Selection markers are essential tools in gene editing, the utility of such systems is inherently constrained by species-specific limitations, governed by divergent host genetic backgrounds and metabolic compatibility. To address this limitation, we leveraged the CRISPR/Cas9 system to develop a universal counter-selection tool. We designed and introduced an sgRNA expression cassettes as counter-selection markers, which directs the Cas9 protein to target and cleave genomic DNA, allowing for the selection of the strains where the sgRNA expression cassette has been replaced. Optimized to target multiple copy sites with sgRNA, this system significantly enhances cell lethality, boosting counter-selection efficiency to over 85.00%. This counter-selection tool is not limited to single strains and is suitable for various scenarios, including multi-copy plasmid assembly and plasmid editing, demonstrating broad application potential.
CRISPR-Cas Systems/genetics* ; Gene Editing/methods* ; RNA, Guide, CRISPR-Cas Systems/genetics* ; Plasmids/genetics*

Objective:To construct a VP8-mRNA vaccine using human rotavirus spike protein VP8 domain as the immunogen and analyze its immunogenicity in mice.Methods:The VP8-mRNA sequence was designed, optimized, and synthesized. The VP8 gene of rotavirus G1P[8] type was used to construct the plasmid pUC57-VP8-Kan-SapⅠ, which was then sequenced. The plasmid confirmed by sequencing was subjected to large-scale amplification and extraction, followed by linearization, in vitro transcription, and capping. The purified capped products were encapsulated with lipid nanoparticles using a microfluidic control apparatus. The encapsulated VP8-mRNA vaccine was administered intramuscularly to mice at 10, 15, and 20 μg. Serum samples were collected for antibody detection by ELISA. Cellular immune responses were detected by flow cytometry and ELISPOT. Statistical analysis was performed using one-way or two-way analysis of variance and Tukey-Kramer test. Results:The encapsulated VP8-mRNA vaccine was rounded and spherical, with a particle size of about 100 nm, a polymer dispersion index of 0.088, and an encapsulation rate of 92.3%. Two doses of VP8-mRNA vaccine immunization could induce a good immune response in mice. The level of IgG antibody induced after immunization in the 15 μg group was comparable to that of the 20 μg group, and there was no statistical difference ( P>0.05), but the antibody levels in the two groups were significantly higher than that in the 10 μg group ( P<0.000 1). VP8-mRNA vaccine could induce neutralizing antibodies against rotavirus G1 and G9 types. The highest level of neutralizing antibodies against rotavirus type G1 was observed in the 15 μg group, which was significantly higher than that in the 10 μg group ( P<0.05). All immunization groups exhibited good neutralizing ability against rotavirus G9 type. The results of ELISPOT showed that lymphocytes from mice in each vaccine group were able to secrete IFN-γ when stimulated with VP8 peptide. Flow cytometry showed that the proportions of CD8 + T cell subsets in the vaccine groups were higher than that in the control group. Conclusion:The VP8-mRNA vaccine has good immunogenicity in mice and can induce good humoral and T-cell immune responses.

This report summarizes the nursing experience of a patient with acute myocardial infarction complicated by cardiogenic shock and frequent electrical storms.The key nursing interventions are outlined as follows.Formation of a multidisciplinary management team:a collaborative treatment plan was developed by assembling a multidisciplinary team of healthcare professionals.Activation of the hospital emergency response system:the hospital's emergency response protocols were promptly activated to ensure efficient emergency nursing care.Dynamic hemodynamic monitoring:circulatory support strategy guided by hemodynamic monitoring.Establish-ment of a malignant arrhythmia warning system:a warning system for malignant arrhythmias was implemented to standardize the emergency procedures for electrical cardioversion and defibrillation,thereby maximizing the time available for life support.Sequential antiplatelet and anticoagulation management:a sequential approach to antiplatelet and anticoagulation therapy was employed to effectively prevent the risks of thrombosis and bleeding.Focus on psychological care and rehabilitation training:emphasis was placed on psychological support and rehabilitation exercises to enhance the patient's comfort and exercise tolerance.The patient was hospitalized for 33 days and was discharged in improved condition.This translation is structured to reflect the conventions of academic writing,making it suitable for inclusion in a clinical report or research paper.

