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<journal-id journal-id-type="publisher-id">Front. Microbiol.</journal-id>
<journal-title>Frontiers in Microbiology</journal-title>
<abbrev-journal-title abbrev-type="pubmed">Front. Microbiol.</abbrev-journal-title>
<issn pub-type="epub">1664-302X</issn>
<publisher>
<publisher-name>Frontiers Media S.A.</publisher-name>
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<article-meta>
<article-id pub-id-type="doi">10.3389/fmicb.2025.1651602</article-id>
<article-categories>
<subj-group subj-group-type="heading">
<subject>Microbiology</subject>
<subj-group>
<subject>Opinion</subject>
</subj-group>
</subj-group>
</article-categories>
<title-group>
<article-title>Earthworm breeding techniques and their roles in microbial regulation and soil carbon sequestration</article-title>
</title-group>
<contrib-group>
<contrib contrib-type="author">
<name><surname>Wu</surname> <given-names>Jiahui</given-names></name>
<xref ref-type="aff" rid="aff1"><sup>1</sup></xref>
<xref ref-type="aff" rid="aff2"><sup>2</sup></xref>
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<contrib contrib-type="author">
<name><surname>Yu</surname> <given-names>Yu</given-names></name>
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<name><surname>Deng</surname> <given-names>Xueqing</given-names></name>
<xref ref-type="aff" rid="aff1"><sup>1</sup></xref>
<xref ref-type="aff" rid="aff2"><sup>2</sup></xref>
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<contrib contrib-type="author" corresp="yes">
<name><surname>Wang</surname> <given-names>Fuwei</given-names></name>
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<contrib contrib-type="author">
<name><surname>Yuan</surname> <given-names>Xianfu</given-names></name>
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<contrib contrib-type="author" corresp="yes">
<name><surname>Wu</surname> <given-names>Hongbao</given-names></name>
<xref ref-type="aff" rid="aff1"><sup>1</sup></xref>
<xref ref-type="aff" rid="aff2"><sup>2</sup></xref>
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<contrib contrib-type="author" corresp="yes">
<name><surname>Wang</surname> <given-names>Jianfei</given-names></name>
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<aff id="aff1"><sup>1</sup><institution>College of Resource and Environment, Anhui Science and Technology University</institution>, <addr-line>Fengyang</addr-line>, <country>China</country></aff>
<aff id="aff2"><sup>2</sup><institution>Anhui Province Agricultural Waste Fertilizer Utilization and Cultivated Land Quality Improvement Engineering Research Center</institution>, <addr-line>Fengyang</addr-line>, <country>China</country></aff>
<author-notes>
<fn fn-type="edited-by"><p>Edited by: Bin Zhou, Chinese Academy of Tropical Agricultural Sciences, China</p></fn>
<fn fn-type="edited-by"><p>Reviewed by: Lin Zhang, Henan University of Science and Technology, China</p>
<p>Dongming Wu, Chinese Academy of Tropical Agricultural Sciences, China</p></fn>
<corresp id="c001">&#x0002A;Correspondence: Fuwei Wang <email>fuwei_wang&#x00040;hotmail.com</email></corresp>
<corresp id="c002">Hongbao Wu <email>wuhb1120&#x00040;163.com</email></corresp>
<corresp id="c003">Jianfei Wang <email>wangjf&#x00040;ahstu.edu.cn</email></corresp>
</author-notes>
<pub-date pub-type="epub">
<day>03</day>
<month>09</month>
<year>2025</year>
</pub-date>
<pub-date pub-type="collection">
<year>2025</year>
</pub-date>
<volume>16</volume>
<elocation-id>1651602</elocation-id>
<history>
<date date-type="received">
<day>22</day>
<month>06</month>
<year>2025</year>
</date>
<date date-type="accepted">
<day>19</day>
<month>08</month>
<year>2025</year>
</date>
</history>
<permissions>
<copyright-statement>Copyright &#x000A9; 2025 Wu, Yu, Deng, Wang, Yuan, Wu and Wang.</copyright-statement>
<copyright-year>2025</copyright-year>
<copyright-holder>Wu, Yu, Deng, Wang, Yuan, Wu and Wang</copyright-holder>
<license xlink:href="http://creativecommons.org/licenses/by/4.0/"><p>This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.</p></license>
</permissions>
<kwd-group>
<kwd>earthworm</kwd>
<kwd>vermicompost</kwd>
<kwd>breeding techniques</kwd>
<kwd>microbial regulation</kwd>
<kwd>carbon sequestration</kwd>
</kwd-group>
<counts>
<fig-count count="2"/>
<table-count count="0"/>
<equation-count count="0"/>
<ref-count count="56"/>
<page-count count="5"/>
<word-count count="4117"/>
</counts>
