Dynamics of seed-borne rice endophytes on early plant growth stages

Abstract

Bacterial endophytes are ubiquitous to virtually all terrestrial plants. With the increasing appreciation of studies that unravel the mutualistic interactions between plant and microbes, we increasingly value the beneficial functions of endophytes that improve plant growth and development. However, still little is known on the source of established endophytes as well as on how plants select specific microbial communities to establish associations. Here, we used cultivation-dependent and -independent approaches to assess the endophytic bacterrial community of surface-sterilized rice seeds, encompassing two consecutive rice generations. We isolated members of nine bacterial genera. In particular, organisms affiliated with Stenotrophomonas maltophilia and Ochrobactrum spp. were isolated from both seed generations. PCR-based denaturing gradient gel electrophoresis (PCR-DGGE) of seed-extracted DNA revealed that approximately 45% of the bacterial community from the first seed generation was found in the second generation as well. In addition, we set up a greenhouse experiment to investigate abiotic and biotic factors influencing the endophytic bacterial community structure. PCR-DGGE profiles performed with DNA extracted from different plant parts showed that soil type is a major effector of the bacterial endophytes. Rice plants cultivated in neutral-pH soil favoured the growth of seed-borne Pseudomonas oryzihabitans and Rhizobium radiobacter, whereas Enterobacter-like and Dyella ginsengisoli were dominant in plants cultivated in low-pH soil. The seed-borne Stenotrophomonas maltophilia was the only conspicuous bacterial endophyte found in plants cultivated in both soils. Several members of the endophytic community originating from seeds were observed in the rhizosphere and surrounding soils. Their impact on the soil community is further discussed.

Figure 1. Dynamics of rice endophytes as revealed by PCR-DGGE profiles of seed, three- and five-week-old rice plants.

Rice endophyte PCR-DGGE patterns of surface-sterilized dehulled seeds and five-day-old shoot, root and remainder of the seeds from two consecutive generations are shown (panel A). PCR-DGGE patterns of root and shoot endosphere community of three- B) and five- C) week-old rice plants cultivated in two soil types. Six replicates per treatments are shown. Arrow heads indicate identified communities from excised PCR-DGGE bands (only numbers) and strains with identical motility (preceded by letter R; see Table 1 and 2), M – marker with a selection of 15 endophyte ribotypes (panel A).

Figure 2. Heat map composition of selected bacterial communities.

Distribution of select endophytic bacterial communities (rows) from two soil types (K and V) and four different habitats (root-free and rhizosphere soil, root and shoot endosphere) is shown. Cells are coloured in spectrum of grey that correlates with percentage of observed bacterium in a given habitat. Habitat from which the assessed bacterium was most likely to be originated from ‘artificial’ soil community is labelled with “inoculated”. Unlabelled cells are most likely represented by assessed bacterium originated from rice seeds.

Figure 3. Biplot ordination diagrams of rice shoot and root bacterial endophytes.

RDA diagrams generated from PCR-DGGE profiles of endophytic bacterial community sampled from shoot (A and B) and root (C and D) tissues of plants cultivated on K (A and C) and V (B and D) soils are shown. Squares and circle represent PCR-DGGE patterns of bacterial communities from plants submitted to, respectively, flooded and unflooded regimes and exposed to low- (empty symbol) and high- (full symbol) BID. Triangles (control treatment) represent PCR-DGGE patterns of bacterial communities from plants submitted to unflooded regime and cultivated in uninoculated soils. Six replicates of each treatment are shown. Stars represent nominal environmental variables. Arrows represent PCR-DGGE bands in which only the most descriptive communities are shown.

Figure 4. Biplot ordination diagrams of rice rhizosphere and bulk soil bacterial communities.

RDA diagrams generated from PCR-DGGE profiles of bacterial community sampled from rhizosphere (A and B) and bulk (C and D) soil of plants cultivated in K (A and C) and V (B and D) soils are shown. See Fig. 3 for symbol description.