Expression and lateral transfer of nitrogen fixation genes in the gut of earthworms.

研究了一种蚯蚓（Amynthas vittatus）对土壤氮含量增加的贡献。将3只蚯蚓成虫标本放入300 g添加1%羧甲基纤维素的土壤（灰色低地土，粉质粘土）中，在22℃黑暗条件下培养32 d。与蚯蚓孵育后，土壤中全氮、氨氮和硝态氮含量及好氧固氮细菌数量显著增加，土壤中^<15>N （δ^<15>N）的自然丰度降低。蚯蚓体内的氮含量在孵育过程中没有减少。蚯蚓的引入显著提高了土壤中^<15>N的还原活性和大气^<15>N向土壤的吸收，但土壤中氮的观测增量远高于根据固氮活性估计的增量。通过从土壤中分离NFBs和分析土壤中nifH的PCR-DGGE模式，估计了蚯蚓生活的土壤中好氧固氮细菌的群落结构。生理特征和16S rDNA序列分析表明，分离得到的nifH可分为茎状固氮根瘤菌、巴西固氮螺旋菌、a.l lipoferum、氧化克雷伯菌、假单胞菌、etrhizobium etli、Rosemonas faurise、Sinorhizobium morelens和Xanthobacter sp。蚯蚓培养土壤中nifH的DGGE条带数量和模式与蚯蚓粪便和非蚯蚓土壤中nifH的DGGE条带不同。本研究结果表明，蚯蚓增加了土壤的氮含量，可能是由于蚯蚓从微观上增强了土壤的固氮活性。

The contribution of an earthworm species (Amynthas vittatus) to the increase of the nitrogen content of soil was examined. Three specimens of adult earthworms were introduced into 300 g of soil (Gray Lowland soil, silty clay) supplemented with 1% carboxymethyl cellulose in a container and incubated for 32 days at 22℃ in the dark. The contents of total-N, NH_4-N and NO_3-N, and the population of aerobic nitrogen-fixing bacteria in soil significantly increased after incubation with the earthworms, while the natural abundance of ^<15>N (δ^<15>N) in soil decreased. The amount of nitrogen in the earthworms did not decrease during the incubation in the microcosm. Both acetylene reduction activity of the microcosm and incorporation of ^<15>N to soil from atmospheric ^<15>N_2 were significantly enhanced by the introduction of the earthworms into soil, though the observed increment of nitrogen in soil was much higher than the estimated one based on the nitrogen-fixing activity.The community structure of aerobic nitrogen-fixing bacteria in the soils where the earthworms had lived was estimated by both isolating NFBs from the soils and the analysis of PCR-DGGE patterns of nifH from the soils. Physiological characteristics and 16S rDNA sequence analysis showed that the isolated NFBs were classified into Azorhizobium caulinodans, Azospirillum brasilense, A.lipoferum, Klebsiella oxytoca, Pseudomonas sp., Rhizobium etli, Rosemonas faurise, Sinorhizobium morelens, and Xanthobacter sp. The number and the pattern of DGGE bands of nifH from the soil incubated with earthworms were different from those of nifH from earthworm casts and the soil in which earthworms did not live.The results obtained in the present study indicated that the earthworms increased the nitrogen content of soil, presumably due to the enhancement of the nitrogen-fixing activity of the soil from the microcosm by the earthworms.