基于T-RFLP技术的不同水位梯度植物根际细菌群落多样性特征分析

Analysis of the characteristics of rhizosphere bacterial diversity from plants with different water level gradients based on T-RFLP

为了解水位梯度控制下不同湿地植物根际细菌群落多样性,以北京市奥林匹克公园植物氧化塘人工湿地为例,采用末端限制性片段长度多态性(T-RFLP)技术结合ANOVA分析,比较了水位梯度控制下三棱草、芦苇、香蒲、睡莲4种植物根际细菌群落的多样性。研究结果显示:随着水位梯度加深,根际细菌群落多样性呈现减少趋势,HhaⅠ、MspⅠ、RsaⅠ3种不同酶切所得结果一致,且综合3种酶切的PAT比对中RFs组合数量随水位梯度加深也呈相应变化趋势。进一步分析发现,随水位梯度加深植物根际可培养细菌RFs数量变化较显著。不同水位梯度下各植物根际细菌群落中差异显著的菌群为Beta变形杆菌纲,水位梯度加深可能导致不同生态型植物根际泌氧能力降低,进而影响植物根际好氧细菌生存,从而出现三棱草根际属水平菌群丰富度最高,其次为芦苇、香蒲,最少为睡莲。各植物根际细菌群落的优势属多数为变形杆菌门,是脱氮除磷的功能性菌门,其中产碱杆菌属和黄杆菌属为四种植物根际细菌的共同优势属,在碳氮循环中起重要作用。

To elucidate rhizosphere bacterial diversity from plants with different water level gradients, we examined bacterial diversity in Beijing Olympic Park plant oxidation pond, an artificial wetland. The rhizosphere bacterial diversities for Carex phacota spr., Phragmites communis, Typha orientalis, and Nymphaea tetragona grown in plants with different water levels were compared based on terminal restriction fragment length polymorphism （T-RFLP） analysis combined with analysis of variance （ANOVA）. Our results showed that rhizosphere bacterial diversity decreased with increasing water levels. Different restriction enzymes, i.e.,HhaI, MspI, and RsaI, yielded the same digestion results. By combing results from digestion with these three restriction enzymes, RF combination numbers from PAT alignment also decreased with increasing water levels, consistent with the results of single restriction enzyme digestion. Further analysis showed that RF numbers in cultured rhizosphere bacteria changed significantly with the increase in water level; in contrast, the RF numbers of uncultured bacteria did not change significantly. Under different water gradients, β-Proteobacteria exhibited the most significant variations among the four plants, possibly because the increase in water level may have reduced the abilities of the different plants to secrete oxygen, thereby affecting the survival of plant rhizosphere aerobic bacteria. The triangular had the highest number of rhizosphere bacterial genera, followed by reeds, cattails, and finally water lilies. The abundances of the aerobic bacteria Alcaligenes sp., Burkholderia sp., Achromobacter sp., Delftia sp., Janthinobacterium sp., Herbaspirllum sp., Ralstonia sp., and Pelomonas sp. were probably related to the capacity of plant roots to secrete oxygen. Moreover, the oxygen-secreting capacity of rhizospheres and root exudates varied because wetland plants with different water level gradients exhibited different growth conditions and metabolic activities, affecting rhizosphere bacteria growth and reproduction. The most dominant organisms in the rhizosphere bacterial communities of these four plants were Proteobacteria, which function in the removal of nitrogen and phosphorus. The common dominant genera among these rhizosphere bacteria were Alcaligenes and Flavobacterium, which play important roles in carbon and nitrogen cycles and contribute to wetland water purification.