Exploring the potential of endophytes and their metabolites for bio-control activity

Abstract

In the current scenario, extensive use of synthetic chemicals in agriculture is creating notable problems such as disease and pest resistance, residues, yield loss, and soil unproductiveness. These harmful chemicals are eventually reaching our food plate through bioaccumulation and biomagnification in a crop. As a result, beneficial microorganisms are regularly being explored as a safer option in the agriculture sector for their ability to produce valuable bioactive secondary metabolites, particularly for crop protection. Such natural (bio) products are harmless to plants, humans, and the environment. In our quest for the search of the sources of bioactive constituents from the microorganisms, endophytes are the front-runner. They mutually reside inside the plant providing support against phytopathogens by releasing an array of bioactive secondary metabolites building climate reliance of the host plant. The purpose of this review is to examine the biocontrol potential of endophytes against bacterial and fungal pathogens in sustainable agriculture. We also attempt to explain the structure and activity of the secondary metabolites produced by bacterial and fungal endophytes in conjunction with their biocontrol function. Additionally, we address potential future research directions for endophytes as biopesticides.

Introduction

Biotic stresses are the primary drivers of agricultural losses. The Food and Agriculture Organization (FAO) assesses that diseases and pests cause 20–40% of global crop production loss each year. An estimated $220 billion is lost annually to plant diseases and $70 billion to pest insects (FAO 2019). Farmers employ synthetic pesticides because they are inexpensive and convenient to apply, and they've had decades to become used to administer them. Nearly, 2 million tons of pesticides are used every year around the world, where China is the leading user country, followed by the USA and Argentina (De et al. 2014; Sharma et al. 2019). Nonetheless, in 2020, worldwide pesticide use has been assessed to get an increment up to 3.5 million tons (Sharma et al. 2019). What's heartbreaking is that the benefits of using pesticides would not outweigh the threats to the crop and the surrounding ecosystem. The influence of synthetic pesticides in agriculture has raised disagreement about their disproportionate use since it risks non-target species, soil and groundwater contamination, and bioaccumulation (Aktar et al. 2009; Khan 2016). Aside from that, pesticide use results in greenhouse gas emissions, a key contributor to climate change. De facto, this significantly influences the growth, distribution, and population density of agricultural insect pests, notably insect-herbivores. Thus, it has exacerbated attempts at pest management for these economically important pests.

Agriculture and beneficial microorganisms have had a long and fruitful relationship, with the latter playing an important part in crop-integrated pest control systems all over the world (Orr and Lahiri 2014). Endophytes (bacteria or fungi) that are antagonistic to insects and pests have positioned themselves as effective warriors capable of holistically protecting the plant (Berg and Hallmann 2006). The host plant and endophytic microorganism interaction are symbiotic and stable. Though there are negative interactions with plants that particular endophytes exhibit, we will focus on the symbionts that interact with plants in a mutual way. The plants show no evident signs of disease, despite being colonized by these endophytes. Remarkably, they use their treasure trove of metabolites in bolstering plant development and defense mechanisms to reclaim the trust of the plant (Hardoim et al. 2015; Malfanova et al. 2013). A recently discovered study on metabolomics indicates endophyte genes known to promote plant development via phytohormones such as auxins, gibberellic acid, and indole-3-acetic acid (Dubey et al. 2020). Additionally, they are believed to generate certain regulatory compounds that improve the availability of nutrients to plants, such as nitrogen (N), potassium (K), and phosphorus (P) (Hashem et al. 2017). Endosymbiont Sinorhizobium meliloti, isolated from Medicago Lupulina, demonstrated plant growth-promoting effects and increased nitrogen uptake (Kong et al. 2015). The inoculation of the endophyte Piriformospora indica has had favorable results on growth for a diverse array of plant species, including increased weight, blooming, and seed production (Fakhro et al. 2010). In addition to facilitating plant development, endophytes can potentially manage diseases, insects, and nematodes (Jaber and Ownley 2018; Hong and Park 2016). As an active ingredient of bio-pesticides, endophytes or their metabolites are the novel agents that carry out notable plant protection methods, thus, emerging as an alternative to chemicals (Nieuwesteeg 2015).

In the first section of the review, we offer a synopsis of the distribution of endophytes (bacteria and fungi) in a host plant and their unique mode of action during biotic stress prevention. We then sought to decipher the structure and functionality of their bioactive secondary metabolites, which have been effective in stymieing bacterial and fungal phytopathogens. Additionally, future research directions in the realm of endophyte-based pesticides are discussed.

