Roche公司灵活式自动化在实验室中的应用

自动化之所以成为实验室发展的首选方案,在于它可以改善工作流程从而提高工作效率,同时减少人工介入环节,保证结果的准确性。一些研究机构对实验室流程比例进行研究发现,样品前处理占了整个实验室一半的资源,样品分析中和分析后占了实验室工作人员15%～20%的时间和精力，

环境友好的无氟制冷剂——Roche诊断公司研发成功使用天然制冷剂的冷冻干燥设备

尽管已经有一些制冷剂能够替代产生温室气体效应的氟里昂，但它们总是不能可靠地避免引起温室气体效应的可能性。第一种无氟制冷剂的问世才真正地减轻了制冷技术给环境带来的危害，这种无氟制冷剂要求采用特殊的制冷技术。在Roche诊断公司的驻地Penzberg市，诞生了一套新型的、真正做到了最佳的环境保护的冷冻干燥设备。

IMpower210:A phase Ⅲ study of second-line atezolizumab vs. docetaxel in East Asian patients with non-small cell lung cancer

Objective: IMpower210(NCT02813785) explored the efficacy and safety of single-agent atezolizumab vs.docetaxel as second-line treatment for advanced non-small cell lung cancer(NSCLC) in East Asian patients.Methods: Key eligibility criteria for this phase Ⅲ, open-label, randomized study included age ≥18 years;histologically documented advanced NSCLC per the Union for International Cancer Control/American Joint Committee on Cancer staging system(7th edition);Eastern Cooperative Oncology Group performance status of 0 or 1;and disease progression following platinum-based chemotherapy for advanced or metastatic NSCLC. Patients were randomized 2:1 to receive either atezolizumab(1,200 mg) or docetaxel(75 mg/m^(2)). The primary study endpoint was overall survival(OS) in the intention-to-treat(ITT) population with wild-type epidermal growth factor receptor expression(ITT EGFR-WT) and in the overall ITT population.Results: Median OS in the ITT EGFR-WT population(n=467) was 12.3 [95% confidence interval(95% CI),10.3-13.8] months in the atezolizumab arm(n=312) and 9.9(95% CI, 7.8-13.9) months in the docetaxel arm[n=155;stratified hazard ratio(HR), 0.82;95% CI, 0.66-1.03]. Median OS in the overall ITT population was 12.5(95% CI, 10.8-13.8) months with atezolizumab treatment and 11.1(95% CI, 8.4-14.2) months(n=377) with docetaxel treatment(n=188;stratified HR, 0.87;95% CI, 0.71-1.08). Grade 3/4 treatment-related adverse events(TRAEs) occurred in 18.4% of patients in the atezolizumab arm and 50.0% of patients in the docetaxel arm.Conclusions: IMpower210 did not meet its primary efficacy endpoint of OS in the ITT EGFR-WT or overall ITT populations. Atezolizumab was comparatively more tolerable than docetaxel, with a lower incidence of grade3/4 TRAEs.

Predicting short-term thromboembolic risk following Roux-en-Y gastric bypass using supervised machine learning

BACKGROUND Roux-en-Y gastric bypass(RYGB)is a widely recognized bariatric procedure that is particularly beneficial for patients with class III obesity.It aids in significant weight loss and improves obesity-related medical conditions.Despite its effectiveness,postoperative care still has challenges.Clinical evidence shows that venous thromboembolism(VTE)is a leading cause of 30-d morbidity and mortality after RYGB.Therefore,a clear unmet need exists for a tailored risk assessment tool for VTE in RYGB candidates.AIM To develop and internally validate a scoring system determining the individualized risk of 30-d VTE in patients undergoing RYGB.METHODS Using the 2016–2021 Metabolic and Bariatric Surgery Accreditation Quality Improvement Program,data from 6526 patients(body mass index≥40 kg/m^(2))who underwent RYGB were analyzed.A backward elimination multivariate analysis identified predictors of VTE characterized by pulmonary embolism and/or deep venous thrombosis within 30 d of RYGB.The resultant risk scores were derived from the coefficients of statistically significant variables.The performance of the model was evaluated using receiver operating curves through 5-fold cross-validation.RESULTS Of the 26 initial variables,six predictors were identified.These included a history of chronic obstructive pulmonary disease with a regression coefficient(Coef)of 2.54(P<0.001),length of stay(Coef 0.08,P<0.001),prior deep venous thrombosis(Coef 1.61,P<0.001),hemoglobin A1c>7%(Coef 1.19,P<0.001),venous stasis history(Coef 1.43,P<0.001),and preoperative anticoagulation use(Coef 1.24,P<0.001).These variables were weighted according to their regression coefficients in an algorithm that was generated for the model predicting 30-d VTE risk post-RYGB.The risk model's area under the curve(AUC)was 0.79[95%confidence interval(CI):0.63-0.81],showing good discriminatory power,achieving a sensitivity of 0.60 and a specificity of 0.91.Without training,the same model performed satisfactorily in patients with laparoscopic sleeve gastrectomy with an AUC of 0.63(95%CI:0.62-0.64)and endoscopic sleeve gastroplasty with an AUC of 0.76(95%CI:0.75-0.78).CONCLUSION This simple risk model uses only six variables to assist clinicians in the preoperative risk stratification of RYGB patients,offering insights into factors that heighten the risk of VTE events.

