Use of plant growth promoting bacteria as an efficient biotechnological tool to enhance the biomass and secondary metabolites production of the industrial crop Pelargonium graveolens L'Hér. under semi-controlled conditions

https://doi.org/10.1016/j.indcrop.2020.112721Get rights and content

Highlights

  • Three PGP bacteria were tested as bioelicitors for Pelargonium graveolens L'Hér growth.

  • Positive effects on growth parameters and photosynthetic pigments were observed.

  • Inoculation with P. rhizophila S211 + O. iheyensis E9 gave the highest leaf biomass.

  • The consortium was the most efficient to enhance phenolic and flavonoid production.

  • The dual inoculations are highly efficient to improve essential oils accumulation.

Abstract

Pelargonium graveolens L'Hér. well known as rose-scented geranium is used worldwide in perfume, cosmetic, pharmaceutical and food industries. The essential oil and phenolic concentrations for this species were reported to be very low which requires their improvement. The effect of three plant growth promoting bacteria namely Pseudomonas rhizophila S211, Halomonas desertis G11 and Oceanobacillus iheyensis E9 on P. graveolens transplants were evaluated with single, dual and consortium modalities under semi controlled conditions. Significant positive effects on the growth related parameters, leaf and root biomass, photosynthetic pigments and secondary metabolites production were observed with inoculum-specific response. The highest root biomass was recorded with the dual inoculation [H. desertis G11+ O. iheyensis E9] while the inoculum [P. rhizophila S211 + O. iheyensis E9] showed the highest ability to enhance fresh leaf biomass (12.27 ± 0.61 g) leading to an improvement rate of 43.34 % over the control. A significant increase in the amounts of total phenolics and flavonoids contents in the inoculated plants over the control was revealed for all the treatments. The consortium was the most efficient to enhance the contents of total phenolics (105.62 ± 2.95 mg GAE/g DW) and flavonoids (48.55 ± 1.25 mg RE/g DW) resulting in an increase over the control of 67.35 % and 53.59 %, respectively. The essential oil synthesis was enhanced significantly for all the inoculated transplants compared with the control conditions. The dual inoculations resulted in the highest levels of essential oil contents in P. graveolens leaves, followed by the consortium than individual inoculants. The dual inoculation [H. desertis G11 + O. iheyensis E9] gave the highest essential oil amount (0.49 ± 0.03 %) with an increase of 68.96 % over the control (0.29 ± 0.01 %). The GC–MS analyses of the essential oils resulted in the identification of 23 compounds. In all treatments, citronellol (25.84–40.02 %), geraniol (9.67–11.96 %), citronellyl formate (5.35–10.14 %), 2-phenylethyl tiglate (4.58–7.13 %), epi-γ-eudesmol (4.38–5.87 %) and linalool (2.56–4.07 %) are the main compounds with qualitative and quantitative variation. Interestingly, the three dual inoculations gave the highest amounts of citronellol, citronellyl formate and citronellyl tiglate compounds. The highest concentration of the high-value citronellol (40.02 %) which defines the commercial value of P. graveolens was recorded with the inoculum [H. desertis G11+ O. iheyensis E9] resulting in an increase rate of 54.87 % over the control (25.84 %). This study suggests the efficiency of the studied inoculums as promising biofertilizers with significant potentials to enhance P. graveolens biomass and secondary metabolites production which present an alternative biotechnological tool to limit the excessive use of chemical fertilizers.

Introduction

Plant secondary metabolites are a useful array of natural products including essential oils, phenolic acids, flavonoids, alkaloids, tannins among others with increasing industrial interest as pharmaceuticals, bio-agrochemicals, flavouring agents and food additives (Gonçalves and Romano, 2018; Thakur et al., 2019). The accumulation levels of secondary metabolites in plants are generally very low, less than 1 % of dry weight with significant fluctuations depending on endogenous and exogenous factors (Riahi et al., 2013; Thakur et al., 2019). Thus, the industrial exploitation requires high plant biomass yields for the extraction of these high-value bioactive molecules (dos Santos Marques et al., 2018). Consequently, finding efficient practical strategies to increase the contents of high-value secondary metabolites and modulate their composition in addition to plant biomass improvement is required and poses a continuous challenge for researchers worldwide (Yang et al., 2018; Riahi et al., 2019a).

The plant biomass and secondary metabolites components present the main target traits for improvement and selection in aromatic and medicinal crops (Sarrou et al., 2017). The use of non conventional biotechnological tools based on genetic engineering methods or the abundant application of chemical fertilizers and pesticides to ameliorate the plant biomass yields and the production of bioactive molecules of interest is often confronted with ethical issues and oppositions by the consumers given their potential negative impact on human health and on environment (Hussain et al., 2012).

The excessive use of chemical fertilizers to improve the yields and quality of plant species is becoming a serious threat to natural ecosystems and present high negative environmental impact (Wang et al., 2018). The long time applications of chemical fertilizers deteriorate the pH, nutritional status, microbial community and increased heavy metals reactivity in soils (Ning et al., 2017; Lin et al., 2019). Furthermore, the chemical fertilization causes continuous degradation of water resources and have negative incidence on human and animal health (Farhadinejad et al., 2014; Prashar and Shah, 2016).

