Genetic Diversity of Porcine Circovirus 2 in Wild Boar and Domestic Pigs in Ukraine

Porcine circovirus type 2 (PCV2) is responsible for a number of porcine circovirus-associated diseases (PCVAD) that can severely impact domestic pig herds. For a non-enveloped virus with a small genome (1.7 kb ssDNA), PCV2 is remarkably diverse, with eight genotypes (a–h). New genotypes of PCV2 can spread through the migration of wild boar, which are thought to infect domestic pigs and spread further through the domestic pig trade. Despite a large swine population, the diversity of PCV2 genotypes in Ukraine has been under-sampled, with few PCV2 genome sequences reported in the past decade. To gain a deeper understanding of PCV2 genotype diversity in Ukraine, samples of blood serum were collected from wild boars (n = 107) that were hunted in Ukraine during the November–December 2012 hunting season. We found 34/107 (31.8%) prevalence of PCV2 by diagnostic PCR. For domestic pigs, liver samples (n = 16) were collected from a commercial market near Kharkiv in 2019, of which 6 out of 16 (37%) samples were positive for PCV2. We sequenced the genotyping locus ORF2, a gene encoding the PCV2 viral capsid (Cap), for 11 wild boar and six domestic pig samples in Ukraine using an Oxford Nanopore MinION device. Of 17 samples with resolved genotypes, the PCV2 genotype b was the most common in wild boar samples (10 out of 11, 91%), while the domestic pigs were infected with genotypes b and d. We also detected genotype b/d and b/a co-infections in wild boars and domestic pigs, respectively, and for the first time in Ukraine we detected genotype f in a wild boar from Poltava. Building a maximum-likelihood phylogeny, we identified a sublineage of PCV2 genotype b infections in both wild and domestic swine, suggesting a possible epizootic cluster and an ecological interaction between wild boar and domestic pig populations in northeastern Ukraine.     

Molecular Epidemic Characteristics and Genetic Evolution of Porcine Circovirus Type 2 (PCV2) in Swine Herds of Shanghai,China

Porcine Circovirus 2 (PCV2) is a crucial swine pathogen and considered a primary causative agent of porcine circovirus-associated diseases (PCVADs), posing a serious economic threat to the swine industry across globe. The world’s biggest agricultural conglomerates have teamed up to create giant commercial pig farms across Shanghai due to the proximity of this region to more affluent lean-pork markets. Since its discovery, PCV2 has displayed extraordinary genetic diversity, and its genome is swiftly evolving through a series of mutations and recombinations. However, limited information on epidemiology, molecular characteristics, vaccine cross-protection, and the co-infection rate of PCV2 with other lethal swine diseases can adversely impact the pig production in the region. To investigate the molecular epidemic characteristics and genetic evolution of PCV2, pigs with doubtful symptoms of PCVADs were sampled from various commercial pig farms with a history of PWMS and/or PDNS across Shanghai from 2014 to 2018. Our results revealed the coexistence of multiple PCV2 genotypes (PCV2b, PCV2e, and PCV2d) among Shanghai pig herds and dominance of PCV2d among them. We also found critical amino acid substitutions in epitope regions of important capsid proteins in PCV2 isolates involved in viral replication and host immune escape. Spotted mutations may favor the prevalence and survival of various PCV2 genotypes despite availability of commercial vaccines. This study also provides insight into the co-infection status of PCV2 with major lethal swine viral diseases such as PPV and PPRSV. Collectively, these investigations will contribute to understanding the molecular epidemiology and evolution of PCV2 across the region.     

