Clinical Value of Hepatitis B Virus RNA Detection in Patients with Chronic Hepatitis B Infection

Objective:To study the clinical value of hepatitis B virus pregenomic RNA(HBV-pgRNA)detection in the treatment of hepatitis B.Methods:60 patients with hepatitis B were included in the study.Serum HBV-pgRNA and HBV DNA levels in different phases of infection and during treatment were detected,and serum hepatitis B surface antigen(HbsAg)titer was detected by chemiluminescent immunoassay.DNA was extracted from liver biopsy tissue,and covalently closed circular DNA was detected to predict the therapeutic value in patients.Results:At the initial stage of treatment,the level of HBV-pgRNA in phase I,II,III,and IV showed a gradual decrease.Comparing the levels of HBV-pgRNA before and after treatment,we found that the level of HBV-pgRNA was significantly lower after treatment(P<0.05).Among the indicators for predicting HBsAg seroconversion,the accuracy of HBV-pgRNA level was 85.0%(51/60).Conclusion:The clinical value of HBV-pgRNA detection in the treatment of hepatitis B is high.

Biosensors for hepatitis B virus detection

A biosensor is an analytical device used for the detection of analytes,which combines a biological component with a physicochemical detector.Recently,an increasing number of biosensors have been used in clinical research,for example,the blood glucose biosensor.This review focuses on the current state of biosensor research with respect to efficient,specific and rapid detection of hepatitis B virus(HBV).The biosensors developed based on different techniques,including optical methods(e.g.,surface plasmon resonance),acoustic wave technologies(e.g.,quartz crystal microbalance),electrochemistry(amperometry,voltammetry and impedance) and novel nanotechnology,are also discussed.

Comparison of ligase detection reaction and real-time PCR for detection of low abundant YMDD mutants in patients with chronic hepatitis B

AIM: To compare the ligase detection reaction (LDR) and real-time PCR for detection of low abundant YMDD mutants in patients with chronic hepatitis B infection.METHODS: Mixtures of plasmids and serum samples from 52 chronic hepatitis B patients with low abundant lamivudine-resistant mutations were tested with LDR and real-time PCR. Time required and reagent cost for both assays were evaluated.RESULTS: Real-time PCR detected 100, 50, 10, 1 and 0.1% of YIDD plasmid, whereas LDR detected 100, 50, 10, 1, 0.1, and 0.01% of YIDD plasmid, in mixtures with YMDD plasmid of 106 copies/mL. Among the 52 clinical serum samples, completely concordant results were obtained for all samples by both assays, and 39 YIDD, 9 YVDD, and 4 YIDD/YVDD were detected. Cost and time required for LDR and real-time PCR are 60/80 CNY (8/10.7 US dollars) and 4.5/2.5 h, respectively.CONCLUSION: LDR and real-time PCR are both sensitive and inexpensive methods for monitoring low abundant YMDD mutants during lamivudine therapy in patients with chronic hepatitis B. LDR is more sensitive and less expensive, while real-time PCR is more rapid.

HBV感染孕妇早期胆汁酸谱检测在妊娠期肝内胆汁淤积症诊断中的应用

目的探讨HBV感染孕妇早期胆汁酸谱检测在妊娠期肝内胆汁淤积症(ICP)诊断中的应用价值。方法回顾性分析2020年1月—2022年12月无锡市妇幼保健院收治的186例HBV感染孕妇,根据ICP诊断标准将其分为单纯HBV组(124例)和HBV并发ICP组(62例),收集两组孕妇一般资料及妊娠早期围保肝生化指标和胆汁酸谱检测结果,采用单因素分析、多因素logistic回归分析HBV感染孕妇并发ICP的主要影响因素,ROC曲线分析预测诊断效能。结果与HBV组比,HBV并发ICP组AST[(46.60±38.98)U/L比(30.97±31.49)U/L,P=0.004]、ALT[(50.80±36.81)U/L比(40.32±29.45)U/L,P=0.037]、DBil[(6.07±2.34)μmol/L比(4.73±1.83)μmol/L,P<0.001]、TBA[(16.98±2.48)μmol/L比(6.01±2.34)μmol/L,P=0.010]明显升高,CA[(0.59±0.49)μmol/L比(0.40±0.34)μmol/L,P=0.007]、GCA[(2.41±1.04)μmol/L比(1.52±0.70)μmol/L,P<0.001]、GDCA[(0.92±0.35)μmol/L比(0.67±0.37)μmol/L,P<0.001]、GCDCA[(2.14±0.89)μmol/L比(1.67±0.56)μmol/L,P<0.001]也升高,且AST、DBil、GCA、GDCA、GCDCA为HBV并发ICP的主要危险因素(P<0.05);AST、DBil、GCA对HBV并发ICP诊断价值较高(分别AUC 0.747、0.725、0.761);GCDCA、GDCA的诊断价值一般(AUC 0.667、0.688)。结论胆汁酸谱GCA、GCDCA、GDCA亚型升高及AST、DBil升高均是HBV感染孕妇并发ICP发生的主要危险因素,且GCA的预测诊断价值最高,临床应结合肝功能指标综合诊断。

