高通量测序技术在按蚊共生微生物研究中的应用

按蚊是疟疾的传播媒介,又称疟蚊。在按蚊体内,尤其是肠道内定殖着大量的共生微生物。共生微生物直接或间接地影响蚊虫的营养、发育、繁殖和免疫等重要生理活动。研究表明,肠道共生菌还能影响疟原虫的入侵、发育和传播。按蚊对常用杀虫剂逐渐产生抗性,疟原虫也对抗疟药产生了抗药性,严重阻碍了消除疟疾进程,迫切需要新的疟疾传播阻断战略。利用共生微生物控制和阻断疟疾传播是有潜力的方法之一,因此研究按蚊的共生微生物组成对理解按蚊、共生微生物以及病原体三者间的互作关系,识别用于防控疟疾的候选细菌有重要意义。高通量测序技术的应用有效克服了传统方法的局限性,极大地提高了对按蚊共生微生物多样性的认识。本文将从高通量测序技术在按蚊共生微生物组成、多样性和影响因素及其在疟疾防控中的应用进展进行综述。     

蚊肠道微生物调节蚊媒传染病传播的研究进展

蚊是一种吸血昆虫,在人群中通过吸血叮咬的方式感染并传播疟原虫、登革病毒、寨卡病毒等多种病原体,给全球公共卫生安全带来重大威胁。中肠组织是蚊虫抵抗病原体感染的第一道防线,蚊的中肠组织内栖息着大量的肠道微生物,当被感染病原体的血液进入到蚊的中肠组织后,某些肠道微生物通过分泌蛋白质、小分子等多种方式直接或间接影响病原体感染蚊中肠上皮层的过程,进而调控了病原体在蚊虫体内的复制传播。本综述总结了有关蚊肠道微生物调控蚊媒传染病传播的情况,阐述了不同种类的蚊肠道微生物影响病原体在蚊体内复制传播的机制。     

肠道菌群与益生菌

人类大肠中，栖息着数百种类多达100万亿个亦即重约1公斤的细菌。长期以来，就以探讨这些细菌对人类（宿主）的健康或病灾影响为研究课题。迄至最近，复以免疫学的进步与保健食品热的兴起，在有关肠道菌群及益生菌（probioties）应用于疾病防治的研究领域取得了很多的成果。所谓益生菌，是同抗生素（antibiotics）相对而言的来源于生物间共生关系（probiosis）的新词。其概念为“依宿主肠道菌群之调整而予自己以有益影响的微生物或其活性物质”。     

克罗恩病病因学研究进展与难治原因的剖析

克罗恩病（Crohn disease，CD）是肠粘膜上皮细胞对微生物抗原的异常免疫反应引起的慢性肠道炎症，在易感宿主甚至正常肠茵也可致病，正常肠腔内细菌产物可以启动肠道炎症并使之持久化。对鼠模型注射细菌胞壁成分可致慢性肉芽肿性肠炎，在一些鼠结肠炎模型中，肠茵己成为发生急、慢性炎症的必需。识别微生物的机制以及它与肠上皮的相互反应，对了解CD的发病很重要。由于肠上皮细胞经常暴露于细菌及其成分之下，它不仅是防御微生物的一种结构，同时也是一功能屏障，     

书讯

正由池肇春、段钟平教授主编的《肠道微生物与消化系统疾病》已由上海科学技术出版社出版,是国内首部有关肠道微生物与消化系统疾病的编著。新近几年在国内外对肠道微生物与消化系统疾病的相关性开展了基础和临床的研究,并取得了长足的进展。全书分上下两篇,上篇为总论,介绍肠道微生物研究现状与进展、细菌学、细菌生理功能、肠道屏障生理功能和屏障功能障碍、肠道细菌生态平衡和生态失调、细菌诊断、肠道微生物与食物消化和营养吸收、肠道微生物与药物代谢、肠道微生物与免疫、肠道微生物与炎症。下篇为肠道     

蚊虫肠道共生菌功能的研究进展

蚊虫肠道共生菌是指在蚊虫肠道内与其长期共同生活、维持肠道微生态稳定的菌群,其参与调控蚊虫生理功能的多个方面。本文简要综述了肠道共生菌的影响因素和其在蚊虫营养、免疫、繁殖以及抗药性等方面的功能,为进一步的研究提供参考。     

