Antigenic and Genetic Variations among Japanese Encephalitis Virus Strains Belonging to Genotype 1

Hyperimmune antisera against four Japanese encephalitis (JE) virus strains, ThCMAr4492 and ThCMAr6793 from Thailand and Nakayama and JaGAr01 from Japan, were used to analyze the antigenic relationships among 12 Thai strains belonging to genotype 1, and two Japanese strains and one Chinese strain belonging to genotype 3. The antiserum for ThCMAr6793 significantly neutralized nine of the 12 Thai strains, none of which was significantly neutralized by antisera for the Nakayama and JaGAr01 strains. The antiserum for ThCMAr4492 neutralized only its homologous strain; therefore, ThCMAr4492 was antigenically different from all other strains. Two Thai strains (Subin and KE-093/83) were significantly less neutralized by all four of the antisera tested. In the deduced amino-acid sequence of the E protein, the 12 Thai strains revealed 100 to 98.2% identity among them and 90.0 to 98.8% identity with the published strains, respectively. Among significant amino-acid substitutions, three residues at positions E-222, E-327 and E-366 were shared by all of the Thai strains, whereas residues at E-89, E-123, E-131, E-178, E-293, E-351 and E-373 seemed to be strain-specific. The amino acids at positions E-178, E-327, E-351, E-373 and E-366 are found either in the peptides with functional T-helper cell epitopes or in the ectodomain of the E protein of other flaviviruses. These amino acids may therefore be responsible for determining the antigenic heterogeneity of these strains.     

Detection of Japanese Encephalitis Virus Genotype V in Culex orientalis and Culex pipiens (Diptera: Culicidae) in Korea

Japanese encephalitis virus (JEV) causes significant viral encephalitis and is distributed throughout the Asian countries. The virus is known to be transmitted by Culex tritaeniorhynchus, which mainly breeds in rice paddies in Korea. In this study, we investigated the presence of other mosquito species that can transmit JEV as a second or regional vector. We selected five cities where patients have experienced JE in the last 5 years as mosquito-collecting locations and subdivided them into four collection sites according to the mosquito habitats (cowshed, downtown area, forest, and swamp). Mosquitoes were caught using the BG-Sentinel trap, CDC black-light trap, Fay-Prince trap, and Gravid trap. A total of 993 pools from 22,774 mosquitoes were prepared according to their species, collection date, and site. We performed a SYBR Green 1-based real-time RT-PCR assay to detect JEV from the mosquito pools. A total of six JEV-positive pools were detected from Culex orientalis and Culex pipiens caught in the Gangwon-do and Gyeonngi-do provinces. All the detected JEVs were revealed as genotype V by phylogenetic analysis of the envelope gene. Our findings confirm that a new genotype of JEV was introduced in Korea and suggest that two mosquito species may play a role in JEV transmission.     

Virus-Specific Cytolytic Antibodies to Nonstructural Protein 1 of Japanese Encephalitis Virus Effect Reduction of Virus Output from Infected Cells