不同核酸提取方法对HBV-DNA检测性能验证情况分析

目的评估2种乙型肝炎病毒(HBV)-DNA提取方法及2家检测试剂的性能,有助于选择优化提取试剂和检测试剂。方法2023年4月采用达安全自动核酸提取仪提取法(磁珠法)和手工提取法(一步法),并用达安和圣湘2种HBV-DNA试剂检测,对其进行精密度、正确度、线性范围、检出限及抗干扰能力等性能进行验证和评价。结果达安全自动核酸提取仪提取达安试剂检测、手工提取达安试剂检测和手工提取圣湘试剂检测在精密度、正确度、线性范围、检出限方面验证结果均达标;达安全自动核酸提取仪提取圣湘试剂检测在低值检测中变异系数大于5%,最低检测限验证不合格;抗干扰能力方面,全自动核酸提取仪提取的2.0 g/dL血红蛋白浓度的样本用达安和圣湘试剂检测结果均不受影响。手工提取甘油三酯浓度达3000 mg/dL的样本用达安和圣湘试剂检测的结果均不受影响。结论不同厂家的提取和检测试剂避免混用,达安和圣湘试剂对HBV-DNA定量检测的结果均符合要求。

Survey of hepatitis B virus infection for liver cancer screening in China:A population-based,cross-sectional study

Background:Hepatitis B virus(HBV)infection is the primary cause of hepatocellular carcinoma(HCC)in China.The target population for HCC screening comprises individuals who test positive for hepatitis B surface antigen(HBsAg).However,current data on the prevalence of HBV infection among individuals who are eligible for HCC screening in China are lacking.We aimed to assess the seroepidemiology of HBV infection among Chinese individuals eligible for HCC screening to provide the latest evidence for appropriate HCC screening strategies in China.Methods:Questionnaires including information of sex,age,ethnicity,marital status,educational level,source of drinking water,as well as smoking and alcohol consumption history and serum samples were collected from females aged 45-64 years and males aged 35-64 years in 21 counties from 4 provinces in eastern and central China between 2015 and 2023.Enzyme-linked immunosorbent assay methods were used to detect the serum HBV marker HBsAg.Results:A total of 603,082 individuals were enrolled,and serum samples were collected for analysis from January 1,2015 to December 31,2023.The prevalence of HBsAg positive in the study population was 5.23%(31,528/603,082).The prevalence of HBsAg positive was greater in males than in females(5.60%[17,660/315,183]vs 4.82%[13,868/287,899],χ^(2)=187.52,P<0.0001).The elderly participants exhibited a greater prevalence of HBV infection than younger participants(χ^(2)=41.73,P<0.0001).Birth cohort analysis revealed an overall downward trend in HBV prevalence for both males and females.Individuals born in more recent cohorts exhibited a lower prevalence of HBV infection as compared to those born earlier.Conclusions:The current prevalence of HBV infection remains above 5%in populations eligible for HCC screening in China.Further efforts should be made to increase the accessibility of HCC screening among individuals with HBV infection.

