1.Study on quality standard of ethnic medicine Diplazium lanceum
Liyuan HU ; Chuhui ZHOU ; Limei LIANG ; Yong CHEN ; Jiajia ZHONG ; Jiangcun WEI
China Pharmacy 2026;37(15):1986-1991
OBJECTIVE To study the quality standard of the ethnic medicine Diplazium lanceum, and to provide reference for its quality control.METHODS The medicinal material of D. lanceum was identified by properties, microscopy, and thin-layer chromatography(TLC), and its moisture, ash, and extract contents were determined. The contents of chlorogenic acid and rutin in 10 batches of D. lanceum were determined by high-performance liquid chromatography . RESULTS The rhizome of D. lanceum is slender and creeping, the leaf is leathery and shriveled, and yellow sporangia are borne on the abaxial surface of the leaf. The transverse section of the leaf shows inclusions, sclerenchyma, phloem cells, etc.; the transverse section of the petiole shows ground tissue, sclerenchyma, endodermis, etc.; the transverse section of the rhizome shows ground tissue, tracheids, vascular bundles, etc.; and the powder shows spiral vessels and brown inclusions. The TLC identification results showed that the chromatogram of the test sample exhibited spots of the same color at the corresponding positions as the chromatogram of the chlorogenic acid reference substance. The content ranges of moisture, total ash, acid-insoluble ash, extract, chlorogenic acid, and rutin in the 10 batches of samples were 10.18%-12.43%, 7.08%-19.12%, 0.95%-10.62%, 14.13%-26.05%, 0.113 1-1.015 9 mg/g, and 0.009 9-0.208 9 mg/g, respectively. CONCLUSIONS In the quality standard of D. lanceum, the moisture content should be ≤12.5%, total ash content should be ≤20.0%, acid-insoluble ash content should be ≤11.0%, extract content should be ≥14.0%, and chlorogenic acid content should be ≥0.11 mg/g. The established method is simple, specific, and reproducible, and can provide a scientific basis for the quality control of D. lanceum.
2.Comparison of the efficacy of continuous VA chemotherapy and I/HDAC consolidation in postremission therapy for acute myeloid leukemia fit for standard chemotherapy
Li SUN ; Pengpeng ZHANG ; Simei REN ; Nan ZHOU ; Liyuan LI ; Zhenzhen WANG ; Weiguang CUI ; Fan YANG ; Jianmin LUO ; Lin YANG
Chinese Journal of Hematology 2025;46(4):343-348
Objective:To compare the efficacy and safety of continuous venetoclax combined azacitidine (VA) chemotherapy and intermedium/high-dose cytarabine (I/HDAC) consolidation in patients with acute myeloid leukemia (AML) fit for standard chemotherapy (transform from UNFIT) .Methods:Clinical data of patients who were fit for standard chemotherapy were collected among those with AML who underwent VA induction in the Department of Hematology, the Second Hospital of Hebei Medical University. The overall survival (OS), relapse-free survival (RFS), event-free survival (EFS), and incidence of adverse events were analyzed retrospectively.Results:This study enrolled 69 patients, consisting of 46 cases in the VA group and 23 cases in the I/HDAC group. We revealed the following. ① The median OS, RFS, EFS were 26.18, 24.69, 20.34 months in the VA group, and 34.14, 30.99, 28.42 months in the I/HDAC group, respectively, with no statistically significant difference (all P>0.05). Median OS of patients who underwent I/HDAC consolidation with European Leukemia Net (ELN) favorable-risk, positive measurable residual disease (MRD), wild type FLT3, or IDH1/2 mutation was significantly longer than those who received VA ( P<0.05). ②Adverse events rate of grade 3 - 4 neutropenia, grade 3 - 4 thrombocytopenia, and bacteremia were significantly lower in the VA group than in the I/HDAC group ( P<0.05) . Conclusions:I/HDAC consolidation was more likely to help get survival benefits for patients with ELN favorable-risk, positive MRD, wild type FLT3, or IDH1/2 mutation. Continuous VA chemotherapy exhibited superior safety than I/HDAC consolidation.