Synergistic Pathogenicity by Coinfection and Sequential Infection with NADC30-like PRRSV and PCV2 in Post-Weaned Pigs

Porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus (PCVs) are two major viruses that affect pigs. Coinfections between PRRSV and PCV2 are frequently reported in most outbreaks, with clinical presentations involving dyspnea, fever, reduced feed intake, weight loss, and death in fattening pigs. The NADC30-like PRRSV and PCV2d are the main circulating virus strains found in China. This study determines the impact of NADC30-like PRRSV and PCV2d mono-infection and coinfection on the immune system, organ pathology, and viral shedding in five-week-old post-weaned pigs. Pigs were randomly divided into six groups: PBS, PRRSV, PCV2, PRRSV-PCV2 coinfection (co), and PRRSV-PCV2 or PCV2-PRRSV sequential infections. Fever, dyspnea, decreased feed intake, weight loss, and pig deaths occurred in groups infected with PRRSV, Co-PRRSV-PCV2, and PRRSV-PCV2. The viral load was higher in Co-PRRSV-PCV2, PRRSV-PCV2, and PCV2-PRRSV than those mono-infected with PRRSV or PCV2. Additionally, cytokines (IFN-γ, TNF-α, IL-4, and IL-10) produced by pigs under Co-PRRSV-PCV2 and PRRSV-PCV2 groups were more intense than the other groups. Necropsy findings showed hemorrhage, emphysema, and pulmonary adhesions in the lungs of pigs infected with PRRSV. Smaller alveoli and widened lung interstitium were found in the Co-PRRSV-PCV2 and PRRSV-PCV2 groups. In conclusion, PRRSV and PCV2 coinfection and sequential infection significantly increased viral pathogenicity and cytokine responses, resulting in severe clinical signs, lung pathology, and death.     

Characterization of a Novel Double-Stranded RNA Virus from Phytophthora pluvialis in New Zealand

A new dsRNA virus from the oomycete Phytophthora pluvialis has been characterized and designated as Phytophthora pluvialis RNA virus 1 (PplRV1). The genome of the PplRV1 reference genome is 6742 bp that encodes two predicted open reading frames (ORFs). ORF1 and ORF2 overlap by a 47 nt “slippery” frameshift sequence. ORF1 encodes a putative protein of unknown function. ORF2 shows high similarity to the RNA-dependent RNA polymerase (RdRp) of other dsRNA viruses. Phylogenetic analysis of the putative PplRV1 RdRp and its most closely related viruses showed PplRV1 is distinct from other known viruses (below 33% amino acid similarity), which indicates this virus may belong to a new virus family. Analyses of the geographical distribution of PplRV1 in relation to two genetically distinct classes of its host revealed two corresponding genotypes of the PplRV1 (termed a and b), which share 92.3% nt identity. The reference genome for the second genotype is 6760 bp long and a prediction of its genetic organization shows three ORFs, with ORF2 being split into two ORFs, ORF2a and ORF2b, that is conserved in seven of eleven genotype b isolates. Additionally, a quick and simple diagnostic method using qPCR has been developed, which is suitable for large scale screens to identify PplRV1 in Phytophthora.     

Porcine TRIM21 Enhances Porcine Circovirus 2 Infection and Host Immune Responses,But Inhibits Apoptosis of PCV2-Infected Cells

Tripartite motif protein 21 (TRIM21) is an interferon-inducible E3 ligase, containing one RING finger domain, one B-box motif, one coiled-coil domain at the N-terminal, as well as one PRY domain and one SPRY domain at the C-terminal. TRIM21 is expressed in many tissues and plays an important role in systemic autoimmunity. However, TRIM21 plays different roles in different virus infections. In this study, we evaluate the relationship between porcine TRIM21 and PCV2 infection as well as host immune responses. We found that PCV2 infection modulated the expression of porcine TRIM21. TRIM21 can enhance interferons and proinflammatory factors and decrease cellular apoptosis in PCV2-infected cells. These results indicate that porcine TRIM21 plays a critical role in enhancing PCV2 infection, which is a promising target for controlling and developing the treatment of PCV2 infection.     

