UV-irradiated rotifers for the maintenance of gnotobiotic zebrafish larvae.

IF 3.7 2区 生物学 Q2 MICROBIOLOGY
mSphere Pub Date : 2025-03-25 Epub Date: 2025-02-24 DOI:10.1128/msphere.00698-24
Susana Márquez Rosales, Peter I Bouchard, Emily M Olmstead, Raghuveer Parthasarathy
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引用次数: 0

Abstract

Host-associated microbial communities profoundly impact the health of humans and other animals. Zebrafish have proven to be a useful model for uncovering mechanisms of host-microbe interactions, but the difficulty of maintaining germ-free or gnotobiotic zebrafish beyond 1 week post-fertilization has limited their utility. To address this, we have developed a simple protocol using UV irradiation of rotifers, a common and nutrient-rich prey species for larval zebrafish, to reduce the bacterial load associated with the rotifers by several orders of magnitude while maintaining their motility and viability. We find that though feeding with UV-treated rotifers does not preserve the sterility of germ-free fish, it enables the maintenance of pre-existing bacterial communities. Normal feeding, in striking contrast, leads to the near-total depletion of these prior populations. We measure the abundance of single- and three-species consortia of zebrafish-commensal bacteria inoculated into initially germ-free larvae in a series of experiments extending to 8 days of feeding, or 13 days post-fertilization. We find, in fish-fed UV-treated rotifers, the persistence of bacterial populations on timescales of days, together with strong species-specific variation. In addition, re-inoculation of differently labeled strains of the same zebrafish-commensal species alongside feeding leads to colonization by the new bacteria without displacement of earlier microbes. Our method will facilitate the use of gnotobiotic zebrafish for investigations of phenomena that emerge later in animal development and for studies that probe microbiome composition fluctuations and stability over extended timescales.IMPORTANCEAll animals, including humans, are host to vast microbial communities that contribute to health and disease through mechanisms that remain largely mysterious. These microbiomes are challenging to study, spurring the use of various model organisms, including zebrafish. Zebrafish, however, are difficult to raise beyond 1 week post-fertilization under gnotobiotic conditions, in other words, germ free or with known microbial constituents, a consequence of normally feeding on live prey that brings their own, generally unknown, microbes. Therefore, we developed a simple protocol in which UV irradiation of rotifers, a widely used small-animal food for larval zebrafish, facilitates the maintenance of gnotobiotic larvae. We show that pre-existing bacterial communities in larvae are minimally affected by feeding on UV-treated rotifers, in strong contrast to feeding on untreated rotifers. We demonstrate that this feeding method allows investigations of zebrafish-associated bacterial community stability over several days, allowing investigation of previously intractable questions about microbiome stability.

紫外线照射下的轮虫对零生斑马鱼幼虫的维持作用。
与宿主相关的微生物群落深刻地影响着人类和其他动物的健康。斑马鱼已被证明是揭示宿主-微生物相互作用机制的有用模型,但在受精后1周内维持无菌或无生物斑马鱼的困难限制了它们的应用。为了解决这个问题,我们开发了一种简单的方案,使用紫外线照射轮虫(斑马鱼幼虫常见且营养丰富的猎物),以减少与轮虫相关的细菌负荷几个数量级,同时保持它们的活力和活力。我们发现,虽然用紫外线处理的轮虫喂养并不能保持无菌鱼的不育性,但它可以维持原有的细菌群落。与之形成鲜明对比的是,正常的喂食会导致这些原有种群几乎全部灭绝。在一系列实验中,我们测量了斑马鱼-共生细菌的单一和三种联合体的丰度,这些细菌接种到最初的无菌幼虫中,持续到饲养8天或受精后13天。我们发现,在鱼喂养的紫外线处理的轮虫中,细菌种群在天的时间尺度上持续存在,同时具有很强的物种特异性变异。此外,在饲养的同时,重新接种同一斑马鱼共生物种的不同标记菌株会导致新细菌的定植,而不会取代早期的微生物。我们的方法将有助于利用非生斑马鱼研究动物发育后期出现的现象,以及在较长时间尺度上探索微生物组组成波动和稳定性的研究。所有动物,包括人类,都是大量微生物群落的宿主,这些微生物群落通过很大程度上仍然神秘的机制促进健康和疾病。这些微生物组的研究具有挑战性,刺激了各种模式生物的使用,包括斑马鱼。然而,斑马鱼很难在受精后1周内在非生菌条件下饲养,换句话说,无菌或含有已知微生物成分,这是通常以活体猎物为食的结果,这些猎物通常会带来自己的微生物,通常是未知的。因此,我们开发了一种简单的方案,即紫外线照射轮虫(一种广泛用于斑马鱼幼虫的小动物食物),以促进生物幼虫的维持。我们表明,喂食紫外线处理的轮虫对幼虫中预先存在的细菌群落的影响最小,与喂食未经处理的轮虫形成强烈对比。我们证明,这种喂养方法可以在数天内调查斑马鱼相关细菌群落的稳定性,从而调查以前关于微生物组稳定性的棘手问题。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
mSphere
mSphere Immunology and Microbiology-Microbiology
CiteScore
8.50
自引率
2.10%
发文量
192
审稿时长
11 weeks
期刊介绍: mSphere™ is a multi-disciplinary open-access journal that will focus on rapid publication of fundamental contributions to our understanding of microbiology. Its scope will reflect the immense range of fields within the microbial sciences, creating new opportunities for researchers to share findings that are transforming our understanding of human health and disease, ecosystems, neuroscience, agriculture, energy production, climate change, evolution, biogeochemical cycling, and food and drug production. Submissions will be encouraged of all high-quality work that makes fundamental contributions to our understanding of microbiology. mSphere™ will provide streamlined decisions, while carrying on ASM''s tradition for rigorous peer review.
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