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Metagenomic analyses of bacterial endophytes associated with the phyllosphere of a Bt maize cultivar and its isogenic parental line from South Africa

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Abstract

Genetic modification of maize with Bacillus thuringiensis (Bt) cry proteins may predispose shifts in the bacterial endophytes’ community associated with maize shoots. In this study, the diversity of bacterial endophytes associated with a Bt maize genotype (Mon810) and its isogenic non-transgenic parental line were investigated at pre-flowering (50 days) and post-flowering (90 days) developmental stages. PCR-DGGE and high throughput sequencing on the Illumina MiSeq sequencer were used to characterize bacterial 16S rRNA gene diversity in leaves, stems, seeds and tassels. PCR-DGGE profile revealed similarity as well as differences between bacterial communities of shoots in both cultivars and at both developmental stages. A total of 1771 operational taxonomic units (OTUs) were obtained from the MiSeq and assigned into 14 phyla, 27 classes, 58 orders, 116 families and 247 genera. Differences in alpha and beta diversity measures of OTUs between the phyllospheres of both genotypes were not significant (P > .05) at all developmental stages. In all cultivars, OTU diversity reduced with plant development. OTUs belonging to the phyla Proteobacteria were dominant in all maize phyllospheres. The class Gammaproteobacteria was dominant in Bt maize while, Alphaproteobacteria and Actinobacteria were dominant in non-Bt maize phyllospheres. Differences in the abundance of some genera, including Acidovorax, Burkerholderia, Brachybacterium, Enterobacter and Rhodococcus, whose species are known beneficial endophytes were observed between cultivars. Hierarchical cluster analysis further suggests that the bacterial endophyte communities of both maize genotypes associate differently (are dissimilar). Overall, the results suggest that bacterial endophytes community differed more across developmental stages than between maize genotypes.

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Acknowledgements

This study was funded by the National Research Foundation (NRF) through the Thuthuka grant (Grant No. 84168) awarded to RA Adeleke. The authors acknowledge the Agricultural Research Council’s sponsorship of Mashiane RA through the Professional development program. We sincerely thank Owen Rhode and members of the Microbiology and Environmental Biotechnology Research Group ARC-ISCW for their technical assistance.

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Authors and Affiliations

  1. Microbiology and Environmental Biotechnology Research Group, Agricultural Research Council-Institute for Soil, Climate and Water, Pretoria, 0001, South Africa

    Ramadimetja A. Mashiane, Obinna T. Ezeokoli & Rasheed A. Adeleke

  2. Unit for Environmental Sciences and Management, North-West University, Potchefstroom, 2520, South Africa

    Ramadimetja A. Mashiane, Obinna T. Ezeokoli, Rasheed A. Adeleke & Cornelius C. Bezuidenhout

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  1. Ramadimetja A. Mashiane
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  2. Obinna T. Ezeokoli
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Correspondence to Rasheed A. Adeleke.

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Ramadimetja A. Mashiane and Obinna T. Ezeokoli have contributed equally to this work.

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11274_2017_2249_MOESM1_ESM.tif

Supplementary material 1 Fig S1. Rarefaction curve of OTUs species in maize phyllospheres. Rarefaction curve was constructed in R software by using the vegan package. (TIF 133 KB)

11274_2017_2249_MOESM2_ESM.tif

Supplementary material 2 Fig. S2. Heatmap of unweighted and weighted Bray-Curtis dissimilarity matrix between samples (a) Weighted (b) Unweighted. (TIF 45 KB)

Supplementary material 3 (TIF 52 KB)

11274_2017_2249_MOESM4_ESM.tif

Supplementary material 4 Fig. S3. Taxonomic diversity of OTUs in maize phyllospheres at order taxa level. “Others” indicate OTUs which were unclassified into phylum. Unassigned include OTUs which failed to align with any reference sequence. (TIF 103 KB)

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Mashiane, R.A., Ezeokoli, O.T., Adeleke, R.A. et al. Metagenomic analyses of bacterial endophytes associated with the phyllosphere of a Bt maize cultivar and its isogenic parental line from South Africa. World J Microbiol Biotechnol 33, 80 (2017). https://doi.org/10.1007/s11274-017-2249-y

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