Metagenomic analysis of 16S sequences of nitrifying bacteria and archaea inhabiting maize rhizosphere

Abstract

The global nitrogen cycle has been disrupted, mainly as a result of nitrogen-based synthetic fertilizers and other agrochemicals used in plant cultivation. Microbial inoculation is fast becoming an environmentally friendly choice of biofertilizer. Nitrifying bacteria and archaea are the chief contributors of nitrogen available for plant use in the soil. The growing demand for maize has intensified its cultivation. Hence, there is a need to identify nitrifying microorganisms associated with maize plants, understand their relationship with the soil environmental factors, other microbes, and the soil community level physiological profile. This would enable efficient management of the studied group of organisms to maximize their potential for the growth of maize. Soil samples from the rhizosphere were obtained at various phases (pretassling, tassling and fruiting) of maize growth at North-West University Farm Molelwane, South Africa. Bulk soil was also collected. The nitrifying bacteria, archaea, and other microorganisms found in the maize rhizosphere were identified using 16S amplicon sequencing. The DNA was isolated from the soil samples using the nucleospin soil DNA extraction kit and sequenced on the Illumina Miseq. The acquired sequences were examined and processed using MG-RAST. The physical and chemical parameters of the rhizosphere were determined, and their impact on the nitrifying community was assessed. Also, the community level physiological profile was carried out using the Microresp Technique. The result revealed 9 genera of nitrifying bacteria; Nitrospira, Nitrosospira, Nitrobacter, Nitrosovibrio, Nitrosomonas, Nitrosococcus, Nitrococcus, unclassified (derived from Nitrosomonadales), unclassified (derived from Nitrosomonadaceae) and 1 archaeon, Candidatus Nitrososphaera. The Nitrospirae phyla group, which had most of the nitrifying bacteria, was more abundant at the tasselling stage (67.94%). Alpha diversity showed no significant difference. However, the Beta diversity showed a significant difference (P = 0.01, R = 0.58) across the growth stages. Although growth stages had no effect on nitrifying

bacteria and archaea diversity, there was variation in microbial structure as it related to maize growth stages. The bulk of the microorganisms were detected during the fruiting stage, whereas the nitrifying bacteria were most abundant during the tasselling stage. The pH values for soil chemical characteristics obtained varied from 5.35 to 6.22, with a mean of 5.93. The carbon-nitrogen ratio is around 9:1. The NH4 to NO3 ratio is about 1:1.4. There was a significant correlation between some of the parameters and the nitrifying microorganisms. The relationship between nitrifying bacteria, archaea, and other microbial groups revealed a significant negative and positive correlation. The Pearson correlation further showed a positive relationship between unclassified Nitrosomonadales and Bacillus (r = 0.59), unclassified Nitrosomonadales and Azospirillum (r = 0.52), Nitrobacter and Azospirillum (r = 0.54), Nitrosomonas and Stenotrophomonas (r = 0.54), Candidatus Nitrosphaera and Rhizobium (r = 0.68), Nitrospira and alanine (r = 0.52), and Lysine and Nitrobacter (r = 0.54). This study found previously known and undiscovered nitrifying bacteria and archaea linked with the maize rhizosphere. It has also demonstrated the relationship between the identified nitrifiers and soil chemical properties. The findings of this study will aid in the improvement of maize growth and development by altering the structure of the rhizosphere microbial community.