3.Characteristics of different metabolites in lower res piratory tract of patients with coal workers pneumoconiosis
Jine DAI ; Xin ZHANG ; Tao ZHOU ; Jiyin ZHANG ; Liyuan XU ; Shaoying LI
Chinese Journal of Industrial Hygiene and Occupational Diseases 2025;43(5):372-378
Objective:To study the characteristics of metabolites in lower resPiratory tract between coal workers' pneumoconiosis patients and dust exposure patients, and compare the differences of metabolites and their main metabolic pathways.Methods:From December 2020 to February 2021, through a prospective cross-sectional study, a total of 26 patients with coal workers' pneumoconiosis (metabolic group of coal workers' pneumoconiosis) were selected from the bronchoalveolar lavage treatment of coal workers' pneumoconiosis and dust exposure in the Respiratory and Critical Care Medicine Department of the 920th Hospital of the Joint Logistics Support Force during the same period. With 19 cases of dust exposure as the control group (dust exposure metabolic group), samples of alveolar lavage fluid were collected from 2 groups. Metabolites of the two groups were quantitatively analyzed by metabonomics technology, and the characteristics of metabolites and their metabolic pathways were compared. The metabolites with potential predictive value were screened by receiver operating characteristic curve (ROC curve) .Results:Through metabolomic analysis of alveolar lavage fluid in the coal workers' pneumoconiosis group and the dust contact group, a total of 28 different metabolites were screened, including trihydroxybutyric acid, alanine, ethanolamine, L-osan, proline (carboxyl), leucine, 2-hydroxyglutaric acid, proline, lactic acid, serine, valine and threonine in the coal workers' pneumoconiosis group. The levels of differential metabolites such as ornithine, isoleucine, threitol, glucose and lysine were higher ( P<0.05). The levels of different metabolites such as sarcoine, pelanoic acid, palmitic acid, heptadecanoic acid, n-butylamine, tetradecanoic acid, isobutylamine, aminoadipic acid, phosphate, uracil and cytosine were higher in the dust exposure group ( P<0.05). Two major metabolic pathways include glycine, serine and threonine metabolism, arginine and proline metabolism, biotin metabolism, and aminoacyl biosynthesis metabolism. Among the 17 metabolites increased in the coal workers' pneumoconiosis group, the AUC of threitol and lactic acid was greater than 0.8, and the specificity and sensitivity of the working characteristic curves of the two metabolites were 80% and 70%, respectively. Conclusion:There were significant differences in the metabolites of lower respiratory tract between patients with coal workers' pneumoconiosis and those exposed to dust, and the differences were related to multiple metabolic pathways. Threitol and lactic acid may have potential predictive value for pneumoconiosis.
4.Phase Ⅲ, multicenter, randomized comparative study of LY01005 and Zoladex ? for patients with premenopausal breast cancer
Xiying SHAO ; Qingyuan ZHANG ; Zhaofeng NIU ; Man LI ; Jingfen WANG ; Zhanhong CHEN ; Ruizhen LUO ; Guangdong QIAO ; Jianguo WANG ; Liyuan QIAN ; Ronghua YANG ; Zhendong CHEN ; Jian WANG ; Yumin YAO ; Jianghua OU ; Tao SUN ; Qiao CHENG ; Yongsheng WANG ; Jian HUANG ; Hongying ZHAO ; Wuyun SU ; Zhong OUYANG ; Yu DING ; Lilin CHEN ; Sumei YANG ; Mengsheng CUI ; Aimin ZANG ; Enxiang ZHOU ; Peizhi FAN ; Jing ZHANG ; Qiang LIU ; Yuee TENG ; Hui LI ; Jianyun NIE ; Jin YANG ; Xiaojia WANG ; Zefei JIANG
Chinese Journal of Oncology 2025;47(4):340-348
Background:To compare the efficacy and safety of monthly administrations of gonadotropin releasing hormone (GnRH) agonists LY01005 and Zoladex ? in Chinese patients with premenopausal breast cancer. Methods:From October 2020 to November 2021, 188 premenopausal breast cancer patients were enrolled in 34 hospitals and randomized 1:1 to receive either LY01005 or Zoladex ? every 28 days for a total of three injections. All patients concomitantly received oral tamoxifen (TAM). The primary efficacy endpoint was cumulative probability of maintaining menopausal level [oestradiol (E2) ≤30 pg/ml] from day 29 to day 85. The second efficacy endpoint included changes in E2, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) compared with the baseline. Pharmacokinetics (PK), pharmacodynamics (PD), and safety were analyzed. The study also evaluated the pharmacokinetic and pharmacodynamic characteristics of LY01005. Results:A total of 188 patients were randomised and 187 patients received either LY01005 or Zoladex ?. Cumulative probabilities of maintaining menopausal level (E2≤30 pg/ml) from day 29 to day 85 were 93.1% for LY01005 and 86.3% for Zoladex ?. The between-group difference was 6.8% (95% CI: -2.3%, 15.9%) and primary efficacy in the LY01005 group was not inferior to that in the Zoladex ? group. Changes in E2, LH, and FSH levels compared with the baseline were equivalent between the two groups (E2: 89.34% to 90.23% vs. 82.11% to 85.02%; LH: 88.89% to 95.52% vs. 89.70% to 97.02%; FSH: 75.36% to 80.85% vs.73.07% to 80.24%, respectively). After three consecutive doses of LY01005, the LH and FSH levels of the subjects showed a transient increase after the first dose, reached a peak on the second day and then started to decrease. The LH and FSH reached a lower level and remained at or below that level until the 85th day. Both treatments were well-tolerated. Conclusion:LY01005 is as effective as Zoladex ? in suppressing E2 to menopausal levels in Chinese patients with premenopausal breast cancer, with a similar safety profile.
