Whole genome analysis of endophytic strain PM1 reveals promising plant Growth-Promoting mechanisms in pomegranate

Abstract

The plant ecosystem harbours diverse symbiotic microorganisms with plant growth promoting and biocontrol activities. The gram- negative endophytic bacterium PM1 strain, isolated from the nodal region of pomegranate. The strain PM1 was studied through whole-genome sequencing, functional annotation, and plant growth-promoting trait (PGPT) gene analysis. Phylogenetic tree analysis and 16S rDNA sequencing confirmed its classification within the genus Brucella. The assembled genome size was 5,200,895 bp with a G + C content of 56.4 %. The average nucleotide identity (ANI) analysis revealed a 97.62 % similarity between PM1 and B. anthropi ATCC 49188 T, a type strain derived from human clinical samples, indicating a close relationship with Brucella anthropi. The functional annotation revealed 2,945 PGPT-related genes, including 32 % linked to direct effects (phytohormone signal production, biofertilization, and bioremediation processes) and 67 % to indirect effects (plant colonization, biocontrol, and competitive exclusion). KEGG analysis revealed genes involved in nitrogen metabolism, phosphate solubilization, siderophore production, hormone biosynthesis (gibberellin, cytokinin, and auxin), root colonization, and stress mitigation. Virulence factor database (VFDB) data revealed the absence of complete virulence gene assemblies, indicating limited pathogenic potential. Furthermore, secondary metabolite analysis predicted the potential production of ochrobactin compounds, which are potent siderophores that are important traits associated with PGPTs. The complete genome analysis of Brucella sp. PM1 provides new insights into plant-bacteria interactions, laying a foundation for advanced postgenomic studies and facilitating the development of bioeffective strategies such as biofertilizers or biocontrol agents for sustainable improvement in crop yields.

Keywords: Brucella anthropi; Endophyte; Plant growth promoting traits (PGPTs); Pomegranate; Secondary metabolites.

Graphical abstract

Fig. 1

Representative image of endophytic bacteria strain PM1 isolated from pomegranate nodal region (a), isolation of endophytic bacteria on LB plate (b) and Gram’s staining of bacteria (c). Scale – 1 cm (a and b) and 10 µm (c).

Fig. 2

Phylogenetic tree of PM1 was inferred by using the Maximum Likelihood method and Tamura-Nei model based on 16S rDNA sequences. Branch support was evaluated through bootstrap analysis with 1000 replications, and the resulting bootstrap values are displayed at the respective nodes.

Fig. 3

Genome based taxonomy of Brucella strain PM1 using Type strain genome server results whole genome based (a) and 16S rDNA based (b). Tree was predicted with FastME 2.1.456 from GBDP distances calculated from genome sequences. Branch lengths are scaled in terms of GBDP distance formula d5; numbers above branches are GBDP pseudo-bootstrap support values from 100 replications. Leaf labels are annotated by affiliation to species (1) and subspecies (2) clusters, genomic G + C content (3), δ values (4), overall genome sequence length (5), number of proteins (6), and the kind of strain (7). User-provided GenBank accession IDs are shown in parentheses; master record accessions are truncated.

Fig. 4

A circular representation of the B. anthropi PM1 genome (5,200,895 bp). This includes, from outer to inner rings, the contigs, CDS on the forward strand, CDS on the reverse strand, RNA genes, CDS with homology to known antimicrobial resistance genes, CDS with homology to know virulence factors, GC content and GC skew (a). The colors of the CDS on the forward and reverse strand indicate the subsystem that these genes belong to. A subsystem is a set of proteins that together implement a specific biological process or structural complex and annotation includes an analysis of the subsystems unique to each genome (b).

Fig. 5

The genome annotation information of the predicted gene functions of PM1 strain based on COG (a) and Interproscan (b) and KEGG (c) databases.

Fig. 5

The genome annotation information of the predicted gene functions of PM1 strain based on COG (a) and Interproscan (b) and KEGG (c) databases.

Fig. 6

The Krona plot depicting the predicted PGPT (plant-Brucella interaction) from the PGPT tool (PLabase). BlastP and HMMER annotation against the PGPT-BASE were used to identify PGPT. Level three of six annotation depth is presented here.

Fig. 7

Secondary metabolite gene cluster prediction based on antiSMASH analysis: total regions of gene cluster (a) and potential siderophore region (b).