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幽门螺杆菌空泡细胞毒素A蛋白的生物信息学分析

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寄生虫病与感染性疾病 2024年第2期22卷 86-93页

作者：李桂玲张东军李娜万秀菊山东省寄生虫病防治研究所山东第一医科大学附属消化病医院山东济宁272033

目的了解幽门螺杆菌空泡细胞毒素A(vaca)的结构和功能,并对其抗原表位进行分析。方法采用生物信息学软件ORF Finder、Protpapram、ProtScale、Cell-PLoc 2.0、SignalP-6.0、TMHMM-2.0、NetNGlyc-1.0、NetPhos-3.1、SOPMA、SWISSMODEL、IEDB、SYFPEITHI、STRING等对vaca蛋白的理化性质及亚细胞定位、二级结构、三级结构、抗原蛋白B细胞、细胞毒性T淋巴细胞表位等进行预测。结果 vaca蛋白由747个氨基酸组成,分子式为C_(3549)H_(5597)N_(1009)O_(1111)S_(9),原子总数为11 275,其编码基因全长3 867 bp,等电点9.33,不稳定指数18.53,脂肪族氨基酸指数82.40,平均疏水性为-0.342。该蛋白有跨膜区无信号肽,有12个糖基化位点和71个磷酸化位点。该蛋白以无规则卷曲(占44.03%)为主,属于稳定的亲水性蛋白。vaca蛋白含有9个B细胞表位,多个T细胞表位。结论 vaca能够磷酸化,并与多种蛋白相互作用,可作为抗幽门螺旋杆菌疫苗的候选蛋白。

关键词：幽门螺旋杆菌 vaca 生物信息学结构

幽门螺杆菌毒力基因数字PCR检测体系的建立

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质量安全与检验检测 2024年第1期34卷 36-41页

作者：范宏博蔡永洪李月玥梁志坚张力玲马婷婷胡松青胡良勇华南理工大学食品科学与工程学院广东广州510630 广州计量检测技术研究院

建立幽门螺杆菌(Helicobacter pylori)毒力基因数字PCR检测体系,为Hp诊疗提供一个精准、灵敏的毒力定量检测方法。根据Hp毒力基因vaca和cagA的序列设计特异性引物和探针,采用质控菌评估检测特异性;以梯度法设置引物浓度和退火温度,对反... 详细信息

建立幽门螺杆菌(Helicobacter pylori)毒力基因数字PCR检测体系,为Hp诊疗提供一个精准、灵敏的毒力定量检测方法。根据Hp毒力基因vaca和cagA的序列设计特异性引物和探针,采用质控菌评估检测特异性;以梯度法设置引物浓度和退火温度,对反应体系及反应条件进行优化;以梯度稀释的DNA为模板,评估检测灵敏度;采用不同浓度模板开展重复性检测,评估检测精密度。结果显示,所设计的引物和探针可特异性地检测vaca和cagA基因,不受大肠杆菌等细菌干扰。vaca和cagA基因的最佳反应温度分别为55.0℃和57.9℃,最佳引物工作浓度分别为650 nmol/L和550 nmol/L,线性分析的拟合度R^(2)分别为0.9991和0.9995,不同浓度样品的变异系数(CV)值均小于10%。本研究建立的Hp毒力基因vaca和cagA数字PCR检测体系具有特异、灵敏和重复性好等优点,可为Hp病理诊疗和科学研究提供精准、可靠的技术支持。

关键词：幽门螺杆菌 vaca cagA 数字PCR 精密度重复性

Prevalence of vaca, cagA and babA2 genes in Cuban Helicobacter pylori isolates

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World Journal of Gastroenterology 2009年第2期15卷 204-210页