Objective:To investigate the incidence and influencing factors of optic disc changes in children and adolescents with high myopia.Methods:A clinical cross-sectional study. A total of 162 children and adolescents with high myopia (162 eyes) who visited Department of Ophthalmology, Northern Jiangsu People's Hospital Affiliated to Yangzhou University from January to April 2025 were included in this study. Myopia refractive error ≥6.00 D and/or axial length (AL) ≥26 mm. All participants underwent best-corrected visual acuity, refraction, fundus color photography, swept-source optical coherence tomography (SS-OCT), and AL measurement. Subfoveal choroidal thickness (ChT) was measured within 1 mm using SS-OCT. Optic disc changes assessed included tilt, rotation, peripapillary atrophy (PPA), and peripapillary hyperreflective ovoid mass-like structures (PHOMS). The patients were divided into the children group (4-11 years old) and the adolescents group (12-18 years old) based on age, with 63 (38.9%, 63/162) and 99 (61.1%, 99/162) cases respectively. The incidence of ocular features and optic disc morphology changes in the two groups was compared and observed. According to the myopia diopter, the patients were divided into the high diopter long axial group (myopia diopter ≥6.00 D, AL≥26 mm) and the low diopter long axial group (myopia diopter <6.00 D, AL≥26 mm), with 85 (52.5%, 85/162) and 77 (47.5%, 77/162) eyes respectively. The incidence of optic disc morphological changes in the two groups was compared and observed. The comparison of quantitative data between groups was conducted using the Mann-Whitney U test. Multivariate logistic regression was used to analyze the correlations between PPA, optic disc tilt, PHOMS occurrence and gender, age, diopter, AL, and ChT. Results:Among the 162 patients, 103 were male and 59 were female. Age was 12 (10.5, 13.5) years old. Among the 162 eyes, the optic disc morphology changed in 152 eyes (93.8%, 152/162). Among them, the PPA, optic disc tilt, PHOMS, and optic disc rotation were 148 (91.4%, 148/162), 95 (58.6%, 95/162), 62 (38.3%, 62/162), and 35 (21.6%, 35/162) eyes respectively. Myopic macular degeneration in 137 eyes. There were 56 eyes with peripheral retinopathy. There was no statistically significant difference in myopia diopter, AL and ChT between the children group and the adolescent group ( Z=-1.201, -1.934, ?0.761; P=0.230, 0.053, 0.447). There was no statistically significant difference in the incidences of PPA, optic disc tilt and optic disc rotation ( χ2=0.293, 2.618, 0.398; P>0.05). There was no statistically significant difference in the incidence of optic disc morphological changes between the low diopter long axial group and the high diopter long axial group ( χ2=0.000, P>0.05). The results of multivariate logistic regression analysis showed that the thinner the ChT, the higher the risk of PPA [odds ratio (OR) =0.98, 95% confidence interval ( CI) 0.97-0.99, P<0.001]. Female ( OR=2.3, 95% CI 1.04-5.07, P=0.039), older age ( OR=1.17, 95% CI 1.01-1.37, P=0.043), thinner ChT ( OR=0.99, 95% CI 0.99-1.00, P=0.012), the higher the risk of optic disc tilt. The older the age, the higher the risk of developing PHOMS ( OR=1.22, 95% CI 1.06-1.40, P=0.006). Conclusions:Optic disc morphology changes may be the most common fundus alterations in children and adolescents with high myopia. The influencing factors of optic disc morphological changes (including PPA, optic disc tilt, and PHOMS) are female sex, advanced age, and ChT thinning.

