tool family analysis. His chromosomes 15 were of maternal UPD with homology/heterology. Patient 2 was diagnosed with deletion type PWS by combined CMA and MS-PCR. CONCLUSION Correct selection of laboratory methods based on the advantages and limitations of various molecular techniques can help with diagnosis of genomic imprinting disorders and enable better treatment and prognosis through early intervention.