Colonization and distribution of endophytes

A significant percentage of both wild and cultivated plants are host to endophytic microorganisms (Hardoim et al. 2015). The key point worth noting here is that plants may contain the amalgam of endophytes and not just a single species. Colonization occurs when fungal or bacterial inoculum enters plants either forcefully breaking the tissue or passively through wounds or natural openings such as stomata or hydathodes. Endophytes predominantly colonize the intercellular spaces that are rich in minerals, sugars, and non-carbohydrate metabolites (Bacon and Hinton 2007). Endophytes are classified into three main categories based on plant-colonizing strategies, i.e., obligate, facultative, and passive. Obligate ones are unable to propagate outside of plant tissue and are likely transmitted via seed rather than originating from the rhizosphere (Frank et al. 2017). Facultative endophytes are opportunistic. They are free-living in soil but colonize the plants when the condition demands. The third group, passive endophytes, does not necessarily colonize the plant throughout its life span; however, they occasionally colonize the plants (Hardoim et al. 2008).

However, it is also imperative to consider the question: Does an endophyte have a pass, which would allow it easy access to a host because obviously pathogens do not get the same welcome and are regularly eradicated? Plants put a lot of faith in their advanced defence mechanisms, which they use to protect themselves from diseases. To defend against infection, plants have developed the ability to distinguish harmful pathogens by their effectors and activate their microbe-associated molecular pattern-triggered immunity. Salicylic acid (SA) and Jasmonic acid (JA) are the crucial components of two distinct signalling pathways activated during pathogen attack. Surprisingly, the same rapid defense response is suppressed or manipulated when it comes to endosymbionts. There is considerable evidence that endophytes are thought to evolve their molecular patterns or type III secretion systems to limit plant immune responses and prevent antagonistic consequences. For example, Trdá et al. (2014) found that the grapevine flagellin sensing system distinguishes between Burkholderia phytofirmans and pathogens such as Pseudomonas aeruginosa and Xanthomonas campestris. This could be because endophytic bacteria's flagellin evolved to avoid detection by the plant's immune system. Consequently, instead of eliminating beneficial endophytes, the plant develops a mutualistic relationship with them (Xu et al. 2018; Liu et al. 2017; Vandenkoornhuyse et al. 2015; Pieterse et al. 2012).

Some endophytes propagate systemically through vascular tissues to different plant parts, such as endosperm (Johnston-Monje and Raizada 2011). A study discovered the proof of transfer of carbon cactus seed-borne rock-degrading endophytes, indicating that these endophytes play a key role in their host's establishment on rock surfaces. Endophytic bacteria were found in seedlings grown from disinfected seeds' root cortex and vascular system, as well as in surface-disinfected fruit (Frank et al. 2017). Due to this intrinsic advantage, seed priming with endophytes has emerged as a viable method for delivering these bioagents to protect plants from soil-borne diseases. There is also an implication of continuous movement of organisms all through the plant parts (Gaiero et al. 2013). In some species, the sowing of seed and consecutive germination allows the vertical transmission of endophytic microorganisms. Endophyte systemic transport across the host plant is often studied using green fluorescent protein (GFP) tagging as this technique work with live cells (Johnston-Monje and Raizada 2011). But studying the colonization of endophytes (and not the movement), FISH, GUS, and fluorogenic dye staining are reasonable alternatives (Aswani et al. 2020; Kandel et al. 2017; Vági et al. 2014). The environment inside the plants provides effective living space for endophytes, due to which they are potentially active to control the pathogens as they are contained and are not exposed to the immediate environment. As the plant grows, they multiply within the intercellular spaces, there by potentially colonizing the entire plant axis (Bacon and Hinton 2007). A barrier effect is created by the endophytes as they extensively colonize the plant tissue, where these endophytes compete with external pathogens for space and nutrition (Wahla and Shukla 2017).

Mechanism of protection of plants by endophytes from plant pathogens

Various endophytes have been isolated from crop plants which have been proven to protect the plant from pathogens (Fig. 1). The general mechanisms used by endophytic bacteria and fungi are described here. Direct and indirect mechanisms are among the forms of actions used by endophytes against pathogens. Direct mechanism includes the endophyte-pathogen interaction and is sensitive to species-specific antagonism (Fadiji and Babalola 2020). Endophytes can directly limit the pathogen damage by producing antimicrobial secondary metabolites, often termed as antibiotics (Kusari et al. 2012). However, stimulation of Induced Systemic Resistance (ISR) in plants is one of the possible indirect ways for the biological control of phytopathogens by endophytes (Malfanova et al. 2013).

