The assembly of foot-and-mouth disease virus (FMDV) particles is poorly understood. In addition, there are important differences in the antigenic and receptor binding properties of virus assembly and dissociation intermediates, and these also remain unexplained. We have established an experimental model in which the antigenicity, receptor binding characteristics, and in vitro assembly of capsid precursor can be studied entirely from purified components. Recombinant capsid precursor protein (P1 region) was expressed in Escherichia coli as myristoylated or unmyristoylated protein. The protein sedimented in sucrose gradients at 5S and reacted with monoclonal antibodies which recognize conformational or linear antigen determinants on the virion surface. In addition, it bound the integrin ␣ v ␤ 6 , a cellular receptor for FMDV, indicating that unprocessed recombinant capsid precursor is both structurally and antigenically similar to native virus capsid. These characteristics were not dependent on the presence of 2A at the C terminus but were altered by N-terminal myristoylation and in mutant precursors which lacked VP4. Proteolytic processing of myristoylated precursor by recombinant FMDV 3C pro in vitro induced a shift in sedimentation from 5S to 12S, indicating assembly into pentameric capsid subunits. Nonmyristoylated precursor still assembled into higher-order structures after processing with 3Cpro , but these particles sedimented in sucrose gradients at approximately 17S. In contrast, mutant precursors lacking VP4 were antigenically distinct, were unable to form pentamers, and had reduced capacity for binding integrin receptor. These studies demonstrate the utility of recombinant capsid precursor protein for investigating the initial stages of assembly of FMDV and other picornaviruses.Foot-and-mouth disease virus (FMDV) is the etiological agent of a highly infectious disease of cattle and other clovenhoofed animals. It is of economic importance, because the presence of the disease results in severe restrictions of international trade. In many parts of the developing world, the disease is endemic, and it continues to pose a serious threat to livestock industries globally, as exemplified by recent major outbreaks in the United Kingdom, Argentina, and Uruguay.FMDV is a small, nonenveloped, positive-strand RNA virus belonging to the genus Aphthovirus within the family Picornaviridae. Other members of the family include poliovirus (PV; genus Enterovirus) and hepatitis A virus (genus Hepatovirus). The mature picornavirus comprises 60 copies each of four structural proteins, termed VP1, VP2, VP3, and VP4, which encapsidate a single, positive-sense RNA genome. The proteins form a pseudo Tϭ3 icosahedral capsid with VP1 located close to the fivefold axes of symmetry and VP2 and VP3 alternating around the threefold axes (24). VP4, which is myristoylated at the N terminus, is an internal component of the capsid (13,48).Although the morphogenesis of picornaviral particles is not completely understood, it is known that capsid...

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Free lipid A of Helicobater pylori was characterized with regard to chemical composition, reactivity with anti-lipid A antibodies, and activity in a Limulus lysate assay. The predominant fatty acids of H. pylori lipid A were 3-OH-18:0, 18:0, 3-OH-16:0, 16:0, and 14:0. Hexosamine was present in amounts similar to those in Campylobacter jejuni or SalmoneUla typhimunium lipid A. The lipopolysaccharide of H. pylori contained 2-keto-3-deoxyoctonic acid, a common constituent of enterobacterial and C. jejuni lipopolysaccharides. In the enzyme-linked immunosorbent assay, the doses of lipid A required to inhibit anti-lipid A by 50%o (EI5 values) by absorption of the immune (rabbit) serum were 7.9, 1.2, and 1.4 ,ug of 0-deacylated lipid A's from H. pylori, C.jejuni, and S. typhimunium per ml, respectively. The lower reactivity ofH. pylori lipid A compared with those of the other two lipid A preparations (as shown by the higher EIso value) was underscored by the use of a murine monoclonal anti-lipid A antibody in the inhibition assay. An EIso value was not obtained at the concentrations tested for H. pylori lipid A; the corresponding figures for C. jejuni and S. typhimunium lipid A's were 13 and 14 ,ug/ml, respectively. No inhibition was obtained with H. pylori lipopolysaccharide, which showed a low-molecular-weight profile on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The activity of H. pylori lipid A in the Limulus assay was approximately 71 and 650 times lower than those of C. jejuni and S. typhimurium lipid A's, respectively. These findings suggest that lipid A is an integral part of the outer cell wall of H. pylori. The lower reactivity of H. pylori lipid A with anti-lipid A antibodies and in the Limulus assay compared with that of C. jejuni or S. typhimunium lipid A may be explained by a different composition of the fatty acids, especially the 3-hydroxy fatty acids, and a possible deviating phosphorylation pattern.Preparation of LPS and lipid A. For H. pylon and C. jejuni, the dried bacteria were first extracted with a chloroformmethanol mixture to remove the phospholipids. LPS was extracted by the phenol-water method of Westphal and Jann 4383 on August 1, 2020 by guest http://iai.asm.org/ Downloaded from

Detection of Helicobacter pylori in specimens obtained by endoscopy requires the gastroenterologist to select suitable patients for endoscopy, and to take an adequate number of biopsy specimens; there must also be correct cleaning of the biopsy forceps.