We demonstrate the presence of nonstructural protein 1 (NS1)-specific antibodies in a significant proportion of convalescent-phase human serum samples obtained from a cohort in an area where Japanese encephalitis virus (JEV) is endemic. Sera containing antibodies to NS1 but not those with antibodies to other JEV proteins, such as envelope, brought about complement-mediated lysis of JEV-infected BHK-21 cells. Target cells infected with a recombinant poxvirus expressing JEV NS1 on the cell surface confirmed the NS1 specificity of cytolytic antibodies. Mouse anti-NS1 cytolytic sera caused a complement-dependent reduction in virus output from infected human cells, demonstrating their important role in viral control. Antibodies elicited by JEV NS1 did not cross lyse West Nile virus- or dengue virus-infected cells despite immunoprecipitating the NS1 proteins of these related flaviviruses. Additionally, JEV NS1 failed to bind complement factor H, in contrast to NS1 of West Nile virus, suggesting that the NS1 proteins of different flaviviruses have distinctly different mechanisms for interacting with the host. Our results also point to an important role for JEV NS1-specific human immune responses in protection against JE and provide a strong case for inclusion of the NS1 protein in next generation of JEV vaccines.The genus Flavivirus, many of whose more than 70 members are arthropod-borne human pathogens, such as dengue virus (DENV), West Nile virus (WNV), yellow fever virus (YFV), tick-borne encephalitis virus, and Japanese encephalitis virus (JEV), has assumed increasing public health importance in recent years. The single-strand, positive-sense RNA genomes of flaviviruses encode a single polyprotein, which is cotranslationally cleaved to produce three structural proteins (capsid [C], membrane [M], and envelope [E]) and seven nonstructural (NS) proteins (NS1, NS2a, NS2b, NS3, NS4a, NS4b, and NS5). NS1, a glycoprotein, is perhaps the most versatile among these, being involved both in vital processes such as viral RNA synthesis and in multiple interactions with the host, in ways that appear to benefit both pathogen and host. Following translocation into the lumen of the endoplasmic reticulum through a hydrophobic signal sequence that is encoded by the carboxyl terminus of E (17), NS1 undergoes glycosylation followed by rapid dimerization (44, 52). In DENV infection of cultured mammalian cells, extracellular NS1 was additionally detected as hexamers (19, 51). Despite the apparent absence of a canonical hydrophobic membrane anchor domain, the NS1s from JEV, Kunjin virus, DENV, and YFV have all been shown to be present on the surface of virus-infected cells (8, 23, 50). The mechanistic details of this membrane anchor still remain uncertain.The ability of DENV NS1 to bind host complement (9, 49) pointed to a role for this protein in DENV pathogenesis. Serum NS1 levels in both DENV and WNV patients correlate directly with disease severity (1, 36). Promotion of immune complex formation (54), ability to elicit autoantibodies with reactivity to platelets and extracellular matrix (10), and damage inflicted on endothelial cells (34) are some of the mechanisms proposed to explain pathogenesis mediated by DENV NS1. Recent studies with WNV NS1 demonstrated its ability to bind human complement factor H, suggesting a role in reducing the host''s ability to bring about complement-mediated control of early virus replication (11).Critical differences between the functions of NS1s encoded by different pathogenic flaviviruses and their contributions to pathology are evident from the published reports, with DENV NS1 believed to be involved in complement activation and the consequent capillary leak syndrome of dengue hemorrhagic fever (6), while WNV NS1 appears relatively more benign and has more to do with modulation of the host innate immune response (11). We have not encountered reports of adverse impacts of JEV NS1 in infected individuals.Paradoxically, several studies have pointed to a role for flavivirus NS1-specific immune responses in protection against flaviviruses. Passive immunization studies using monoclonal antibodies (MAbs) (24, 28, 29, 55) as well as immunization of mice using naked DNA constructs expressing NS1 (35, 40) revealed that antibodies directed to prM or E of DENV and NS1 of DENV and JEV are protective. Studies by different groups have shown that active immunization with purified NS1 or passive immunization with MAbs against YFV and DENV NS1 provides protection from lethal viral challenge in the absence of neutralizing antibodies (24, 45, 48). A panel of anti-WNV NS1 MAbs revealed multiple antibody-mediated mechanisms for protection, some mediated through complement and others via the Fc receptor (12). Those authors went on to show that anti-NS1 MAbs that facilitate phagocytosis and clearance of WNV-infected cells through Fc-γ receptors I and/or IV belonged to the IgG2a subclass and bound to cell surface-associated NS1 (13).Earlier studies also pointed to the cytolytic potential of NS1 antibodies, a property that might contribute significantly to their protective ability. Passive immunization experiments using a panel of anti YFV NS1-specific MAbs showed a significant correlation between protection and in vitro complement-mediated cytolysis of YFV-infected mouse neuroblastoma cells (47). Additionally, immunization of mice with a DNA vaccine construct carrying JEV NS1 induced a strong antibody response exhibiting complement-mediated cytolysis of JEV-infected cells (35), but no neutralizing activity, and resulted in protection against subsequent challenge with virus. Cell-mediated immune responses directed to NS1 of JEV have also been reported to play a role in cytotoxic T-lymphocyte-mediated killing of JEV-infected murine target cells (41). Thus, NS1 appears to contribute to protection in the murine model by inducing both humoral and cell-mediated arms of the immune response.It was therefore of interest to query whether NS1-specific antibodies in humans exposed to JEV exhibit cytolytic activity and to determine if these antibodies are capable of reducing virus production by infected cells. In this study we report for the first time the existence of detectable levels of anti-NS1 antibodies in a significant proportion of sera from humans infected with JEV and demonstrate their ability to induce antibody-dependent complement-mediated cytolysis of cells expressing JEV NS1 on the surface. These sera failed to cause lysis of cells infected with WNV or DENV, both of which cocirculate with JEV in the Indian subcontinent and have been reported in the region where we enrolled our volunteers, revealing stringent specificity and absence of flaviviral cross-reactivity for these cytolytic antibodies. Furthermore, we demonstrate the ability of NS1-specific antibodies elicited in mice to limit virus production in infected human SW-13 cell monolayers, which may explain, at least in part, the widely reported protective ability of flavivirus NS1. Significantly, we found no evidence for the ability of NS1 from JEV to bind human complement factor H, in contrast to the case for WNV NS1 (11). Taken together, these findings suggest that JEV NS1 may positively and significantly affect virus-specific protective immune responses.     