Fig. 1.

Bioactivity mechanisms opted by endophytes to protect plants from biotic stress

A single plant contains many microbial species and association encourages the endophytes to release bioactive compounds as well as the host plant to prevent the proliferation of harmful microbes in the same plant (Kusari et al. 2012). Endophytes can produce siderophores to make the availability of iron for the plant and meanwhile making it unavailable for pathogens (Yadav 2018). The virulent phytopathogens Ralsotonia solanacearum and Xanthomonas oryzae pv. Oryzae, which causes bacterial wilt in groundnut and bacterial blight disease in rice, were suppressed by the solvent-extracted siderophore from endophytic Penicillium chrysogenum (Chowdappa et al. 2020). Some of the siderophores that endophytes are known to produce can confer biocontrol activities such as types of hydroxamates, phenolates and/or catecholates (Rajkumar et al. 2010). Competition is an important tool used by endophytes to avoid the colonization of host tissue by pathogens (Martinuz et al.2012). Phytophthora sp. Symptoms, when treated with mixtures of endophytes from leaves of cacao tree leaves through a foliar application, were successfully reduced, thus demonstrating competition as one mechanism of disease suppression in a plant (Fadiji and Babalola 2020). Many endophytic bacteria have been reported to inhibit the pathogenic fungal growth with the production of several antifungal metabolites such as Iturin (Elkahoui et al. 2011). Such metabolites tend to deform the hyphae, and eventually killing the pathogenic fungus (Massawe et al. 2018). Endophytes are also known to produce antagonistic volatile organic compounds (VOCs) (Elkahoui et al. 2015). VOCs spread over a wide range of distances with a high space effect. These VOCs belong to several chemical groups such as alcohols, terpenes, esters, ketones, hydrocarbons, nitrogen-containing heterocycles, sulfur derivatives, and carboxylic acids (Selim et al. 2017).

To colonize the surface of plants, they produce several enzymes that aid in the hydrolysis of the plant cell wall. These enzymes help to indirectly reduce phytopathogens and also support the degradation of the fungal cell wall (Gao et al. 2010). Endophytes that colonize the plant interior can induce distinct modifications of the cell wall, such as accumulation of papillae formation, callose and phenolic compounds (Wang et al. 2016). These depositions can lead to the construction of the structural barrier at the site where a potential attack can be possible by phytopathogen (Pitzschke 2018). It is possible to secrete more than 1300 compounds by these endophytes; cellulose, hemicelluloses, proteins and chitin are among them (Fadiji and Babalola 2020). Application of genetic manipulation to the genetic makeup of 1,3-glucanase present in a strain of Lysobacter enzymogenes lowered the biological control action towards the damping-off disease of sugar beet caused by Pythium spp. (Gao et al. 2010). Streptomyces’ lytic enzymes have a great effect on antagonizing the witches broom disease of cacao (Macagnan et al. 2008). The pectinase enzyme had also been reported to aid in reducing pathogenesis in plants (Fadiji and Babalola. 2020). Cell-wall degrading enzymes produced by bacterial endophytes are known to be engaged in the process of reducing defense pathways in plants as their cell wall contains many different proteins that are involved in plant defense and repair mechanism (Norman-Setterblad et al. 2000). Stimulation of such responses consistently results in declining the spread of pathogens inside the plants (Iniguez et al. 2005). According to Gao et al. (2010), the production of phytoalexins can be induced by endophytes, and this might be a critical reason for plants to fight phytopathogens.

Endophytes as biocontrol agents

While it is essential to understand the fundamental characteristics of endophytes linked to pest control, our opinion is that building a sustainable system of plant protection will be feasible only if they can be delivered in the field and performed. For this reason, we discuss a few noteworthy discoveries that would help us apprehend how productive endophytes are as bio-control agents and to what extent we may count on them as alternative pesticides.

