主机表型的考试<em>食蚊鱼</em>继抗生素治疗
Summary
这项研究涉及到的方法来揭示继抗生素皮肤和肠道微生物群落组成的改变模型鱼主机上的影响。
Abstract
The commonality of antibiotic usage in medicine means that understanding the resulting consequences to the host is vital. Antibiotics often decrease host microbiome community diversity and alter the microbial community composition. Many diseases such as antibiotic-associated enterocolitis, inflammatory bowel disease, and metabolic disorders have been linked to a disrupted microbiota. The complex interplay between host, microbiome, and antibiotics needs a tractable model for studying host-microbiome interactions. Our freshwater vertebrate fish serves as a useful model for investigating the universal aspects of mucosal microbiome structure and function as well as analyzing consequential host effects from altering the microbial community. Methods include host challenges such as infection by a known fish pathogen, exposure to fecal or soil microbes, osmotic stress, nitrate toxicity, growth analysis, and measurement of gut motility. These techniques demonstrate a flexible and useful model system for rapid determination of host phenotypes.
Introduction
它已经确定，抗生素可以破坏人类微生物导致生态失调，这意味着微生物群落的不平衡。抗生素治疗后的微生物群的组成的改变已经显示降低社会的多样性，减少主要成员，并改变社区代谢，特别是在肠道1,2。肠道微生物的抗生素干扰可以减少定植抗力艰难梭菌 3,4和沙门氏菌 5。
此外，该微生物群的破坏已与许多综合征和人类疾病的发展（ 例如，抗生素相关性小肠结肠炎，炎性肠病，代谢紊乱， 等等 ）。抗生素也被广泛的农业实现为在生长促进畜禽生产6。这些功能强大的工具的使用也不是没有附带影响，这是抗生素耐药性的迅速崛起，以及一个破坏微生物的作用明显有它的有人居住的主机。许多研究表明，广谱抗生素的使用具有长期持久的后果的微生物群的结构和功能，然而，从抗生素被破坏的微生物影响宿主生理副作用仅具有尚待支持推测。
主机，微生物和抗生素之间的相互作用是远远以简洁的方式理解。因此，一个简单而更容易处理模式有利于高度复杂的哺乳动物系统上脱落光。在人体黏膜表面，包括肠道，怀有最高的密度和微生物的多样性，也是最亲密的微生物 – 宿主相互作用。鱼提供S上的粘膜皮肤微生物everal优势作为模型系统。的真骨鱼类 （硬骨鱼）是最早谱系的脊椎动物的含义，硬骨鱼既有先天和后天已经共同发展与共生的细菌群落7的关系的免疫系统内的分歧之一。鱼皮股哺乳动物1型粘膜表面，如生理功能，免疫组件和黏液产生细胞8的排列许多特性。鱼皮粘膜表面的外部位置提供了一个微生物易于操纵实验和样品。
西方蚊， 食蚊（G. 慈），是已经在过去已经用于研究交配和毒理学9，10，11的模型的鱼。由于体积小，种群数量在野外为入侵物种，男 inimal保健费用，和顽强的本性，我们已经开发G.慈竹作为粘膜微生物模式。此外， 食蚊鱼分享生下活年轻，胎生哺乳动物，这是鱼的种类罕见的生理。我们完成了鱼皮正常菌群的时间用16S最广泛的研究， 食蚊鱼 12剖析。进一步的工作表明以下使用广谱抗生素13的皮肤和肠道菌群破坏宿主三个负面影响。
在鱼以下抗生素暴露检查五个不同的效果。该微生物的最完善的主机的好处是病原体竞争排斥。鱼类病原爱德华氏菌是已知会导致肠道败血症的爆发商业鲶鱼养殖场14。 E.氏菌也被证明致死感染斑马鱼类=“外部参照”> 15，16和17 食蚊鱼 。从水柱这种病菌的一个挑战可以作为排斥的措施。作为比较，以易感性的个体的病原体，暴露于混合生物的高密度时存活率也进行了。粪便和富含有机物的土壤被用作微生物群落的普遍遇到的来源。
细菌肠社区执行另一建立角色是养分处理和能量收获，从而影响该主机的整体营养摄取。作为营养的严重测量，鱼体重的被喂食标准饮食前一个月和后进行比较。抗生素治疗鱼的平均体重下降，而平均控制鱼的体重增加超过一个月。这种缺乏体重增加的机制尚不清楚。一个可能的因素是在肠的食物的运送时间。地理标志莫蒂lity方法从斑马鱼（Adam Rich在，纽约州立大学布鲁克波特，个人通信）适用于确定运输时间。它没有如抗生素治疗鱼类具有改变的过境时间尚未确定。
所有的生物体，特别是鱼类，在自然的环境中遇到的一个共同挑战是渗透胁迫。 食蚊鱼已被证明，当在高浓度的盐分18敏锐地强调快速适应。出人意料的是，鱼表现出抗生素的微生物改变生存下降到一个高盐胁迫。对于这种新颖的表型的机制正在调查中。对水生动物，尤其是在水族箱另一种常见的应力，是氮的毒性形式（氨，硝酸盐，亚硝酸盐和）。针对硝酸盐生存不是抗生素治疗和控制鱼之间显著不同。在这个手稿提出的方法可与食蚊或类似鱼模式生物，如斑马使用鱼和青鳉，测量下列实验操作中的鱼的表型。
Protocol
Representative Results
Discussion
一些挑战需要抗生素治疗的药物在鱼组织被耗尽后，清洁APW休息时间。如果暂停时间段被跳过然后抗生素存在可混淆的结果，特别是当测定涉及暴露于细菌。为了研究而无需在主机上，初步实验监控微生物组成（16S分析或全基因组测序）和人口密度（通过定量PCR 16S定量）对微生物的总数大的变化抗生素暴露期间从改变肠道微生物组成的影响将是需要。而3天在本系统中是最优的，改变宿主和/或抗?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
This project was partially funded by a FAST (Faculty and Student Team) Award to TPP and JMC from EURECA (Center for Enhancing Undergraduate Research Experiences and Creative Activities) at Sam Houston State University.
Materials
| Rifampicin | Calbiochem | 557303-1GM | |
| Sodium Nitrate | Sigma Aldrich | S5506 | |
| Fluorescein-labeled 70 kDa anionic dextran | ThermoFisher Scientific | D1823 | |
| PBS tablets | Calbiochem | 6500-OP | tablets dissolve in water to make phosphate-buffered saline |
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