流行性乙型脑炎病毒GSS株prM、E和NS1蛋白基因的克隆及在非复制型痘苗病毒中的表达

克隆流行性乙型脑炎(乙脑)病毒野毒(JEV)GSS株前膜蛋白信号序列、前膜蛋白(prM)、包膜蛋白(E)、非结构蛋白-1(NSl)和非结构蛋白NS2a的编码基因,并与非复制型痘苗病毒载体NTV进行同源重组,构建了乙脑病毒非复制型重组痘苗病毒疫苗株NTVA(E／L)JEV。通过：PCR和Southern blot检测证明,在非复制型痘苗病毒中有乙暗病毒prM信号序列、prM、E、NS1和NS2a基因的插入：Western blot检测证明,重组病毒可以在细胞内成功地表达prM、E和NSl蛋白,并可将prM、E和NSl蛋白分泌到细胞培养上清中;免疫荧光检测证明,E和NSl蛋白主要分布在细胞膜上。电镜下可见分泌到细胞外的病毒样颗粒。     

我国新分离乙型脑炎病毒株毒力特征研究

将分离自不同年代的17株乙脑毒株在小鼠脑内传代,然后将病毒在BHK21细胞单层上观察不同毒株的空斑形成大小形态,小鼠脑内和皮下途径接种观察病毒的毒力,结果显示不同毒株在BHK21细胞上形成的噬斑大小不尽相同,减毒株形成的噬斑最小。所有毒株对小鼠的脑内毒力都很强,病毒滴度高达lg8.0/mL以上,毒株间无明显差异。毒株对9～11g小日龄小鼠的皮下毒力有一定差异但不明显,但对14～16g较大日龄小鼠则差别明显。PFU/LD50的对数值差异除一株(M47株)为8.44外,其余各株差别在3.94～0.45间。本研究结果证明自然界乙脑毒株存在明显的神经外毒力差异,毒力差异与分离年代和病毒基因型无关,但从人体分离到的毒株毒力表现较强。     

Inhibition of Japanese Encephalitis Virus Infection by the Sulfated Polysaccharide Extracts from Ulva lactuca