The endophytic fungus Beauveria bassiana from grapevine activated the host plant’s defense genes that prevented Plasmopara viticola downy mildew infection (Rondot and Reineke 2019). During plant assays, Pseudomonas viridiflava, an endophytic bacterium strain, reduced X. campestris proliferation. Inoculation with this isolate also reduced Sclerotinia sclerotiorum necrotic lesions on canola leaves (Romero et al. 2019). Research showed that the tomato plant-derived bacteria, Bacillus velezensis, may decrease Verticillium wilt incidence in tomato plants by 70% when administered in open field circumstances because the bacterium produces compounds like lipopeptide (bacillomycin, surfactin, and fengycin) and volatile antifungal compounds (Dhouib et al. 2019). When B. tequilensis was applied to Fusarium-infected tomato plants, the disease severity and vascular browning were reduced by 76% and 83%, respectively, compared to the control (Aydi-Ben-Abdallah et al. 2016). A fungal root endophyte, Penicillium brevicompactum, completely suppressed germinated and ungerminated barley infections produced by Gaeumannomyces graminis (Murphy et al. 2015). Plant protecting Bacillus subtilis HC8, an isolate of Heracleum sosnowskyi (hogweed) promoted, and protected the growth of the tomato plant against tomato root rot disease (Malfanova et al. 2011). In the presence of Pseudomonas sp. B16E, the fungal pathogen Fusarium culmorum produced 97% fewer macroconidia, and the proportion of Fusarium head blight seed infection in wheat was decreased by around 90% (Mnasri et al. 2017). The endophytic P. indica decreased the severity of Verticillium dahliae disease by more than 30% on tomato plants (Fakhro et al. 2010). Additionally, it was discovered that P. indica affected the Pepino mosaic virus concentration in tomato shoots grown hydroponically (Fakhro et al. 2010). The bacteria Burkholderia cepacia (B3) and P. aeruginosa (P3), both isolated from symptomless oil palm root tissues, have been demonstrated to block Ganoderma boninense from spreading. Compared to control, the bacteria, both alone and in combination, inhibited the pathogen’s spread, with an epidemic rate of 0.10–0.24 units (Sapak et al. 2008). The intensity of disease symptoms was substantially decreased after 72 h of treatment with Bacillus amyloliquefaciens strain Ar10 (100 and 85.05% decrease in necrosis region and weight loss, respectively) (Azaiez et al. 2018). Table 1 outlines the endophytes employed as biocontrol agents to control phytopathogens in various agricultural plants. Various bioactive substances have been investigated to reduce a range of phytopathogens which are thoroughly reviewed in the upcoming section.

Table 1.

Bio-control activities of endophytes against target pathogens of field crops

S. No	Host plant	Endophytes	Target pathogens	Bio-control activity	References
1	Theoba cacao (Cocoa bean)	Pseudomonas aeruginosa and Chryseobacterium proteolyticum	Phytophthora palmivora	100% inhibition of Black pod rot disease on cocoa pods	Alsultan et al. (2019)
2	Vitis vinifera (Grapevine)	Beauveria bassiana	Plasmopara viticola	Significant reduction in Grapevine Downy mildew	Rondot and Reineke (2019)
3	Brassica napus (Rapeseed)	Pseudomonas viridiflava	Xanthomonas campestris; Sclerotinia sclerotiorum	Significant reduction in the area of Black rot infection in leaves of Canola at 24 and 36 h representing approximately 40 and 21%, respectively	Romero et al. (2019)
4	Solanum lycopersicum (Tomato)	Bacillus velezensis	Verticillium dahliae	Reduced significantly the Verticillium wilt incidence in tomato plants by 70.43 ± 7.08%	Dhouib et al. (2019)
5	Solanum elaeagnifolium (Silver leaf Nightshade)	Bacillus tequilensis	Fusarium oxysporum lycopersici	Significant decreases (77–83%) in Fusarium wilt severity and vascular browning extent (76%) in tomato	Aydi-Ben-Abdallah et al. (2016)
6	Hordeum murinum spp. murinum L. (Barley)	Penicillium brevicompactum	Gaeumannomyces graminis	100% suppression of seed-borne barley infections on both germinated and ungerminated barley seed	Murphy et al. (2015)
7	Saccharum officinarum (Sugarcane)	Epicoccum nigrum	Fusarium verticillioides, Colletotrichum paradoxa	Reduced radial growth of target fungal pathogens by more than 50%	Fávaro et al. (2012)
8	Jatropha curcas (Physic nut)	Colletotrichum truncatum	Sclerotinia sclerotiorum	70% antifungal activity	Kumar and Kaushik (2013)
9	Triticum aestivum (Wheat)	Bacillus subtilis SG6	Fusarium graminearum	Reduced DI of Fusarium head blight by 72.6%, and FHB index by 77.5%	Zhao et al. (2014)
10	Triticum durum L. cv. Karim (Wheat)	Pseudomonas sp. B16E	Fusarium culmorum	The strain B16E caused 100% and 97% reduction in macroconidia production of Fc2 and Fc3 fungal strains of F. Culmorum, respectively. In general, the percentage of seed infection of Fusarium head blight in wheat was reduced by about 90% and 70%	Mnasri et al. (2017)
11	Solanum lycopersicum (Tomato)	Piriformospora indica	Verticillium dahliae	Limitation of Verticillium wilt disease severity in tomato by more than 30%	Fakhro et al. (2010)
12	Elaeis guineensis (Oil palm root)	Burkholderia cepacia and Pseudomonas aeruginosa	Ganoderma boninens	Basal stem rot incidence was reduced by 76% in Oil palm seedlings pre-inoculated with P. aeruginosa (P3). B. cepacia (B3) reduced incidence by 42% and the mixture of P. aeruginosa and B. cepacia by 54%	Sapak et al. (2008)
13	Solanum tuberosum (Potato)	Bacillus amyloliquefaciens strain Ar10	Pectobacterium carotovorum	Reduced Bacterial soft rot of potato by 100 and 85.05% reduction of necrosis deep /area and weight loss, respectively	Azaiez et al. (2018)

