QUANTIFYING THE BIOMOLECULAR FATE OF DIETARY C AND N IN EARTHWORM TISSUES AND CASTS

蚯蚓在处理土壤有机质（OM）方面的重要性得到了亚里士多德和查尔斯达尔文等杰出学者的认可，亚里士多德将蚯蚓描述为“地球的肠子”，查尔斯达尔文将蚯蚓的概念引入了“自然的犁”。如今，蚯蚓通过挖掘土壤来改变有机化合物分布的能力使它们被描述为“土壤工程师”。它们在土壤中的摄食活动被广泛认为是有益的，并通过向矿物质土壤层添加植物营养素和改善保水性、通气性、稳定性和可加工性来提高土壤质量。这些改善促进了根系生长，增加了土壤动物的生物多样性。蚯蚓利用土壤中的有机物质，如死亡的植物材料，作为食物来源。蚯蚓把有机物拉到洞穴里，或者干脆把它吃掉，然后把它存款在土壤表面或下面的蠕虫粪便中。在进食过程中，它们将食物分解成小块。其中一些被消化，并有助于身体的质量，其余的被排泄在蠕虫铸件和粘液分泌物。令人惊讶的是，目前还不知道蚯蚓吃的有机物质的哪些成分，有多少被吸收到它们的身体质量中，以及它们的饮食中有多少和哪些部分被排泄回土壤。为了找出蚯蚓吃，吸收和排泄的有机物的部分，我们将在土壤中生态系统中用13 C-和15 N-标记的黑麦草喂养一个普通的草地，凋落物喂养的蚯蚓物种。13 C和15 N是碳和氮的天然存在的非放射性稳定同位素，其在大气中的丰度比最丰富的同位素12 C和14 N低得多。13 C：12 C和15 N：14 N之间的比率可以使用同位素比率质谱法计算。用于实验的黑麦草将在密封的生长室中生长：含有高比例13 C的CO2气体将用于光合作用，含有高比例15 N的硝酸盐（NO3-）将以溶液形式供应到根部，产生13 C和15 N标记的植物材料。当蚯蚓吃草时，它们吸收到体内的营养物质，然后以呼吸气体、粘液和排泄物的形式排出，也会被标记为13 C和15 N。我们会从蚯蚓的角质层、肌肉组织、粘液和粪便中，以及从剩余的黑麦草和土壤中提取不同的物质，以计算蚯蚓利用了多少标记物质。我们还将从中型生态系统中取样二氧化碳，看看有多少食物被用于能源生产。通过测量有多少13 C和15 N标签被纳入所有这些样品中，我们将能够计算出有多少蛋白质，碳水化合物和脂肪从草已被处理的生物。重要的是，我们还将能够精确地确定蚯蚓是否以及如何从土壤中的有机物质中释放（引发）或储存（封存）更多的碳和氮。总的来说，这项研究将为理解蚯蚓如何影响有机物的分解，它们可以处理多少有机物，以及它们的活动是否增加或减少土壤中储存的碳和氮的量增加了一个新的维度。

The importance of earthworms in the processing of soil organic matter (OM) was recognised by such illustrious scholars as Aristotle, who described them as 'the intestines of the Earth', and Charles Darwin who introduced the concept of earthworms as 'Nature's plough'. Nowadays, the capacity of earthworms to modify the distribution of organic compounds through the excavation of soils has led them to be described as 'soil engineers'. Their feeding activity in soils is widely recognised as beneficial, and results in increased soil quality through the addition of plant nutrients to mineral soil horizons and improved water retention, aeration, stability and workability. These improvements enhance root growth and increase biodiversity of soil fauna. Earthworms use soil organic matter, such as dead plant material, as a food source. Earthworms pull organic matter down into burrows, or simply eat it and subsequently deposit it in worm casts (their faeces) at or below the soil surface. During ingestion they transform their diet into small fragments. Some of these are digested and contribute to the earthworm's body mass, the rest being excreted in worm casts and mucus excretions. Surprisingly, it is unknown which components of organic matter earthworms eat, how much is absorbed into their body mass, and how much and which parts of their diet are excreted back into the soil. To find out which parts of organic matter earthworms eat, absorb and excrete, we are going to feed a common grassland, litter-feeding species of earthworm with 13C- and 15N-labelled ryegrass in soil mesocosms. 13C and 15N are naturally occurring non-radioactive stable isotopes of carbon and nitrogen that exist in much lower abundances in the atmosphere than the most abundant isotopes, 12C and 14N. The ratio between 13C:12C and 15N:14N can be calculated using isotope ratio mass spectrometry. The ryegrass for the experiment will be grown in sealed growth chambers: CO2 gas containing a high proportion of 13C will be supplied for photosynthesis, and nitrate (NO3-) containing a high proportion of 15N will be supplied in solution to the roots, producing 13C- and 15N-labelled plant material. When the earthworms eat the grass, the nutrients they absorb into their bodies, and then excrete as respired gas, mucus and casts, will also be labelled with 13C and 15N. We will extract different substances from the cuticle, muscle tissue, mucus and casts of the earthworms, and also from the remaining ryegrass and soil, to work out how much of the labelled substances have been utilised by the earthworms. We will also sample CO2 from the mesocosms to see how much of the diet has been used for energy production. By measuring how much of the 13C and 15N label has been incorporated into all of these samples, we will be able to calculate how much protein, carbohydrate and fat from the grass has been processed by the earthworm. Importantly, we will also be able to determine precisely if and how earthworms cause greater release (priming) or storage (sequestration) of carbon and nitrogen from organic matter in soils. Overall, this research will add a new dimension to the understanding of how earthworms affect the breakdown of organic matter, how much organic matter they can process, and whether their activity increases or decreases the amount of carbon and nitrogen stored in the soil.