摘要： Background: Picornaviruses, such as the structurally related polioviruses and rhinoviruses, are important human pathogens which have been the target of major drug development efforts. Receptor-mediated uncoating and thermal inactivation of poliovirus and rhinovirus are inhibited by agents that bind to each virus by inserting into a pocket in the beta barrel of the viral capsid protein, VP1. This pocket, which is normally empty in human rhinovirus-14 (HRV14), is occupied by an unknown natural ligand in poliovirus. Structural studies of HRV14-drug complexes have shown that drug binding causes large, localized changes in the conformation of VP1. Results: We report the crystal structures of six complexes between poliovirus and capsid-binding, antiviral drugs, including complexes of four different drugs with the Sabin vaccine strain of type 3 poliovirus, and complexes of one of these drugs with two other poliovirus strains that contain sequence differences in the drug-binding site. In each complex, the changes in capsid structure associated with drug binding are limited to minor adjustments in the conformations of a few side chains lining the binding site. Conclusions: The minor structural changes caused by drug binding suggest a model of drug action in which it is the conformational changes prevented by the bound drug, rather than obvious conformational changes induced by drug binding, which exert the biological effect. Our results, along with additional structures of rhinovirus-drug complexes, suggest possible improvements in drug design, and provide important clues about the nature of the conformational changes that are involved in the uncoating process.

关键词： Animals   Antibodies, Monoclonal/immunology   Binding Sites   Cell Adhesion Molecules/metabolism   Intercellular Adhesion Molecule-1   Mice   Models, Molecular   Mutation   Poliovirus/metabolism   Receptors, Virus/chemistry   Receptors, Virus/physiology   Structure-Activity Relationship  

摘要： The N-terminal domain 1 of the human poliovirus receptor (hPVR), a three-domain, immunoglobulin-like molecule, was previously shown to be necessary and sufficient to confer poliovirus (PV) susceptibility to mouse cells. However, studies with truncated versions of hPVR suggested that the C-terminal hPVR domains may contribute to receptor function. We describe sets of hybrid receptors, constructed between hPVR and hlCAM-1 (human intercellular adhesion molecule-1) that were tested in mouse cells for hPVR functionality. Whereas the context in which hPVR is expressed is of minor importance, all three domains of hPVR are required to reach wild-type function. Single and multiple amino acid exchanges were introduced into the first hPVR domain in order to localize regions that were involved in virus-receptor interaction. The mutations were analyzed for their ability to bind PV1 (Mahoney) or monoclonal antibodies as well as their ability to support viral replication in either the hPVR alpha or hybrid hPVR-hlCAM-1 receptor context. When placed into a model of the V domain of hPVR, the effect of the mutations indicated that the C'C''D as well as the DE region harbored amino acids that contacted the PV1(M) surface in the process of receptor-virus complex formation. The binding of the virus to the receptor and subsequent uptake into the cells were linked;no hPVR mutants were observed that bound the virus but blocked infection. N-glycosylation of the four sites in domains 1 and 2 is not required for hPVR function, but glycosylation in domain 1 has a greater effect on receptor function than that of domain 2. (C) 1994 Academic Press, Inc.

