Abstract:This study aimed to investigate the changes of microbial community structure and their antibiotics resistance genes (ARGs) abundances in the intestinal tract of Eisenia fetida, via detecting the sludge passing through different functional areas in earthworms' gut. Prior to the DNA extraction, all fresh samples were pretreated with propidium monoazide (PMA) to characterize active functional microorganisms. The results showed that the abundances of bacterial 16S rDNA and eukaryotic 18S rDNA significantly increased (P<0.05) by 28.2 and 42.2 times in stomach, and they significantly decreased (P<0.05) in the gizzard and hindgut, after 5 days of earthworm digestion. In addition, the sludge digested by earthworms changed dominant proteobacteria to tenericutes in gizzard, and modified dominant bacteroidetes to firmicutes in stomach. However, the digestion process of earthworms had little effect on bacterial community structure in hindgut. For the ARGs, the abundances of ermF, tetX and sul2 significantly increased (P<0.05) by 1.9×102, 8.4×105 and 25.9 times in stomach of earthworm, respectively. While the total abundances of ARGs in gizzard and hindgut significantly decreased (P<0.05) by 11.0 and 45.2 times, respectively. Moreover, network analysis revealed that the feeding behavior of earthworms for sludge could indicate effects on the structural diversity of host bacteria of ARGs in their intestine.
彭兰生, 关孟欣, 黄魁, 夏慧, 桑春雷. 蚯蚓摄食污泥对其肠道功能区微生物种群及耐药基因的影响[J]. 中国环境科学, 2022, 42(1): 465-473.
PENG Lan-sheng, GUAN Meng-xin, HUANG Kui, XIA Hui, SANG Chun-lei. Effects of excess sludge fed by earthworms on microbial community and antibiotic resistance genes in their intestinal functional area. CHINA ENVIRONMENTAL SCIENCECE, 2022, 42(1): 465-473.
Wei L, Zhu F, Li Q, et al. Development, current state and future trends of sludge management in China: Based on exploratory data and CO2-equivaient emissions analysis [J]. Environment International, 2020, 144: 106093.
[2]
薛重华, 孔祥娟, 王胜, 等. 我国城镇污泥处理处置产业化现状、发展及激励政策需求[J]. 净水技术, 2018, 37(12): 41-47. Xue C, Kong X, Wang S, et al. Industrialization status, development analysis and incentive policy demands of municipal sludge treatment and disposal industry in china [J]. Water Purification Technology, 2018, 37(12): 41-47.
[3]
Marcoux M, Matias M, Olivier F, et al. Review and prospect of emerging contaminants in waste – Key issues and challenges linked to their presence in waste treatment schemes: General aspects and focus on nanoparticles [J]. Waste Manage, 2013, 33(11): 2147-2156.
[4]
Calero-Cáceres W, Melgarejo A, Colomer-Lluch M, et al. Sludge as a potential important source of antibiotic resistance genes in both the bacterial and bacteriophage fractions [J]. Environmental Science & Technology, 2014, 48(13): 7602-7611.
[5]
Liu Z, Klümper U, Liu Y, et al. Metagenomic and metatranscriptomic analyses reveal activity and hosts of antibiotic resistance genes in activated sludge [J]. Environment International, 2019, 129: 208-220.
[6]
Cui G, Lü F, Zhang H, et al. Critical insight into the fate of antibiotic resistance genes during biological treatment of typical biowastes [J]. Bioresource Technology, 2020, 317: 123974.
[7]
Su J, Wei B, Ouyang W, et al. Antibiotic resistome and its association with bacterial Communities during sewage sludge composting [J]. Environmental Science & Technology, 2015, 49(12): 7356-7363.
[8]
Tong J, Lu X, Zhang J, et al. Factors influencing the fate of antibiotic resistance genes during thermochemical pretreatment and anaerobic digestion of pharmaceutical waste sludge [J]. Environmental Pollution, 2018, 43: 1403-1413.
[9]
Xia H, Wu Y, Chen X, et al. Effects of antibiotic residuals in dewatered sludge on the behavior of ammonia oxidizers during vermicomposting maturation process [J]. Chemosphere, 2019, 218: 810-817.
[10]
Aira M, Olcina J, Pérez-Losada M, et al. Characterization of the bacterial communities of casts from Eiseniaandrei fed with different substrates [J]. Applied Soil Ecology, 2016, 98: 103-111.
[11]
Hu J, Zhao H, Wang Y, et al. The bacterial community structures in response to the gut passage of earthworm (Eisenia fetida) feeding on cow dung and domestic sludge: Illumina high-throughput sequencing-based data analysis [J]. Ecotoxicology and Environmental Safety, 2020, 190: 110-149.