This report summarizes the nursing experience of a patient with acute myocardial infarction complicated by cardiogenic shock and frequent electrical storms.The key nursing interventions are outlined as follows.Formation of a multidisciplinary management team:a collaborative treatment plan was developed by assembling a multidisciplinary team of healthcare professionals.Activation of the hospital emergency response system:the hospital's emergency response protocols were promptly activated to ensure efficient emergency nursing care.Dynamic hemodynamic monitoring:circulatory support strategy guided by hemodynamic monitoring.Establish-ment of a malignant arrhythmia warning system:a warning system for malignant arrhythmias was implemented to standardize the emergency procedures for electrical cardioversion and defibrillation,thereby maximizing the time available for life support.Sequential antiplatelet and anticoagulation management:a sequential approach to antiplatelet and anticoagulation therapy was employed to effectively prevent the risks of thrombosis and bleeding.Focus on psychological care and rehabilitation training:emphasis was placed on psychological support and rehabilitation exercises to enhance the patient's comfort and exercise tolerance.The patient was hospitalized for 33 days and was discharged in improved condition.This translation is structured to reflect the conventions of academic writing,making it suitable for inclusion in a clinical report or research paper.

Objective:To construct a VP8-mRNA vaccine using human rotavirus spike protein VP8 domain as the immunogen and analyze its immunogenicity in mice.Methods:The VP8-mRNA sequence was designed, optimized, and synthesized. The VP8 gene of rotavirus G1P[8] type was used to construct the plasmid pUC57-VP8-Kan-SapⅠ, which was then sequenced. The plasmid confirmed by sequencing was subjected to large-scale amplification and extraction, followed by linearization, in vitro transcription, and capping. The purified capped products were encapsulated with lipid nanoparticles using a microfluidic control apparatus. The encapsulated VP8-mRNA vaccine was administered intramuscularly to mice at 10, 15, and 20 μg. Serum samples were collected for antibody detection by ELISA. Cellular immune responses were detected by flow cytometry and ELISPOT. Statistical analysis was performed using one-way or two-way analysis of variance and Tukey-Kramer test. Results:The encapsulated VP8-mRNA vaccine was rounded and spherical, with a particle size of about 100 nm, a polymer dispersion index of 0.088, and an encapsulation rate of 92.3%. Two doses of VP8-mRNA vaccine immunization could induce a good immune response in mice. The level of IgG antibody induced after immunization in the 15 μg group was comparable to that of the 20 μg group, and there was no statistical difference ( P>0.05), but the antibody levels in the two groups were significantly higher than that in the 10 μg group ( P<0.000 1). VP8-mRNA vaccine could induce neutralizing antibodies against rotavirus G1 and G9 types. The highest level of neutralizing antibodies against rotavirus type G1 was observed in the 15 μg group, which was significantly higher than that in the 10 μg group ( P<0.05). All immunization groups exhibited good neutralizing ability against rotavirus G9 type. The results of ELISPOT showed that lymphocytes from mice in each vaccine group were able to secrete IFN-γ when stimulated with VP8 peptide. Flow cytometry showed that the proportions of CD8 + T cell subsets in the vaccine groups were higher than that in the control group. Conclusion:The VP8-mRNA vaccine has good immunogenicity in mice and can induce good humoral and T-cell immune responses.

