Indicators of Air Pollution-derived Stress in Birds

项目亮点：这是第一个比较颗粒物暴露对原位鸟类皮质酮水平影响的项目。本研究将评估已确定的空气污染物与作为健康指标的禽类肠道微生物群之间的关系。这是一个多学科项目，利用广泛的实地调查和实验室技术来调查鸟类污染应激的不同生物指标。概述：人为活动的增加（图1）导致了空气污染物的大量增加1。其中一些物质，即使处于亚致死水平，也会对人类和非人类的健康产生不利影响。因此，寻找可重复使用的方法来监测环境中的污染物水平是很重要的。鸟类是很好的生物指示物，因为它们无处不在，而且相对容易收集观测数据。我们将测试鸟类通过皮质酮水平和肠道微生物群的变化对空气污染做出反应的假设。这也将使我们能够开发基于性状的生物指标。我们把空气污染导致的压力称为暴露在环境中，降低了个体的达尔文适应度。不是在受控的实验室环境中进行这项研究，我们将在原地监测野生大山雀（Parus major），以研究在原地暴露于空气污染的情况。为了获得当地污染程度的精确指示，我们将收集拉利（即没有离开巢的有环的雏鸟）的羽毛。从拉利收集的血液和粪便样本也将有助于确定空气污染的局部影响。采用单粒子电感耦合等离子体质谱（spICP-MS）研究了大山雀羽毛化学中金属污染物在城乡梯度上的空间变化。我们将使用spICP-MS和共聚焦显微镜将化合物分离成颗粒和溶解形式。为监测空气污染而对鸟类进行的有限研究往往局限于重金属元素，重点是海鸟。据我们所知，目前还没有研究试图将颗粒物暴露与原位皮质酮水平联系起来。这项研究的另一个新颖元素是使用鸟类肠道微生物组作为压力的指标。最终，这项工作可以在广泛分布并在整个城市广泛移动的生物指标中确定空气污染衍生压力的新指标。它还将加强羽毛作为一种组织的使用，这种组织可以进行非侵入性采样，以测量城市的空气污染。这两个结果对鸟类保护和人类健康都具有积极意义。大山雀巢箱将沿着城市到农村的梯度安装，使用百分比绿化覆盖来定义梯度。巢箱将被放置在靠近现有大学和议会空气质量监测站的地方，因为这些监测站将提供氮氧化物和颗粒物的基线读数。我们将把每只小鸡放在巢箱里，并取样一根轮廓羽毛。羽毛将使用spICP-MS分析污染物。共聚焦显微镜和spICP-MS将使我们能够区分微粒和溶解物质。每窝3只带环的雏鸡将采集血样，用放射免疫法评估皮质酮水平。处理后3分钟内采集基线样本，30分钟后采集应力诱导样本10。雀鸟会被抓起来放在袋子里收集粪便样本。我们将从粪便样本中提取DNA，并使用16S核糖体rRNA测序来鉴定和比较细菌组合11。

Project Highlights: This is the first project to compare the effects of particulate matter exposure on corticosterone levels in birds in situ. This study will assess the relationship between identified air pollutants and avian gut microbiome as an indicator of health. This is a multidisciplinary project using a wide variety of fieldwork and laboratory techniques to investigate different bioindicators of pollution-derived stress in birds.Overview: Increased anthropogenic activity (Fig. 1) has led to a large increase in air pollutants1. Some of these substances, even at sub-lethal levels, can have adverse health effects on non-humans, as well as humans2. It is therefore important to find repeatable methods for monitoring pollutant levels in the environment3. Birds are excellent bioindicators as they are ubiquitous and relatively easy to collect observational data from4. We will test the hypothesis that birds respond to air pollution through changes in their corticosterone levels and gut microbiome. This will allow us to develop trait-based bioindicators too. We refer to air pollution-derived stress as exposure that reduces the Darwinian fitness of an individual`5. Rather than conducting this research in a controlled laboratory environment, we will monitor wild great tits (Parus major) in situ to study exposure to air pollution in situ. In order to obtain a precise indication of local pollution levels, feathers will be collected from pulli (i.e. ringed young birds that have not left the nest). Blood and faecal samples collected from pulli will also help to identify the localised impacts of air pollution. Single-particle inductively coupled plasma mass spectrometry (spICP-MS) will be used to investigate the spatial variation of metal contaminants in the feather chemistry of great tit individuals on an urban to rural gradient. We will use spICP-MS and confocal microscopy to separate the compounds into particulate and dissolved forms.The limited research on birds that has been conducted to monitor air pollution has often been confined to heavy metal elements, with a focus on seabirds7. To the best of our knowledge, no study has attempted to link particulate exposure with corticosterone levels in situ. Another novel element of this research is the use of the bird gut microbiome as an indicator of stress. Ultimately, this work could identify new indicators of air pollution-derived stress in bioindicators that are widely distributed and move extensively throughout cities. It will also reinforce the use of feathers as a tissue that can be non-invasively sampled to measure air pollution in cities. Both outcomes have positive implications for avian conservation and human health. Great tit nestboxes will be installed along an urban to rural gradient, using percent green cover to define the gradient. Nestboxes will be placed in close proximity to existing university and council air quality monitoring stations, as these will provide baseline NOx and particulates readings. We will ring each chick in nestboxes and sample one contour feather. Feathers will be analysed for pollutants using spICP-MS. Confocal microscopy and spICP-MS will enable us to distinguish between particulate and dissolved substances9. Blood samples will be taken from 3 ringed chicks per brood for corticosterone assessment by radioimmunoassay. A baseline sample will be taken within 3 minutes of handling, followed by a stress-induced sample obtained after 30 minutes10. Birds will be caught and placed inside bags to collect faecal samples. We will extract DNA from the faecal samples, and use 16S ribosomal rRNA sequencing to identify and compare the bacterial assemblages11.