[12]
Huang K, Xia H, Wu Y, et al. Effects of earthworms on the fate of tetracycline and fluoroquinolone resistance genes of sewage sludge during vermicomposting [J]. Bioresource Technology, 2018, 259: 32-39.
[13]
Cui G, Li F, Li S, et al. Changes of quinolone resistance genes and their relations with microbial profiles during vermicomposting of municipal excess sludge [J]. Science of the Total Environment, 2018, 644: 494-502.
[14]
黄魁, 夏慧, 陈景阳, 等. 蚯蚓对城市污泥蚯蚓堆肥过程中微生物特征变化的影响[J]. 环境科学学报, 2018, 38(8): 3146-3152. Huang k, Xia H, Chen J, et al. Effects of earthworms on changs of microbial feature during vermicomposting of municipal sludge [J]. Acta Scientiae Circumstantiae, 2018, 38(8): 3416-3152.
[15]
HJ 535-2009 水质氨氮的测定纳式试剂分光光度法[S]. HJ 535-2009 Water quality-Determination of ammonia nitrogen-Nessler's reagent spectrophotometry[S].
[16]
Gómez-Brandón M, Aira M, Lores M, et al. Epigeic earthworms exert a bottleneck effect on microbial communities through gut associated processes [J]. The Public Library of Science One, 2011, 6(9): e24786.
[17]
Gómez-Brandón M, Lores M, Domínguez J, et al. Species-specifice effects of epigeic earthworms on microbial community structure during first stages of decomposition of organic matter [J]. The Public Library of Science One, 2012, 7(2): e31895.
[18]
Gómez-Brandón M, Lores M, Domínguez J. Changes in chemical and microbiological properties ofrabbit manure in a continuous-feeding vermicomposting system [J]. Bioresource Technology, 2013, 128(1): 310-316.
[19]
晁会珍, 孙明明, 朱国繁, 等. 蚯蚓肠道细菌生态功能及毒理学研究进展[J]. 生态毒理学报, 2020, 15(5): 35-48. Chao H, Sun M, Zhu G, et al. Ecological functioning of the earthworm intestinal bacteria and their role in toxicology research [J]. Asian Journal of Ecotoxicology, 2020, 15(5): 35-48.
[20]
Zheng R, Liu R, Shan Y, et al. Characterization of the first cultured free-living representative of Candidatus Izemoplasma uncovers its unique biology [J]. The ISME Journal, 2021, 15: 2676-2691.
[21]
Aira M, Monroy F, Domínguez J. Changes in microbial biomass and microbial ac-tivity of pig slurry after the transit through the gut of the earthworm Eudrilus eugeniae [J]. Biology and Fertility of Soils, 2006, 42(4): 371–376.
[22]
Wang N, Wang W, Jiang Y, et al. Variations in bacterial taxonomic profiles and potential functions in response to the gut transit of earthworms (Eisenia fetida) feeding on cow manure [J]. Science of the Total Environment, 2021, 787: 147392.
[23]
陈景阳, 夏慧, 黄魁, 等. 四环素对污泥蚯蚓粪中微生物种群和抗性基因的影响[J]. 环境科学, 2019, 40(7): 3263-3269. Chen J, Xia H, Huang K, et al. Effects of tetracycline on microbial communities and antibiotic resistance genes of vermicompost from dewatered sludge [J]. Environmental Science, 2019, 40(7): 3263-3269.
[24]
De Gannes V, Eudoxie G, Hickey W. Prokaryotic successions and diversity in composts as revealed by 454-pyrosequencing [J]. Bioresource Technology, 2013, 133: 573-580.
[25]
Gasparich G E. The Family Entomoplasmataceae [M]. The Prokaryotes, 2014: 505-514.
[26]
Lund M B, Holmstrup M, Lomstein B A. et al. Beneficial effect of Verminephrobacter nephridial symbionts on the fitness of the earthworm aporrectodea tuberculat [J]. Applied and Environmental Microbiology, 2010, 76(14): 4738-4743.
[27]
Liu D, Lian B, Wu C, et al. A comparative study of gut microbiota profiles of earthworms fed in three different substrates [J]. Symbiosis, 2018, 74: 21-29.
[28]
王洪涛,丁晶,邵元虎,等.4种蚯蚓肠道微生物对砷毒性响应差异研究 [J]. 生态学报, 2022,DOI:10.5846/stxb202010132612. Wang H, Ding J, Shao Y, et al. Comparative study on the responses of gut microbiota of four species of earthworms to arsenic toxicity [J]. Acta Ecologica Sinica. 2022,DOI:10.5846/stxb202010132612.