作者： Lino E Torres Karelia Melián Arlenis Moreno Jordis Alonso Carlos A Sabatier Mayrín Hernández Ludisleydis Bermúdez Boris L Rodríguez Department of Microbiology and Immunology Biotechnology Division National Centre for Scientif ic Research Ciudad de La Habana Cuba Department of Gastroenterology Medical and Chirurgic Research Centre (CIMEQ) Ciudad de La Habana Cuba Anatomopathology Department Medical and Chirurgic Research Centre (CIMEQ) Ciudad de La Habana Cuba

AIM: To investigate the prevalence of vacuolating cytotoxin (vaca), cytotoxin associated gene A (cagA) and blood adhesion binding antigen (babA2) genotypes of Helicobacter pylori (H pylori) isolates from Cuban dyspeptic patients. METHODS: DNA was extracted from H pylori-positive cultures taken from 130 dyspeptic patients. Genotyping was performed by PCR, using specifi c primers for vaca (s1, s2, m1, m2), cagA and babA2 genes. Endoscopic observations and histological examinations were used to determine patient pathologies. RESULTS: vaca alleles s1 , s2 , m1 and m2 were detected in 96 (73.8%), 34 (26.2%), 75 (57.7%) and 52 isolates (40%), respectively, while the cagA gene was detected in 95 isolates (73.2%). One hundred and seven isolates (82.3%) were babA2-positive. A signif icant correlation was observed between vacas1m1 and cagA and between vacas1m1 and babA2 genotypes (P vacas1 and babA2 genotypes. Eighty fi ve (65.4%) and 73 (56.2%) strains were type 1 (vacas1-cagA-positive) and "triple- positive" (vacas1-cagA-babA2-positive), respectively, and their presence was significantly associated with duodenal ulcer (P < 0.01 and P < 0.001, respectively). CONCLUSION: The distribution of the main virulence factors in the Cuban strains in this study resembled that of the Western-type strains, and the more virulent H pylori isolates were signifi cantly associated with duodenal ulcer, ulcer disease being the worst pathology observed in the group studied.

关键词：古巴消化系统疾病基因 vaca cagA babA

Helicobacter pylori in dental plaque and stomach of patients from Northern Brazil

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World Journal of Gastroenterology 2010年第24期16卷 3033-3039页

作者： Mnica Baraúna Assumpo Luisa Caricio Martins Hivana Patricia Melo Barbosa Katarine Antonia dos Santos Barile Sintia Silva de Almeida Paulo Pimentel Assumpo Tereza Cristina de Oliveira Corvelo Endoscopy Department Joo de Barros Barreto University Hospital Immunopathology Laboratory Nucleus of Tropical Medicine Federal University of Pará Immunogenetics Laboratory Federal University of Pará Paulo Pimentel Assumpo Surgery Department Joo de Bar-ros Barreto University Hospital

AIM: To establish whether virulence factor genes vaca and cagA are present in Helicobacter pylori (H. pylori) retrieved from gastric mucosa and dental plaque in pa-tients with dyspepsia. METHODS: Cumulative dental plaque specimens and gastric biopsies were submitted to histological exami-nation, rapid urease test and polymerase chain reac-tion (PCR) assays to detect the presence of cagA and vaca polymorphisms.RESULTS: Detection of H. pylori from dental plaque and gastric biopsy samples was greater by PCR com-pared to histological examination and the rapid ure-ase test. DNA from H. pylori was detected in 96% of gastric mucosa samples and in 72% of dental plaque samples. Sixty-three (89%) of 71 dental plaque sam-ples that were H. pylori-positive also exhibited identical vaca and cagA genotypes in gastric mucosa. The most common genotype was vacas1bm1 and cagA positive, either in dental plaque or gastric mucosa. These viru-lent H. pylori isolates were involved in the severity of clinical outcome.CONCLUSION: These pathogenic strains were found simultaneously in dental plaque and gastric mucosa, which suggests that gastric infection is correlated with the presence of H. pylori in the mouth.

关键词： Helicobacter pylori Gastric mucosa Dental plaque cagA vaca

Helicobacter pylori and gastric cancer in the Middle East: A new enigma?