Comparative Characterization and Pathogenicity of a Novel Porcine Epidemic Diarrhea Virus (PEDV) with a Naturally Occurring Truncated ORF3 Gene Coinfected with PEDVs Possessing an Intact ORF3 Gene in Piglets

Coinfection caused by various genotypes of porcine epidemic diarrhea virus (PEDV) is a new disease situation. We previously reported the coexistence of PEDV strains containing different ORF3 genotypes in China. In this study, the PEDV strains 17GXCZ-1ORF3d and 17GXCZ-1ORF3c were isolated and plaque-purified from the same piglet, which had a natural large deletion at the 172–554 bp position of the ORF3 gene or possessed a complete ORF3 gene, respectively. Meanwhile, 17GXCZ-1ORF3d had >99% nt identity with 17GXCZ-1ORF3c in the 5′UTR, ORF1a/1b, S, E, M, N and 3′UTR regions but only demonstrated low nucleotide identities (80.5%) in the ORF3 gene. To elucidate the pathogenicity, 7-day-old piglets were infected. Piglets infected with these two PEDV strains exhibited severe clinical signs and shed the virus at the highest level within 96 hpi. Compared with the piglets inoculated with the 17GXCZ-1ORF3c strain, the piglets inoculated with the 17GXCZ-1ORF3d strain had higher mortality rates (75% vs. 50%), an earlier onset of clinical signs with a significantly higher diarrhea score, lower VH:CD ratios and a higher percentage of PEDV-positive enterocytes. This study is the first to report PEDV coinfections with different ORF3 genotypes, and a PEDV strain with a large deletion in the ORF3 gene might have the advantage of a potential genetic marker, which would be useful during vaccine development.     

NS2a-NS2b sequence analysis of DEN-4 virus strains

To study the genetic variability of DEN-4 Chinese isolates, and to trace the origin of DV4 Chinese isolates, we cloned and sequenced the NS2a-NS2b junction of 5 isolates and prototype DV4 (H-241). Our results show that isolates from the 1990 Guangdong epidemic, which were isolated in the early, middle, and late periods of the epidemic, share the same sequence in the NS2a-NS2b junction. The sequence similarity between isolates from the Guangdong epidemic in 1990 and DV4 H-241 is 96%; these isolates can be grouped into genotype I. The sequence similarity between the isolate from the Guangdong epidemic in 1987 and Dominica strain 814669 is 96%; this isolate can be grouped into genotype II. For the first time, our results show that there are also 2 DV4 genotypes in the Guangdong area of southern China, and these isolates perhaps were introduced from other epidemic areas outside of China.     

Pegylated interferon alfa-2b plus ribavirin for treatment of chronic hepatitis C

AIM: To study the safety and efficacy of pegylated interferon alfa-2b, indigenously developed in India, plus ribavirin in treatment of hepatitis C virus(HCV). METHODS: One-hundred HCV patients were enrolled in an open-label, multicenter trial. Patients were treated with pegylated interferon alfa-2b 1.5 μg/kg per week subcutaneously plus oral ribavirin 800 mg/d for patients with genotypes 2 and 3 for 24 wk. The same dose of peginterferon plus weight-based ribavirin(800 mg/d for ≤ 65 kg; 1000 mg/d for 65-85 kg; 1200 mg/d for 85-105 kg; 1400 mg/d for 105 kg body weight) was administered for 48 wk for patients with genotypes 1 and 4. Serological and biochemical responses of patients were assessed.RESULTS: Eighty-two patients(35 in genotypes 1 and 4 and 47 in 2 and 3), completed the study. In genotype 1, 25.9% of patients achieved rapid virologic response(RVR): while the figures were 74.1% for early virologic response(EVR) and 44.4% for sustained virologic response(SVR). For genotypes 2 and 3, all patients bar one belonged to genotype 3, and of those, 71.4%, 87.5%, and 64.3% achieved RVR, EVR, and SVR, respectively. In genotype 4, 58.8%, 88.2%, and 52.9% of patients achieved RVR, EVR, and SVR, respectively. The majority of patients attained normal levels of alanine aminotransferase by 4-12 wk of therapy. Most patients showed a good tolerance for the treatment, although mild-to-moderate adverse events were exhibited; only two patients discontinued the study medication due to serious adverse events(SAEs). Eleven SAEs were observed in nine patients; however, only four SAEs were related to study medication.CONCLUSION: Peginterferon alfa-2b, which was developed in India, in combination with ribavirin, is a safe and effective drug in the treatment of HCV.     