Bioactive secondary metabolites from endophytes

It is hardly an exaggeration to assert that secondary metabolites are endophytes’ lead performers. Any activity ascribed to these microsymbionts is primarily due to their ability to synthesize various bioactive compounds, including alkaloids, steroids, terpenoids, peptides, polyketones, and flavonoids, quinols, and phenols (Singh et al. 2017). Hence, studies of these metabolites may provide us with more insight into detecting endophytes for commercial purposes (Astuti et al. 2017). A variety of products based on secondary metabolites of endophytic microbes are already available on the market or near completion (Lugtenberg et al. 2016). For instance, Epichlo endophytes have been successfully marketed in New Zealand, Australia, South America, and the United States in perennial ryegrass and tall fescue with various characteristics and agricultural advantages (Young et al. 2013). Similarly, Kusagamycin and Blasticidin-S isolated from Streptomyces kasugaensis and Streptomyces griseochromogenes respectively, are the recommended fungicides to control rice blast caused by Pyricularia oryzae (Saxena 2014).

Many endophytic microbes have been found to produce a wide range of structurally diverse antagonistic lipopeptides (Sharma et al. 2021; Fira et al. 2018). Lipopeptides have the potential to be drugs with their unique mechanism of action. Lipopeptides are gaining popularity due to their antibacterial, antiviral, anticancer, and immunosuppressive properties, as well as the fact that they have a unique mechanism that impacts membrane permeability, ultimately leading to cell disruption (Mondol et al. 2013; Tareq et al. 2014; Zhou et al. 2019). Antimicrobial compounds produced by endophytes are also environmentally beneficial, pathogen-toxic, and do not harm humans (Singh et al. 2017). Endophytic inoculation of plants promotes plant growth while also modulating the production of bioactive molecules with high medicinal potential, such as capsaicin, a bioactive compound, abundantly found in red and chilli peppers also produced by Alternaria alternata, an endophytic fungus isolated from Capsicum annum (Clark and Lee 2016).

However, it is critical to recognize that endophytes do not always choose which metabolites to produce. Occasionally, the host plant stimulates the endophyte’s metabolism, producing a specific class of compounds (Ludwig-Müller 2015; Heinig et al. 2013). Figure 2 illustrates several essential phytopathogen-controlling metabolites’ chemical structures, while the next section details how they operate. Table 2 summarizes the pesticidal activity of secondary metabolites isolated from endophytic microorganisms.

Fig. 2.

Structures of important and biologically active compounds isolated from various endophytes

Table 2.

Secondary metabolites of endophytic microorganisms and their bio-control activities against phytopathogens