<custom-meta-wrap>
<custom-meta>
<meta-name>section-at-acceptance</meta-name>
<meta-value>Terrestrial Microbiology</meta-value>
</custom-meta>
</custom-meta-wrap>
</article-meta>
</front>
<body>
<sec id="s1">
<title>1 Introduction</title>
<p>Earthworms, as a common soil animal, play an important role in the material cycle and nutrient transformation process of many terrestrial ecosystems (<xref ref-type="bibr" rid="B31">Ngo et al., 2012</xref>; <xref ref-type="bibr" rid="B17">Hoeffner et al., 2018</xref>). Earthworms can promote the mineralization of nitrogen and phosphorus, and thereby enhance the availability of soil nutrients (<xref ref-type="bibr" rid="B28">Medina-Sauza et al., 2019</xref>; <xref ref-type="bibr" rid="B7">Bhadauria and Saxena, 2010</xref>). Earthworms enhance soil porosity through their burrowing activities, facilitating soil aeration and drainage while improving soil structure, promoting plant root growth, and enhancing root access to water and nutrients (<xref ref-type="bibr" rid="B13">Ganault et al., 2024</xref>). Moreover, enhancing earthworm activity may increase microbial diversity and activity in the soil, accelerating litter decomposition and contributing to the carbon (C) sequestration in soil (<xref ref-type="bibr" rid="B24">Liu et al., 2019</xref>).</p>
<p>Vermicompost is the excrement of earthworms, which provides a more stable habitat for microorganisms by regulating pH and cation exchange capacity (<xref ref-type="bibr" rid="B21">Lim et al., 2015</xref>). Vermicomposting takes advantage of earthworms during composting to generate an organic material that may be physically, nutritionally, and biochemically improved compared to compost (<xref ref-type="bibr" rid="B2">Akhila and Entoori, 2022</xref>). This process converts nutrients in organic matter (such as nitrogen and phosphorus) into forms that are easily absorbed by plants, thereby increasing soil fertility (<xref ref-type="bibr" rid="B21">Lim et al., 2015</xref>; <xref ref-type="bibr" rid="B17">Hoeffner et al., 2018</xref>; <xref ref-type="bibr" rid="B20">Li et al., 2024</xref>).</p>
<p>The role of earthworms in shaping soil microbial communities and regulating C cycling has emerged as a central focus in soil ecological research (<xref ref-type="bibr" rid="B1">Ahmed and Al-Mutairi, 2022</xref>; <xref ref-type="bibr" rid="B42">Thomas et al., 2020</xref>). However, in recent years, the population of earthworms in the soil has gradually decreased due to intensive agricultural activities. So, how to adopt scientific and reasonable earthworm breeding techniques has become a key issue (<xref ref-type="bibr" rid="B32">Pelosi et al., 2013</xref>). Thus, this article aims to provide a theoretical basis and technical guidance for earthworm breeding techniques, and deeply analyze the roles of earthworms in microbial regulation and soil C sequestration.</p></sec>
<sec id="s2">
<title>2 Earthworm breeding techniques</title>
<p>The selection of a suitable earthworm species is important for breeding (<xref ref-type="bibr" rid="B9">Butt, 2008</xref>). Currently, there are over 6,000 species of earthworms in the world (<xref ref-type="bibr" rid="B38">Singh et al., 2020</xref>). Ecologically, earthworms can be classified into three categories: epigeic, endogeic, and anecic species, based on the living behavior (<xref ref-type="bibr" rid="B47">Wasunan et al., 2023</xref>). Briefly, epigeic earthworms survive on the surface of soil and consume surface organic matter, endogeic earthworms burrow horizontal galleries to feed on soil organic matter, and anecic earthworms burrow vertical galleries to feed on a mixture of surface and deep soil organic matter (<xref ref-type="bibr" rid="B17">Hoeffner et al., 2018</xref>). Among them, epigeic earthworms are the most suitable for converting organic matter, while anecic earthworms are more suitable for use as high-protein feed for livestock and poultry (<xref ref-type="bibr" rid="B35">Qiu, 1999</xref>; <xref ref-type="bibr" rid="B36">Rong et al., 2020</xref>). The <italic>Eisenia Fetida</italic> belongs to the epigeic earthworm, which has a strong ability to decompose organic wastes such as livestock manure and urban sludge, and is suitable for breeding with manure (<xref ref-type="bibr" rid="B51">Yadav and Garg, 2011</xref>). In addition, the <italic>Ohira</italic> II earthworms can withstand low temperatures and can still move at &#x02212;10 &#x000B0;C, making them suitable for high-density breeding in cold regions like Northeast China (<xref ref-type="bibr" rid="B23">Liu et al., 2021</xref>). The <italic>Pheretima Guillelmi</italic> has a relatively large body size, prefers organic wastes and fertile soil, and is suitable for breeding in vegetable fields and areas with sufficient feed (<xref ref-type="bibr" rid="B22">Lin et al., 2024</xref>).</p>
