大蜡螟和黄粉虫肠道菌中聚乙烯地膜降解细菌的筛选及其降解性能

Screening of polyethylene film-degrading bacteria from gut microbiota of Galleria mellonella and Tenebrio molitor

摘要:

【背景】在我国农业生产中大量使用的聚乙烯(polyethylene，PE)地膜难以降解，在土壤中长期累积影响农作物生长并破坏生态环境，发掘微生物资源，寻求聚乙烯生物降解途径对治理“白色污染”具有重要意义。【目的】以不同来源的啮食塑料昆虫大蜡螟、黄粉虫为材料，从肠道菌群中分离筛选出对PE具有降解能力的细菌菌株，研究其降解农用地膜的效能。【方法】饲喂PE膜片驯化大蜡螟、黄粉虫幼虫，采集肠道液富集培养、共代谢驯化、选择培养基筛选等方法从肠道细菌中分离出以PE为唯一碳源的细菌菌株。将菌株接种到以PE膜片为唯一碳源的培养基中共培养，通过测定菌体生长量，定期检测膜片失重率，结合高分辨场发射扫描电子显微镜观察、红外扫描分析和膜片力学性能测定，评价菌株对聚乙烯地膜的降解效果。对筛选出的降解性能良好的菌株通过16S rRNA基因扩增和序列分析进行菌株鉴定。【结果】从新疆蜜蜂蜂巢中的土著大蜡螟肠道分离获得的聚乙烯降解菌菌株最多，其聚乙烯的降解效率高于其他来源的分离菌株。从中筛选出具有较高降解能力的3个菌株XJDLM-3、XJDLM-8和XJDLM-12，它们能利用PE膜片生长，扫描电镜观察经过30 d降解的PE膜片表面出现明显的侵蚀孔洞和裂痕，红外扫描图谱发生改变，拉伸强度、断裂伸长率和弹性模量等力学性能显著下降，膜片失重率分别达到了8.06%、5.66%和5.39%。从新疆蜜蜂蜂巢中的土著大蜡螟肠道分离出降解效果较好的细菌菌株，经鉴定XJDLM-8和XJDLM-12为Bacillus cereus，XJDLM-3为Enterobacter bugandensis。【结论】证明了新疆蜜蜂蜂巢中的土著大蜡螟肠道存在对PE具有较高降解能力的菌株，丰富了PE降解菌的菌种资源，在PE地膜降解中具有开发应用的潜力。

Abstract:

[Background] Polyethylene mulch film is widely used for agricultural production in China. Because it is very difficult to be naturally degraded in the fields, it finally accumulates in the soil and negatively affects the growth of crops and the ecological environment. Therefore, it is of great significance to explore the microbial resources for biodegradation of such “white pollution”. [Objective] Bacteria were isolated and screened from the intestinal flora of plastic-eating insects such as Galleria mellonella and Tenebrio Molitor from different sources, and their degradation efficiency of agricultural plastic film was characterized. [Methods] The larvae of Galleria mellonella and Tenebrio molitor were domesticated by polyethylene membrane, and the bacteria with polyethylene as the sole carbon source were isolated from intestinal bacteria by collecting intestinal fluid, enriching culture, co-metabolic domestication and selecting culture medium. The strain was inoculated into the medium with polyethylene membrane as the only carbon source for co-culture. The degradation effect of the strain on polyethylene mulch film was evaluated by measuring cell growth, regularly detecting the weight loss rate of the film, combined with high-resolution field emission scanning electron microscope observation, infrared scanning analysis and determination of mechanical properties of the film. The strains with good degradability were identified by 16S rRNA gene amplification and sequence analysis. [Results] Most polyethylene-degrading bacteria were isolated from the intestines of the indigenous Galleria mellonella in the honeybee hive in Xinjiang, and the degradation efficiency of polyethylene was higher than that of the isolates from other sources. Three strains XJDLM-3, XJDLM-8 and XJDLM-12, with high degradation ability were selected to grow using polyethylene membrane. scanning electron microscope observation showed that obvious erosion holes and cracks appeared on the surface of degraded polyethylene film after 30 days, and the infrared scanning pattern changed. The mechanical properties such as tensile strength, elongation at break and elastic modulus decreased significantly, and the weight loss rate of polyethylene diaphragm reached 8.06%, 5.66% and 5.39%, respectively. A bacterial strain with good degradation effect was isolated from the intestines of the native Galleria mellonella in the honeybee hive in Xinjiang. XJDLM-8 and XJDLM-12 were identified as Bacillus cereus, XJDLM-3 and Enterobacter bugandensis. [Conclusion] Strains were isolated with high ability to degrade polyethylene in the intestines of the native Galleria mellonella in the honeybee hive in Xinjiang, to provide the potential of development and application in the degradation of polyethylene mulch film.