Multiple Plant Growth-Promotion Traits in Endophytic Bacteria Retrieved in the Vegetative Stage From Passionflower

Abstract

Bacteria exhibiting beneficial traits like increasing the bioavailability of essential nutrients and modulating hormone levels in plants are known as plant growth promoting (PGP) bacteria. The occurrence of this specific group of bacteria in the endophytic environment may reflect the decisive role they play in a particular condition. This study aimed to determine the taxonomical diversity of the culturable bacterial endophytes, isolated in the vegetative stage of passionflower (Passiflora incarnata), and assess its potential to promote plant growth by phenotypic and genotypic approaches. The sequencing and phylogenetic analysis of the 16S rRNA gene allowed us to classify 58 bacterial endophytes into nine genera. Bacillus (70.7%) was the most dominant genus, followed by Pseudomonas (8.6%) and Pantoea (6.9%). A few isolates belonged to Rhodococcus and Paenibacillus, whereas the genera Lysinibacillus, Microvirga, Xanthomonas, and Leclercia were represented by only one isolate. The strains were tested for nitrogen fixation, phosphate solubilization, indole-acetic-acid synthesis, and siderophore production. Moreover, PGP related genes (nifH, ipdC, asb, and AcPho) were detected by PCR-based screening. Most of the isolates (94.8%) displayed a potential for at least one of the PGP traits tested by biochemical assays or PCR-based screening. Nine strains were selected based on results from both approaches and were evaluated for boosting the Cape gooseberry (Physalis peruviana) germination and growth. All tested isolates improved germination in vitro, and the majority (78%) increased growth parameters in vivo. The results suggested that most of culturable bacteria inhabiting P. incarnata in the vegetative stage could be used as probiotics for agricultural systems. Besides, their occurrence may be associated with specific physiological needs typical of this development stage.

Keywords: Cape gooseberry; PGP bacteria; PGP genes; Passiflora incarnata; inoculant.

Figure 1

Neighbor-joining phylogenetic tree based on partial 16S rRNA sequences of endophytic bacteria isolated from Passiflora incarnata leaves and reference sequences from EzBioCloud. The branch colors indicate different bacterial genera. Only bootstrap values equal and greater than 60% are displayed as circles with increasing size up to 100%. Accession numbers from references sequences are in parentheses.

Figure 2

Diversity of PGP traits among endophytic bacteria according to their taxonomic affiliations inferred from phylogenetic analysis of 16S rRNA gene sequences. Growth on a N-free media (); detection of the nifH gene(); formation of halo on a NBRIP medium ();detection of the AcPho gene (); IAA-like compound production (); detection of the ipdC gene (); formation of orange halo on a CAS Agar (); detection of the asb gene (); and unspecific amplification of a gene involved iron metabolism (tatA; ).

Figure 3

Relative abundance of PGP strains in the five most dominant genera: (A) PGP activities tested by biochemical assays and (B) PGP activities detected by PCR-based approach.

Figure 4

UpSet plot of PGP traits detected by biochemical assays and by PCR. The bar chart on the left indicates the total number of isolates that exhibit each PGP trait. The upper bar chart indicates the intersection size between sets of isolates with one or more PGP traits. Dark connected dots indicate which PGP trait is considered for each intersection.

Figure 5

Effect of bacterial strains isolated from P. incarnata on the Cape gooseberry germination: (A) Representative pictures on sprouted seeds at 10 days and (B) Bar chart of germination percentage (%G) and vigor index (VI) exhibited by each bacterial strain. The vigor index (VI) is graphed with black and white colored rhombuses. Values represent the arithmetic mean ± SD. Treatments with different letters, within each tested parameter, are significantly different according to the Tukey statistical test (p < 0.05).

Figure 6

Effect of bacterial inoculants isolated from P. incarnata on the Cape gooseberry growth. Values represent the arithmetic mean ± SD. Treatments with different letters, within each tested parameter, are significantly different according to the Tukey statistical test (p < 0.05).