<p>Apart from species selection, earthworm breeding techniques involve inoculation density, breeding substrate, breeding environment, etc. (<xref ref-type="fig" rid="F1">Figure 1</xref>). Different densities of earthworms are closely related to their growth and development. The appropriate density of earthworms is 1.5 kg/m<sup>2</sup> (<xref ref-type="bibr" rid="B46">Wang et al., 2023</xref>; <xref ref-type="bibr" rid="B49">Xiao et al., 2023</xref>). Appropriate temperature, humidity, and sufficient food can reduce the occurrence of earthworm escape (<xref ref-type="bibr" rid="B34">Presley et al., 1996</xref>; <xref ref-type="bibr" rid="B10">Chen and Zhong, 2022</xref>). To prevent natural enemies, 5% lime can be scattered around the farm to form a defense line (<xref ref-type="bibr" rid="B52">Yi, 2018</xref>).</p>
<fig position="float" id="F1">
<label>Figure 1</label>
<caption><p>Earthworm breeding techniques.</p></caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fmicb-16-1651602-g0001.tif">
<alt-text>Diagram detailing earthworm breeding methods, divided into four sections: site selection, substrate selection, feeding management, and temperature and humidity. Site selection includes proximity to livestock, a wide area, cool, damp conditions, and avoiding main roads. Substrate selection involves using cow, pig, or sheep dung with a pH of 6.5 to 7.5 and a C/N ratio of 25:1. Feeding management suggests releasing 1.5 kilograms of earthworms per square meter, regular watering, escape prevention, enemy prevention, and feeding 15 to 20 centimeters of material weekly. Temperature and humidity are ideal at 60%-70% and 15&#x000B0;C to 25&#x000B0;C.</alt-text>
</graphic>
</fig>
<p>Livestock and poultry manure are commonly used as the substrate for earthworm breeding (<xref ref-type="bibr" rid="B4">Bakayoko et al., 2009</xref>). The breeding site should be close to livestock and poultry farms or in an open area that is convenient for storing raw materials (<xref ref-type="bibr" rid="B29">Meng, 2018</xref>; <xref ref-type="bibr" rid="B48">Xiao et al., 2025</xref>), which facilitates breeding substrate acquisition. Cow manure is more beneficial for the growth of earthworms, the production of vermicompost, and the optimal maturation of the substrate compared to pig manure and sheep manure (<xref ref-type="bibr" rid="B44">Vodounnou et al., 2016</xref>; <xref ref-type="bibr" rid="B45">Wang et al., 2022</xref>). The C/N ratio and pH value in the substrate are important factors affecting the growth and reproduction of earthworms. The optimal C/N ratio is 25:1, and the pH value typically ranges from 6.5 to 7.5 (<xref ref-type="bibr" rid="B39">Sonia et al., 2016</xref>; <xref ref-type="bibr" rid="B6">Behera, 2018</xref>). During the breeding process, feed is usually added once a week, with a cover depth of 15&#x02013;20 cm (<xref ref-type="bibr" rid="B25">Liu, 2023</xref>).</p>
<p>Temperature and humidity are also key factors for the growth and reproduction of earthworms. Earthworms prefer to grow in cool and humid environments (<xref ref-type="bibr" rid="B15">Grant, 1955</xref>; <xref ref-type="bibr" rid="B27">Ma et al., 2019</xref>). To provide the most suitable environment for the growth and reproduction of earthworms, the moisture content of the substrate should be monitored daily (<xref ref-type="bibr" rid="B33">Perreault and Whalen, 2006</xref>; <xref ref-type="bibr" rid="B16">Han et al., 2024</xref>). The temperature range most suitable for earthworm growth is typically 15&#x02013;25 &#x000B0;C, and the humidity is approximately 60&#x02013;70% (<xref ref-type="bibr" rid="B5">Baker and Whitby, 2003</xref>; <xref ref-type="bibr" rid="B19">Li et al., 2020</xref>; <xref ref-type="bibr" rid="B53">Yin et al., 2022</xref>).</p></sec>
<sec id="s3">
<title>3 The roles of earthworms in microbial regulation and soil C sequestration</title>