Genetic variability of CYP2B6 polymorphisms in four southern Chinese populations

AIM： To investigate the genotype and allelic frequencies of Cytochrome P450 2B6 polymorphisms in four southern Chinese populations.
METHODS： DNA was obtained from blood samples from Han Chinese from Hong Kong and three minority groups,the Wa, Bulang and Lahu from Yunnan in southern China. Genotyping was performed using real-time PCR and confirmed by direct sequencing.
RESULTS： A total of 507 subjects from southern China were studied. Results showed there is a high prevalence of 516G 〉 T （34.5%） in ethnic Chinese compared to literature reports on other Asian populations and Caucasians. The frequency of the 516TT genotype is higher in the Hah majority （23.1%） than in three other ethnic minority groups （i.e., 7.4%, 9.1% and 15.8%） in southern China.
CONCLUSION： This was the first study to document the spectrum of CYP2B6 allelic variants and genotypes in a southern Chinese population. The 516G 〉 T allele is associated with a defective metabolism of efavirenz （EFV）, which therefore may predispose to drug toxicity.Treatment regimens for human immunodeficiency virus （HIV） and heroin addiction may need to be optimized in different populations because of the marked variability of the key metabolizing enzyme.     

Hepatitis C virus genotype and subtype distribution among ethnic minorities in Liaoning Province of China

To provide information and a basis for improved hepatitis C prevention and treatment, we aimed to determine the distribution of hepatitis C virus (HCV) genotypes among patients with hepatitis C from 4 ethnic minorities in Liaoning Province of China over the past 8 years and analyze and explore the virus’ genotype evolution and possible clinical significance.For gene-sequencing, we collected peripheral blood samples of HCV-infected patients belonging to the Korean, Hui, Mongol, and Manchu ethnic minorities in Liaoning Province who were diagnosed at the Second Hospital of Dalian Medical University, Anshan Central Hospital, and the Second People''s Hospital of Fuxin City between November 2011 and November 2019. To analyze genotype evolution and possible influencing factors, we determined the ratio of various genotypes. Among the 102 HCV-infected patients from 4 ethnic minorities in Liaoning Province, 46 had gene typing (GT)1b (45.10%), 15 had GT2a (14.71%), 14 had GT3a (13.73%), 13 had GT6a (12.75%), 3 had GT1a (2.94%), and 11 had an unclassified genotype (10.78%). The distribution of various genotypes in the Korean, Mongol, and Manchu ethnic minorities was significantly different (χ² = 10.788, P = .029; χ² = 7.846, P = .049; and χ² = 22.400, P = .000, respectively). All ethnic minorities exhibited >40% of GT1b. In the Korean (14/33) and Manchu (14/30) ethnic minorities, the proportion of GT1b was significantly higher than those of other genotypes (P < .05). The ethnic Koreans had a high proportion of GT3a (18.18%, 6/33), whereas the ethnic Mongolians had a high proportion of GT6a (23.08%, 6/26). GT1a was only found in the Korean (6.06%, 2/33) and Manchu (3.33%, 1/30) ethnic minorities; in the Hui ethnic minority, only 3 genotypes were prevalent: GT1b, GT2, and GT3a. The ethnic minorities in Liaoning Province currently have diverse HCV genotypes; the most prevalent genotype is GT1b, followed by GT2a and GT3a, and the prevalence of GT3 and GT6 has increased. The distribution of HCV genotypes varies across different ethnic minorities. The Korean and Manchu ethnic minorities have the most prevalent genotypes, whereas the Hui ethnic minority has a relatively single distribution of the HCV genotype.     

