"Duckweed-Microbe Co-Cultivation Method" for Isolating a Wide Variety of Microbes Including Taxonomically Novel Microbes.

We herein described a new microbial isolation method using the interaction between the floating aquatic plant, duckweed, and microbes. We harvested microbial cells from Japanese loosestrife roots and co-cultivated these cells with aseptic duckweed using artificial inorganic medium for the plant for four weeks. During the co-cultivation, some duckweeds were collected every week, and the roots were used for microbial isolation using a low-nutrient plate medium. As a result, diverse microbial isolates, the compositions of which differed from those of the original source (Japanese loosestrife root), were obtained when the roots of duckweed were collected after 2 weeks of cultivation. We also successfully isolated a wide variety of novel microbes, including two strains within the rarely cultivated phylum, Armatimonadetes. The present study shows that a duckweed-microbe co-cultivation approach together with a conventional technique (direct isolation from a microbial source) effectively obtains more diverse microbes from a sole environmental sample.

Quantifying the associations between fungal endophytes and biocontrol-induced herbivory of invasive purple loosestrife (Lythrum salicaria L.).

Fungal endophytes are one of several groups of heterotrophic organisms that associate with living plants. The net effects of these groups of organisms on each other and ultimately on their host plants depend in part on how they facilitate or antagonize one another. In this study we quantified the associations between endophyte communities and herbivory induced by a biological control in the invasive Lythrum salicaria at various spatial scales using a culture-based approach. We found positive associations between herbivory damage and endophyte isolation frequency and richness at the site level and weak, positive associations at the leaf level. Herbivory damage was more strongly influenced by processes at the site level than were endophyte isolation frequency and community structure, which were influenced by processes at the plant and leaf levels. Furthermore, endophytic taxa found in low herbivory sites were nested subsets of those taxa found at high herbivory sites. Our findings suggest that endophyte communities of L. salicaria are associated with, and potentially facilitated by, biocontrol-induced herbivory. Quantifying the associations between heterotrophic groups ultimately may lead to a clearer understanding of their complex interactions with plants.

Microbial community analysis in the roots of aquatic plants and isolation of novel microbes including an organism of the candidate phylum OP10.

A number of molecular ecological studies have revealed complex and unique microbial communities in various terrestrial plant roots; however, little is known about the microbial communities of aquatic plant roots in spite of their potential use for water quality improvement in aquatic environments (e.g. floating treatment wetland system). Here, we report the microbial communities inhabiting the roots of emerged plants, reed (Phragmites australis) and Japanese loosestrife (Lythrum anceps), collected from a floating treatment wetland in a pond by both culture-independent and culture-dependent approaches. Culture-independent analysis based on 16S rRNA gene sequences revealed that the microbial compositions between the two aquatic plant roots were clearly different (e.g. the predominant microbe was Betaproteobacteria for reed and Alphaproteobacteria for Japanese loosestrife). In comparisons of microbial communities between the plant roots and pond water taken from near the plants, the microbial diversity in the plant roots (e.g. 4.40-4.26 Shannon-Weiner index) were higher than that of pond water (e.g. 3.15 Shannon-Weiner index). Furthermore, the plant roots harbored 2.5-3.5 times more phylogenetically novel clone phylotypes than pond water. The culture-dependent approach also revealed differences in the microbial composition and diversity among the two plant roots and pond water. More importantly, compared to pond water, we succeeded in isolating approximately two times more novel isolate phylotypes, including a bacterium of candidate phylum OP10 (recently named Armatimonadetes) from the plant roots. These findings suggest that aquatic plants roots are significant sources for a variety of novel organisms.

Relationships among three pathways for resource acquisition and their contribution to plant performance in the emergent aquatic Plant Lythrum salicaria (L.).

Three pathways for resource acquisition exist in the emergent aquatic plant, Lythrum salicaria (L.); a subterranean root system, a free-floating adventitious root system, and arbuscular mycorrhiza (AM) fungal hyphae colonizing subterranean roots. This study examined the relationship(s) among these pathways and their contribution to plant performance. If the free-floating adventitious root system and/or AM fungi contribute to plant growth in wetland habitats, we predicted that their absence would result in a significant reduction in plant performance. Furthermore, if a reduction in resource uptake, effected by an absence of free-floating adventitious roots and/or AM fungi, is compensated for by increased allocation to remaining pathway(s) for resource uptake, we predicted altered patterns of resource allocation among shoots and the remaining pathway(s) for resource uptake. Contrary to our predications, plants experiencing adventitious root removal and/or grown in the absence of AM fungi generally had greater biomass and total shoot height than controls. Similarly, while levels of AM colonization and subterranean root biomass displayed a treatment effect, the observed responses did not correspond with our predictions. This was also true for shoot : subterranean root dry weight ratios. Our results indicate that there is interaction among the 3 pathways for resource acquisition in L. salicaria and an effect on plant performance. The adaptive significance of these characteristics is unclear, highlighting the potential difficulties in extrapolating from terrestrial to aquatic plant species and among aquatic plant species with potentially different life history strategies.

Phosphorus, arbuscular mycorrhizal fungi and performance of the wetland plant Lythrum salicaria L. under inundated conditions.

The role of arbuscular mycorrhizal (AM) fungi in aquatic and semi-aquatic environments is poorly understood, although they may play a significant role in the establishment and maintenance of wetland plant communities. We tested the hypothesis that AM fungi have little effect on plant response to phosphorus (P) supply in inundated soils as evidenced by an absence of increased plant performance in inoculated (AM+) versus non-inoculated (AM-) Lythrum salicaria plants grown under a range of P availabilities (0-40 mg/l P). We also assessed the relationship between P supply and levels of AM colonization under inundated conditions. The presence of AM fungi had no detectable benefit for any measures of plant performance (total shoot height, shoot dry weight, shoot fresh weight, root fresh weight, total root length or total root surface area). AM+ plants displayed reduced shoot height at 10 mg/l P. Overall, shoot fresh to dry weight ratios were higher in AM+ plants although the biological significance of this was not determined. AM colonization levels were significantly reduced at P concentrations of 5 mg/l and higher. The results support the hypothesis that AM fungi have little effect on plant response to P supply in inundated conditions and suggest that the AM association can become uncoupled at relatively high levels of P supply.