Food Animal Health Research Program, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691.
Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.
Avian Dis. 2020 Sep 1;64(3):277-285. doi: 10.1637/aviandiseases-D-19-00200.
Although poultry microbiome discoveries are increasing due to the potential impact on poultry performance, studies examining the poultry respiratory microbiome are challenging because of the low microbial biomass and uniqueness of the avian respiratory tract, making it difficult to sample enough material for microbial analysis. Invasive sampling techniques requiring euthanasia are currently used to increase microbial mass for the analysis, thus making it impossible to sample individual birds longitudinally. In this study, we compared invasive (nasal wash, upper tracheal wash, lower tracheal wash, and lower respiratory lavage) and noninvasive (tracheal and choanal swabs) respiratory sampling techniques in two independent experiments by using 4-wk-old chickens. We first established the experimental baseline of respiratory microbiota by using invasive techniques to enable reasonable comparisons between sampling methods and between experiments. Although noninvasive sampling (live-bird swabs) resulted in lower 16S ribosomal RNA gene copy numbers compared with invasive sampling, live swabs were able to detect the dominant microbes captured by invasive techniques. Nevertheless, swabs from euthanatized birds were more reflective of the microbiota captured through invasive methods than live swab. Furthermore, from two separate experiments, we also demonstrated that respiratory microbiota sampling is highly reproducible, especially in the trachea and lower respiratory tract. Our study provides new insights and perspectives on decision making when sampling and studying poultry respiratory microbiota.
尽管禽类微生物组的发现因可能对禽类生产性能产生影响而在不断增加，但由于禽类呼吸道微生物的生物量低且具有独特性，研究禽类呼吸道微生物组极具挑战性，这使得采集足够的微生物分析材料变得困难。目前，为了增加微生物质量以进行分析，常采用侵入性采样技术（需要安乐死），从而无法对个体禽类进行纵向采样。在这项研究中，我们通过两个独立的实验，使用 4 周龄的鸡比较了侵入性（鼻腔冲洗、上气管冲洗、下气管冲洗和下呼吸道灌洗）和非侵入性（气管和鼻后腔拭子）呼吸采样技术。我们首先使用侵入性技术建立了呼吸微生物组的实验基线，以便在采样方法之间和实验之间进行合理的比较。虽然非侵入性采样（活体禽鸟拭子）的 16S 核糖体 RNA 基因拷贝数低于侵入性采样，但活体拭子仍能够检测到侵入性技术捕获的主要微生物。然而，安乐死禽鸟的拭子比活体拭子更能反映通过侵入性方法捕获的微生物群。此外，通过两个独立的实验，我们还表明，呼吸微生物组采样具有高度的可重复性，尤其是在气管和下呼吸道。我们的研究为禽类呼吸微生物组的采样和研究提供了新的见解和思路。