Innovative biotechnology approaches such as the application of beneficial associations between plants and bacteria called plant growth promoting bacteria were recently proposed as sustainable and environmentally respectful methods which avoids or minimize the use of synthetic fertilizers and pesticides and rely on bio-fertilization (Ibort et al., 2018; Tennakoon et al., 2019). These PGP bacteria have been shown able to increase nutrient uptake, promote plant growth and protection against soil-borne pathogens which present effective and promising alternative biotechnological tools (Pagnani et al., 2018). The use of plant growth promoting bacteria as bio-elicitors to improve crop yields is recognized as a clean and efficient strategy in both in vitro and in vivo plant systems through various mechanisms under normal as well as adverse conditions (Thakur et al., 2019). The application of this strategy to major agronomic species was widely elucidated (Ambreetha et al., 2018; Rozier et al., 2019). However, investigations interested in aromatic and medicinal plants and targeting the impact of these bacteria on the production of secondary metabolites is still limited and is recently in progress (Cappellari et al., 2019).

Pelargonium graveolens L'Hér. (Geraniaceae), well known as rose-scented geranium is a valuable industrial crop commonly cultivated worldwide for its high-value secondary metabolites used as ingredients in high grade perfumery, pharmaceutical and cosmetic industries with increasing demand in the international markets (Shawl et al., 2006). Previous scientific reports interested in rose-scented geranium essential oils and organic extracts showed antioxidant (Cavar and Maksimovic, 2012), anti-inflammatory (Ghanizadeh et al., 2015), antimicrobial (Ben Hsouna and Hamdi, 2012), hypoglycemic (Boukhris et al., 2012), DNA damage protective effects (Boukhris et al., 2015) and therapeutic potential in metabolic diseases (Neagu et al., 2018).

In Tunisia, rose-scented geranium is propagated mainly by stem cuttings and is growing under different bioclimatic stages with both rainfall and irrigated systems. In addition to its ornamental use, this species is used in Tunisian perfume industry and to produce flavouring flower-water used in traditional pastries, as antispasmodic agent and to treat hyperglycemia, headaches and angina (Ghrabi-Gammar et al., 2009; Boukhris et al., 2012, 2015).

Based on the previous considerations, the main aim of the present study is to evaluate the plant growth promoting potentials of three bacterial strains namely Pseudomonas rhizophila S211, Halomonas desertis G11 and Oceanobacillus iheyensis E9 in single, dual and consortium inoculation modalities on transplants of Pelargonium graveolens L'Hér. under semi-controlled conditions. The effects of the inoculations were assessed based on various quantitative growth parameters, the photosynthetic pigments contents and their abilities to enhance the production of industrially valuable essential oils and phenolic secondary metabolites.

Section snippets

Bacterial strains and plant material

Three bacterial strains (Pseudomonas rhizophila S211 (S1), Halomonas desertis G11 (S2) and Oceanobacillus iheyensis E9 (S3)) provided by the Laboratory of Biotechnology and Bio-Geo Resources Valorisation LR11ES31 at the Higher Institute of Biotechnology of Sidi Thabet in Tunisia were used in this investigation as bio-elicitors to improve the biomass and secondary metabolites production for the studied plant species. The three selected bacterial strains were isolated, identified and

Effect of bacterial inoculation on the plant growth parameters

After three months of cultivation under greenhouse conditions, the obtained results showed that the applied PGPBs have the capacity to enhance P. graveolens growth. The transplants subjected to single, dual or consortium inoculations exhibited a remarkable visual modification of their roots and aerial parts compared with the non inoculated plants (Fig. 1). Except for the inoculation [P. rhizophila S211 + O. iheyensis E9], the plant height values of the treated plants increased significantly as

Conclusion

Based on this investigation, the inoculation of P. graveolens transplants with three plant growth promoting bacteria with single and mixture modalities under semi-controlled conditions resulted in significant positive effects on the growth related parameters and enhanced the production of phenolic, flavonoid and essential oil metabolites over the control. Significant variations in plant responses depending both on the studied parameter and the used inoculum were observed. The inoculation with

CRediT authorship contribution statement

Leila Riahi: Investigation, Methodology, Formal analysis, Writing - original draft, Writing - review & editing. Hanene Cherif: Investigation, Methodology, Resources, Writing - review & editing. Sahar Miladi: Investigation, Writing - review & editing. Mohamed Neifar: Methodology, Resources, Writing - review & editing. Bilel Bejaoui: Resources, Writing - review & editing. Habib Chouchane: Resources, Writing - review & editing. Ahmed Slaheddine Masmoudi: Resources, Writing - review & editing.

Declaration of Competing Interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Acknowledgments

The authors acknowledge financial support from the Tunisian Ministry of Higher Education and Scientific Research in the ambit of the laboratory project LR11ES31 and the PhosAgro/UNESCO/IUPAC partnership in the ambit of the Green Chemistry for Life project “Development of biofertilizers for a sustainable agriculture in Tunisia”. The authors are also grateful to Dr. Wafa Hassen and Dr. Darine El-Hidri for supplying plant growth promoting bacterial strains.

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