Pulsed export of carbon in the north-western Mediterranean Sea

The short term(hourly scale)variability of heterotrophic prokaryote(HP)vertical distribution and respiratory activity,was investigated in the north-western(NW)Mediterranean Sea.HP vertical distribution was determined on board by flow cytometry analysis of seawater samples collected by series of CTD casts.Cell counts and viability were determined for all samples.HP respiratory rates were determined later in the laboratory from filtered seawater samples(23 dm^(3))from 300-1150-m depth.The average cell viability was 94.8%±2.2%(n=240).There was no accumulation of dead cells,due to quick decay of damaged cells.In the epipelagic layer,three HP groups were distinguished,two(HNA1,HNA2)who se cells exhibited a high nucleic acid content and one(LNA)with low nucleic acid content cells.HNA2 was most populated at 50 m but not detected at 90 m and below,presumably aerobic anoxygenic photoheterotrophic bacteria(AAPs).The variability in HP abundance was mainly confined in the upper 80 m.A few secondary peaks of HP abundance were observed(80-150 m)in connection with abundance troughs in the surface layer.HP cells were continuously present in a wide layer around 500 m(mean 191×10^(3)cells/cm^(3)).Below this layer,HP abundance randomly exhibited peaks,coupled to respiratory rate peaks.The HP abundance and variability in the water column was suppressed during a strong wind event.The observed sporadic variability was tentatively interpreted through a pulsed carbon-export mechanism induced by the microorganism production of dissolved poly saccharide s,followed by flocculation and rapid sinking.This mechanism would thus contribute to(ⅰ)preventing organic matter accumulation in the epipelagic layer,(ⅱ)seeding the water column with live HP cells,and(ⅲ)supplying the aphotic water column with fre sh and labile organic matter.This important vertical flux mechanism needs further observations and modelling.

Evidence of bisphosphonate-conjugated sitafloxacin eradication of established methicillin-resistant S.aureus infection with osseointegration in murine models of implant-associated osteomyelitis

Eradication of MRSA osteomyelitis requires elimination of distinct biofilms.To overcome this,we developed bisphosphonateconjugated sitafloxacin(BCS,BV600072)and hydroxybisphosphonate-conjugate sitafloxacin(HBCS,BV63072),which achieve“target-and-release”drug delivery proximal to the bone infection and have prophylactic efficacy against MRSA static biofilm in vitro and in vivo.Here we evaluated their therapeutic efficacy in a murine 1-stage exchange femoral plate model with bioluminescent MRSA(USA300LAC::lux).Osteomyelitis was confirmed by CFU on the explants and longitudinal bioluminescent imaging(BLI)after debridement and implant exchange surgery on day 7,and mice were randomized into seven groups:1)Baseline(harvested at day7,no treatment);2)HPBP(bisphosphonate control for BCS)+vancomycin;3)HPHBP(hydroxybisphosphonate control for HBCS)+vancomycin;4)vancomycin;5)sitafloxacin;6)BCS+vancomycin;and 7)HBCS+vancomycin.BLI confirmed infection persisted in all groups except for mice treated with BCS or HBCS+vancomycin.Radiology revealed catastrophic femur fractures in all groups except mice treated with BCS or HBCS+vancomycin,which also displayed decreases in peri-implant bone loss,osteoclast numbers,and biofilm.To confirm this,we assessed the efficacy of vancomycin,sitafloxacin,and HBCS monotherapy in a transtibial implant model.The results showed complete lack of vancomycin efficacy while all mice treated with HBCS had evidence of infection control,and some had evidence of osseous integrated septic implants,suggestive of biofilm eradication.Taken together these studies demonstrate that HBCS adjuvant with standard of care debridement and vancomycin therapy has the potential to eradicate MRSA osteomyelitis.