[29]
Yin J, Zhang X, Wu B, et al. Metagenomic insights into tetracycline effects on microbial community and antibiotic resistance of mouse gut [J]. Ecotoxicology, 2015, 24(10): 2125-2132.
[30]
Zhu D, An X, Chen Q, et al. Antibiotics disturb the microbiome and increase the incidence of resistance genes in the gut of acommon soil collembolan [J]. Environmental Science & Technology, 2018, 52(5): 3081-3090.
[31]
Zhou G, Qiu X, Wu X, et al. Horizontal gene transfer is a key determinant of antibiotic resistance genes profiles during chicken manure composting with the addition of biochar and zeolite [J]. Journal of Hazardous Materials, 2021, 408: 124883.
[32]
李建辉, 张莹莹, 黄魁, 等. 宏基因组学解析蚯蚓粪中微生物种群及耐药基因的组成[J]. 中国环境科学, 2020, 40(12): 5375-5382. Li J, Zhang Y, Huang K, et al. Composition of microbial community and antibiotic resistance genes in vermicomposts revealed by metagenomic analysis [J]. China Environmental Science, 2020, 40(12): 5375-5382.
[33]
Boucher Y, Labbate M, Koenig J, et al. Integrons: mobilizable platforms that promote genetic diversity in bacteria [J]. Trends in Microbiology, 2007, 15(7): 301-309.
[34]
Ramsden S, Ghosh S, Bohl L, et al. Phenotypic and genotypic analysis of bacteria isolated from three municipal wastewater treatment plants on tetracycline-amended and ciprofloxacin-amended growth media [J]. Journal of Applied Microbiology, 2010, 109(5): 1609-1618.
[35]
Byzov B, Khomyakov N, Kharin S, et al. Fate of soil bacteria and fungi in the gut of earthworms [J]. European Journal of Soil Biology, 2007, 43(1): 149-156.
[36]
Khomyakov N, Kharin S, Nechitailo T, et al. Reaction of microorganisms to the digestive fluid of earthworms [J]. Microbiology, 2007, 76(1): 45-54.
[37]
Zhang J, Liu J, Wang Y, et al. Profiles and drivers of antibiotic resistance genes distribution in one-stage and two stage sludge anaerobic digestion based on microwave-H2O2 pretreatment [J]. Bioresource Technology, 2017, 241: 573–581.
[38]
卢文强, 孙昊宇, 王雅娟, 等. 抗生素的胁迫与抗生素抗性基因产生与传播关系的研究[J]. 生态毒理学报, 2020, 15(4): 129-138. Lu W, Sun H, Wang Y, et al. The relationship of antibiotic stress with emergence and dissemination of antibiotic resistance genes [J]. Asian Journal of Ecotoxicology, 2020, 15(4): 129-138.
[39]
Tian B, Fadhil N, Powell J, et al. Long-term exposure to antibiotics has caused accumulation of resistance determinants in the gut microbiota of honeybees [J]. Combining Chemistry and Biology, 2012, 3(6): e00377-12.
[40]
Ding J, Zhu D, Hong B, et al. Long-term application of organic fertilization causes the accumulation of antibiotic resistome in earthworm gut microbiota [J]. Environment International, 2019, 124: 145-152.
[41]
Li B, Yang Y, Ma L, et al. Metagenomic and network analysis reveal wide distribution and co-occurrence of environmental antibiotic resistance genes[J]. The ISME Journal, 2015, 9(11): 2490-2502.
[42]
Zhu B, Chen Q, Chen S, et al. Does organically produced lettuce harbor higher abundance of antibiotic resistance genes than conventionally produced? [J]. Environment International, 2017, 98: 152-159.
[43]
Huang K, Xia H, Zhang Y, et al. Elimination of antibiotic resistance genes and human pathogenic bacteria by earthworms during vermicomposting of dewatered sludge by metagenomic analysis [J]. Bioresource Technology, 2020, 297: 122451.
[44]
程晨, 张璐璐, 赵晓祥. 重金属铜对养殖场周边土壤菌群及其大环内酯类抗生素抗性水平相关基因的影响[J]. 东华大学学报(自然科学版), 2019, 45(6): 943-950. Chen C, Zhang L, Zhao X, et al. Effects of copper on microflora and macrolide resistance genes in soil surrounding the farm [J]. Journal of Donghua University (Natural Science), 2019, 45(6): 943-950.