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World Journal of Gastroenterology 2010年第26期16卷 3226-3234页

作者： Nawfal R Hussein Institute of Infection Immunity and Inflammation and the Centre for Biomolecular Sciences University of Nottingham Nottingham NG7 2RD United Kingdom

The Middle East is the home of ethnic groups from three main backgrounds: Semitic (Arabs and Jews), Indo-European (Persians and Kurdish) and Turkic (Turkish and Turkmens). Its geographic location, which has been under continuous influences from Asia, Europe and Africa, has made it an ideal site for epidemiological studies on Helicobacter pylori (H. pylori) infection and genotyping. The gastric cancer rate differs in this region from very high in Iran (26.1/105) to low in Israel (12.5/105) and very low in Egypt (3.4/105). Epidemiological studies showed that the prevalence of H. pylori is almost similar in those countries with a high level of infection in childhood. Importantly, the frequency of vaca s1 and m1 regions and cagA+ genotypes were higher in non Semitic populations who inhabit the North than Semitic populations, the inhabitants of Southern parts of the Middle East. H. pylori infection prevalence, distribution pattern of virulence factors, diet and smoking could not have explained the difference in cancer rate. This reflects the multifactorial aetiology of gastric cancer and suggests that H. pylori infection does not always directly correlate with the risk for gastrointestinal disease, such as gastric cancer. Further detailed investigations and international comparative studies of each risk factor needto be performed to investigate whether this represents a true enigma.

关键词： Helicobacter pylori Middle East Gastric cancer dupA cagA vaca iceA

Can bacterial virulence factors predict antibiotic resistant Helicobacter pylori infection?

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World Journal of Gastroenterology 2018年第9期24卷 971-981页

作者： Denise E Brennan Colin Dowd Colm O'Morain Deirdre McNamara Sinéad M Smith Department of Clinical Medicine School of Medicine Trinity College Dublin

AIM To evaluate the association between virulence factor status and antibiotic resistance in Helicobacter pylori(H. pylori)-infected patients in Ireland. METHODS DNA was extracted from antral and corpus biopsies obtained from 165 H. pylori-infected patients. Genotyping for clarithromycin and fluoroquinolone-mediating mutations was performed using the Genotype Helico DR assay. cag A and vac A genotypes were investigated using PCR. RESULTS Primary, secondary and overall resistance rates for clarithromycin were 50.5%(n = 53/105), 78.3%(n = 47/60) and 60.6%(n = 100/165), respectively. Primary, secondary and overall resistance rates for fluoroquinolones were 15.2%(n = 16/105) and 28.3%(n = 17/60) and 20%(n = 33/165), respectively. Resistance to both antibiotics was 12.4%(n = 13/105) in treatment-na?ve patients, 25%(n = 15/60) in those previously treated and 17%(n = 28/165) overall. A cag A-positive genotype was detected in 22.4%(n = 37/165) of patient samples. The dominant vac A genotype was S1/M2 at 44.8%(n = 74/165), followed by S2/M2 at 26.7%(n = 44/165), S1/M1 at 23.6%(n = 39/165) and S2/M1 at 4.8%(n = 8/165). Primary clarithromycin resistance was significantly lower in cag A-positive strains than in cag A-negative strains [32%(n = 8/25) vs 56.3%(n = 45/80); P = 0.03]. Similarly, in patients infected with more virulent H. pylori strains bearing the vac A s1 genotype, primary clarithromycin resistance was significantly lower than in those infected with less virulent strains bearing the vac A s2 genotype, [41%(n = 32/78) vs 77.8%(n = 21/27); P = 0.0001]. No statistically significant association was found between primary fluoroquinolone resistance and virulence factor status.CONCLUSION Genotypic H. pylori clarithromycin resistance is high and cag A-negative strains are dominant in our population. Less virulent(cag A-negative and vac A S2-containing) strains of H. pylori are associated with primary clarithromycin resistance.