Hepatitis associated with hepatitis B virus in broilers

Infection by hepatitis B virus （HBV） results in acute and chronic liver damages in humans. Liver products of broilers as a primary food consumed in our daily life have a close connection with public health. The prevalence of the virus in livers and serum of broilers is of great significance, owning to the potential transmission between chickens and humans. Liver tissues and serum samples were tested to investigate the prevalence of hepatitis B virus infection in slaughtered broilers, for expression of HBV antigens and antibodies. The distribution and positive rate of hepatitis B surface antigen （HBsAg）, hepatitis B core antigen （HBcAg） and hepatitis B e antigen （HBeAg） in liver samples were examined using immunohistochemistry. HBsAg was mainly located in the cytoplasm of hepatocytes with a positivity of 81.61% whereas HBeAg and HBcAg were primarily located in the nucleus of hepatocytes with a positivity of 40.13 and 49.10%, respectively. Enzyme-linked immunosorbent assay （ELISA） analysis of serum for HBV serological markers demonstrated a high prevalence of hepatiits B surface antibody （HBsAb, 54.91%） and hepatitis B core antibody （HBcAb, 27.68%）, whereas HBeAb, HBsAg and HBeAg were rarely detectable. Classic hepatitis pathological changes, including swollen hepatocytes, focal parenchymal necrosis, lymphocytic infiltration and hyperplasia of fibrous connective tissues were observed using histopathological analysis. Some of the liver samples were found positive for HBV DNA using nested PCR. Sequence comparison confirmed that all sequences shared 97.5-99.3% identity with human HBV strains. These results demonstrated the existence of HBV in livers and serums of broilers. Animals or animal products contaminated with HBV could raise an important public health concern over food safety and zoonotic risk.

HBV DNA与乙肝五项定量在不同年龄段乙肝病毒感染患者病情进展中的意义

目的探讨乙肝病毒(HBV)DNA与乙肝五项定量[包括乙肝表面抗原(HBsAg)、乙肝表面抗体(HBsAb)、乙肝e抗原(HBeAg)、乙肝e抗体(HBeAb)、乙肝核心抗体(HBcAb)]在HBV感染进程中各个阶段的差异,从而为慢性乙型肝炎、乙肝肝硬化、肝癌不同阶段的诊治提供参考依据。方法选择2020年1月至2021年12月收治的295例HBV感染患者为研究对象,根据年龄将其分为青年组(≤45岁,105例)、中年组(46~59岁,128例)和老年组(≥60岁,62例)。比较三组慢性乙型肝炎、乙肝肝硬化、肝癌患者的HBV DNA及乙肝五项定量。结果三组慢性乙型肝炎、乙肝肝硬化和肝癌患者的年龄比较,差异无统计学意义(P>0.05)。青年组和中年组中,慢性乙型肝炎、乙肝肝硬化和肝癌患者的HBV DNA、HBsAg、HBeAg水平比较,差异具有统计学意义(P<0.05);慢性乙型肝炎患者的HBV DNA、HBsAg、HBeAg水平明显高于乙肝肝硬化和肝癌患者(P<0.05)。老年组中,慢性乙型肝炎、乙肝肝硬化和肝癌患者的HBV DNA、HBsAg、HBsAb、HBeAg、HBeAb、HBcAb水平比较,差异无统计学意义(P>0.05)。结论青中年患者中,随着慢性乙型肝炎向乙肝肝硬化、肝癌的病程进展,HBV DNA、HBsAg、HBeAg水平均呈现下降趋势,监测相关指标趋势对于提示疾病发展具有一定意义。

乙型肝炎病毒(HBV)的ELISA检测及化学发光法检测的诊断价值

目的 本研究探讨化学发光法与酶联免疫吸附法在(HBV)中的诊断价值。方法 收集本院2020年6月—2023年4月收治的300例乙型病毒性肝炎病例的血标本为观察组,另随机选择同期住院的300例非乙型病毒性肝炎病例的血标本为对照组。两组均采用HBV的ELISA检测及化学发光法检测,对比两组检测结果。结果化学发光法对HBV的标准曲线相关系数R2=0.998,平均批内变异系数2.98%,批间变异系数4.20%,优于ELISA检测的平均批内变异系数6.27%及平均批间变异系数7.74%;化学发光法的最低检出浓度为2.5ng/mL,ELISA检测乙型肝炎病毒最低检出浓度为25ng/mL,化学发光法灵敏度优于ELISA检测法。ELISA检测阳性率明显低于化学发光法检测结果,阴性率高于化学发光法。结论 化学发光法有利于早期诊断乙型病毒性肝炎,较ELISA检测具有很高的灵敏度与特异度。

Serum hepatitis B core-related antigen as a surrogate marker of hepatitis B e antigen seroconversion in chronic hepatitis B

