Conjugate-induced Polysaccharide Antibodies

革兰氏阴性肠道病原菌的表面多糖，以被膜或脂多糖的形式存在，既是重要的毒力因子，又是保护性抗原。这些多糖通过与载体蛋白结合而增强了免疫原性。在伤寒发病率较高的越南，成人和儿童的连续临床研究表明，与重组铜绿假单胞菌外蛋白A(REPA)结合的胶囊多糖(Vi)可引起对伤寒的高应答。在第三阶段的试验中，大约12,000名2-5岁的儿童注射了结合疫苗，没有表现出明显的副作用。Vi-REPA主动监测27个月有效率为91%，被动监测28~48个月有效率为85%。Pacebo组儿童在2.5年后接种结合疫苗。甲型副伤寒沙门氏菌是东南亚第二常见的肠道热病因，结合疫苗被发现对成人、青少年和2-4岁的儿童是安全和免疫原性的。在中国东南部，甲型副伤寒病例已超过斑疹伤寒。计划进行第三阶段临床试验。大肠杆菌O157是一种新出现的病原体，会导致幼儿溶血性尿毒症综合征。O157 O特异性多糖-REPA结合物在成年志愿者中的第一阶段研究证明了其安全性和免疫原性。对2-5岁儿童的O157结合物的第二阶段研究正在进行中。给儿童注射一两次疫苗表明疫苗是安全的。免疫原性数据将被测量。无毒志贺毒素I和II是从突变的E.ColiO157中提纯出来的，并将与O特异的多糖偶联制成双价疫苗。O157大肠杆菌的主要宿主是牛。脂多糖-蛋白结合物在牛身上显示出免疫原性，并计划进行一项攻击性研究，以证明牛体内携带状态的清除。霍乱弧菌O1和O139是霍乱感染的主要血清型。O139的包膜多糖与O1的脂多糖合成的偶联物能诱导小鼠产生杀弧菌抗体。稻叶和小川两种血清型的脱毒脂多糖已被用作结合疫苗的基础。在动物实验中，结合物诱导的抗体水平高于单独使用内毒素的水平。将制备一种化学合成的小川O-特异性多糖，并将其偶联到蛋白质载体上。比较用天然纯化和化学合成的O-特异多糖制备的霍乱疫苗的免疫原性。这些结合物的临床试验正在计划中。在发达国家最常见的沙门氏菌感染之一是鼠伤寒沙门氏菌。耐药菌株在疾病分离株中很常见。我们比较了最常见和耐药菌株的O-特异性多糖结构。我们发现，O-乙酰基是不同菌株的主要抗原决定簇：O-乙酰阳性菌株与O-乙酰阴性菌株的交叉反应很差。因此，疫苗制备菌株的选择将取决于流行病学调查和细菌学鉴定。弯曲杆菌感染是美国最常见的肠道感染之一。关于引起疾病的流行类型或保护性抗原，人们知之甚少。我们已经开始收集流行病学数据，并在各种生长条件下进行实验。

Surface polysaccharides of Gram-negative enteric pathogens, in the form of capsule or lipopolysaccharide, are both essential virulence factors and protective antigens. The immunogenicity of these polysaccharides were enhanced by binding to carrier proteins. Sequential clinical studies in adults and in children in Vietnam, an area with a high attack rate of typhoid, showed that the capsular polysaccharide (Vi) conjugates bound to the recombinant Pseudomonas aeruginosa exoprotein A (rEPA) elicited high responses against typhi. In a Phase III trial, About 12,000 2-5 years old children injected with the conjugate vaccine showed no significant side reaction. The efficacy of Vi-rEPA was 91% after 27 months active and 85% after 28-48 months passive surveillance. Chidren in the pacebo group were given the conjugate vaccine 2.5 years later. Salmonella paratyphi A, the second most common cause of enteric fever in Southeast Asia, conjugate vaccine was found to be safe and immunogenic in adults, teenagers and then 2-4 year old children. In southeast China, cases of paratyphi A has exceeded those of typhi. A phase III clinical trial is planned. Escherichia coli O157, an emerging pathogen, causes hemolytic uremic syndrome in young children. Phase 1 study of E. coli O157 O-specific polysaccharide-rEPA conjugate demonstrated safety and immunogenicity in adult volunteers. The phase II study of E. coli O157 conjugate in 2-5 years old children is underway. Children injected once or twice showed that the vaccine was safe. Immunogenicity data are to be measured. Non-toxic shiga toxin I and II are purified from mutant E. coli O157 and will be conjugated with O-specific polysaccharide for a bivalent vaccine. The major reservoir of E.coli O157 is cattle. LPS-protein conjugate showed to be immunogenic in cattle and a challenge study is planned to demonstrate the clearance of the carriage state in cattle. Vibrio cholera O1 and O139 are the major sero types in cholera infections. Conjugates synthesized with capsular polysaccharide of O139 and O-specific polysaccharide of LPS of O1 elicited vibriocidal antibodies in mice. Detoxified LPS from both Inaba and Ogawa serotypes have been used as a base for conjugate vaccine. In animal study, the conjugates elicited higher antibody level than LPS alone. A chemical synthesized O-specific polysaccharide for Ogawa will be prepared and conjugated to protein carrier. The immunogenicity of cholera vaccine prepared with naturally purified or chemically synthesized O-specific polysaccharide will be compared. Clinical trials of these conjugates are planned. One of the most common Salmonella infection in developed country is Salmonella typhimurium. Antibiotic resistant strains have been common in disease isolates. We have compared the O-specific polysaccharide structure of the most common and antibiotic resistant strains. We found that the O-acetyl group on the abequose is a major antigenic determinant for various strains: O-acetyl positive strains cross react with O-acetyl negative strains poorly. Selection of strains for vaccine preparation will therefore rely on the epidemiology survey and bacteriology identification. Campylobacter infection is one of the most common enteric infection in the US. Little is known about the prevalent type that causes disease or the protective antigens. We have started to collect epidemiology data and experimenting with various growth conditions.