<p>Earthworms constitute the dominant biomass of invertebrates in soil and are known as the &#x0201C;engineers of the soil ecosystem&#x0201D; (<xref ref-type="bibr" rid="B8">Blouin et al., 2013</xref>). Earthworm activities create dynamic microenvironments that significantly alter the composition and function of soil microorganisms, ultimately affecting C sequestration processes (<xref ref-type="bibr" rid="B11">Dempsey et al., 2013</xref>; <xref ref-type="bibr" rid="B28">Medina-Sauza et al., 2019</xref>). A previous study showed that inoculating different ecological types of earthworms increased the diversity of soil fungal communities, thereby enhancing soil quality (<xref ref-type="bibr" rid="B55">Zhang et al., 2025</xref>). Earthworms have a promoting effect on enzyme activity and increase the diversity and uniformity of bacterial communities, thereby effectively improving soil quality (<xref ref-type="bibr" rid="B50">Xu et al., 2021</xref>). The vermicompost is rich in abundant microorganisms, which can be applied to the soil as a biological fertilizer, thereby increasing soil enzyme activity and improving soil fertility (<xref ref-type="bibr" rid="B14">Gao et al., 2015</xref>; <xref ref-type="bibr" rid="B56">Zhao et al., 2023</xref>). Many studies have shown that the addition of vermicompost can increase the richness of soil microorganisms (<xref ref-type="bibr" rid="B21">Lim et al., 2015</xref>; <xref ref-type="bibr" rid="B38">Singh et al., 2020</xref>; <xref ref-type="bibr" rid="B40">Tan et al., 2021</xref>).</p>
<p>Earthworms play a pivotal role in regulating soil microbial communities through multiple pathways. First, earthworms increase soil porosity by creating channels through their digging, allowing oxygen to penetrate more easily and improving the water infiltration and retention capacity (<xref ref-type="bibr" rid="B28">Medina-Sauza et al., 2019</xref>). The balance between ventilation and water retention provides a more suitable environment for microorganisms, thereby regulating the composition and function of microbial communities (<xref ref-type="bibr" rid="B21">Lim et al., 2015</xref>). Second, earthworms promote the decomposition of organic matter and nutrient cycling, enhance soil fertility, and enable plants to obtain more nutrients during the process of burrowing and feeding (<xref ref-type="bibr" rid="B41">Thejesh, 2020</xref>; <xref ref-type="bibr" rid="B1">Ahmed and Al-Mutairi, 2022</xref>). These alterations in soil fertility and plant growth may regulate the microbial community. Third, through the digestive processes of earthworms, the excreted vermicompost can elevate the content of nitrogen, phosphorus, and potassium and increase the soil water retention capacity, which may affect microbial communities (<xref ref-type="bibr" rid="B21">Lim et al., 2015</xref>; <xref ref-type="bibr" rid="B43">Turab et al., 2023</xref>).</p>
<p>Earthworms play a fundamental role in soil C sequestration through their unique biological activities that transform organic matter into more stable forms (<xref ref-type="bibr" rid="B12">Don et al., 2008</xref>; <xref ref-type="bibr" rid="B54">Zhang et al., 2013</xref>; <xref ref-type="bibr" rid="B30">Meng, 2022</xref>). Their contribution to C storage operates through multiple pathways, primarily involving the physical breakdown of plant residues, microbial stimulation, and the formation of organo-mineral complexes (<xref ref-type="bibr" rid="B54">Zhang et al., 2013</xref>; <xref ref-type="bibr" rid="B3">Angst et al., 2017</xref>; <xref ref-type="bibr" rid="B42">Thomas et al., 2020</xref>). As earthworms consume and digest organic materials, they accelerate decomposition while simultaneously creating microenvironments that favor C stabilization (<xref ref-type="bibr" rid="B26">Lubbers et al., 2017</xref>; <xref ref-type="bibr" rid="B3">Angst et al., 2017</xref>). The gut passage of organic matter introduces enzymes and gut-associated microbes that chemically modify C compounds, often leading to the formation of humic substances with greater resistance to microbial degradation (<xref ref-type="bibr" rid="B3">Angst et al., 2017</xref>; <xref ref-type="bibr" rid="B42">Thomas et al., 2020</xref>). The mucus secreted by the earthworm intestines combines with soil particles and organic matter to form more stable aggregates (<xref ref-type="bibr" rid="B18">Lavelle et al., 1997</xref>; <xref ref-type="bibr" rid="B3">Angst et al., 2017</xref>). Studies have shown that the adhesive properties of polysaccharides and fungal hyphae largely determine the formation of large aggregates, where the fungal hyphae can prevent certain compounds in the soil aggregates from being utilized by microorganisms, thereby making the C more stable (<xref ref-type="bibr" rid="B37">Samuel et al., 2008</xref>; <xref ref-type="bibr" rid="B42">Thomas et al., 2020</xref>).</p>