关键词： Helicobacter pylori Antibiotic resistance Fluoroquinolone Clarithromycin Virulence factor vaca CagA

Genomic variability of Helicobacter pylori isolates of gastric regions from two Colombian populations

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World Journal of Gastroenterology 2017年第5期23卷 800-809页

作者： Andrés Jenuer Matta Alvaro Jairo Pazos Javier Andrés Bustamante-Rengifo Luis Eduardo Bravo Registro Poblacional de Cáncer de Cali Department of PathologySchool of MedicineUniversidad del Valle Department of Biology Universidad de Narino

AIM To compare the genomic variability and the multiple colonization of Helicobacter pylori(H. pylori) in patients with chronic gastritis from two Colombian populations with contrast in the risk of developing gastric cancer(GC): Túquerres-Nari?o(High risk) and Tumaco-Nari?o(Low risk).METHODS Four hundred and nine patients from both genders with dyspeptic symptoms were studied. Seventy-two patients were included in whom H. pylori was isolated from three anatomic regions of the gastric mucosa,(31/206) of the high risk population of GC(Túquerres) and(41/203) of the low risk population of GC(Tumaco). The isolates were genotyped by PCR-RAPD. Genetic diversity between the isolates was evaluated by conglomerates analysis and multiple correspondence analyses. RESULTS The proportion of virulent genotypes of H. pylori was 99% in Túquerres and 94% in Tumaco. The coefficient of similarity of Nei-Li showed greater genetic diversityamong isolates of Túquerres(0.13) than those of Tumaco(0.07). After adjusting by age, gender and type of gastritis, the multiple colonization was 1.7 times more frequent in Túquerres than in Tumaco(P = 0.05).CONCLUSION In Túquerres, high risk of GC there was a greater probability of multiple colonization by H. pylori. From the analysis of the results of the PCR-RAPD, it was found higher genetic variability in the isolates of H. pylori in the population of high risk for the development of GC.

关键词： Helicobacter pylori 致病力小岛 cag cagA vaca 多重殖民 PCR-RAPD

Helicobacter pylori vac A genotype is a predominant determinant of immune response to Helicobacter pylori CagA

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World Journal of Gastroenterology 2017年第26期23卷 4712-4723页

作者： Alexander Link Cosima Langner Wiebke Schirrmeister Wiebke Habendorf Jochen Weigt Marino Venerito Ina Tammer Dirk Schlüter Philipp Schlaermann Thomas F Meyer Thomas Wex Peter Malfertheiner Department of Gastroenterology Hepatology and Infectious DiseasesOtto-von-Guericke University Institute of Medical Microbiology Otto-von-Guericke University Helmholtz Center for Infection Research Department of Molecular Biology Max Planck Institute for Infection Biology Department of Molecular Genetics Medical Laboratory for Clinical ChemistryMicrobiology and Infectious Diseases

AIM To evaluate the frequency of Helicobacter pylori(H. pylori) Cag A antibodies in H. pylori infected subjects and to identify potential histopathological and bacterial factors related to H. pylori Cag A-immune response.METHODS Systematic data to H. pylori isolates, blood samples, gastric biopsies for histological and molecular analyses were available from 99 prospectively recruited subjects. Serological profile(anti-H. pylori, anti-Cag A) was correlated with H. pylori isolates(cag A, EPIYA, vac A s/m genotype), histology(Sydney classification) and mucosal interleukin-8(IL-8) m RNA and protein expression. Selected H. pylori strains were assessed for H. pylori Cag A protein expression and IL-8 induction in co-cultivation model with AGS cells.RESULTS Thirty point three percent of microbiologically confirmed H. pylori infected patients were seropositive for Cag A. Majority of H. pylori isolates were cag A gene positive(93.9%) with following vac A polymorphisms: 42.4% vac A s1m1, 23.2% s1m2 and 34.3% s2m2. Anti-Cag AIg G seropositivity was strongly associated with atrophic gastritis, increased mucosal inflammation according to the Sydney score, IL-8 and cag A m RNA expression. V a c A s a n d m p o l y m o r p h i s m s w e r e t h e m a j o r determinants for positive(vac A s1m1) or negative(vac A s2m2) anti-Cag A serological immune response, which also correlated with the in vitro inflammatory potential in AGS cells. In vitro co-cultivation of representative H. pylori strains with AGS cells confirmed functional Cag A translocation, which showed only partial correlation with Cag A seropositivity in patients, supporting vac A as major co-determinant of the immune response.CONCLUSION Serological immune response to H. pylori cag A + strain in H. pylori infected patients is strongly associated with vac A polymorphism, suggesting the crucial role of bacterial factors in immune and clinical phenotype of the infection.