Japanese encephalitis virus (JEV), a neurotropic flavivirus, is one of the major causes of acute encephalitis in humans. After infection, inflammatory reactions and neurological diseases often develop. Still there are no effective drugs available against virus infection. Recently, extracts of algae have been shown to possess a broad range of biological activities including antivirus activity. In this study, we identified that the sulfated polysaccharide extracts from Ulva lactuca can inhibit JEV infection in Vero cells. Mechanistic studies further revealed that the Ulva sulfated polysaccharide extracts can block virus adsorption and thus make the virus unable to enter cells. The Ulva sulfated polysaccharide extracts also effectively decrease the production of pro-inflammatory cytokines in the JEV-infected primary mixed glia cells. In an animal study, the JEV-infected C3H/HeN mice appeared to have neurobehavioral abnormalities on the fifth day and died on the seventh day post infection. However, the JEV-infected mice pretreated with the Ulva sulfated polysaccharide extracts can delay the onset of hind limb paralysis and thereby prevent mice from death.     

乙型脑炎病毒减毒中间株SA14—12—1—7基因组全序列的测定

本研究通过对乙型脑炎活疫苗减毒过程中间株SA14 12 1 7株进行全序列测定和分析 ,进一步了解乙脑活疫苗减毒及其稳定性的分子机制。根据已发表的SA14 14 2株及SA14 株的序列 ,设计 6对重叠引物 ,涵括整个乙脑病毒的基因组 ,通过RT PCR扩增出SA14 12 1 7株的各cDNA片段 ,分别克隆到pGEM T载体 ,转化至TG1受体菌中 ,挑取阳性克隆进行鉴定后测序。结果表明SA14 12 1 7株基因组全序列长 10 976个核苷酸 ,从 96到 10 394为一个长开放读码框 ,编码 3432个氨基酸。与野毒株SA14 和疫苗株SA14 14 2的核苷酸序列和氨基酸序列相比 ,同源性均在 99%以上 ,突变位点分散于各个区域 ,E区有 5个位点与疫苗株一致而与野毒株不同 ,3个位点与野毒株一致而与疫苗株不同 ,推测与其容易产生回复突变、恢复毒力有关。此外 ,NS3、NS5和 3′NTR的几个位点可能与病毒毒力稳定性相关。综上所述 ,乙脑病毒减毒中间株的基因组全序列基本类似于已发表的序列 ,若干突变位点影响病毒的弱毒性及毒力的稳定性。全序列的测定对于研究疫苗株的减毒机理具有重要意义     

不同苗种来源的中国虾夷扇贝群体的遗传多样性分析

虾夷扇贝(Mizuhopecten yessoensis)于1982年从日本引入中国并展开规模化养殖.由于引入的亲贝数目有限,使虾夷扇贝在人工育苗养殖过程中群体遗传多样性水平下降.本研究使用7对微卫星引物对日本原种贝(♀、♂)自交后的子代群体(RZ)、国内种贝(♀、♂)自交后的子代群体(DZ)、日本原种贝(♂)与国内种贝(♀)的杂交群体(ZJ)和国内自然海区(中国旅顺月亮湾)天然繁殖群体(HC)4个不同的虾夷扇贝群体的遗传多样性进行了研究.实验结果表明,4个群体的平均有效等位基因数为3.2～3.8,平均期望杂合度为0.6718～0.7017,日本野生群体做为种贝繁殖的苗种(KZ)与中国养殖群体相比,遗传多样性水平较高,除了DZ群体外其他群体的遗传多样性并无显著的变化.     