<p>Based on the roles of earthworms in microbial regulation and soil C sequestration, we propose a conceptual framework of earthworms on straw decomposition and soil organic C accumulation in farmland soils (<xref ref-type="fig" rid="F2">Figure 2</xref>). Earthworms stimulate microbial activity by secreting mucus, break down straw to increase the contact area, and mix organic matter with the soil, thereby accelerating straw decomposition and promoting the combination of microbial residue C with soil minerals, ultimately significantly increasing soil C accumulation (<xref ref-type="fig" rid="F2">Figure 2</xref>).</p>
<fig position="float" id="F2">
<label>Figure 2</label>
<caption><p>Conceptual framework of earthworms on straw decomposition and soil organic C accumulation.</p></caption>
<graphic mimetype="image" mime-subtype="tiff" xlink:href="fmicb-16-1651602-g0002.tif">
<alt-text>Diagram illustrating how earthworms promote straw decomposition and soil organic carbon accumulation. On the left, an image of an earthworm. The process involves microbial catabolism releasing CO2, microbial anabolism, and formation of microbial residue carbon. Arrows depict pathways leading to microbial residue carbon formation and stabilization. These contribute to carbon accumulation in soil as aggregate-occluded and mineral-adsorbed carbon. Prominent labels emphasize enhancing straw decomposition, and promoting microbial residue carbon formation and stabilization.</alt-text>
</graphic>
</fig>
</sec>
<sec id="s4">
<title>4 Conclusions and prospects</title>
<p>Earthworms and their products play an important role in the improvement of soil fertility in terms of the physical and chemical properties of the soil. We summarized the optimal earthworm breeding techniques involve species selection, inoculation density, breeding substrate, breeding environment, etc. These scientific earthworm breeding techniques provide the foundation for the production of earthworms and vermicompost efficiently in the future. In addition, we synthesized the roles of earthworms in microbial regulation and soil C sequestration to provide a theoretical basis for the application of earthworms in agriculture. Although earthworms can promote the activity of soil microbes and organic matter decomposition, however, whether this decomposed C can persist stably in the soil for a long time remains largely uncertain. Therefore, future research can focus on the potential and mechanisms of earthworms on soil C stability in the long term.</p></sec>
</body>
<back>
<sec sec-type="author-contributions" id="s5">
<title>Author contributions</title>
<p>JWu: Writing &#x02013; original draft, Writing &#x02013; review &#x00026; editing. YY: Writing &#x02013; review &#x00026; editing. XD: Writing &#x02013; review &#x00026; editing. FW: Conceptualization, Funding acquisition, Supervision, Writing &#x02013; review &#x00026; editing. XY: Writing &#x02013; review &#x00026; editing. HW: Writing &#x02013; review &#x00026; editing. JWa: Conceptualization, Funding acquisition, Supervision, Writing &#x02013; review &#x00026; editing.</p>
</sec>
<sec sec-type="funding-information" id="s6">
<title>Funding</title>
<p>The author(s) declare that financial support was received for the research and/or publication of this article. This study was supported by the National Key R &#x00026; D Program of China (2022YFD2301402), Anhui Science and Technology University Key Construction High Altitude Discipline Project (XK-XJGY001), the Anhui Agricultural University High-Level Talent Project (ZHYJ202303), Science Key Research Program of Anhui Province university (2023AH051854), the Open Project Funding Plan of the Key Laboratory of Organic Fertilizer Creation, Ministry of Agriculture and Rural Affairs, and Investigation and Sampling of Surface Samples for the Third National Soil Census in Nanling County (WH11CG2023FW0866).</p>
</sec>
<sec sec-type="COI-statement" id="conf1">
<title>Conflict of interest</title>
<p>The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.</p>
</sec>
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<title>Generative AI statement</title>
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