The role of CFA/I in adherence and toxin delivery by ETEC expressing multiple colonization factors in the human enteroid model.
The role of CFA/I in adherence and toxin delivery by ETEC expressing multiple colonization factors in the human enteroid model.
复制标题
CFA/I在人肠模型中表达多种定植因子的ETEC的粘附和毒素递送中的作用。
DOI：
10.1371/journal.pntd.0010638
复制
发表时间：
2022-07
影响因子：
3.8
通讯作者：
中科院分区：
文献类型：
--
作者：
产肠毒素大肠杆菌（ETEC）是中低收入国家旅行者和幼儿腹泻的主要病原体。ETEC通过定殖因子（CFs）粘附于肠上皮，分泌热稳定毒素（ST）和/或热不稳定毒素（LT），导致细胞离子运输和水分分泌失调。ETEC分离物通常含有编码多个CF的基因，这些CF是作为疫苗抗原的靶标。CFA/I是与ETEC相关的主要CF，可引起中重度腹泻，在发病机制中起重要作用。全球肠道多中心研究发现，78%表达CFA/ i的ETEC也编码较小的CF CS21，这促使人们对这两种CF的联合作用进行了研究。Western blots和电镜显示，CFA/I和CS21的表达存在生长介质依赖性和菌株依赖性差异。通过人类肠道模型和一系列CFA/I和CS21特异性突变体，证明了CFA/I在表达CFA/I和CS21的ETEC菌株粘附过程中的关键作用。此外，只有抗CFA/I抗体抑制表达CFA/I和CS21的全球ETEC分离株的粘附。在感染肠道单层的上清液中测量ST的递送和由此产生的cGMP分泌，并观察菌株特异性ST的递送和时间依赖性cGMP的产生。有趣的是，cGMP水平在野生型和cf缺乏菌株之间相似，反映了这种静态有氧感染模型的局限性。尽管有ETEC的粘附和ST的递送，肠样单分子层的完整性没有被破坏，这表明在感染期间上皮电阻没有减少，IL-8细胞因子也没有产生。综上所述，这些数据表明，在全球临床CFA/I- cs21菌株中靶向CFA/I足以抑制粘附性，支持专注于阻断主要CFA的疫苗策略。此外，人类肠道模型在研究ETEC发病机制和评估疫苗诱导的功能性抗体反应方面具有重要的实用价值。产肠毒素大肠杆菌（ETEC）是中低收入国家旅行者和幼儿腹泻的主要原因。ETEC通过附着在小肠表面的称为定植因子（CFs）的粘附素感染小肠，并将毒素输送到肠道。这些CFs是开发ETEC疫苗诱导定植阻断反应的最佳靶点。在目前的研究中，我们利用人类肠道模型研究了两种cf，即CFA/I和CS21，当它们在地理上不同的ETEC临床分离株中同时表达时，在感染中的作用。该系统是研究肠道病原体最具人类相关性和敏感性的模型。我们确定CFA/I对表达CFA/I和CS21的ETEC菌株粘附人肠道很重要。菌株特异性的毒素传递水平被量化，但野生型和缺乏cf的突变株之间没有差异。这一结果强调了在模型中考虑肠道运动和氧水平影响的机会。受感染的肠道没有破坏细胞单层或产生炎症信号。这项研究支持了在ETEC疫苗开发中靶向CFA/I的策略，并强调了肠道是一个重要的模型系统。
Enterotoxigenic Escherichia coli (ETEC) is a primary causative agent of diarrhea in travelers and young children in low-to-middle-income countries (LMICs). ETEC adhere to intestinal epithelia via colonization factors (CFs) and secrete heat-stable toxin (ST) and/or heat-labile toxin (LT), causing dysregulated cellular ion transport and water secretion. ETEC isolates often harbor genes encoding more than one CF that are targets as vaccine antigens. CFA/I is a major CF that is associated with ETEC that causes moderate-to-severe diarrhea and plays an important role in pathogenesis. The Global Enteric Multicenter Study finding that 78% of CFA/I-expressing ETEC also encode the minor CF CS21 prompted investigation of the combined role of these two CFs. Western blots and electron microscopy demonstrated growth media-dependent and strain-dependent differences in CFA/I and CS21 expression. The critical role of CFA/I in adherence by ETEC strains expressing CFA/I and CS21 was demonstrated using the human enteroid model and a series of CFA/I- and CS21-specific mutants. Furthermore, only anti-CFA/I antibodies inhibited adherence by global ETEC isolates expressing CFA/I and CS21. Delivery of ST and resulting cGMP secretion was measured in supernatants from infected enteroid monolayers, and strain-specific ST delivery and time-dependent cGMP production was observed. Interestingly, cGMP levels were similar across wildtype and CF-deficient strains, reflecting a limitation of this static aerobic infection model. Despite adherence by ETEC and delivery of ST, the enteroid monolayer integrity was not disrupted, as shown by the lack of decrease in transepithelial electrical resistance and the lack of IL-8 cytokines produced during infection. Taken together, these data demonstrate that targeting CFA/I in global clinical CFA/I-CS21 strains is sufficient for adherence inhibition, supporting a vaccine strategy that focuses on blocking major CFs. In addition, the human enteroid model has significant utility for the study of ETEC pathogenesis and evaluation of vaccine-induced functional antibody responses. Enterotoxigenic Escherichia coli (ETEC) is a major cause of diarrhea in travelers and in young children in low-to-middle-income countries (LMIC). ETEC infects the small intestine using surface located adhesins called colonization factors (CFs) and delivers toxins to the intestine. These CFs are optimal targets for ETEC vaccine development for the induction of colonization-blocking responses. In the current study we investigated the role of 2 CFs, namely CFA/I and CS21 when they are simultaneously expressed in geographically diverse ETEC clinical isolates, in infection using the human enteroid model. This system is the most human relevant and sensitive model to study enteric pathogens. We determined that CFA/I was important for adherence to the human enteroid by ETEC strains expressing CFA/I and CS21. Strain-specific levels of toxin delivery were quantified but there was no difference between wildtype and CF-deficient mutant strains. This result highlights the opportunity to consider the effects of intestinal movement and oxygen levels in the model. Infected enteroids did not have disrupted cell monolayers or produce inflammatory signals. This study supports the strategy to target CFA/I in ETEC vaccine development and underscores the enteroid as a significant model system.
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DOI：
10.1093/infdis/jiab359
发表时间：
2021-12-20
期刊：
The Journal of infectious diseases
影响因子：
--
作者：
Fleckenstein JM;Sheikh A
通讯作者：
Sheikh A
肠毒素大肠杆菌临床分离株的表型和基因型表征来自北美北部。
影响因子：
--
作者：
Guerra JA;Romero-Herazo YC;Arzuza O;Gómez-Duarte OG
通讯作者：
Gómez-Duarte OG
影响因子：
6.4
作者：
Freedman, DJ;Tacket, CO;Crabb, JH
通讯作者：
Crabb, JH
影响因子：
34.3
作者：
Anderson, John D.;Bagamian, Karoun H.;Rheingans, Richard
通讯作者：
Rheingans, Richard
影响因子：
3.5
作者：
Giron, JA;GomezDuarte, OG;Kaper, JB
通讯作者：
Kaper, JB