肠道菌群在门静脉高压发病机制中的作用

肠道微生物是人体中最大的共生微生物集合,而肝脏由于其解剖位置及独特的血管系统易暴露于肠道的微生物之中并受其代谢产物的影响。正常状态下由于肠上皮屏障功能和肝脏的清洁解毒功能,可防止微生物或微生物产物转移到肝脏和体循环中。然而,当肠屏障功能被破坏时,大量细菌产物可进入肝脏和全身循环,并刺激机体的免疫防御及炎症反应,而这些扰动在肝硬化中尤为突出,增加了临床上显著门静脉高压的发展。整合并总结了肠道菌群在门静脉高压发病机制中的作用,从而为临床治疗提供新的策略。     

书讯

正本刊讯由池肇春与段钟平教授主编的《肠道微生物与消化系统疾病》已由上海科学技术出版社出版,是国内首部有关肠道微生物与消化系统疾病的编著.近几年,国内外学者对肠道微生物与消化系统疾病的相关性开展了基础和临床的研究,并取得了长足的进展.此书分上下两篇:上篇为总论,介绍肠道微生物研究现状与进展、细菌学、细菌生理功能、肠道屏障生理功能和屏障功能障碍、肠道细菌生态平衡和生态失调、细菌诊断、肠道微生物与食物消化和营养吸收、     

乌干达齐卡林区未成熟非洲伊蚊和两种捕食巨蚊幼虫之间的相互作用

巨蚊属蚊虫在热带非洲广泛分布,其成虫吃花蜜、不吸血。而它们的幼虫能吞食其它蚊种幼虫,故有生物防制的作用。两种巨蚊为乌干达林区很普通的蚊种,其幼虫与传播森林黄热病的重要媒介非洲伊蚊共生于树洞积水。巨蚊幼虫以后者为食。作者曾提示这可能是非洲伊蚊虫群在该林区波动的一个因素。本文报告在有或无巨蚊存在条件下,对孳生场所中非洲伊蚊幼虫数量进行观察,评价林区巨蚊与伊蚊虫群波动的状况。 1974年1月至1975年12月作者在乌干达齐卡林区,随机选择30个自然树洞和30个人造竹罐,对非洲伊蚊幼虫、蛹以及两种巨蚊每2周1次逐一进行检查并记录。将每一孳生地的代表性幼虫和蛹带回实验室饲养至成虫,以便鉴定,余下水体部份仍倒回各自容     

书讯

正本刊讯由池肇春与段钟平教授主编的《肠道微生物与消化系统疾病》已由上海科学技术出版社出版,是国内首部有关肠道微生物与消化系统疾病的编著.近几年,国内外学者对肠道微生物与消化系统疾病的相关性开展了基础和临床的研究,并取得了长足的进展.此书分上下两篇:上篇为总论,介绍肠道微生物研究现状与进展、细菌学、细菌生理功能、肠道屏障生理功能和屏障功能障碍、肠道细菌生态平衡和生态失调、细菌诊断、肠道微生物与食物消化和营养吸收、肠道微生物与药物     

共生菌在控制蚊媒传染病中的研究进展

蚊子作为疟疾、黄热病等传染病的主要传播媒介,严重威胁着全球流行地区的人类健康。目前,蚊媒控制措施除了使用化学杀虫剂外,遗传控制也是一种新的尝试。但考虑到生态安全性,科学家又提出了一种新型控蚊策略——共生控制。由于蚊子体内拥有多种共生菌群,且这些共生菌存在于包括消化道在内的各种组织中,因此可以通过影响宿主的寿命、繁殖能力、媒介能力等来影响蚊媒传染病传播。本文对蚊体内共生菌之间的相互作用进行简述,并对蚊体内共生菌在控制蚊媒传染病中的研究进展进行综述和展望。     

Ecological safety of mosquitocidal biocides based on Bacillus thuringiensis israelensis