Complete Bovine Leukemia Virus (BLV) Provirus Is Conserved in BLV-Infected Cattle throughout the Course of B-Cell Lymphosarcoma Development

Bovine leukemia virus (BLV) and human T-cell leukemia virus types 1 and 2 (HTLV-1 and HTLV-2) belong to the same subfamily of oncoviruses. Defective HTLV-1 proviral genomes have been found in more than half of all patients with adult T-cell leukemia examined. We have characterized the genomic structure of integrated BLV proviruses in peripheral blood lymphocytes and tumor tissue taken from animals with lymphomas at various stages. Genomic Southern hybridization with SacI, which generates two major fragments of BLV proviral DNA, yielded only bands that corresponded to a full-size provirus in all of 23 cattle at the lymphoma stage and in 7 BLV-infected but healthy cattle. Long PCR with primers located in long terminal repeats clearly demonstrated that almost the complete provirus was retained in all of 27 cattle with lymphomas and in 19 infected but healthy cattle. However, in addition to a PCR product that corresponded to a full-size provirus, a fragment shorter than that of the complete virus was produced in only one of the 27 animals with lymphomas. Moreover, when we performed conventional PCR with a variety of primers that spanned the entire BLV genome to detect even small defects, PCR products were produced that specifically covered the entire BLV genome in all of the 40 BLV-infected cattle tested. Therefore, it appears that at least one copy of the full-length BLV proviral genome was maintained in each animal throughout the course of the disease and, in addition, that either large or small deletions of proviral genomes may be very rare events in BLV-infected cattle.     

Estimating the Burden of Japanese Encephalitis Virus and Other Encephalitides in Countries of the Mekong Region

Diverse aetiologies of viral and bacterial encephalitis are widely recognized as significant yet neglected public health issues in the Mekong region. A robust analysis of the corresponding health burden is lacking. We retrieved 75 articles on encephalitis in the region published in English or in French from 1965 through 2011. Review of available data demonstrated that they are sparse and often derived from hospital-based studies with significant recruitment bias. Almost half (35 of 75) of articles were on Japanese encephalitis virus (JEV) alone or associated with dengue. In the Western Pacific region the WHO reported 30,000–50,000 annual JEV cases (15,000 deaths) between 1966 and 1996 and 4,633 cases (200 deaths) in 2008, a decline likely related to the introduction of JEV vaccination in China, Vietnam, or Thailand since the 1980s. Data on dengue, scrub typhus and rabies encephalitis, among other aetiologies, are also reviewed and discussed.Countries of the Mekong region are undergoing profound demographic, economic and ecological change. As the epidemiological aspects of Japanese encephalitis (JE) are transformed by vaccination in some countries, highly integrated expert collaborative research and objective data are needed to identify and prioritize the human health, animal health and economic burden due to JE and other pathogens associated with encephalitides.     

Herpes Simplex Virus 1 DNA Is in Unstable Nucleosomes throughout the Lytic Infection Cycle,and the Instability of the Nucleosomes Is Independent of DNA Replication

Herpes simplex virus 1 (HSV-1) DNA is chromatinized during latency and consequently regularly digested by micrococcal nuclease (MCN) to nucleosome-size fragments. In contrast, MCN digests HSV-1 DNA in lytically infected cells to mostly heterogeneous sizes. Yet HSV-1 DNA coimmunoprecipitates with histones during lytic infections. We have shown that at 5 h postinfection, most nuclear HSV-1 DNA is in particularly unstable nucleoprotein complexes and consequently is more accessible to MCN than DNA in cellular chromatin. HSV-1 DNA was quantitatively recovered at this time in complexes with the biophysical properties of mono- to polynucleosomes following a modified MCN digestion developed to detect potential unstable intermediates. We proposed that most HSV-1 DNA is in unstable nucleosome-like complexes during lytic infections. Physiologically, nucleosome assembly typically associates with DNA replication, although DNA replication transiently disrupts nucleosomes. It therefore remained unclear whether the instability of the HSV-1 nucleoprotein complexes was related to the ongoing viral DNA replication. Here we tested whether HSV-1 DNA is in unstable nucleosome-like complexes before, during, or after the peak of viral DNA replication or when HSV-1 DNA replication is inhibited. HSV-1 DNA was quantitatively recovered in complexes fractionating as mono- to polynucleosomes from nuclei harvested at 2, 5, 7, or 9 h after infection, even if viral DNA replication was inhibited. Therefore, most HSV-1 DNA is in unstable nucleosome-like complexes throughout the lytic replication cycle, and the instability of these complexes is surprisingly independent of HSV-1 DNA replication. The specific accessibility of nuclear HSV-1 DNA, however, varied at different times after infection.     