BACKGROUND Quantitative hepatitis B core-related antigen(qHBcrAg)has a better correlation with intrahepatic hepatitis B virus(HBV)covalently closed circular DNA(cccDNA)than HBV DNA or hepatitis B e antigen(HBeAg),but data are still lacking for its clinical application.AIM The aim was to investigate serum qHBcrAg levels in patients with chronic hepatitis B and assess the correlation of serum qHBcrAg with pregenomic RNA(pgRNA),cccDNA,and HBeAg seroconversion.METHODS This study was a secondary analysis of patients who underwent percutaneous liver biopsy between July 2014 and June 2019 in two multicenter randomized controlled clinical trials of peginterferon vs nucleos(t)ide analog(NUC)-based therapy(NCT03509688 and NCT03546530).Serum qHBcrAg,pgRNA,HBV DNA,hepatitis B core antigen,HBeAg,liver cccDNA,and HBV DNA were measured.The correlations of serum qHBcrAg with other biomarkers were analyzed.RESULTS A total of 139 patients were included.The mean qHBcrAg levels were 5.32±1.18 log10 U/mL at baseline and decreased during treatment(all P<0.0001).Serum qHBcrAg levels were positively correlated with pgRNA(r=0.597,P<0.0001)and cccDNA(r=0.527,P<0.0001)levels.The correlation of serum qHBcrAg level and intrahepatic HBV DNA levels at baseline was weak but significant(r=0.399,P<0.0001).HBcrAg predicted HBeAg seroconversion,with areas under the receiver operating characteristics curve of 0.788 at 24 wk and 0.825 at 48 wk.Log HBcrAg at wk 24 and 48 was independently associated with HBeAg seroconversion[odds ratio(OR)=2.402,95%confidence interval(CI):1.314-4.391,P=0.004;OR=3.587,95%CI:1.315-9.784,P=0.013].CONCLUSION Serum HBcrAg levels were correlated with HBV virological markers and could be used to predict HBeAg seroconversion.

Chronic hepatitis B liver disease in patients living in the Amazon region: S gene mutations and genotypes characterization

The Amazon region is considered to be a high endemic area for Hepatitis B Virus (HBV) infections, Rond&ocirc;nia state having the highest prevalence. The aim of this study was to identify molecular genotypes and mutations in the S gene region of HBV viral genomes from 20 patients using a DNA microarray. Results: Serological tests showed that 88% of patients were HBeAg negative, 82% had anti-HBe antibodies and 33% were co-infected with Hepatitis Delta Virus. Sixteen percent of the patients were considered cirrhotic, and 11% have been transfused. The microarray technique identified the genotypes A (4 patients), D (7 patients) and F (7 patients) in 18 samples. Mutations were detected in all 3 genotypes and, overall, A159G, which has been associated with a reduced antigenicity of the virus, was detected most frequently. In genotype A, G119E was the most frequently detected mutation followed by mutations A159G, F134Y, W172C, Y161F and T143S. A159G was detected in all genotype D and F samples followed by mutations T143S, Y161F, N131T, T114S and G119E in genotype D and mutations T143S, Y161F, N131T, T114S and G119E in genotype F. Conclusion: The analysis of mutations repartition among genotypes suggests that some of them are preferentially or exclusively associated with genotype A, D or F. This type of tool is adapted for clinical and therapy monitoring of patient as well as for molecular epidemiology research on HBV.

On-treatment quantitative hepatitis B e antigen predicted response to nucleos(t)ide analogues in chronic hepatitis B

AIM To investigate potential predictors for treatment response to nucleos(t)ide analogues(NAs) in hepatitis B e antigen(HBe Ag)-positive chronic hepatitis B(CHB) patients.METHODS Seventy-six HBeA g-positive CHB patients received 96-wkNAs optimized therapy(lamivudine and adefovir dipivoxil) were studied retrospectively. Serum hepatitis B surface antigen, HBeA g, hepatitis B core antibody, hepatitis B virus(HBV) DNA and alanine aminotransferase levels were quantitatively measured before and during the treatment at 12 and 24 wk. Stepwise logistic regression analyses were performed to identify predictors for treatment response, and areas under the receiver operating characteristic curves(AUROC) of the independent predictors were calculated.RESULTS Forty-three CHB patients(56.6%) achieved virological response(VR: HBV DNA ≤ 300 copies/mL) and 15 patients(19.7%) developed HBeA g seroconversion(SC) after the 96-wk NAs treatment. The HBe Ag level(OR = 0.45, P = 0.003) as well as its declined value(OR = 2.03, P = 0.024) at 24-wk independently predicted VR, with the AUROC of 0.788 and 0.736, respectively. The combination of HBe Ag titer < 1.3 lg PEIU/mL and its decreased value > 1.6 lg PEIU/mL at 24-wk predicted VR with a sensitivity, specificity, positive predictive value(PPV), negative predictive value(NPV) of 85%, 100%, 100% and 83%, respectively, and the AUROC increased to 0.923. The HBeA g level(OR = 0.37, P = 0.013) as well as its declined value(OR = 2.02, P = 0.012) at 24-wk also independently predicted HBeA g SC, with the AUROC of 0.828 and 0.814, respectively. The HBe Ag titer <-0.5 lg PEIU/mL combined with its declined value > 2.2 lg PEIU/mL at 24-wk predicted HBeA g SC with a sensitivity, specificity, PPV, NPV of 88%, 98%, 88% and 98%, respectively, and the AUROC reached 0.928.CONCLUSION The combination of HBeA g level and its declined value at 24-wk may be used as a reference parameter to optimize NAs therapy.