关键词： HEPATITIS B   NEONATE   HEPATITIS B IMMUNE GLOBULIN  

摘要： The need for an additional dose of hepatitis B immune globulin (HBIg) was studied by comparing infants receiving 1 ml HBIg at birth followed by hepatitis B immunization, concomitant with DTP-polio vaccine, at 3, 4, 5 and 11 months (schedule E), with infants receiving the same schedule with additional HBIg at 3 months (schedule F). The immune response to recombinant hepatitis B vaccine (20 mu g) was evaluated in 195 infants born to HBsAg-positive mothers allocated to groups E and F and compared with historic controls who received plasma vaccine (10 mu g) according to schedule F. Blood samples were drawn at 0, 3, 4, 6, 11, 12 and 24 months of age. No difference in efficacy between the two schedules was observed;8 and 6% of infants born to HBeAg-positive carrier mothers in groups E and F, respectively, became HBsAg carriers. Passively acquired antibodies at birth remained present for about 5 months in most infants. The seroprotection rates (anti-HBs greater than or equal to 10 IU l(-1)) were over 90% at all time points and similar for groups E and F. The titres of anti-HBs attained during the first 6 months were statistically lower (p less than or equal to 0.02) for group E than for group F but similar. thereafter. Anti-HBs titres in infants receiving the recombinant vaccine were significantly lower than in infants receiving the plasma vaccine (p<<0.001). Supplemental doses of HBIg in infants receiving a high dose of HBIg (>200 IU) at birth and the first dose of vaccine at the age of 3 months are not advised.

关键词： POLIOVIRUS VACCINE   TISSUE CULTURE   VIRULENCE  

摘要： The three Sabin strains of poliomyelitis seed virus were serially passaged in either secondary monkey kidney or Vero cell cultures and the tenth passage of each virus harvest compared to non-passaged Sabin reference virus of the same type using the monkey neurovirulence test. All three types were further attenuated by passage in Vero cells, whereas only type 2 became further attenuated after passage in secondary monkey kidney cells. After passage in Vero cells, type 3 poliomyelitis virus became more heat stable, as measured by its replicative capacity at 40 degrees C.

关键词： PCR-RFLP法   脊髓灰质炎病毒   临床分析  

摘要： 用ＰＣＲ－ＲＦＬＰ法对近年来从麻痹病人分离到的２６株脊灰炎病毒（ＰＶ）作了型内分析研究。１６株ＰＶ１的ＲＦＬＰ图式呈现多样性，毒株的图式可与Ｓａｂｉｎ１型疫苗株完全相同或不同，某些毒株亦可无电泳条带出现。７株ＰＶ２与３株ＰＶ３的ＲＦＬＰ图式则分别与疫苗株Ｓａｂｉｎ２型和３型完全相同。全面分析这些ＲＦＬＰ后，我们认为１６株ＰＶ１为野毒株，而７株ＰＶ２及３株ＰＶ３则可能是疫苗相似株。鉴于近年来广西的麻痹病例仍主要由ＰＶ１所致。故此，加强突击普服ＯＰＶ运动及作好分离毒株的型内分析已成为消灭脊灰炎的关键。

关键词： IRES   PICORNAVIRUS   POLIOVIRUS   REPLICATION SIGNALS   RNA SYNTHESIS  

摘要： The 5' untranslated region of poliovirus RNA has been reported to possess two functional elements: (i) the 5' proximal 88 nucleotides form a cloverleaf structure implicated in positive-strand RNA synthesis during viral replication, and (ii) nucleotides 134 to at least 556 function as a highly structured internal ribosome entry segment (IRES) during cap-independent, internal initiation of translation. We show here that the IRES itself is bifunctional and contains sequences necessary for viral RNA synthesis per se. For this purpose, we used a dicistronic poliovirus RNA in which the translation of the viral non-structural (replication) proteins is uncoupled from the poliovirus IRES. In this system, RNA synthesis is readily detectable in transfected cells, even when the poliovirus IRES is inactivated by point mutation. However, deletion of the major part of the poliovirus IRES renders viral-specific RNA synthesis undetectable. Using the same system, we show that a three nucleotide deletion at position 500 in the 5' untranslated region drastically affects both translation efficiency and RNA synthesis. Furthermore, disruption of the secondary structure of the IRES around nucleotide 343 has minimal effects on IRES function, but dramatically reduces viral RNA replication. Taken together, these results provide direct evidence that sequences essential for viral RNA synthesis are located in the 3' region of the poliovirus IRES.