S.No	Endophyte	Host plant	Plant pathogen/Pests	Secondary metabolites	References
(A)	Bacterial endophytes
1	Bacillus amyloliquefaciens	Bacopa monnieri	Rhizoctonia sp., Sclerotium sp., and Pythium sp.	Bacillomycin (1); Surfactin (2)	Jasim et al. (2016)
2	Bacillus amyloliquefaciens; Bacillus velezensis	Zea mays	Sclerotinia sclerotiorum	1,3- butadiene (3); N, N-dimethyldodecylamine (4); 2-undecanone (5); Benzothiazole (6)	Massawe et al. (2018)
3	Paenibacilluspolymyxa	Zea (teosinte)	Fusarium graminearum	Fusaricidin C (7) & D (8)	Mousa et al. (2015)
4	Micromonosporachalcea	Cucumis sativus	Pythium aphanidermatum	β‐1,6‐glucanases	El-Tarabily et al. (2009)
5	Bacillus subtilis	Sugarcane	Saccharicola bicolor	Surfactin (2)	Hazarika et al. (2019)
6	Bacillus halotolerans	Phoenix dactylifera	Fusarium oxysporum f. sp. albedinis	Inthomycin-A (9); 5-deoxybutirosamine (10); PlipastatinA1(11)	Slama et al. (2019)
7	Bacillus amyloliquefaciens	Solanum tuberosum	Pectobacteriumcarotovorum	Glycolipid (12)	Azaiez et al. (2018)
8	Bacillus subtilis	Oryza sativa	Xanthomonas oryzaepv. oryzae	Surfactin (2)	Kumar et al. (2020)
(B)	Fungal endophytes
9	Chaetomium globosum	Ginkgo biloba	Fusarium graminearum	1,2-benzenedicarboxaldehyde-3,4,5-trihydroxy-6-methyl (flavipin) (13)	Xiao et al. (2013)
10	Chaetomium globosum	Panax notoginseng	Phomaherbarum; Epicoccum nigrum	Chaetomugilin A (14) & D (15)	Li et al. (2016)
11	Cochliobolus sp.	Sapindus saponaria	Fusarium solani	Curvularin (16)	Ataides et al. (2018)
12	Xylaria spp.	Paulliniacupana	Colletotrichum gloeosporioides	Piliformic acid (17); Cytochalasin D (18)	Elias et al. (2018)
13	Acremonium sp.	Mentha piperita	Sclerotinia sclerotiorum, Botrytis cinerea, Fusarium oxysporum, and Rhizoctonia solani	1-heptacosanol (19); 1-nonadecane (20)	Chowdhary and Kaushik (2018)
14	Peziculasporulosa	Picearubens	Fungus (Not specified)	Crytosporiopsin (21)	McMullin et al. (2017)
15	Penicillium chrysogenum	Eleusine coracana	Fusarium graminearum	5-hydroxy benzofuranone (22); Dehydrocostus lactone (23); Iridoide glycoside (Harpagoside) (24)	Mousa et al. (2016)
16	Trichoderma brevicompactum	Allium sativum	Rhizoctonia solani	Trichodermin (25)	Shentu et al. (2014)
17	Trichoderma koningiopsis	Vinca sp.	Pyriculariaoryzae, Aspergillus fumigatus, and Botrytis cinera	Trichodermin (25)	Leylaie and Zafari (2018)
18	Chaetomium globosum	Withania somnifera	Sclerotinia sclerotiorum	Antibiotic Sch 210,971 (26)	Kumar et al. (2013)
19	Chloridium sp.	Azadirachta indica	Pseudomonas aeruginosa	Javanicin (27)	Kharwar et al. (2008)
20	Eupenicillium sp. LG41	Xanthium sibiricum	Acinetobacter sp.	(2S)-Butylitaconic acid; (2S)-hexylitaconic acid (28)	Li et al. (2014)
21	Trichoderma longibrachiatum	Dendrobium nobile	Bacillus mycoides	Dendrobine (29)	Sarsaiya et al. (2020)
22	Phomopsis prunorum	Hypericum ascyron	Pseudomonas syringaepv. lachrymans	Phomoterpene (30); Phomoisocoumarins C (31) & D (32)	Qu et al. (2020)
23	Trichoderma sp. EFI 671	Laurus sp.	Fusarium graminearum, Rhizoctonia solani, Sclerotinia sclerotiorum, and Botrytis cinerea	Eburicol (33); β-sitostenone (34); Ergosterol (35); Ergosterol peroxide (36)	Kaushik et al. (2020)