血清HBsAg和HBV DNA预测慢性乙型肝炎肝组织病理状态的评价

目的 探讨血清乙型肝炎表面抗原（HBsAg）水平和乙型肝炎病毒DNA（HBV DNA）载量预测慢性乙型肝炎（CHB）肝组织炎症活动度和纤维化程度的效能.方法 472例经肝组织活检的CHB患者入选本研究,其中HBeAg阳性279例,HBeAg阴性193例.肝组织病理学分级≥G2、≥G3、≥G4分别被定义为显著炎症、严重炎症和进展期炎症,病理学分期≥S2、≥S3和≥S4分别被定义为显著纤维化、严重纤维化和进展期纤维化.结果 HBeAg阳性患者血清HBsAg在G1与G3、G2与G3、S1与S4、S2与S4、S3与S4之间的差异均有统计学意义（P均＜0.05）,血清HBV DNA载量在S1与S4、S2与S4、S3与S4之间的差异均有统计学意义（P均＜0.05）;HBeAg阴性患者血清HBsAg在肝组织不同病理学分级和分期之间的差异无统计学意义（P＞0.05）,血清HBV DNA载量在G1与G3、S1与S2、S1与S3、S1与S4之间的差异均有统计学意义（P均＜0.05）.HBeAg阳性患者血清HBsAg诊断严重炎症和进展期纤维化的ROC曲线下面积分别为0.711 （95％ CI：0.647～0.775）和0.765（95％ CI：0.707～0.823）,血清HBV DNA诊断严重炎症和进展期纤维化的ROC曲线下面积分别为0.589（95％ CI：0.519 ～0.659）和0.700（95％ CI：0.632 ～0.769）;HBeAg阴性患者血清HBV DNA诊断非显著炎症和非显著纤维化的ROC曲线下面积分别为0.644（95％ CI：0.565 ～0.723）和0.684（95％ CI：0.606～0.761）.结论 血清HBsAg对HBeAg阳性患者肝组织严重炎症和进展期纤维化有一定的预测价值;血清HBV DNA对HBeAg阳性患者肝组织严重炎症和进展期纤维化和对HBeAg阴性患者肝组织非显著炎症和非显著纤维化有一定的预测价值.

化学发光分析法定量检测住院患者HBV血清标志物及临床意义

目的获得西安地区住院患者血清乙型肝炎病毒血清标志物的流行病学资料,为医院院内感染管理以及医护人员的防护提供依据。方法利用Abott Architecti 4000SR(i4000SR)化学发光分析仪定量检测2015年10 593例住院病人血清中乙型肝炎病毒表面抗原(HBsAg)、乙型肝炎病毒表面抗体(HBsAb)、乙型肝炎病毒e抗原(HBeAg)、乙型肝炎病毒e抗体(HBeAb)和乙型肝炎病毒核心抗体(HBcAb),其中男性5 248例,女性5 345例。结果 HBV感染率为7.01%(743/10 593),受检者血清中HBV五项检测结果共有14种模式,其中感染期模式HBsAg,HBeAb和HBcAb均为阳性(模式3)占5.17%(548/10 593),HBsAg,HBeAg和HBcAb均为阳性(模式2)占1.34%(142/10 593),HBsAg和HBcAb均为阳性(模式4)占0.25%(27/10 593),其他非常见感染期模式占0.25%(26/10 593);恢复期模式中HBsAb阳性占21.02%(2227/105 93),HBsAb,HBeAb和HBcAb均阳性(模式6)占13.71%(1 452/10 593),HBsAb和HBcAb均阳性占15.07%(1 596/10 593);五项指标皆阴性的占31.38%(3 324/10 593)。不同性别和不同年龄段的患者中HBV血清标志物结果模式阳性率差异均有统计学意义(P<0.05);感染模式2比例最高的科室为消化内科7.39%(36/487),感染模式3比例最高的科室为消化内科16.43%(80/487),HBsAb阳性比例最高的科室为胸外科89.23%(58/65)。结论了解住院患者乙肝感染特点为管理和控制院内HBV传播和推广乙肝疫苗接种提供了必要的临床资料数据,同时提示需要进一步采取有效措施来降低西安地区HBV的感染率。