Objective:To investigate the incidence and influencing factors of optic disc changes in children and adolescents with high myopia.Methods:A clinical cross-sectional study. A total of 162 children and adolescents with high myopia (162 eyes) who visited Department of Ophthalmology, Northern Jiangsu People's Hospital Affiliated to Yangzhou University from January to April 2025 were included in this study. Myopia refractive error ≥6.00 D and/or axial length (AL) ≥26 mm. All participants underwent best-corrected visual acuity, refraction, fundus color photography, swept-source optical coherence tomography (SS-OCT), and AL measurement. Subfoveal choroidal thickness (ChT) was measured within 1 mm using SS-OCT. Optic disc changes assessed included tilt, rotation, peripapillary atrophy (PPA), and peripapillary hyperreflective ovoid mass-like structures (PHOMS). The patients were divided into the children group (4-11 years old) and the adolescents group (12-18 years old) based on age, with 63 (38.9%, 63/162) and 99 (61.1%, 99/162) cases respectively. The incidence of ocular features and optic disc morphology changes in the two groups was compared and observed. According to the myopia diopter, the patients were divided into the high diopter long axial group (myopia diopter ≥6.00 D, AL≥26 mm) and the low diopter long axial group (myopia diopter <6.00 D, AL≥26 mm), with 85 (52.5%, 85/162) and 77 (47.5%, 77/162) eyes respectively. The incidence of optic disc morphological changes in the two groups was compared and observed. The comparison of quantitative data between groups was conducted using the Mann-Whitney U test. Multivariate logistic regression was used to analyze the correlations between PPA, optic disc tilt, PHOMS occurrence and gender, age, diopter, AL, and ChT. Results:Among the 162 patients, 103 were male and 59 were female. Age was 12 (10.5, 13.5) years old. Among the 162 eyes, the optic disc morphology changed in 152 eyes (93.8%, 152/162). Among them, the PPA, optic disc tilt, PHOMS, and optic disc rotation were 148 (91.4%, 148/162), 95 (58.6%, 95/162), 62 (38.3%, 62/162), and 35 (21.6%, 35/162) eyes respectively. Myopic macular degeneration in 137 eyes. There were 56 eyes with peripheral retinopathy. There was no statistically significant difference in myopia diopter, AL and ChT between the children group and the adolescent group ( Z=-1.201, -1.934, ?0.761; P=0.230, 0.053, 0.447). There was no statistically significant difference in the incidences of PPA, optic disc tilt and optic disc rotation ( χ2=0.293, 2.618, 0.398; P>0.05). There was no statistically significant difference in the incidence of optic disc morphological changes between the low diopter long axial group and the high diopter long axial group ( χ2=0.000, P>0.05). The results of multivariate logistic regression analysis showed that the thinner the ChT, the higher the risk of PPA [odds ratio (OR) =0.98, 95% confidence interval ( CI) 0.97-0.99, P<0.001]. Female ( OR=2.3, 95% CI 1.04-5.07, P=0.039), older age ( OR=1.17, 95% CI 1.01-1.37, P=0.043), thinner ChT ( OR=0.99, 95% CI 0.99-1.00, P=0.012), the higher the risk of optic disc tilt. The older the age, the higher the risk of developing PHOMS ( OR=1.22, 95% CI 1.06-1.40, P=0.006). Conclusions:Optic disc morphology changes may be the most common fundus alterations in children and adolescents with high myopia. The influencing factors of optic disc morphological changes (including PPA, optic disc tilt, and PHOMS) are female sex, advanced age, and ChT thinning.

Objective To assess the prevalence of mycoplasma genitalium(MG)in patients attending sexually transmitted disease(STD)clinic in Guangzhou,and to provide an epidemiological foundation for clinical treatment and laboratory diagnostics.Methods Utilizing real-time polymerase chain reaction(PCR),we analyzed MG DNA in 2,749 clinical specimens collected from 2,722 outpatients in the Dermatology Hospital of Southern Medical University from July 2019 to December 2021.Concurrent testing for MG,Chlamydia trachomatis(CT),and Neisseria gonorrhoeae(NG)was performed on 2,382 of these specimens.Patient data extracted from medical records were used to investigate the correlation between STD symptoms and MG infection.Results The investigation revealed that the overall prevalence of MG infection was 4.4%among the sampled patients(120 out of 2,722),with a higher prevalence in males(4.9%,or 87 out of 1,790)compared to in females(3.5%,or 33 out of 932).Notably,the prevalence decreased with increasing age.The highest incidence of MG infection was observed in females aged 18～25 years(6.4%,or 18 out of 281),while the lowest was in males aged 46 years and above(1.5%,or 5 out of 342),showing a statistically significant variation across age groups(P<0.05).Among males with urethritis symptoms,MG positive rate was significantly higher at 7.3%(42 out of 574).The rate of single MG infection was prominent,accounting for 89.9%(71 out of 79)in MG-positive male patients and 61.5%(16 out of 26)in MG-positive female patients.Co-infection rate of MG with CT was 1.2%in females and 0.3%in males,indicating a significant dif-ference(P<0.05).Conclusion The findings suggest a relatively high prevalence of MG infection and co-infection with CT among STD clinic attendees in Guangzhou,particularly in the younger demographic.The study underscores the need for early screening and vigilant surveillance of MG to mitigate its transmission among sexually active popula-tions at high risk.