Bacteria may live as endophytes in a variety of plants, although their metabolic abilities are poorly studied. Latest advancements in in-situ metabolite analysis may open new avenues for identifying and defining bacterial metabolites without isolating them. In addition to genuine plant hormones, plant-associated bacteria often generate metabolites that mimic the impact of natural plant hormones as structural analogues (Brader et al. 2014). Direct in-situ metabolite analysis was performed to investigate antimicrobial lipopeptides and various other molecules (i.e., pyrrolnitrin, 2, 4-diacetylphloroglucinol, and phenazine-1-carboxylic acid) derived from B. subtilis and P. fluorescens strains (Raaijmakers and Mazzola 2012). Jasim et al. (2016) evaluated the capability of a bacterial endophyte B. amyloliquefaciens to produce bacillomycin (1) and surfactin (2) with antifungal activity. This strain has been reported to play a significant role in induced systemic resistance in Bacopa monnieri. GC–MS analysis of Bacillus sp. Endophytes isolated from maize seeds revealed the presence of volatile bioactive compounds viz., 1, 3- butadiene (3), N, N-dimethyl-dodecyl amine (4), 2-undecanone (5) and Benzothiazole (6) (Jasim et al. 2016). All four VOCs possessed strong antifungal properties against necrotrophic fungus, S. sclerotiorum (Massawe et al. 2018). Paenibacillus polymyxa controlled Gibberella Ear Rot in modern maize caused by Fusarium graminearum by releasing toxigenic fusaricidins (fusaricidin C (7) & fusaricidin D (8)) (Mousa et al. 2015). Cell-wall degrading β-1,3, β -1,4, and β-1,6-glucanases from endophytic actinomycetes were exploited against Pythium aphanidermatum, causing damping-off in cucumber across the world (El‐Tarabily et al. 2009). The antifungal potential of the endophyte B. subtilis was demonstrated against several fungal pathogens such as Saccharicola, Cochliobolus, Alternaria, and Fusarium sp. The bio-control activity of the endophyte was related to its potential to produce surfactin (2) (Hazarika et al. 2019). Metabolomic analysis of endophytic B. halotolerans revealed the presence of several bioactive metabolites that served as fungicides (Inthomycin-A (9), 5-deoxybutirosamine (10), and plipastatin A1 (11) (Slama et al. 2019). The endophytic strain Ar10 of B. amyloliquefaciens isolated from potatoes showed strong antagonistic activity against Pectobacterium carotovorum strain II16, the causal agent of soft rot disease in potatoes (Azaiez et al. 2018). The cell-free supernatant of this strain actively exhibited a broad spectrum of antibacterial activity against several human and plant pathogenic bacteria by producing antibacterial compounds of glycolipid (12) nature. This strain significantly decreased the severity of disease symptoms on potato tubers by using either the bacteria vegetative cells or the cell-free supernatant (Azaiez et al. 2018).

B. subtilis, a bacterial isolate of root and stem tissues of Oryza sativa, was found to exhibit strong antibacterial activity against X. oryzaepv. Oryzae, which is the causal organism of leaf blight disease in rice. Polymerase chain reaction-based study of genes revealed the presence of surfactin (2) producing genes in B. subtilis (Kumar et al. 2020). Panax ginseng plant has been said to inhabit Raoultella ornithinolytica.