Expression of the Tick-borne Encephalitis Virus NS1 Gene in Gram-Negative Bacteria from the Mouse Nasopharynx

Bacteria were isolated from the nasopharynx of BALB/c mice and electroporated with pUR290(NS1)₂ containing two copies of tick-borne encephalitis virus (TBEV, strain Sofjin) NS1 under the control of the lac promoter. The plasmid persisted in transformants for at least ten passages. The NS1 gene expression was detected in Gram-negative enterobacteria by immunoblotting with monoclonal antibodies against the TBEV nonstructural glycoprotein NS1. Recombinant NS1 was detected in bacterial cells and in the culture medium. Intranasal immunization with recombinant bacteria caused antibodies against NS1 to appear in the serum of BALB/c mice. However, the humoral immune response to NS1 failed to protect mice from a TBEV challenge.     

Diagnostic Approach for the Differentiation of the Pandemic Influenza A(H1N1)v Virus from Recent Human Influenza Viruses by Real-Time PCR

Background

The current spread of pandemic influenza A(H1N1)v virus necessitates an intensified surveillance of influenza virus infections worldwide. So far, in many laboratories routine diagnostics were limited to generic influenza virus detection only. To provide interested laboratories with real-time PCR assays for type and subtype identification, we present a bundle of PCR assays with which any human influenza A and B virus can be easily identified, including assays for the detection of the pandemic A(H1N1)v virus.

Principal Findings

The assays show optimal performance characteristics in their validation on plasmids containing the respective assay target sequences. All assays have furthermore been applied to several thousand clinical samples since 2007 (assays for seasonal influenza) and April 2009 (pandemic influenza assays), respectively, and showed excellent results also on clinical material.

Conclusions

We consider the presented assays to be well suited for the detection and subtyping of circulating influenza viruses.     

Review of Climate,Landscape, and Viral Genetics as Drivers of the Japanese Encephalitis Virus Ecology

The Japanese encephalitis virus (JEV), an arthropod-born Flavivirus, is the major cause of viral encephalitis, responsible for 10,000–15,000 deaths each year, yet is a neglected tropical disease. Since the JEV distribution area has been large and continuously extending toward new Asian and Australasian regions, it is considered an emerging and reemerging pathogen. Despite large effective immunization campaigns, Japanese encephalitis remains a disease of global health concern. JEV zoonotic transmission cycles may be either wild or domestic: the first involves wading birds as wild amplifying hosts; the second involves pigs as the main domestic amplifying hosts. Culex mosquito species, especially Cx. tritaeniorhynchus, are the main competent vectors. Although five JEV genotypes circulate, neither clear-cut genotype-phenotype relationship nor clear variations in genotype fitness to hosts or vectors have been identified. Instead, the molecular epidemiology appears highly dependent on vectors, hosts'' biology, and on a set of environmental factors. At global scale, climate, land cover, and land use, otherwise strongly dependent on human activities, affect the abundance of JEV vectors, and of wild and domestic hosts. Chiefly, the increase of rice-cultivated surface, intensively used by wading birds, and of pig production in Asia has provided a high availability of resources to mosquito vectors, enhancing the JEV maintenance, amplification, and transmission. At fine scale, the characteristics (density, size, spatial arrangement) of three landscape elements (paddy fields, pig farms, human habitations) facilitate or impede movement of vectors, then determine how the JEV interacts with hosts and vectors and ultimately the infection risk to humans. If the JEV is introduced in a favorable landscape, either by live infected animals or by vectors, then the virus can emerge and become a major threat for human health. Multidisciplinary research is essential to shed light on the biological mechanisms involved in the emergence, spread, reemergence, and genotypic changes of JEV.     