Novel Method to Deal with Interval Quadratic Equations via Sign-Variation Analysis

In this article, analytical results are obtained apparently for the first time in the literature, for the lower and upper bounds of the roots of quadratic equations when two or all three coefficients a, b, c constitute an interval, with a method called the sign-variation analysis. The results are compared with the parametrization technique offered by Elishakoff and Miglis, and with the solution yielded by minimization and maximization commands of the Maple software. Solutions for some interval word problems are also provided to edulcorate the methodology. This article only focuses on the real roots of those quadratic equations, complex solutions being beyond this investigation.

基于CALS及Surpac-FLAC^3D耦合技术的复杂空区稳定性分析

由于受地质条件和探测技术的限制,传统探测方法很难获得复杂空区精确的实际空间分布形状,由此得到的空区用于建立数值分析模型显得过于粗糙,在此基础上进行空区稳定性分析的可靠性程度不高。采用空区激光自动扫描系统(CALS)对复杂空区进行激光扫描,在获得精准三维点云数据的基础上,运用Surpac软件模拟出空区的三维地质模型,通过耦合方法可以在FLAC3D中生成与实际空区空间分布一致的数值模型,提高复杂空区稳定性计算的准确性和可靠性。选取三道庄露天矿的地下空区作为工程实例,利用CALS获得不规则空区的实际空间边界。在此基础上,运用Surpac进行三维块体模拟,研究Surpac与FLAC3D模型耦合技术,成功地将三维模型数据导入FLAC3D中,结合现场实际勘测的围岩力学参数,对空区进行静力计算,分析空区的稳定性。

曲格列酮对大鼠肝细胞氧化性损伤的研究

目的:研究曲格列酮的肝细胞毒性及其可能的机制。方法:在原代大鼠肝细胞模型和其他3种不同来源非肝细胞系,用WST-1法、乳酸脱氢酶释放法、荧光检测法、生物发光法等方法,观察曲格列酮对肝细胞的损伤作用。结果:不同剂量曲格列酮作用后,其在原代培养大鼠肝细胞的IC_(50)值最低,即其对原代肝细胞毒性最大。曲格列酮诱导活性氧(ROS)生成,并引起还原型谷胱甘肽(GSH)和ATP水平比对照细胞显著降低。不同剂量的曲格列酮作用5h后诱发肝细胞凋亡、坏死。结论:曲格列酮对大鼠肝细胞的毒性作用强于对其他非肝细胞来源细胞的毒性作用。曲格列酮可以引起肝细胞氧化性损伤,肝细胞凋亡和坏死。

CYP2D6PCR基因型与DXT表型和基因芯片检测的比较(英文)

目的 :为了评价CYP2D6的基因型和表型的联系以及基因芯片在CYP2D6多基因分析中的应用。方法 :2 4 2健康志愿者 ,口服dextromethorphan后收集尿液测定其代谢率 ,收集 2 0ml血提取DNA ,并通过基因特异性PCR和 / (或 )基因芯片分析CYP2D6 2——— 11, 17和多拷贝CYP2D6基因 ,其中 5个基因 ( 3、 4、 6、 7和 9)用PCR和CYD4 5 0基因芯片同时分析。结果 :CYP2D6基因型比表型更富有信息和更能反映CYP2D6酶的表达。CYP2D6 3、 4、 6、 7和 9的基因检测在CYP4 5 0基因芯片和基因特异性PCR中显示高度的一致性。结论 :基因芯片在检测基因多位点的多基因中是一个有发展前途和可靠的方法。

延长Bench mark VENTANA空气滤清器使用寿命的改进方法

BenchmarkVENTANA是一台由Roche公司推出的集免疫组化和cifsh分子染色的全自动高端染色仪器，全自动免疫组化仪现已在各大医院广泛应用。免疫组化与cifsh分子染色是现代临床病理治疗及预后治疗综合评分的重要指标，