5.A retrospective matching study of partial nephrectomy and radical nephrectomy for pathological T3a stage renal cell carcinoma
Zezhen ZHOU ; Liyuan GE ; Fan ZHANG ; Shaohui DENG ; Ye YAN ; Hongxian ZHANG ; Guoliang WANG ; Lei LIU ; Yi HUANG ; Shudong ZHANG
Journal of Peking University(Health Sciences) 2025;57(4):704-710
Objective:To evaluate the long-term oncological outcomes of partial nephrectomy(PN)in patients with renal cell carcinoma(RCC)who were clinically staged as clinical T1(cT1)preoperatively but upstaged to pathological T3a(pT3a)after surgery.Methods:A total of 427 RCC patients postopera-tively diagnosed as pT3aN0M0 at Peking University Third Hospital from February 2013 to December 2022 were retrospectively reviewed.Among them,33 cT1 patients upstaged to pT3a RCC received PN(PN group),while 394 non-upstaged pT3a RCC patients underwent radical nephrectomy(RN,RN group).Propensity score matching was performed at a 1∶1 ratio based on baseline characteristics.The Kaplan-Meier method was used to assess overall survival(OS),cancer-specific survival(CSS),and disease-free survival(DFS),with Log-rank tests and Cox regression models for multivariate analysis.Results:Before matching,the PN group(n=33)had significantly higher rates of perirenal fat invasion(PFI,45.5%vs.15.2%)and segmental renal vein involvement(42.4%vs.20.8%),but lower rates of renal sinus invasion(RSI,21.2%vs.73.6%)and renal vein tumor thrombus(0%vs.15.2%)compared with the RN group(n=394,all P<0.05).After matching,baseline characteristics were comparable between the PN group(n=33)and RN group(n=33).No significant differences were observed in operative time,blood loss,mean hospital stay,complication rate,positive margin rate,or conversion to open surgery between the two groups(P>0.05).However,the PN group showed significantly higher estimated glomerular filtration rate(eGFR)postoperatively[76.9(55.4,87.3)mL/(min·1.73 m2)vs.61.7(56.8,73.5)mL/(min·1.73 m2),P<0.05],indicating better renal function preserva-tion.No significant differences were found in OS,CSS,or DFS between the groups(P>0.05).Multi-variate ana-lysis identified renal vein invasion(RVI),higher Fuhrman grades(Ⅲ-Ⅳ),and sarcoma-toid differentiation as independent risk factors for DFS and CSS in the pT3a RCC patients(P<0.05).Conclusion:For cT1 RCC patients upstaged to pT3a,PN preserves renal function more effectively while achieving com-parable oncological outcomes to RN.RVI,higher Fuhrmann grade,and sarcomatoid differentiation are independent risk factors for pT3N0M0 RCC patients.