Bacillus thuringiensis israelensis (Bti) has been developed into many products for the biological control of dipteran larvae, including mosquitoes (Culicidae), black flies (Simuliidae), and midges (Chironomidae) in various parts of the World. Bti appears to pose significantly less of a risk than other chemical pesticides used for mosquito control and eradication programs. Bioproducts based on Bti are highly selective with short environmental persistence, and thus they have very little potential to cause damage to populations of non-target organisms. So far, no example of an unexpected pathogenic organism being developed in the field as well as no examples of resistance to Bti both laboratory and field populations of mosquitoes have been documented. There are some indications that large declines in insect biomass can occur after long-term use of Bti in freshwater wetlands. However, no evidence for permanent damage to ecosystem function has been found. Organisms that utilized insects for food, adapted to the declines and either switched to other food sources or migrate (birds) outside of the treated zones to acquire insects. Even though over 40 tons of Bti have been applied in West Africa alone, no indications of human health or non-target effects have been reported.     

16S rRNA phylogenetic analysis of the bacterial endosymbionts associated with cytoplasmic incompatibility in insects.

Bacterial endosymbionts of insects have long been implicated in the phenomenon of cytoplasmic incompatibility, in which certain crosses between symbiont-infected individuals lead to embryonic death or sex ratio distortion. The taxonomic position of these bacteria has, however, not been known with any certainty. Similarly, the relatedness of the bacteria infecting various insect hosts has been unclear. The inability to grow these bacteria on defined cell-free medium has been the major factor underlying these uncertainties. We circumvented this problem by selective PCR amplification and subsequent sequencing of the symbiont 16S rRNA genes directly from infected insect tissue. Maximum parsimony analysis of these sequences indicates that the symbionts belong in the alpha-subdivision of the Proteobacteria, where they are most closely related to the Rickettsia and their relatives. They are all closely related to each other and are assigned to the type species Wolbachia pipientis. Lack of congruence between the phylogeny of the symbionts and their insect hosts suggest that horizontal transfer of symbionts between insect species may occur. Comparison of the sequences for W. pipientis and for Wolbachia persica, an endosymbiont of ticks, shows that the genus Wolbachia is polyphyletic. A PCR assay based on 16S primers was designed for the detection of W. pipientis in insect tissue, and initial screening of insects indicates that cytoplasmic incompatibility may be a more general phenomenon in insects than is currently recognized.     

Heterologous expression in transgenic mosquitoes

Arthropod-borne diseases such as malaria and dengue virus afflict billions of people worldwide imposing major economic and social burdens. Control of such pathogens is mainly performed by vector management and treatment of affected individuals with drugs. The failure of these conventional approaches due to emergence of insecticide-resistant insects and drug-resistant parasites demonstrate the need of novel and efficacious control strategies to combat these diseases. Genetic modification (GM) of mosquito vectors to impair their ability to be infected and transmit pathogens has emerged as a new strategy to reduce transmission of many vector-borne diseases and deliver public health gains. Several advances in developing transgenic mosquitoes unable to transmit pathogens have gained support, some of them attempt to manipulate the naturally occurring endogenous refractory mechanisms, while others initiate the identification of an exogenous foreign gene which disrupt the pathogen development in insect vectors. Heterologous expression of transgenes under a native or heterologous promoter is important for the screening and effecting of the transgenic mosquitoes. The effect of the transgene on mosquito fitness is a crucial parameter influencing the success of this transgenic approach. This review examines these two aspects and describes the basic research work that has been accomplished towards understanding the complex relation between the parasite and its vector and focuses on recent advances and perspectives towards construction of transgenic mosquitoes refractory to vector-borne disease transmission.     

按蚊中肠细菌及其应用的研究进展

按蚊中肠细菌能够在控制传疟按蚊种群数量和抑制疟原虫在按蚊体内的发育等方面发挥重要作用。本文从细菌与按蚊的食物关系、按蚊幼虫和成蚊中肠细菌及其应用、细菌在按蚊体内的经发育期传递、细菌与按蚊成蚊的免疫反应,以及利用细菌防控疟疾流行的可行性等方面进行综述。     

Bacterial gut symbionts are tightly linked with the evolution of herbivory in ants

