Background Casuarina equisetifolia is one of the most important artificially planted protective forests along the coast in southern China for windbreaks, soil erosion, and sand dune stabilization. Self-renewing of C. equisetifolia is very limited, which might be caused by low soil nutrient levels and reduced microbial activity. Methods Use of high-throughput sequencing of the 18S rDNA to investigate the microbial communities from the rhizosphere and root endosphere of C. equisetifolia in young-aged, intermediate-aged, and mature-aged forests. Results Our results indicate that the diversity of rhizosphere fungal microbiomes in field-grown C. equisetifolia is much lower than that of the endosphere microbiomes. Bioinformatic analysis showed that rhizocompartments produce the strongest differentiation of rhizosphere and endosphere communities. Notably, the distribution of rhizosphere fungi communities was significantly influenced by the environmental factors, not by forest ages. Conclusions The presented study suggests that the rhizocompartments and environmental factors, rather than forest ages, determine the diversities of fungal community.

We determined the role of endophytic bacteria of C. equietifolia, involved in synthesis of allelochemicals. The root endophytic bacteria were isolated and their biological activity was studied to determine their allelopathic potential in fermentation broth. Twenty one species of endophytic bacteria were isolated from the C. equisetifolia roots, of which Bacillus were the dominant genus, Acinetobacter and Staphylococcus were other dominant genera. These 21 endophytic bacteria inhibited the seeds germination of C. equisetifolia, among which Bacillus amyloliquefaciens was most inhibitory. Main allelopathic components of fermentation broth of 3-endophytic bacteria (Bacillus amyloliquefaciens, Bacillus aryabhattai and Paenibacillus glycanilyticus) with strongest allelopathic potential were rich in phenols, esters, organic acids, aldehydes, alcohols, ketones etc. Among them, two allelopathic substances [2,2'-methylenebis[6-(1,1-dimethylethyl)-4-methyl-Phenol and 1,2,3,4-Butanetetrol] were present in both endophytic bacteria fermentation broth and in root and soil extracts. These results confirmed that the secondary metabolites of C. equisetifolia root endophytic bacteria were allelopathic.

Productivity decline of Casuarina equisetifolia plantation and difficulty in natural regeneration remains a serious problem because of allelopathy. Previous studies have confirmed that 2,4-di-tert-butylphenol (2,4-DTBP) are the major allelochemicals of the C. equisetifolia litter exudates. The production of these allelochemicals may derive from decomposition of litter or from the litter endophyte and microorganisms adhering to litter surfaces. In the present study, we aimed to evaluate the correlation between allelochemicals in litter and endophytic and epiphytic fungi and bacteria from litter. A total of 100 fungi and 116 bacteria were isolated from the interior and surface of litter of different forest ages (young, half-mature, and mature plantation). Results showed that the fermentation broth of fungal genera Mycosphaerella sp. and Pestalotiopsis sp., and bacterial genera Bacillus amyloliquefaciens, Burkholderia-Paraburkholderia, and Pantoea ananatis had the strongest allelopathic effect on C. equisetifolia seeds. Allelochemicals, such as 2,4-DTBP and its analogs were identified in the fermentation broths of these microorganisms using GC/MS analysis. These results indicate that endophytic and epiphytic fungi and bacteria in litters are involved in the synthesis of allelochemicals of C. equisetifolia. To further determine the abundance of the allelopathic fungi and bacteria, Illumina MiSeq high-throughput sequencing was performed. The results showed that bacterial genera with strong allelopathic potential were mainly distributed in the young and half-mature plantation with low abundance, while the abundance of fungal genera Mycosphaerella sp. and Pestalotiopsis sp. were higher in the young and mature plantations. In particular, the abundance of Mycosphaerella sp. in the young and mature plantations were 501.20% and 192.63% higher than in the half-mature plantation, respectively. Overall, our study demonstrates that the litter fungi with higher abundance in the young and mature plantation were involved in the synthesis of the allelochemical 2,4-DTBP of C. equisetifolia. This finding may be important for understanding the relationship between autotoxicity and microorganism and clarifying the natural regeneration problem of C. equisetifolia.

Sugarcane ratoon performance declines with increasing age due to the intergenerational accumulation and release of allelochemicals by old sugarcane roots. We aimed to clarify the effects of these allelochemicals on sugarcane seedling growth under continuous sugarcane ratoon cropping. We investigated the allelopathic effects of treatment with root aqueous extracts from sugarcane ratoons of different ages on the osmoregulatory substance content, antioxidant enzyme activity, membrane lipid peroxidation product content, photosynthetic physiological characteristics, and root exudate enzymatic characteristics of sugarcane seedlings. The root aqueous extracts exerted allelopathic effects on sugarcane seedlings. The relative electrolyte leakage, proline content, soluble protein content, soluble sugar content, malondialdehyde content, and catalase activity of the leaves were more sensitive than those of the roots. Conversely, the superoxide dismutase and peroxidase activities of the roots (positive response) were more sensitive than those of the leaves (negative response). The total chlorophyll content and net photosynthetic rate of the leaves exhibited relatively sensitive negative responses. Urease activity negatively responded but sucrase activity positively responded. We concluded that the root aqueous extracts of sugarcane ratoons may exert their allelopathic effects by affecting the level of osmoregulatory substances and causing peroxidative damage to the cell membranes of sugarcane seedlings and altering the activities of various enzymes in the rhizosphere.

Casuarina equisetifolia, an important pioneer tree with resistance to typhoon and stress tolerance, is mainly cultivated in the coastal areas of tropical and subtropical zones. Casuarina equisetifolia forests have high litter accumulation rates and slow litter decomposition rates. The accumulation of litter may affect the secondary regeneration of Casuarina equisetifolia, and the decomposition of litter is largely controlled by microorganisms such as bacteria and fungi. While the characteristics of the microbial diversity in the litter of Casuarina equisetifolia forests have not yet been resolved. In order to explore the microbial distribution characteristics and community dynamic changes in the litter of a Casuarina equisetifolia forest, we used Illumina Miseq high-throughput sequencing technology to analyze the endophytic and exophytic bacterial and fungal communities of Casuarina equisetifolia forests of three ages (young forest, half-mature forest, and mature forest). The results showed that the young and half-mature forests had the highest bacterial diversity, and the half-mature and mature forests had the highest fungal diversity. The community structure and endophytic and exophytic communities of different forest ages were also different. Exophytic bacteria of the phylum Proteobacteria had a gradually decreasing abundance with stand age, while endophytic bacteria of the phylum Actinomycetes had a gradually increasing abundance with stand age; in the half-mature forest, the abundance of nitrogen-fixing microorganisms, such as the genera Rhizobium, Bradyrhizobium, and Actinoplanes, were increased relative to all of the other forest age types. The fungal community structure was different than the bacterial structure. Specifically, the fungal communities of both the young and mature forests were dominated by the class Dothideomycetes of the phylum Ascomycota, while phylum Basidiomycota was dominant in the half-mature forest. The correlation with the physicochemical properties of the litter showed that exophytic bacteria were related to the organic carbon and water content of the litter, endophytic bacteria were related to litter pH and nitrogen content, and fungi were correlated tonitrogen and phosphorus contents. In summary, the community structures of bacteria and fungi in the half-mature forest were more uniform, which was beneficial to the growth of Casuarina equisetifolia plants, and the changes observed in the physical and chemical factors of litter were one of the key factors driving microbial community structure.