6.Characteristics of different metabolites in lower res piratory tract of patients with coal workers pneumoconiosis
Jine DAI ; Xin ZHANG ; Tao ZHOU ; Jiyin ZHANG ; Liyuan XU ; Shaoying LI
Chinese Journal of Industrial Hygiene and Occupational Diseases 2025;43(5):372-378
Objective:To study the characteristics of metabolites in lower resPiratory tract between coal workers' pneumoconiosis patients and dust exposure patients, and compare the differences of metabolites and their main metabolic pathways.Methods:From December 2020 to February 2021, through a prospective cross-sectional study, a total of 26 patients with coal workers' pneumoconiosis (metabolic group of coal workers' pneumoconiosis) were selected from the bronchoalveolar lavage treatment of coal workers' pneumoconiosis and dust exposure in the Respiratory and Critical Care Medicine Department of the 920th Hospital of the Joint Logistics Support Force during the same period. With 19 cases of dust exposure as the control group (dust exposure metabolic group), samples of alveolar lavage fluid were collected from 2 groups. Metabolites of the two groups were quantitatively analyzed by metabonomics technology, and the characteristics of metabolites and their metabolic pathways were compared. The metabolites with potential predictive value were screened by receiver operating characteristic curve (ROC curve) .Results:Through metabolomic analysis of alveolar lavage fluid in the coal workers' pneumoconiosis group and the dust contact group, a total of 28 different metabolites were screened, including trihydroxybutyric acid, alanine, ethanolamine, L-osan, proline (carboxyl), leucine, 2-hydroxyglutaric acid, proline, lactic acid, serine, valine and threonine in the coal workers' pneumoconiosis group. The levels of differential metabolites such as ornithine, isoleucine, threitol, glucose and lysine were higher ( P<0.05). The levels of different metabolites such as sarcoine, pelanoic acid, palmitic acid, heptadecanoic acid, n-butylamine, tetradecanoic acid, isobutylamine, aminoadipic acid, phosphate, uracil and cytosine were higher in the dust exposure group ( P<0.05). Two major metabolic pathways include glycine, serine and threonine metabolism, arginine and proline metabolism, biotin metabolism, and aminoacyl biosynthesis metabolism. Among the 17 metabolites increased in the coal workers' pneumoconiosis group, the AUC of threitol and lactic acid was greater than 0.8, and the specificity and sensitivity of the working characteristic curves of the two metabolites were 80% and 70%, respectively. Conclusion:There were significant differences in the metabolites of lower respiratory tract between patients with coal workers' pneumoconiosis and those exposed to dust, and the differences were related to multiple metabolic pathways. Threitol and lactic acid may have potential predictive value for pneumoconiosis.
7.Correlation of GPRC6A Gene Polymorphism and Its Expression with Pulmonary Infection in Elderly Patients with Chronic Heart Failure
Qiufeng WANG ; Wenping JIAO ; Lin GAO ; Weicheng LI ; Liyuan ZHOU ; Nina BIAN ; Sen LIANG ; Menglin WANG ; Dongbo GAO
Journal of Modern Laboratory Medicine 2025;40(6):62-67
Objective To investigate the correlation between G protein-coupled receptor family C group 6 member A(GPRC6A)gene polymorphisms and their expression and pulmonary infections in elderly patients with chronic heart failure(CHF).Methods 138 elderly CHF patients admitted to the Xianyang First People's Hospital from January 2021 to January 2024 were selected as the research subjects,and were divided into an infected group(n=42)and an uninfected group(n=96)based on their lung infection status.Polymerase chain reaction(PCR)was used to detect polymorphisms at the rs6901250 and rs1606365 loci of the GPRC6A gene.The allele and genotype frequency distributions of the infected and uninfected groups were compared.Logistic regression modeling was used to analyze the s6901250 and rs1606365 loci under three genetic models(co-dominant,dominant and reces-sive)and lung infections in elderly patients with CHF.Real-time fluorescence quantitative PCR(RT-qPCR)method was used to detect the expression level of GPRC6A gene.The predictive value of the mRNA expression level of the GPRC6A gene for the development of pulmonary infections in elderly patients with CHF was analyzed by applying the receiver operator characteristic(ROC)curve.Results The distribution of genotypes at loci rs6901250 and rs1606365 of the GPRC6A gene in both the infected and uninfected groups of the lungs of elderly CHF patients conformed to the Hardy-Weinberg equilibrium law(χ2=0.199~0.376,all P>0.05),which was representative of the population.Compared with the uninfected group,the frequency of allele A at locus rs6901250(57.14%vs 41.67%)was significantly higher in the infected group,Allele G(54.76%vs.37.50%)and genotype GG(14.06%vs 29.99%)frequencies were significantly higher at locus rs1606365,and the differences were statistically significant(χ2=5.628,7.114,6.849,all P<0.05).At locus rs6901250,in the co-dominant model(GG vs AA)and the dominant model(GA+AA vs GG),the elderly CHF patients with AA genotype the risk of lung infection was higher than that of GG genotype(OR=1.753,1.546,all P<0.05);.rs1606365 locus showed that the risk of lung infection was higher than that of CC genotype in el-derly CHF patients with GG genotype under all three genetic models of co-dominant model(CC vs GG),dominant model(CG+GG vs CC)and recessive model(CG+CC vs GG)(OR=1.833,1.741,0.695,all P<0.05).The mRNA expression level of GPR-C6A gene in the lung-infected group of elderly CHF patients(1.43±0.35)was significantly higher than that in the uninfected group(1.02±0.21),and the difference was statistically significant(t=8.515,P<0.001).The results of the ROC curve analysis showed that the GPRC6A gene expression level predicted lung infection in elderly CHF patients with an AUC value of 0.895,a cut-offvalue of 1.37,and sensitivity and specificity of 85.7%and 66.7%,respectively.Conclusion The AA genotype at the rs6901250 locus and the GG genotype at the rs1606365 locus of the GPRC6A gene increased the risk of developing lung infec-tions in elderly patients with CHF.MRNA expression levels of the GPRC6A gene were elevated in the infected group,and its ex-pression level could be used as a predictive indicator for the development of lung infections in elderly patients with CHF.