VRML在交互式可视化药学教学中的应用

本文探讨了将 VRML应用于交互式可视化药学教学的可行性及用 VRML实现化学结构多样性可视化的手段。尝试在改变我国传统药学教学方式。

在药品研发及商业化全程实施药品安全科学管理规范(三)

为系统阐述和研究药品安全科学,全面引入药品风险管理理念,国家药品不良反应监测中心特邀请国外相关专家共同撰写了《在药品研发及商业化全程实施药品安全科学管理规范》一文,并将其译成中文以共享。

Peginterferon alfa-2a for the treatment of chronic hepatitis C in the era of direct-acting antivirals

BACKGROUND: The availability of novel direct-acting antivirals (DAAs) represents a new era of curative hepatitis C virus (HCV) treatment, with over 95% of patients infected with HCV genotype 1 achieving sustained virological response (SVR). Nevertheless, the majority of patients globally are unable to access these treatments because of cost and infrastructure constraints and, thus, remain untreated and uncured. DATA SOURCE: Relevant articles of peginterferon (PegIFN)-based treatments in HCV and sofosbuvir-based treatments, simeprevir, daclatasvir/asunaprevir, ritonavir-boosted paritaprevir/ombitasvir/dasabuvir, and grazoprevir/elbasvir, were searched in PubMed database, including general population and special population. RESULTS: PegIFN in combination with ribavirin remains an important and relevant option for some patients, achieving SVR rates of up to 79% in genotype 1 and 89% in genotype 2 or 3 infections, which increases for patients with favorable IL28B genotypes. Triple therapy of DAA plus PegIFN/ribavirin is effective in treating difficult-to-cure patients infected with HCV genotype 3 or with resistance-associated variants. Owing to its long history in HCV management, the efficacy, tolerability and long-term outcomes associated with PegIFN alfa-2a are well established and have been validated in large-scale studies and in clinical practice for many populations. Furthermore, emerging data show that IFN-induced SVR is associated with lower incidences of hepatocellular carcinoma compared with DAAs. On the contrary, novel DAAs have yet to be studied in special populations, and long-term outcomes, particularly tumor development and recurrence in patients with cirrhosis and/or hepatocellular carcinoma, and reactivation of HBV in dually infected patients, are still unclear. CONCLUSION: In this interferon-free era, PegIFN-based regimens remain a safe and effective option for selected HCV patients.

Osteopontin as potential biomarker and therapeutic target in gastric and liver cancers

Gastric cancer and liver cancer are among the most common malignancies and the leading causes of death worldwide,due to late detection and high recurrence rates.Today,these cancers have a heavy socioeconomic burden,for which a full understanding of their pathophysiological features is warranted to search for promising biomarkers and therapeutic targets.Osteopontin (OPN) is overexpressed in most patients with gastric and liver cancers.Over the past decade,emerging evidence has revealed a correlation of OPN level and clinicopathological features and prognosis in gastric and liver cancers,indicating its potential as an independent prognostic indicator in such patients.Functional studies have verified the potential of OPN knockdown as a therapeutic approach in vitro and in vivo .Furthermore,OPN mediates multifaceted roles in the interaction between cancer cells and the tumor microenvironment,in which many details need further exploration.OPN signaling results in various functions,including prevention of apoptosis,modulation of angiogenesis,malfunction of tumor-associated macrophages,degradation of extracellular matrix,activation of phosphoinositide 3-kinase-Akt and nuclear factor-κB pathways,which lead to tumor formation and progression,particularly in gastric and liver cancers.This editorial aims to review recent findings on alteration in OPN expression and its clinicopathological associations with tumor progression,its potential as a therapeutic target,and putative mechanisms in gastric and liver cancers.Better understanding of the implications of OPN in tumorigenesis might facilitate development of therapeutic regimens to benefit patients with these deadly malignancies.