关键词： Helicobacter pylori Seropositivity 毒力因素 CagA vaca 有免疫力的反应

Helicobacter pylori Virulence Genes cagA, babA2, and vaca Detection in Dyspeptic Patients from Burkina Faso

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American Journal of Molecular Biology 2023年第3期13卷 141-155页

作者： Tegwinde Rebeca Compaore Kalifou Traore Nômawendé Ines Compaore Lassina Traore Sylvie Zida Serge Theophile Soubeiga Dinanibe Kambire Jean Claude R. P. Ouedraogo Aminata Dickel Sidibe Yasmine Astrid Sana Tani Sagna Wendkuuni Florencia Djigma Henri Gautier Ouedraogo Jacques Simpore Département Biomédical et Santee Publique Institut de Recherche en Sciences de la Santé (IRSS) Ouagadougou Burkina Faso Centre de Recherche Biomoléculaire Pietro Annigoni (CERBA) Ouagadougou Burkina Faso Laboratoire de Biologie et de Génétique Moléculaire (LABIOGENE) Université Joseph KI-ZERBO Ouagadougou Burkina Faso Clinique El Fateh-Suka Service de médecine et de spécialités médicales Ouagadougou Burkina Faso Département de Médecine et de Pharmacopée Traditionnelle Institut de Recherche en Sciences de la Santé (IRSS) Ouagadougou Burkina Faso

The diverse clinical presentation of Helicobacter pylori (H. pylori) infection results from the interaction between bacterial virulence, host genetics, socio-demographic and environmental factors. This study aimed to characterize Helicobacter pylori virulence genes and the associated behavioral factors among dyspeptic patients in Burkina Faso. Two hundred and fifty (250) stool samples were collected from patients with dyspepsia seen at health centers in Ouagadougou, Burkina Faso. Bacterial deoxyribonucleic acid (DNA) was extracted using a commercial kit. Virulence genes were detected using conventional multiplex Polymerase Chain Reaction with specific primers. The overall prevalence of Helicobacter pylori of the 250 participants was 91.20%. CagA virulence gene was present among 20.19% of individuals, while babA2 and vaca were detected respectively among 9.65% and 67.54% of the population positive for Helicobacter pylori. Among vaca subtypes, vacas1 was the most frequent, with 39.04%, followed by vacai1 (19.74%), vacai2 (17.54%), and vacas2 with 10.96%. Regarding vacam1 and vacam2, they were less frequent at 6.14% each. “Handwashing three times or less per day” significantly increased the risk of having vacai2 allele and H. pylori rRNA16s, with p-values of 0.013 and 0.020, respectively. The consumption of non-tap water increases the risk of carrying the cagA virulence gene. Additionally, H. pylori-positive patients living with more than four (4) people in their household had about two times the risk of having the vacas1 allele. The present study shows the detection of Helicobacter pylori cagA, vaca subtypes, and babA2 by stool a PCR method in Burkina Faso. The strong association between sanitary habits and virulence factors depicts the composite interaction between ecological factors, gastric mucosa, and bacteria. Therefore, the synergic action of these factors should be considered when aiming for bacterial eradication and gastric pathology cure.

关键词： Helicobacter pylori Stool cagA babA2 vaca