8.Application of triple pre-rehabilitation strategy in breast cancer patients undergoing neoadjuvant chemotherapy
Qizhou YANG ; Nandi WANG ; Liyuan MA ; Shuping LI ; Jing CHEN ; Jianghong ZHOU
Chinese Journal of Practical Nursing 2025;41(29):2257-2264
Objective:To explore the effect of triple pre-rehabilitation strategy on improving nutritional status, negative emotions and quality of life in breast cancer patients undergoing neoadjuvant chemotherapy, so as to provide reference for clinical practice.Methods:A quasi experimental research was adopted. A total of 133 patients who underwent neoadjuvant chemotherapy for breast cancer at Gansu Provincial Cancer Hospital between December 2023 and December 2024 were recruited as research participants. Patients were stratified into the experimental group ( n = 66) and control group ( n = 67) according to their admission sequence. The control group received routine health guidance, while the experimental group was provided with a triple pre-rehabilitation strategy based on the control group. The serum albumin, body mass index, anxiety, depression scores and quality of life scores were compared between the two groups before intervention, 3 weeks after intervention and 2 weeks post-surgery. Results:Totally 64 patients in each group completed the study. The age of the experimental group was (49.50 ± 10.46) years, and that of the control group was (50.42 ± 12.50) years. All of them were females. After a 3-week intervention, the serum albumin levels in the experimental group were (42.62 ± 1.56) g/L, compared to (40.50 ± 1.31) g/L 2 weeks post-surgery. In the control group, these values were (40.98 ± 1.35) g/L and (40.00 ± 1.67) g/L, respectively. The differences between groups were statistically significant ( t = -6.38, -1.89, both P<0.05). Additionally, body mass index of the experimental group was (24.02 ± 1.35) kg/m 2 after the 3-week intervention and (23.48 ± 1.72) kg/m 2 2 weeks post-surgery, which were significantly higher than those of the control group (22.98 ± 1.72) kg/m 2 and (22.50 ± 1.31) kg/m 2, respectively ( t = - 3.81, - 3.61, both P<0.05). The anxiety scores in the experimental group were (6.00 ± 2.09) of 3 weeks after intervention and (7.00 ± 2.62) 2 weeks post-surgery, both of which were significantly lower than those in the control group (7.00 ± 2.26) and (8.00 ± 2.76), with statistically significant differences ( t = 2.60, 2.10, both P<0.05). Additionally, the depression scores in the experimental group were (5.00 ± 2.20) of 3 weeks after intervention and (6.00 ± 2.40) 2 weeks post-surgery, which were also significantly lower than those in the control group (6.00 ± 2.02) and (7.00 ± 2.09), with statistically significant differences ( t = 2.68, 2.52, both P<0.05). Furthermore, the quality of life scores in the experimental group were (94.84 ± 18.00) of 3 weeks after intervention and (84.02 ± 17.13) 2 weeks post-surgery, which were significantly higher than those in the control group (82.00 ± 18.75) and (72.03 ± 17.10), with statistically significant differences ( t = - 3.95, - 3.96, both P<0.05). Conclusions:The triple pre-rehabilitation strategy plays a crucial role in mitigating the nutritional risk for breast cancer patients undergoing neoadjuvant chemotherapy, alleviating their negative emotions, and enhancing their quality of life. This approach is anticipated to serve as a valuable reference for preoperative health guidance tailored to breast cancer patients undergoing neoadjuvant chemotherapy.