Advances in Crosstalk between Porcine Circoviruses and Host

Porcine circoviruses (PCVs), including PCV1 to PCV4, are non-enveloped DNA viruses with a diameter of about 20 nm, belonging to the genus Circovirus in the family Circoviridae. PCV2 is an important causative agent of porcine circovirus disease or porcine circovirus-associated disease (PCVD/PCVAD), which is highly prevalent in pigs and seriously affects the swine industry globally. Furthermore, PCV2 mainly causes subclinical symptoms and immunosuppression, and PCV3 and PCV4 were detected in healthy pigs, sick pigs, and other animals. Although the pathogenicity of PCV3 and PCV4 in the field is still controversial, the infection rates of PCV3 and PCV4 in pigs are increasing. Moreover, PCV3 and PCV4 rescued from infected clones were pathogenic in vivo. It is worth noting that the interaction between virus and host is crucial to the infection and pathogenicity of the virus. This review discusses the latest research progress on the molecular mechanism of PCVs–host interaction, which may provide a scientific basis for disease prevention and control.     

Cap Is the Protease of the Porcine Circovirus 2

Circoviruses are the smallest single-stranded DNA viruses that infect mammalian species, avian species, fish, and insects. The infections of circoviruses are known to be associated with a series of fatal diseases, but the protease of circovirus still remains unknown. In this research, we identified viral capsid protein (Cap) as the protease of porcine circovirus type 2 (PCV2), to our knowledge the first circoviruses protease to be reported. First, we found that the expression of host proteins is affected due to PCV2 infection in the porcine kidney (PK-15) cells. Then, by proteomic analysis, 253 host proteins that were down regulated were identified due to direct or indirect effects of PCV2. Further, Cap expression, but not other ORFs of PCV2, significantly reduced both JMJD6 (bifunctional arginine demethylase and lysyl-hydroxylase) and CCT5 (the chaperonin containing TCP1 subunit 5) in PK-15 cells. Finally, the results in vitro hydrolysis assays demonstrated that Cap could directly degraded either JMJD6 or CCT5 with different catalytic efficiency. In summary, our study expands repertoire of PCV2 Cap and promotes the development of inhibitors toward the anti-PCV2.     

欧洲型PRRSV、PCV2二联重组痘苗病毒疫苗猪体免疫实验研究

目的评价实验室构建的欧洲型PRRSV和PCV2二联重组痘苗病毒猪体免疫效果。方法分别用重组痘苗病毒rVTT-PCV2-ORF2-EU-ORF3-ORF5和rVTT-EU-ORF3-ORF5进行猪体免疫试验,每周采血,分离血清,采用ELISA检测病毒特异性抗体、中和抗体检测及细胞因子水平,评价免疫效果。结果 rVTT-PCV2-ORF2-EU-ORF3-ORF免疫组35d时,猪血清欧洲型PRRSV GP3和GP5及PCV2ORF2特异性抗体,显著高于野毒组（t=6.61,t=9.21,t=10.34,P〈0.05）;猪血清中和抗体35d时,各免疫组均可产生较高的免疫水平,差异均无统计学意义（t=0.48,P〉0.05）;重组痘苗病毒疫苗rVV-ORF2-EU-ORF3-ORF5免疫可诱导IL-4和IFN-γ产生,增强猪体细胞免疫应答水平。结论欧洲型PRRSV、PCV2二联重组痘苗病毒疫苗能诱导猪体产生体液和细胞免疫应答,为欧洲型PRRSV、PCV2的新型疫苗研究奠定了实验基础。     

Genotyping of hepatitis C virus (HCV) in infected patients from Northeast Mexico

Information about HCV genotypes in infected patients from different regions of Mexico is limited. Objective: To determine the prevalence of HCV genotypes in a group of HCV infected patients who attended a third level Hospital in Northeast of Mexico. Methods: Genotyping analysis was performed using the InnoLiPA-HCV genotype assay in 147 patients (65 males and 82 females, mean age 44 ± 12 years) with positive anti-HCV antibodies and detectable HCV-RNA levels. Results: Infected individuals were more likely to be female (56%). Histological data showed that 63% of the patients had chronic hepatitis, while the remainder presented cirrhosis (37%). The most frequent HCV genotype was 1 (73%). We found the following distribution: genotype 1 (2.7%), 1a (28.6%), 1b (37.4%), 1a/1b (4.1%), 2a (1.4%), 2b (8.8%), 2c (0.7%), 2a/2c (2.7%), 3 (2%), 3a (10.2%), 4 (0.7%) and 4c (0.7%). The most frequent associated risk factor was blood transfusion (72.5%). Conclusion: Prevalence of HCV genotypes in the Northeast of Mexico is similar to those reported previously in other Mexican regions and the most frequent risk factor continues being blood transfusion.     