The Three Subtypes of Tick-Borne Encephalitis Virus Induce Encephalitis in a Natural Host,the Bank Vole (Myodes glareolus)

Tick-borne encephalitis virus (TBEV) infects bank voles (Myodes glareolus) in nature, but the relevance of rodents for TBEV transmission and maintenance is unclear. We infected colonized bank voles subcutaneously to study and compare the infection kinetics, acute infection, and potential viral persistence of the three known TBEV subtypes: European (TBEV-Eur), Siberian (TBEV-Sib) and Far Eastern (TBEV-FE). All strains representing the three subtypes were infective and highly neurotropic. They induced (meningo)encephalitis in some of the animals, however most of the cases did not present with apparent clinical symptoms. TBEV-RNA was cleared significantly slower from the brain as compared to other organs studied. Supporting our earlier findings in natural rodent populations, TBEV-RNA could be detected in the brain for up to 168 days post infection, but we could not demonstrate infectivity by cell culture isolation. Throughout all time points post infection, RNA of the TBEV-FE was detected significantly more often than RNA of the other two strains in all organs studied. TBEV-FE also induced prolonged viremia, indicating distinctive kinetics in rodents in comparison to the other two subtypes. This study shows that bank voles can develop a neuroinvasive TBEV infection with persistence of viral RNA in brain, and mount an anti-TBEV IgG response. The findings also provide further evidence that bank voles can serve as sentinels for TBEV endemicity.     

日本脑炎病毒E基因与流感病毒HA基因的串联表达及应用

根据GenBank公布的日本脑炎病毒(Japanese Encephalitis Virus, JEV) SA14 14 2株的核酸序列和人流感病毒的血凝素基因(ha)序列, 设计一对特异性引物, 用 PCR方法扩增编码 JEV囊膜蛋白主要抗原域基因, 其中含ha基因主要核苷酸序列。将PCR产物定向克隆入原核表达载体 pET 32a( ), 构建原核表达载体 pET EHA。阳性质粒转化BL21(DE3)宿主菌, 经 IPTG诱导获得表达, 重组蛋白以包涵体的形式存在。Western blot分析表明表达产物具有良好的免疫学活性。利用纯化的表达产物与流感病毒血凝素单抗及乳胶建立了诊断日本脑炎病毒抗体水平的乳胶凝集试验。结果表明乳胶凝集方法具有简便快速、敏感性高、特异性强、价格低廉、可现场检测等优点, 是一种适合基层兽医单位用于日本脑炎病毒抗体水平检测的新方法。     

Human Immunodeficiency Virus Type-1 (HIV-1) Continues to Evolve in Presence of Broadly Neutralizing Antibodies More than Ten Years after Infection

Background

The evolution of HIV-1 and its immune escape to autologous neutralizing antibodies (Nabs) during the acute/early phases of infection have been analyzed in depth in many studies. In contrast, little is known about neither the long-term evolution of the virus in patients who developed broadly Nabs (bNabs) or the mechanism of escape in presence of these bNabs.

Results

We have studied the viral population infecting a long term non progressor HIV-1 infected patient who had developed broadly neutralizing antibodies toward all tier 2/3 viruses (6 clades) tested, 9 years after infection, and was then followed up over 7 years. The autologous neutralization titers of the sequential sera toward env variants representative of the viral population significantly increased during the follow-up period. The most resistant pseudotyped virus was identified at the last visit suggesting that it represented a late emerging escape variant. We identified 5 amino acids substitutions that appeared associated with escape to broadly neutralizing antibodies. They were V319I/S, R/K355T, R/W429G, Q460E and G/T463E, in V3, C3 and V5 regions.

Conclusion

This study showed that HIV-1 may continue to evolve in presence of both broadly neutralizing antibodies and increasing autologous neutralizing activity more than 10 years post-infection.