9.Establishment and evaluation of method for rapid detection of Bacillus cereus cytK and nhe toxin genes by dual nucleic acid test strips
Jifei YANG ; Beizhen PAN ; Yan LIU ; Yujiao ZHOU ; Jianyu YANG ; Xianyu ZHANG ; Wenbo DING ; Haoyu LI ; Liyuan SUN
Journal of Jilin University(Medicine Edition) 2025;51(2):516-525
Objective:To establish a method of dual nueleic acid test strips for rapid detection of Bacillus cereus cytK and nhe toxin genes based on polymerase chain reaction(PCR)and colloidal gold technique,and to evaluate its specificity,sensitivity,repeatability and stability.Methods:Bacillus cereus DNA was extracted by boiling method.Specific primers were designed with Bacillus cereus cytK and nhe as the target genes.Clonal transformation was used to identify the PCR products.The optimal labeling amounts of colloidal gold-labeled streptavidin,quality control line(C line),cytK detection line(T1)and nhe detection line(T2)were determined.The nucleic acid test strips were assembled and its specificity,sensitivity,reproducibility and stability were evaluated.Results:The DNA concentration of Bacillus cereus was 248 mg·L-1,and the purities were 1.8-2.0.After cloning and plasmid sequencing,the similarities between the PCR products and the sequences of cytK and nhe registered in the GenBank database were 100%.Under the condition of pH 7.0,the optimal amount of streptavidin labeling per 200 μL of colloidal gold solution was 6.0 μL;the optimal marking amount was 2.00 g·L-1 for the quality control line(C line),0.550 g·L-1 for cytK gene detection line(T1)and 0.2 g·L-1 for nhe gene detection line(T2).In the specificity test,positive result on the test strips was seen only for Bacillus cereus,and no cross-reactivity was observed for Staphylococcus aureus,Escherichia coli,Pseudomonas aeruginosa and Bacillus subtilis,which were consistent with the electrophoresis results.Sensitivity assay showed that even when DNA concentration was reduced to 10-2 mg·L-1,three bands(C line,T1 line and T2 line)could be observed,and the detection limit of the test strip was one-tenth of agarose gel electrophoresis(10-1 mg·L-1).The nucleic acid test strips were verified by different operators in different laboratories,and the results were consistent.The stability of the test strips was verified at the 6th,9th and 12th months,and the results showed good stability.Conclusion:The dual nucleic acid test strip method established in this study can simutaneously detect the cytK and nhe toxin genes of Bacillus cereus with high sensitivity and specificity,achieving short-term visual detection.
10.Establishment of LAMP combined with CRISPR/Cas12a system for detecting tlh gene of Vibrio parahaemolyticus and its evaluation
Yujiao ZHOU ; Jifei YANG ; Yan LIU ; Wenbo DING ; Xianyu ZHANG ; Jianyu YANG ; Linran GAO ; Yundong ZHAO ; Liyuan SUN
Journal of Jilin University(Medicine Edition) 2025;51(5):1399-1406
Objective:To establish a rapid detection method for pathogenic microorganisms by combining loop-mediated isothermal amplification(LAMP)and clustered regularly interspaced short palindromic repeats(CRISPR)/CRISPR-associated protein 12a(Cas12a)(CRISPR-Cas12a)system,and to evaluate its efficacy for detecting the thermolabile hemolysin(tlh)gene of Vibrio parahaemolyticus(Vp).Methods:Using the tlh gene of Vp as the target gene,LAMP primers and CRISPR RNA(crRNA)were designed to construct and optimize the optimal concentration ratio of each component in the LAMP-CRISPR detection system.Bacillus cereus,Staphylococcus aureus,and Escherichia coli were used as control groups,and the specificity,sensitivity,reproducibility and positive conformity rate were verified to establish a rapid LAMP-CRISPR/Cas12a method for detecting the tlh gene of Vp.Results:The method specifically detected Vp,while Bacillus cereus,Staphylococcus aureus,and Escherichia coli yielded negative results.The DNA extraction concentration of Vp was 190.67 mg·L-1 with an A(260)/(A280)ratio of 1.84.Under the reaction conditions of 37℃ with 80 cycles for 40 min using quantitative PCR(qPCR)method,when the concentrations of Cas12a protein and crRNA in the LAMP-CRISPR/Cas12a system were 50 nmol·L-1,the visual brightness and relative fluorescence intensity peaks were high.The sensitivity of LAMP CRISPR/Cas12a for detecting Vp DNA concentration could reach 10-6 mg·L-1.The reproducibility test results showed that different experimenters had consistent results in different experimental environments and times.Conclusion:The established LAMP-CRISPR/Cas12a method can rapidly detect the tlh gene of Vp with high sensitivity and specificity,and can achieve short-term visual detection in the field.

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