Effect of HBB genotype on survival in a cohort of transfusion-dependent thalassemia patients in Cyprus

Initiation of regular transfusion in transfusion-dependent thalassemia (TDT) is based on the assessment of clinical phenotype. Pathogenic HBB variants causing β-thalassemia are important determinants of phenotype and could be used to aid decision-making. We investigated the association of HBB genotype with survival in a cohort study in the four thalassemia centers in Cyprus. HBB genotype was classified as severe (β0/β0 or β+/β0), moderate (β+/β+), or mild (β0/β++ or β+/β++). Risk factors for mortality were evaluated using multivariate Cox proportional- hazards regression. Of the 537 subjects who were followed for a total of 20,963 person-years, 80.4% (95% confidence interval [95% CI]: 76.4-84.7) survived to 50 years of age with increasing rates of liver-, infectionand malignancy-related deaths observed during recent follow-up. We evaluated non-modifiable risk factors and found worse outcomes associated with male sex (hazard ratio 1.9, 95% CI: 1.1-3.0, P=0.01) and milder genotype (hazard ratio 1.6, 95% CI: 1.1-2.3, P=0.02). The effect of genotype was confirmed in a second model, which included treatment effects. Patients with a milder genotype initiated transfusion significantly later and had reduced blood requirements compared to those with moderate or severe genotypes, although pre-transfusion hemoglobin levels did not differ between genotypes. Our results suggest that early treatment decisions to delay transfusion and different long-term treatment strategies in individuals with milder genotypes have led to adverse longterm effects of under-treated thalassemia and worse survival. We propose that HBB genotype determination and use of this information to aid in decision-making can improve long-term outcomes of thalassemia patients.     

贵州地区HCV感染者基因型分布特征

目的 探讨贵州地区丙型肝炎患者HCV基因型分布特征,为HCV感染的防控和个体化治疗提供临床依据。方法 选取2011年9月-2018年10月贵阳市公共卫生救治中心1211例HCV RNA阳性的丙型肝炎患者,采用PCR直接测序法,与GenBank中已知的HCV序列进行对比,获得HCV基因型及亚型,并分析其分布与性别、年龄、民族、地区、感染途径等因素的关系。计数资料组间比较采用χ2检验或Fisher确切概率法。结果 1211例HCV感染者共检出4种基因型和11种基因亚型,其中以1b型(26.84%)、3b型(27.17%)和6a型(24.28%)为主。不同HCV基因型在男女感染者中分布差异有统计学意义(χ2=15.428,P=0.009),其中男性以3b型为主(29.34%),女性以1b型为主(32.21%)。不同HCV基因型分布在各年龄组中差异有统计学意义(χ2=67.439,P<0.001),≤18岁组与≥60岁组以1b型为主(分别为66.67%、58.93%),而19~39岁组以3b型(28.93%)、6型(29.29%)为主,40~59岁组以1b型(29.54%)、3b型(27.33%)和6型(24.28%)为主。各种感染途径的HCV基因型分布差异有统计学意义(χ2=153.916,P<0.001),感染方式以静脉药瘾为主(57.97%),其次是性接触和有创美容(均为8.42%);经静脉药瘾、有创美容感染HCV者均以3b型为主(分别为31.48%、32.35%),性接触感染者多为HCV 6型(36.27%)。不同民族、贵州地区间HCV基因型分布差异均无统计学意义(P值均>0.05)。结论 贵州地区HCV基因型分布呈多样性,3b、1b、6a型为主要流行株,存在HCV 6型中的多种少见基因亚型,且在不同的年龄、性别、感染途径方面HCV基因型分布存在差异。     