关键词： 脊髓灰质炎疫苗   强化免疫   反向被动血凝抑制试验  

摘要： 本文用反向被动血凝抑制试验对65名0～4岁儿童于液体或糖丸脊髓灰质炎疫苗强化免疫前、后进行血清抗体水平测定,结果表明,两种剂型疫苗的强化免疫效果相似,均能极显著地提高Ⅱ型抗体水平,而Ⅰ、Ⅲ 抗体仅略有上升。作者还讨论了强化免疫效果的影响因素,提出南方地区应增加强化免疫的次数。

关键词： INACTIVATED POLIOVIRUS VACCINE   THIMEROSAL   STORAGE TEMPERATURE   COMBINATION VACCINE   POTENCY  

摘要： High-potency inactivated poliovirus vaccine (eIPV) was combined with diptheria-tetanus-pertussis (DTP) vaccine containing thimerosal as a preservative to simulate the performance of a potential tetravalent vaccine. Neither type 1 nor type 3 poliovirus antigens appeared to be affected by thimerosal after exposure for 1 h at 37 degrees C as measured by enzyme-linked immunosorbent assay (ELISA). One epitope on the type 2 antigen was damaged within 5 min of exposure, however, the overall potency was unchanged when measured using a polyclonal antibody preparation. Exposure to thimerosal at 37 degrees C decreased the potency of all three poliovirus types to well below the level caused by heat deterioration alone in 1-2 days and to 0% after 16-17 days. At 25 degrees C, the potency of type 1 poliovirus decreased by 46% in 1 day, whereas poliovirus types 2 and 3 were stable for I week. Storage of eIPV at 4 degrees C in the presence of thimerosal reduced the potency of type 1 poliovirus antigen to undetectable levels after 4-6 months. Type 2 and 3 antigens were less markedly affected by 8 months of exposure to thimerosal at 4 degrees C. The loss of potency of type 1 as measured by ELISA was paralleled by a reduced level of neutralizing antibodies in mice injected with these preparations. The results obtained from testing eIPV in combination with DTP and thimerosal were generally similar to those obtained using eIPV with thimerosal. It remains to be seen to what extent thimerosal will affect the immunogenicity of eIPV in humans when injected as combined eIPY-DTP vaccine.

摘要： Poliovirus (PV)-infected cells undergo extensive proliferation and rearrangement of intracellular smooth membranes to generate vesicles on which viral RNA replication occurs. PV proteins 2C and 2BC are known to be tightly associated with these membranous replication complexes and have been proposed to be involved in the formation of these virus-induced vesicles. We have expressed these proteins, and proteins with mutations in the putative nucleotide (NTP) binding motifs, in human cells using recombinant vaccinia viruses and T7 RNA polymerase-directed transcription. To ascertain the subcellular localization properties of these proteins in the absence of other PV proteins and to determine whether they induced ultrastructural changes, cells expressing 2C and 2BC proteins were examined by immunofluorescence (IF) microscopy, electron microscopy (EM), and immuno-EM (IEM). The cytoplasm of cells expressing either 2C or 2BC exhibited vesicles of 50-350 nm in diameter, which resembled those found in PV-infected cells. Both 2C and 2BC were associated with these vesicles. Mutations in the putative NTP binding motif did not affect vesicle induction by 2C or 2BC, Despite the membrane reorganization and vesicle formation induced by 2C and 2BC proteins, no enhanced synthesis of lipid was observed. Guanidine hydrochloride at a concentration that inhibits PV replication, did not have significant effects on the IF patterns of either 2C or 2BC. An additional prominent alteration in cells expressing 2C, but not 2BC, was the formation of extensive tubular membrane structures with a myelin-like arrangement in the lumen of the rough endoplasmic reticulum. IEM analyses showed that 2C was associated with these structures. In the presence of other PV proteins, the tubular membrane structures induced by 2C were not detected. These structures are not observed in poliovirus-infected cells, but likely indicate a novel property of 2C that induces a complex interaction with intracellular membranes