Tumor necrosis factor-alpha -308G/A polymorphism and risk of hepatocellular carcinoma in hepatitis C virus-infected patients

Tumor necrosis factor-alpha (TNF-α) is an important cytokine in generating an immune response against infection with hepatitis C virus (HCV). The functions of TNF-α may be altered by single-nucleotide polymorphisms (SNPs) in its gene structure. We hypothesized that SNPs in TNF-α may be important in determining the outcome of an HCV infection. To test this hypothesis, we investigated the role of the polymorphism -308G/A, which is located in the promoter region of the TNF-α gene, in the progression of HCV infection in Egyptian patients using a quantitative real-time polymerase chain reaction (qRT-PCR). The distribution of this polymorphism and its impact on the serum level of TNF-α was compared between 90 HCV-infected patients [45 with HCV-induced cirrhosis and 45 with HCV-related hepatocellular carcinoma (HCC)] and 45 healthy Egyptian volunteers without any history of liver disease. Our results showed that at the TNF-α -308 position, the G/G allele was most common (78.5% ) in the study population, with the G/A and A/A alleles occurring less frequently (13.3% and 8.1% , respectively). Frequencies of G/G, G/A, and A/A genotypes were 87%, 7%, and 6% in patients with liver cirrhosis and were 94% , 4% , and 2% in patients with HCC, respectively. Serum levels of TNF-α were significantly higher in HCV-infected patients than in healthy controls, indicating that the TNF-α -308 polymorphism does not influence the production of TNF-α. The serum level of TNF-α was positively correlated with HCV infection. Taken together, these findings suggest that the TNF-α -308 polymorphism may not be a host genetic factor associated with the severity of HCV infection, but may be an independent risk factor for HCC.

Capecitabine treatment patterns in patients with gastroesophageal cancer in the United States

AIM:To assess the use of capecitabine-based therapy and associated complication rates in patients with gastroesophageal cancer(GEC)in a real-world treatment setting. METHODS:Patients with claims between 2004 and 2005 were identified from the Thomson Reuters MarketScan  databases.Capecitabine regimens were compared with 5-fluorouracil(5-FU)and other chemotherapy regimens,and were stratified by treatment setting. RESULTS:We identified 1013 patients with GEC:approximately half had treatment initiated with a 5-FU regimen,whereas 11%had therapy initiated with a capecitabine regimen.The mean capecitabine dose overall was 2382±1118 mg/d,and capecitabine was used as monotherapy more often than in combination. Overall,5-FU regimens were the most common treatment option in neoadjuvant and adjuvant settings, while other non-capecitabine regimens were used more widely in first-and second-line settings.The overall unadjusted complication rate for capecitabine regimens was about half of that seen with 5-FU regimens.In multivariate analyses,capecitabine recipients had a 51%(95%CI:26%-81%)lower risk of developing any complication than 5-FU recipients did.The risk of developing bone marrow,constitutional,gastrointestinal tract,infectious,or skin complications was lowerwith capecitabine therapy than with 5-FU.CONCLUSION:Capecitabine appeared to have a favorable side effect profile compared with 5-FU,which indicates that it may be a treatment option for GEC.

一项在欧洲和中国进行的新型胃泌素释放肽前体(ProGRP)免疫检测多中心评估研究

背景在欧洲和中国进行Elecsys?胃泌素释放肽前体(ProGRP)免疫检测的多中心评估研究。方法在欧洲的3个中心和中国的2个中心,在肺癌中,通过不精密度、稳定性、方法学比较和鉴别诊断能力来评价该检测法。结果 5个分析物浓度的中间不精密度范围为变异系数:2.2%-6.0%。在不同储存条件下,血浆和血清样本均显示出良好的稳定性。在血浆中Elecsys?和ARCHITECT检测(斜率1.02,截距-2.72 pg/m L)之间表现出良好的相关性。同时,Elecsys?检测在血清和血浆样本之间表现出良好的相关性(斜率0.93,截距2.35 pg/m L;相关系数0.97)。ProGRP作为不受种族、年龄、性别或吸烟史相关影响的检测手段,可鉴别小细胞和非小细胞肺癌(NSCLC);截断值为84pg/m L时,曲线下面积为0.90,95%CI:0.87-0.93;敏感性为78.3%,特异性为95%。ProGRP浓度中位数在良性病变(38pg/m L)、其他恶性肿瘤(40 pg/m L)或NSCLC(39 pg/m L)中较低,而在3期以上慢性肾脏疾病中浓度较高(>100pg/m L)。结论 Elecsys?ProGRP检测在血清和血浆中稳定性增加,较现有检测法明显更具优势。ProGRP检测在中国的首次评价在不同种族中显示出相当的鉴别能力。

Detection of hyper-conserved regions in hepatitis B virus X gene potentially useful for gene therapy