丙型肝炎病毒非编码区ABC程序酶切分型研究

目的为进一步了解中国是否存在HCV 3b基因及1a、2b和6a基因型感染，建立HCV 5′端非编码区(5′ NCR)不同基因型的基因库。方法分型方法按ABC程序进行，A应用BHH′(BsrBⅠ、HaeⅡ、HinfⅠ)复介内切酶消化5′NCR cDNA，可将不同基因型划分为5组：1a、1b，6a，2a、2b，3a，3b、4a。B应用BstU Ⅰ内切酶鉴别1a、1b。C应用Hae Ⅲ内切酶鉴别2a、2b、3b、4a及6a。电泳检测片段大小。结果(1)la、1b、2a、2b、3a、3b、4a、6a 8种基因型参比品的ABC分型结果表明，该8种基因型获得良好的分型效果。(2)93份HCV RNA阳性患者ABC分型结果表明，1b型感染率占66．67％，2a型18．28％，1b／2b型、3b型及2b型均为3．23％，2a／2b型和1b／2a型各为2．15％，1a型1．08％。结论结果表明应用HCV 5′-NCR ABC分型技术既保证了HCV RNA检测的灵敏度，又能完成1a-6a型中的8种基因型的鉴别。     

多重PCR HCV基因分型检测方法的建立及初步应用

目的建立多重PCR HCV RNA基因分型的方法,并应用于临床诊断。方法采用型特异性引物建立单管逆转录聚合酶链反应(RT-PCR)和双管引物特异性套式聚合酶链反应(nested-PCR)检测技术,分别对72例HCV RNA阳性的标本,设计HCV的C区特异性引物,采用Trizol提取RNA进行单管巢式PCR分型,1a、1b、2a、2b和3a在一管内扩增。结果统计结果表明,我国丙型肝炎流行主要以1b(Ⅱ)和2a(Ⅲ)2种基因型为主,但有部分1a、2b、3a的HCV感染者检出,其中1a占1.4%(1/72),1b占68.1%(49/72),2a占13.9%(10/72),3a占1.4%(1/72),1b/2a型混合占11.1%(8/72),未分出型或其他型者为4.2%(3/72)。结论本文建立的方法具有良好的特异性和敏感性,省时、省力,方便检测,可同时区分5种HCV基因型;我国北方地区发现有1a和3a基因型感染病例。     

Geographic distribution, virologic and clinical characteristics of hepatitis B virus genotypes in China

The significance of hepatitis B virus (HBV) genotypes for the heterogeneity of chronic HBV infection and severity of liver disease is not well understood. The aim of this study was to determine the distribution and virologic characteristics of HBV genotypes in China and possible association with the diversity of liver disease. The study includes 1096 chronic HBV carriers from nine provinces in China. We collected clinical and laboratory data and analysed the HBV strains in sera by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and nucleotide sequencing techniques. The most common HBV genotypes were B (41%) and C (53%), while genotypes A and D were also found. A North-South divide was identified in genotype B and C distribution - genotype C was predominant in northern China, while genotype B was more prevalent in southern provinces. Patients with genotype B were younger than those with genotype C, and had a lower prevalence of HBeAg - 65%vs 72%, respectively (P = 0.03). However, the severity of liver disease did not differ significantly between patients infected with genotype B or C - neither when comparing liver function tests (1024 patients), nor hepatic inflammation and fibrosis (264 patients). Amongst 47 patients with genotype D (by PCR-RFLP), 37 (79%) were infected with a new subtype (designated Dc), having a recombination fragment from genotype C precore/core region. This is the first large-scale HBV genotype study from China and convincing documentation of the North-to-South gradient of genotypes C vs B in this country. HBV DNA recombination over the surface and precore/core genes increases the diversity of HBV strains and may have diagnostic and clinical implications.