高灵敏化学发光法体外HBV转录与复制水平的检测体系

目的:建立一套稳定的化学发光法体外HBV 转录与复制水平检测体系．方法:将人肝癌细胞株HepG2和可产生复制型HBV颗粒的HepG2．2．15细胞培养3 d,收集细胞均分为两份,分别抽提细胞总RNA 和HBV复制中间体DNA．以地高辛标记的 HBV重组质粒pHBV4．1作为探针,分别进行 Northern及Southern吸印杂交,以化学发光法检测杂交结果．将pHBV4．1以对数级差稀释后分别点样于尼龙膜用作内标,同时进行杂交及检测．结果:HepG2．2．15细胞提取物中检测出较强的HBV转录产物:3．5 kb,2．4／2．1 kb HBV mRNA的信号,及较强的HBV复制产物:HBV 复制中间体DNA的信号．HepG2细胞提取物检测结果均为阴性．内标检测信号强度随点样浓度逐渐递减而减弱,检测的灵敏度可达到 1 Pg,接近同位素法检测的敏感度．整个实验重复3次以上结果均相似．结论:成功建立了一套稳定的高灵敏化学发光法体外转录与复制水平检测体系．

年龄和血清HBsAg、HBV DNA预测慢性乙型肝炎肝组织病理状态的研究

目的构建基于年龄和血清HBs Ag、HBV DNA诊断慢性乙型肝炎肝组织不同病理状态的Logistic回归模型,优化血清HBs Ag、HBV DNA诊断肝组织不同病理状态的效能。方法经肝组织活检的慢性乙型肝炎患者472例,其中HBe Ag阳性279例,HBe Ag阴性193例。血清HBs Ag和HBe Ag采用Abbott Architect I2000及其配套试剂检测,血清HBV DNA采用实时荧光定量PCR检测。统计分析采用SPSS 13.0软件。结果 HBe Ag阳性患者的血清HBs Ag和HBV DNA与病理学分级和分期均呈显著负相关(P<0.05);HBe Ag阴性患者的血清HBV DNA与病理学分级和分期呈显著正相关(P<0.01)。预测HBe Ag阳性和阴性患者不同病理状态的回归模型的预测概率诊断不同病理状态的ROC曲线下面积均显著大于对角参考线下面积(P<0.01)。对HBe Ag阳性患者,预测进展期纤维化的回归模型的预测概率和血清HBs Ag诊断进展期纤维化的最佳截断值分别为≥0.185和≤3.797 log10IU/ml,其对应的灵敏度、特异度、准确度分别为0.886、0.646、0.706和0.800、0.660、0.695;对HBe Ag阴性患者,预测显著纤维化的回归模型的预测概率和血清HBV DNA诊断显著纤维化的最佳截断值分别为≥0.603和≥3.095 log10IU/m L,其对应的灵敏度、特异度、准确度分别为0.636、0.720、0.668和0.669、0.653、0.663。结论基于年龄和血清HBs Ag、HBV DNA构建的Logistic回归模型可提升血清HBs Ag、HBV DNA诊断肝组织不同病理状态的效能。

