毒死蜱降解菌Shingopyxis granuli CP-2土著细菌帮手的筛选
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1.苏省农业科学院农产品质量安全与营养研究所;2.江苏省农业科学院农产品质量安全与营养研究所;3.生态环境部南京环境科学研究所

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Screening of Indigenous Microbial Helpers for the Chlorpyrifos-degrading Bacterium Shingopyxis granuli CP-2
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1.Institute of Food Safety and Nutrition, Jiangsu Academy of Agricultural Sciences;2.Nanjing Institute of Environmental Sciences

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    摘要:

    毒死蜱残留是食品安全中的一个重要问题,应用微生物降解是消除其残留的有效方法。但其微生物降解受降解菌与目的环境中土著微生物互作关系的影响,目前缺少对该微生态过程的研究。以毒死蜱降解菌Shingopyxis granuli CP-2为材料,从毒死蜱降解菌-土著微生物互作的角度,首先从土壤中分离出一批细菌,并对其进行16S rRNA基因全长测序鉴定;然后从中筛选对CP-2生长有促进作用的土著细菌帮手,并结合以上测序数据分析帮手的相关生物信息学结果;最后初步研究CP-2的土著细菌帮手对其降解毒死蜱的影响。结果表明,分离到的109株土著细菌隶属于变形菌门、厚壁菌门、拟杆菌门和放线菌门;其中45株细菌显著促进CP-2的生长,属于CP-2的土著细菌帮手;该帮手库中45株细菌主要隶属于3个门、4个纲、7个目、13个科和20个属;从其中选择一株对CP-2生长促进作用较强的B72菌株,检测发现,该菌及其无菌发酵液显著促进CP-2对毒死蜱的降解。综上,该研究建立由45株土著细菌构成的毒死蜱降解菌CP-2的帮手库,为后续土壤中毒死蜱或者其他污染物的微生物降解相关研究及应用提供丰富的菌种资源,且毒死蜱降解菌CP-2的土著细菌帮手显著促进其降解毒死蜱的能力,为今后毒死蜱降解菌和土著细菌帮手们的共接种修复污染提供理论指导和技术支撑。

    Abstract:

    【Objective】Chlorpyrifos residue poses a significant challenge to food safety. Microbial degradation which is called bioaugmentation is an effective approach for the elimination of such residues. Bioaugmentation often involves an invasion process requiring the establishment and activity of a foreign microbe in the resident community of the target environment. Interactions with resident micro-organisms, either antagonistic or cooperative, are believed to impact invasion. However, few studies have examined how the interactions between the invaded degrading bacteria and resident microorganisms in the target environment can influence microbial degradation. In this study, chlorpyrifos-degrading bacteria Shingopyxis granuli CP-2 was used as material, from the perspective of microbe-microbe interactions, to select resident bacterial helper of CP-2. 【Method】Soils from the field were first collected, a batch of bacteria from the soil was isolated by continuous dilution method, and identified by full-length sequencing of the 16S rRNA gene. The 16S rRNA gene sequences of all isolates were aligned using MUSCLE. Sequences in the alignment were trimmed at both ends to obtain maximum overlap using the MEGA X software, which was also used to construct taxonomic cladograms. A maximum-likelihood (ML) tree was constructed, using a General Time Reversible (GTR) + G + I model, which yielded the best fit to our data set. Bootstrapping was carried out with 100 replicates retaining gaps. A taxonomic cladogram was created using the EVOLVIEW web tool (https://evolgenius.info//evolview-v2/). The taxonomic status (phylum) of each rhizobacterial strain was also added as heatmap rings to the outer circle of the tree. The resident bacterial helper which could promote the growth of CP-2 was then screened by supernatant assay from the isolates isolated from soil, and the bioinformatics results of these helpers were analyzed. At last, a bacterial isolate which well promoted the growth of CP-2 was chosen, and its effect on CP-2's ability to degrade chlorpyrifos was investigated in vitro. 【Result】109 strains of indigenous bacteria were isolated and were classified into four main phyla: Proteobacteria (54.1%), Actinobacteria (14.8%), Firmicutes (15.6%), and Bacteroidetes (15.6%). Among them, 41.3% significantly inhibited the growth of CP-2, 17.4% had no significant effect on CP-2, and 41.3% (45 bacterial strains) significantly enhanced CP-2's growth and were identified as indigenous bacterial helpers of CP-2. The 45 bacterial strains in the helper bank mainly belong to 3 phyla, 4 classes, 7 orders, 13 families and 20 genera. One strain (B72), which exhibited a strong growth-promoting effect on CP-2 was selected to assess its impact on chlorpyrifos degradation by CP-2. The results demonstrated that both the bacterial strain B72 and its supernatant significantly promoted the chlorpyrifos degrading ability of CP-2. 【Conclusion】Together, the strains identified in this study provide valuable resources for future research and applications involving microbial degradation of soil toxicants such as chlorpyrifos or other pollutants. Furthermore, the indigenous bacterial helper of chlorpyrifos degrading bacterium CP-2 significantly promoted its ability to degrade chlorpyrifos, which offers theoretical guidance and technical support for potential co-inoculation strategies involving both chlorpyrifos-degrading bacteria and indigenous bacterial helpers aimed at pollution remediation.

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李梅,王中阳,荆莉莉,侯玉刚,马丽雅,生弘杰,余向阳.毒死蜱降解菌Shingopyxis granuli CP-2土著细菌帮手的筛选[J].土壤学报,,[待发表]
LI MEI, WANG Zhongyang, JING Lili, HOU Yugang, MA Liya, SHENG Hongjie, YU Xiangyang. Screening of Indigenous Microbial Helpers for the Chlorpyrifos-degrading Bacterium Shingopyxis granuli CP-2[J]. Acta Pedologica Sinica,,[In Press]

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  • 收稿日期:2023-11-20
  • 最后修改日期:2024-03-11
  • 录用日期:2024-05-14
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