Ants are a dominant feature of terrestrial ecosystems, yet we know little about the forces that drive their evolution. Recent findings illustrate that their diets range from herbivorous to predaceous, with “herbivores” feeding primarily on exudates from plants and sap-feeding insects. Persistence on these nitrogen-poor food sources raises the question of how ants obtain sufficient nutrition. To investigate the potential role of symbiotic microbes, we have surveyed 283 species from 18 of the 21 ant subfamilies using molecular techniques. Our findings uncovered a wealth of bacteria from across the ants. Notable among the surveyed hosts were herbivorous “turtle ants” from the related genera Cephalotes and Procryptocerus (tribe Cephalotini). These commonly harbored bacteria from ant-specific clades within the Burkholderiales, Pseudomonadales, Rhizobiales, Verrucomicrobiales, and Xanthomonadales, and studies of lab-reared Cephalotes varians characterized these microbes as symbiotic residents of ant guts. Although most of these symbionts were confined to turtle ants, bacteria from an ant-specific clade of Rhizobiales were more broadly distributed. Statistical analyses revealed a strong relationship between herbivory and the prevalence of Rhizobiales gut symbionts within ant genera. Furthermore, a consideration of the ant phylogeny identified at least five independent origins of symbioses between herbivorous ants and related Rhizobiales. Combined with previous findings and the potential for symbiotic nitrogen fixation, our results strongly support the hypothesis that bacteria have facilitated convergent evolution of herbivory across the ants, further implicating symbiosis as a major force in ant evolution.     

Fighting malaria with engineered symbiotic bacteria from vector mosquitoes

The most vulnerable stages of Plasmodium development occur in the lumen of the mosquito midgut, a compartment shared with symbiotic bacteria. Here, we describe a strategy that uses symbiotic bacteria to deliver antimalaria effector molecules to the midgut lumen, thus rendering host mosquitoes refractory to malaria infection. The Escherichia coli hemolysin A secretion system was used to promote the secretion of a variety of anti-Plasmodium effector proteins by Pantoea agglomerans, a common mosquito symbiotic bacterium. These engineered P. agglomerans strains inhibited development of the human malaria parasite Plasmodium falciparum and rodent malaria parasite Plasmodium berghei by up to 98%. Significantly, the proportion of mosquitoes carrying parasites (prevalence) decreased by up to 84% for two of the effector molecules, scorpine, a potent antiplasmodial peptide and (EPIP)(4), four copies of Plasmodium enolase-plasminogen interaction peptide that prevents plasminogen binding to the ookinete surface. We demonstrate the use of an engineered symbiotic bacterium to interfere with the development of P. falciparum in the mosquito. These findings provide the foundation for the use of genetically modified symbiotic bacteria as a powerful tool to combat malaria.     

Field prevalence of Wolbachia in the mosquito vector Aedes albopictus

The endosymbiotic bacteria in the genus Wolbachia have been proposed as a potential candidate to deliver pathogen-blocking genes into natural populations of medically important insects. The successful application of Wolbachia in insect vector control depends on the ability of the agent to successfully invade and maintain itself at high frequency under field conditions. Here, we evaluated the prevalence of Wolbachia infections in a field population of the Wolbachia-superinfected mosquito Aedes albopictus. A field prevalence of 100% (n = 1,016) was found in a single population in eastern Thailand via polymerase chain reaction (PCR) testing of Wolbachia both from individual parent females and their corresponding F1 offspring. This is the first report of accurate Wolbachia prevalence in a field population of an insect disease vector. The prevalence of superinfection was estimated to be 99.41%. All single-infected individual mosquitoes (n = 6) were found to harbor group A Wolbachia. For this particular population, none was found to be single-infected with group B Wolbachia. Our results also show that PCR testing of field materials alone without checking F1 offspring overestimated the natural prevalence of single infection. Thus, the confirmation of infection status by means of F1 offspring was critical to the accurate estimates of Wolbachia prevalence under field conditions.     

Xenodiagnosis: use of mosquitoes for the diagnosis of arboviral infections

The arboviruses have a worldwide distribution and, mosquitoes and ticks contribute principally in their transmission. In the last two decades, arboviral diseases have been recognised due to their resurgence and spread in newer geographic areas. Surveys to determine the prevalence of arboviruses in any region largely depend on the isolation attempts from the arthropods along with the serosurveys. Xenodiagnosis means use of insects for the diagnosis of infectious diseases affecting human being. The present communication discusses the application of mosquitoes for propagation and assays of arboviruses, the technique of mosquito inoculation and importance of xenodiagnosis.