Au纳米颗粒HBV DNA基因探针的制备及其在目视化检测HBV DNA中的应用

制备金(gold,Au)纳米颗粒乙型肝炎病毒脱氧核糖核酸(hepatitis B virus deoxynucleic acid,HBV DNA)基因探针,研究其在目视化检测HBV DNA中的应用。Au纳米颗粒通过金-硫(Au-S)共价键结合烷氢硫醇修饰的寡核苷酸,制备检测探针。用荧光标记法检测Au纳米颗粒表面寡核苷酸的覆盖率和与其互补的寡核苷酸的杂交效率。检测探针与固定在尼龙膜上的捕捉探针构成双探针,用斑点杂交法检测HBV DNA,加入银离子(Ag+)-对苯二酚液染色观察结果。制备的Au纳米颗粒粒径为(12±5)nm,分散良好,在520nm有最大吸收峰,经寡核苷酸修饰后,最大吸收峰改变为524nm。Au纳米颗粒表面寡核苷酸的最大覆盖率为(132±10)条,最大杂交效率为(22±3)%。在尼龙膜上用斑点杂交法可检出低至10fmol的合成靶DNA,可目视化检出乙肝患者血清中HBV DNA的PCR产物。Au纳米颗粒HBV DNA基因探针可用于目视化检测HBV DNA,此方法具有敏感性高、特异性好、简单、价廉的特点,可望在许多领域,尤其是在多基因检测芯片上有潜在的应用价值。

不同方法抽提血清HBVDNA得率的比较

目的 比较不同的HBVDNA抽提方法对PCR产物量的影响。方法 将HBVDNA阳性血清及投入了HBVDNA质粒的HBVDNA阴性血清 ,分别采用 7种不同方法抽提核酸。抽提物作PCR后 ,产物行琼脂糖凝胶电泳 ,并对阳性扩增条带进行密度定量。结果 血清直接煮沸法、经典的蛋白酶裂解加酚 /氯仿抽提法、蛋白酶裂解加酚 /氯仿抽提法省缺乙醇沉淀和柱抽提法的HBVDNA抽提得率分别为 75 .2 %、13.8%、2 1.9%和 31.0 %;用碱变性裂解和蛋白酶裂解后煮沸所得上清液 ,以及血清直接作为模板 ,行PCR后不能得到阳性扩增条带。结论 核酸抽提方法选择不当能直接影响基因检测的灵敏度 ,导致基因定量准确性下降。血清直接煮沸法抽提HBVDNA得率高、操作简便、省时和经济 ,值得推荐。

慢性乙型肝炎患者HBsAg、HBeAg及HBV-DNA定量结果的临床意义与相关性分析

目的研究慢性乙型肝炎患者HBsAg、HBeAg、HBV-DNA、ALT定量检测结果之间的相关性与临床意义。方法采用电化学发光法检测HBsAg、HBeAg,实时荧光定量聚合酶链反应检测HBV-DNA,化学速率法检测ALT。结果 HBeAg阳性组ALT血清浓度、ALT异常率均高于HBeAg阴性组(P<0.05),但HBsAg浓度低于HBeAg阴性组(P<0.05)。HBeAg阳性组组内结果分析:HBeAg血清浓度与HBV-DNA在免疫耐受期成正相关(r=0.54),HBsAg与HBeAg、HBV-DNA成负相关(r=-0.521 2、-0.462 7),HBsAg、HBV-DNA、HBeAg与ALT之间均无相关性(r=0.008、0.02、0.000 6)。HBeAg阴性组组内结果分析:HBV-DNA与HBsAg无相关性(r=-0.05),HBV-DNA与ALT成正相关(r=0.39),HBsAg与ALT无相关性(r=0.008)。结论 HBV-DNA、HBsAg定量、HBeAg定量检测虽然可以反映HBV复制状态,但并不能代表乙肝患者的病情演变,只有定期长时间的监测才有利于患者病情的管理和早期发现病毒状态的变化。

阜阳市人群HBV感染的血清流行病学调查

目的了解某地自然人群乙型肝炎病毒（HBV）感染情况。方法调查2005年3—12月在某疾病预防控制中心进行健康体检人群的HBV血清标志物检测结果。HBV血清标志物检测采用酶联免疫吸附试验。结果单纯检测乙型肝炎表面抗原（HBsAg）的2802人中，阳性255人，阳性率9．10％；各年龄组人群以31～40岁组HBsAg阳性率最高，占12．16％（71／584）。1907例检测HBV 5项血清标志物者中，HBsAg阳性者238例（12．48％），以HBsAg阳性合并抗HBe、抗HBc阳性为主（44．96％），HBsAg阳性合并HBeAg、抗HBc阳性次之（38．24％）；5项全阴性者占总人群的46．88％。结论该地区自然人群HBV感染水平较高，近一半人没有保护性抗体，应加强健康宣教，推广普及乙肝疫苗的接种，改善环境卫生。