用简单分子分析(PCR-RFLP)测定日本大都市河流系统中微生物组的水质

L. Neneng, R. Nugroho, Y. Komai, Naru Takayama, K. Kawamura
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引用次数: 5

摘要

城市化通过污水、有毒化学物质和过量的营养物质污染自然淡水环境,造成藻华、污染和生态系统退化。为了确保自然水的可持续利用,需要适当的监测方法。本研究旨在利用低成本的基于dna的方法,PCR(聚合酶链反应)-RFLP(限制性片段长度多态性),研究日本大都市河流系统中微生物群落的多样性,作为环境变化的潜在生物指标。在一个河流系统的七个地点对地表水进行了采样。测定了水体化学参数和重金属浓度。从样品中提取微生物DNA,用通用引物扩增核糖体RNA,琼脂糖凝胶进行RFLP评分。水化学分析表明,某污水处理厂入海口的地表水有富营养化的迹象。各监测点地表水重金属浓度均较低(< 0.01 ppm)。PCR-RFLP分析显示16S和18S rrna均存在多态性,表明该方法至少可以检测河流系统中部分微生物组的变化。对部分片段进行测序,发现序列与污水处理厂分离的纤毛虫接近,暗示受到污水污染。主成分分析(PCA)鉴定出与水化学参数相关的RFLPs,可作为环境污染的生物指标。我们还发现RFLPs与水质参数无关,这表明这种简单的基于dna的分析也可以检测水生态系统中无法通过水质化学测量量化的生物变化。
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Water Quality Measurements with a Simple Molecular Analysis (PCR-RFLP) of the Microbiome in a Metropolitan River System in Japan
Urbanization has affected natural freshwater environments by contamination with sewage, toxic chemicals, and excess nutrients, which cause algal bloom, pollution, and ecosystem degradation. To ensure sustainable use of natural waters, appropriate monitoring methods are required. This study aims to investigate the diversity of the microbial community in a metropolitan river system in Japan using a low-cost DNA-based approach, PCR (Polymerase Chain Reaction)-RFLP (Restriction Fragment Length Polymorphism), as a potential bioindicator of environmental change. Surface waters were sampled in seven sites in a river system. Water chemical parameters and concentrations of heavy metals were determined. Microbial DNA was extracted from the samples, ribosomal RNA was amplified with universal primers, and RFLP was scored by agarose gels. Water chemical analyses showed that surface water at the inflow point of a sewage treatment plant had signs of eutrophication. Heavy metal concentrations in surface water were low (< 0.01 ppm) in all sites. The PCR-RFLP analysis showed polymorphisms both in 16S and 18S rRNAs, indicating that the method can detect at least a part of the microbiome changes in a river system. Sequencing of some fragments found the sequence close to a ciliate isolated in wastewater treatment plants, implying contamination from sewage. Principal component analysis (PCA) identified the RFLPs associated with chemical water parameters, which could be bioindicators of environmental pollution. We also found the RFLPs independent of water quality parameters, suggesting that this simple DNA-based analysis can also detect biological changes in water ecosystems that are not quantified by chemical measurements of water quality.
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