Reply to Pantaleón García et al.
Reply to Pantaleón García et al.
复制标题
回复 Pantaleón García 等人。
DOI：
10.1152/ajplung.00205.2022
复制
发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Willis,KentA
中科院分区：
文献类型：
--
作者：
Ambalavanan,Namasivayam;Willis,KentA
致编辑：我们要感谢 Pantaleón García 等人 (1) 对之前工作 (2) 的澄清。我们同意他们的观点，即解决微生物群工作中的多种潜在混杂因素是很困难的，因此我们饶有兴趣地阅读了他们在信中对分层随机笼养的解释。我们赞扬他们对先验随机化设计的承诺，这是临床前研究中经常缺乏的严格方法。我们欢迎此类技术不仅在微生物组研究领域而且在整个动物模型研究中得到更广泛的采用——即使微生物组没有被有意研究。尽管实验开始时的分层笼随机化是限制笼效应的极好方法（3），但我们想强调小鼠微生物群研究的两个局限性，这些局限性经常被忽视。首先要考虑的是，在特定的无病原体条件下饲养的小鼠经常会受到其所饲养的动物设施特有的系统性混杂因素的影响 (4-6)。这与微生物群中的非细菌成员（例如真菌）尤其相关，它们饲养的时间越长，群落复杂性可能会稳步降低 (7)。减轻与动物饲养设施相关的系统性影响的一些潜在方法可能包括对微生物组进行连续采样、将小鼠随机分配到不同的位置以及从不同的商业供应商购买小鼠。其次，在特定的无病原体条件下，微生物组的主要来源是母亲 (6, 8)，其程度远远大于野生或野化小鼠或人类。这种“母体效应”因兄弟姐妹同住和相关的食粪行为而变得更加复杂。当目标是询问具有不同基因型的小鼠时，减轻母体效应可能包括使用相同接种物的粪便微生物群转移到无菌小鼠体内、随机交叉寄养或使用混合基因型窝等技术。尽管正如 Lipinski 等人 (2) 所观察到的那样，广泛的同窝确实倾向于使微生物群正常化，但 Robertson 等人 (8) 的研究表明，使用 F2 同窝仔猪可以使微生物群正常化更可靠。
TO THE EDITOR: We would like to thank Pantaleón García et al.(1) for their clarifications regarding their prior work (2). We agree with them that addressing the multiple potential confounders in microbiota work is difficult, and so we read with interest their explanation of stratified random caging in their Letter. We applaud their commitment to an a priori randomization design, a rigorous approach that is often sorely lacking in preclinical research. We would welcome a wider adoption of such techniques across not just the field of microbiome research but throughout animal model research—even when the microbiome is not being intentionally investigated.Although stratified cage randomization at the beginning of an experiment is an excellent way to limit cage effects (3), we would like to highlight two limitations of murine microbiota research that often go unappreciated. The first consideration is that mice housed under specific pathogen-free conditions are often subjected to systemic confounders unique to the animal facility in which they are housed (4–6). This is particularly relevant to the nonbacterial members of the microbiota, such as fungi, that may exhibit a steady reduction in community complexity the longer they are housed (7). Some potential approaches to mitigating systemic effects related to the animal housing facility may include serial sampling of the microbiome, randomizing mice to different locations, and purchasing mice from different commercial vendors. Second, under specific pathogenfree conditions, the primary source of the microbiome is the mother (6, 8), to a far greater extent than occurs in wild or rewilded mice or in humans. This “maternal effect” is compounded by sibling cohousing and related coprophagy. When the goal is to interrogate mice with different genotypes, mitigation of the maternal effect may include such techniques as fecal microbiota transfer with the same inoculum into germ-free mice, randomized cross-fostering, or the use of mixed genotype litters. Although extensive cohousing does tend to normalize microbiota, as observed by Lipinski et al.(2), work by Robertson et al.(8) suggests that the use of F2 littermates produces a more reliable normalization of the microbiota.