AIM To detect hyper-conserved regions in the hepatitis B virus(HBV) X gene(HBX) 5' region that could be candidates for gene therapy.METHODS The study included 27 chronic hepatitis B treatmentnaive patients in various clinical stages(from chronic infection to cirrhosis and hepatocellular carcinoma, both HBeA g-negative and HBeA g-positive), and infected with HBV genotypes A-F and H. In a serum sample from each patient with viremia > 3.5 log IU/m L, the HBX 5' end region [nucleotide(nt) 1255-1611] was PCRamplified and submitted to next-generation sequencing(NGS). We assessed genotype variants by phylogenetic analysis, and evaluated conservation of this region by calculating the information content of each nucleotide position in a multiple alignment of all unique sequences(haplotypes) obtained by NGS. Conservation at the HBx protein amino acid(aa) level was also analyzed.RESULTS NGS yielded 1333069 sequences from the 27 samples, with a median of 4578 sequences/sample(2487-9279, IQR 2817). In 14/27 patients(51.8%), phylogenetic analysis of viral nucleotide haplotypes showed a complex mixture of genotypic variants. Analysis of the information content in the haplotype multiple alignments detected 2 hyper-conserved nucleotide regions, one in the HBX upstream non-coding region(nt 1255-1286) and the other in the 5' end coding region(nt 1519-1603). This last region coded for a conserved amino acid region(aa 63-76) that partially overlaps a Kunitz-like domain.CONCLUSION Two hyper-conserved regions detected in the HBX 5' end may be of value for targeted gene therapy, regardless of the patients' clinical stage or HBV genotype.

Characterization of hepatitis B virus X gene quasispecies complexity in mono-infection and hepatitis delta virus superinfection

Hepatitis delta virus(HDV) seems to strongly suppress hepatitis B virus(HBV)replication, although little is known about the mechanism of this interaction. Both these viruses show a dynamic distribution of mutants, resulting in viral quasispecies. Next-generation sequencing is a viable approach for analyzing the composition of these mutant spectra. As the regulatory hepatitis B X protein(HBx) is essential for HBV replication, determination of HBV X gene(HBX)quasispecies complexity in HBV/HDV infection compared to HBV monoinfection may provide information on the interactions between these two viruses.AIM To compare HBV quasispecies complexity in the HBX 5' region between chronic hepatitis delta(CHD) and chronic HBV mono-infected patients.METHODS Twenty-four untreated patients were included: 7/24(29.2%) with HBeAgnegative chronic HBV infection(CI, previously termed inactive carriers), 8/24(33.3%) with HBeAg-negative chronic hepatitis B(CHB) and 9/24(37.5%) with CHD. A serum sample from each patient was first tested for HBV DNA levels.The HBX 5' region [nucleotides(nt) 1255-1611] was then PCR-amplified for subsequent next-generation sequencing(MiSeq, Illumina, United States). HBV quasispecies complexity in the region analyzed was evaluated using incidencebased indices(number of haplotypes and number of mutations), abundancebased indices(Hill numbers of order 1 and 2), and functional indices(mutation frequency and nucleotide diversity). We also evaluated the pattern of nucleotide changes to investigate which of them could be the cause of the quasispecies complexity.RESULTS CHB patients showed higher median HBV-DNA levels [5.4 logIU/mL,interquartile range(IQR) 3.5-7.9] than CHD(3.4 logIU/mL, IQR 3-7.6)(P = n.s.)or CI(3.2 logIU/mL, IQR 2.3-3.5)(P < 0.01) patients. The incidence and abundance indices indicated that HBV quasispecies complexity was significantly greater in CI than CHB. A similar trend was observed in CHD patients, although only Hill numbers of order 2 showed statistically significant differences(CHB2.81, IQR 1.11-4.57 vs CHD 8.87, 6.56-11.18, P = 0.038). There were no significant differences in the functional indices, but CI and CHD patients also showed a trend towards greater complexity than CHB. No differences were found for any HBV quasispecies complexity indices between CHD and CI patients. G-to-A and C-to-T nucleotide changes, characteristic of APOBEC3 G, were higher in CHD and CI than in CHB in genotype A haplotypes, but not in genotype D. The proportion of nt G-to-A vs A-to-G changes and C-to-T vs T-to-C changes in genotype A and D haplotypes in CHD patients showed no significant differences. In CHB and CI the results of these comparisons were dependent on HBV genotype.CONCLUSION The lower-replication CHD and CI groups show a trend to higher quasispecies complexity than the higher-replication CHB group. The mechanisms associated with this greater complexity require elucidation.