Metagenomic analysis of endophytic microbiome inhabiting maize plant
Abstract
Endophytes are promising microbes which support the plant in many beneficial ways such as growth promotion and health improvement with no harmful effects to their host. However, to have better insights in to important microbes, diverse next-generation sequencing (NGS) methods have been adopted to study the diversity of microbes in the maize plant, with limited understanding of the functions. Presently, shotgun metagenomics is one of the novel sequencing techniques which allows the profiling of the entire microbial communities and their functions in a given environment. Therefore, this study is aimed at profiling the influence of different farming practices on the diversity, community structure, and functions of the endophytic microbiome associated with the root of maize plant (Zea mays L.) using shotgun metagenomic sequencing. The study was carried out on the experimental plots of North-West University School Farm, Molelwane, Mafikeng, North West Province, South Africa. Three experimental sites were adopted within the farm representing different farming practices namely inorganic fertilizer (NK) site, organic fertilizer (FK) site, and no fertilizer (CK) site. These sites have been in operation for more than 15years following standard procedures. Maize seed WEMA (WE 3127) was used in the experiment, these seeds were planted on the 3 sites respectively. Each farming site was divided into three regions representing 3 replicates for each farming site for the sequencing. The plants were harvested at the fruiting stage of growth. The total DNA extraction for each biological replicate was carried out and sequenced using shotgun sequencing approach. The sequenced data obtained in fastq format were uploaded on to an online database called MG-RAST where quality control (QC) assessments were carried out. Our taxonomic annotation results using the Subsystem database revealed a total phyla of 28 endophytic bacteria, 3 endophytic archaea and 2 endophytic fungi in all the samples. The major endophytic bacteria phyla observed were Firmicutes, Bacteroidetes, Actinobacteria, Bacilli, Proteobacteria,
Acidobacteria, Chloroflexi, Verrucomicrobia, Tenericutes, Planctomycetes, Cyanobacteria, and Chlorobi. Bacteroidetes dominated maize from organic fertilizer sites, Firmicutes dominated the no fertilizers site while Proteobacteria dominated inorganic fertilizer sites. The three identified endophytic archaea phyla are Crenarchaeota, Euryarchaeota, and Thaumarchaeota while the identified fungi phyla were Ascomycota and Basidiomycota in the samples across sites. The diversity analysis revealed that the abundance of endophytic bacteria, archaea and fungi in all the sites are in the order organic fertilizer (FK) > no fertilizer (CK) > inorganic fertilizer (NK) with higher abundance in samples from organic fertilizer site. Our functional annotation results further revealed a total of 28 functional groups within the endophytic microbiomes across the farming sites. Some functional groups and metabolisms associated with plant growth promotion such as secondary metabolism, nitrogen metabolism, iron acquisition and metabolism alongside phosphorus metabolism were observed in the endophytes across the sites. Our results further showed the presence of putative functional genes associated with plant growth-promotion (PGP) and endophytic behaviors. Taken together, using the shotgun metagenomic approach, this study showed that organic farming has a positive influence on the diversity, abundance, and functions of the endophytic microbiome in maize plant.