[Objective]To establish the reference intervals of thyroid hormones in pregnant women from Sanming city and compare them with those from other areas in China.[Methods]The study recruited 605 pregnant women and 229 non-pregnant healthy women who visited Sanming First Hospital between March 29 and June 28,2023.Blood samples were se-quentially collected from the participants to determine the serum levels of thyroid stimulating hormone(TSH),free triiodo-thyronine(FT3)and free thyroxine(FT4).The reference intervals of thyroid hormones were established by using a percen-tile range of P2.5 to P97.5 and their changes in different periods of pregnancy were analyzed and compared with those from oth-er areas in China.[Results]There were significant differences in levels of TSH,FT4 and FT3 at different periods of pregnan-cy(all P<0.05).Compared with non-pregnant women,women in first trimester had significantly decreased levels of TSH and FT4,but similar level of FT3.Since the second trimester,TSH level displayed rising tendency,while FT3 and FT4 lev-els showed gradual decrease.The reference intervals of thyroid hormones in the first,second and third trimester of pregnan-cy in Sanming city were TSH(0.068,2.943)μU/mL,FT3(4.302,6.888)pmol/L,FT4(8.240,14.719)pmol/L;TSH(0.419,3.274)μU/mL,FT3(4.074,6.629)pmol/L,FT4(6.726,11.980)pmol/L;TSH(0.422,3.570)μU/mL,FT3(3.741,5.850)pmol/L,FT4(6.103,10.347)pmol/L,respectively.Significant differences were found in the reference in-tervals of thyroid hormones during pregnancy among different areas in China.[Conclusions]The levels of TSH,FT3,FT4 during pregnancy are different from those during non-pregnancy,and also significantly differ in different periods of preg-nancy.Reference intervals of thyroid hormones in pregnant women are affected by various factors such as geographic loca-tion,ethnicity and laboratory test method,etc.Therefore,establishing the population-specific reference intervals of thy-roid hormones will benefit for the diagnosis and treatment of thyroid disease in pregnant women from Sanming city.

BACKGROUND: The clinical features of enthesitis in patients with psoriatic arthritis (PsA) have been reported in some Western countries, but data in China are very limited. This study aimed to describe the characteristics of enthesitis in Chinese patients with PsA and compared them with those in other cohorts. METHODS: Patients with PsA enrolled in the Chinese Registry of Psoriatic Arthritis (CREPAR) (December 2018 to June 2021) were included. Data including demographics, clinical characteristics, disease activity measures, and treatment were collected at enrollment. Enthesitis was assessed by the Spondyloarthritis Research Consortium of Canada (SPARCC), Maastricht ankylosing spondylitis enthesitis score (MASES), and Leeds enthesitis index (LEI) indices. A multivariable logistic model was used to identify factors related to enthesitis. We also compared our results with those of other cohorts. RESULTS: In total, 1074 PsA patients were included, 308 (28.7%) of whom had enthesitis. The average number of enthesitis was 3.3 ± 2.8 (range: 1.0-18.0). More than half of the patients (165, 53.6%) had one or two tender entheseal sites. Patients with enthesitis had an earlier age of onset for both psoriasis and arthritis, reported a higher proportion of PsA duration over 5 years, and had a higher percentage of axial involvement and greater disease activity. Multivariable logistic regression showed that axial involvement (odds ratio [OR] 2.21, 95% confidence interval [CI], 1.59-3.08; P <0.001), psoriasis area and severity index (PASI) (OR: 1.03, 95% CI: 1.01-1.04; P = 0.002), and disease activity score 28-C reactive protein (DAS28-CRP) (OR: 1.25, 95% CI: 1.01-1.55; P = 0.037) were associated with enthesitis. Compared with the results of other studies, Chinese patients with enthesitis had a younger age, lower body mass index (BMI), a higher rate of positive human leukocyte antigen (HLA)-B27, more frequent dactylitis, and a higher proportion of conventional synthetic disease-modifying antirheumatic drugs' (csDMARDs) use. CONCLUSIONS Enthesitis is a common condition among Chinese patients with PsA. It is important to evaluate entheses in both peripheral and axial sites.
Humans ; Arthritis, Psoriatic/drug therapy* ; East Asian People ; Enthesopathy/complications* ; Registries ; Severity of Illness Index ; Spondylarthritis/epidemiology*