Morphology And Structure Of Root Nodules Of Hippophae Rhamnides And Isolation And Identification Of Bacteria In The Nodules

The roots of Hippophae rhamnoides can form a special organ in association Frankia in the soil,called the nodule,in which the free nitrogen(N2)in the air is fixed effectively.However,due to the difficulty in the isolation and culture of Frankia,the relevant basic research is severely limited.In this study,the morphological and anatomical structure of the root nodule of H.rhamnoides,and the different forms of Frankia in the nodule were observed by modern microscopy techniques.Meanwhile,nodules of H.rhamnoide growing in Mafang Forest Farm of Yongshou County,Anshang Loess,Plateau Test Station,Wuliwan,Jingbian County,and Yuyang City,Yulin City,Shaanxi Province were collected.And,bacterium in the root nodules were isolated on three different media using root nodule section and homogenizenized nodule method.The isolated bacterial strains were identified by observation of colony morphology,physiological and biochemical indexes and 16 S rDNA molecular identification,in order to analyse the diversity of nodule bacteria.The results obtained are as follows:1.The distribution of root nodules indicated that the number of the nodulation of H.rhamnoides is not high,only a small number of annual and biennial roots,and occasionally on the perennial roots possessed nodules locally.The primary nodule appears as a single nodule,which develops into a coral-like tumor mass after multiple bifurcations,similar to the eucalyptus nodules.2.The anatomical structure of the single nodule showed that the site of infection by Frankia was mainly concentrated in the cells in the outer region of the outer cortex and the middle cortex,and the invading cells in the medial region and the middle column of the middle cortex were relatively few.The difference in the characteristics of infected and non-infected cells showed that the different host cells of the nodule have different ability to resist the infection of Frankia;and after the Frankia infect the host cells,they exist in three different forms of mycelium,vesicles and sporangia.3.According to the colony morphology,physiological and biochemical indexes of the strains and the results of 16 S rDNA molecular identification,refer to "Common BacterialSystem Identification Manual" and "Berger Bacterial Identification Manual(Eighth Edition)",the 90 strains isolated from the nodules of H.rhamnoides collected in the four areas of Mafang,Ansai,Wuliwan and Zhuanlongwan can be classified into 4 phyla,7 classes,14 orders,20 families and 27 genera.Among them,the 52 strains were isolated from the root nodules of H.rhamnoides in Mafang area,they are belonged to 4 phyla,6 classes,12 orders,19 families and 26 genera,mostly to the genus Micromonospora,Bacillus and Sphingomonas.The five strains isolated from the root nodule of H.rhamnoides in Ansai area belong to 3 phyla,3 classes,3 orders,3 families and 3 genera,and the 3 genera are Streptomyces,Acinetobacter and Bacillus.The five strains isolated from the root nodule of H.rhamnoides in Wuliwan area belonged to 2 phyla,2 classes,2 orders,3 families and 3 genera(Janibacter,Micromonospora and Bacillus).The 28 strains were isolated from the root nodules of H.rhamnoides in the Zhuanlongwan area,they are belonged to 3 phyla,4 classes,5 orders,5 families and 6 genera.The 6 genera were Micromonospora,Plantactinospora,Actinobacter,Hungatella and Bacillus and Sphingomonas.The further understanding of the internal anatomy of nodule and study on the bacterial diversity of root nodules of H.rhamnoides and the internal anatomical structure of nodule will help to further explore the nitrogen fixation mechanism of root nodules and improve the nitrogen fixation capacity of forests of H.rhamnoides,which are widely distributed in the northwestern China.The cultivation of forests of H.rhamnoides with ecologically fragile environment and great ecological significance also has important practical significance.