In the case of fungal endophytes, advanced studies have been undertaken and many fungal metabolites having biocontrol potential have been reported. Flavipin (13) from Chaetomium globosum, an endophytic fungus isolated from Ginkgo biloba, has been found effective against F. graminearum (Xiao et al. 2013). In another study, C. globosum isolated from Panax notoginseng was reported to produce antifungal compounds chaetomugilin A (14), chaetomugilin D (15) against Phoma herbarum and Epicoccum nigrum (Li et al. 2016). Curvularin (16) was established to be a potential antifungal compound isolated from Cochliobus sp. Inhabiting the Sapindus saponaria plant. This compound has been shown to be effective against various pathogenic fungi such as Moniliophthora perniciosa, Didymella bryoniae, and Fusarium solani (Ataides et al. 2018). Piliformic acid (17) and cytochalasin D (18) isolated from two Xylaria spp. Showed fungistatic activity against Colletotrichum gloeosporioides (Elias et al. 2018). The fungal endophytes (Acremonium sp.) isolated from Mentha piperita exhibited antifungal activity against chickpea rot, causing pathogens possibly because of the release of 1-heptacosanol (19) and 1-nonadecene (20) from the endophytes (Chowdhary and Kaushik 2018). Biologically active cryptosporiopsin (21) isolated from Pezicula sporulosa is reportedly antifungal and antibacterial due to the presence of C-5 chlorine in the compound (McMullin et al. 2017). Bioassay-guided purification of the ethyl acetate extract of finger millet endophytes (Penicillium sp.) revealed the identification of anti-fusarium compounds, i.e., 5-hydroxy benzofuranone (22), dehydrocostus lactone (23), and an iridoide glycoside (24) (Mousa et al. 2016). Trichodermin (25) (4β-acetoxy-12, 13-epoxy-Δ⁹-trichothecene) isolated from the endophytic fungus, Trichoderma brevicompactum showed strong inhibitory action against several fungal phytopathogens like F. oxysporum, Colletotrichum lindemuthianum, C. ampelinum, Botrytis cinerea and Rhizoctonia solani (Shentu et al., 2014). 1H-Nuclear magnetic resonance and 13C-Nuclear magnetic resonance spectroscopy revealed the identity of the compound as Trichodermin (25) (Leylaieand Zafari 2018). Chaetomium globosum EF18, an endophytic fungus inhabiting Withania somnifera, was found antagonistic against S. sclerotiorum. Its ethyl acetate extract showed more than 80% growth inhibition of fungal mycelium. NMR and MASS analysis revealed the compound identity as Antibiotic Sch 210,971 (26). Compounds produced by fungal endophytes have also demonstrated antibacterial activity. Chloridium sp., an endophytic fungal isolate of Azadirachta indica, can produce an antibacterial naphthaquinone, Javanicin (27), under the liquid as well as a solid culture medium (Kharwar et al. 2008). This well-functional compound shows significant antibacterial activity with a 2 µl/ml concentration against P. aeruginosa (Kharwar et al. 2008). Similarly, (2S)-Butylitaconic acid and (2S)-hexylitaconic acid (28), isolated from an endophytic fungus Eupenicillium sp. LG41exhibited strong antagonism against Acinetobacter sp… This antagonistic strain is an inhabitant of the roots of Xanthium sibiricum, which validates the fact that root endophytes can be a potential source of plant defense system (Li et al. 2014). The endophytic fungus Trichoderma longibrachiatum MD33 isolated from Dendrobium nobile contained naturally active dendrobine (29) that showed antibacterial action against the bacterium Bacillus mycoides (Sarsaiya et al. 2020).

A combination of sesquiterpenoid enantiomers, phomoterpene (30), and new isocoumarins antibacterial compounds, phomoisocoumarins C (31) & D (32) were isolated from Phomopsis prunorum (Qu et al. 2020). This endophytic fungus, an inhabitant of Hypericum ascyron was shown to antagonize the plant pathogenic bacterial strain Pseudomonas syringae pv. Lachrymans (Qu et al. 2020). Recently, Kaushik et al. (2020) isolated bioactive metabolites triglyceride mixture, eburicol (33), β-sitostenone (34), ergosterol (35), and ergosterol peroxide (36) from the extract of fungal endophyte, Trichoderma sp. Interestingly, these metabolites displayed potential antifungal activity against various fungal pathogens (F. graminearum, R. solani, S. sclerotiorum, and B. cinerea). The pesticidal efficacy of secondary metabolites extracted from fungal endophytes is summarized in Table 2.

Conclusion and future prospects of endophyte-based pesticides

The pathogen resistance, contaminated ecosystems, and other environmental hazards caused by synthetic pesticides have led us to examine the sources of environmentally friendly and sustainable chemicals. Endophytes that colonize inside the higher plants are the reservoir of therapeutic and beneficial bioactive compounds. These microbes provide global advantages to farming communities. Thus, endophytes, which have characteristics for plant benefits, can be a promising biological agent for sustainable agriculture. Studies have suggested that the endophytic microbiome has plant protection and growth promotion mechanisms similar to that of rhizospheric analogues (Dubey et al. 2020). Studies have marked that the organisms and their habitats that are exposed to perpetual metabolic and environmental interactions should produce even more secondary metabolites.

Microbial endophytes are one of the poorly examined groups of microorganisms that have the ability and potential to introduce highly valuable bioactive novel compounds as green pesticides for sustainable agriculture and building climate resilience. Many of them do not have direct applications in these fields. Still, a thorough study may provide an extensive and detailed idea about the development of endophytic-based products and their management in the field. New biosynthetic pathways may be isolated and identified using sophisticated molecular biology methods. As a result, we will be able to discover novel bioactive chemicals in both the industrial and academic setting. In addition, it is possible that endophytes produce bioactive compounds through a number of biochemical processes. Future research should investigate how each one contributes. It will also be essential to demonstrate endophytes' functional and taxonomic diversity by examining their genes in greater depth via the omics approach.

Author contributions

AyS wrote the manuscript. TM finalized the tables. The review has been checked and revised by NK, AS and ND.

Funding

The financial support was provided by the Department of Science and Technology, Government of India under Project No. DST/INT/TUNISIA/P-04/2017 and the Tunisian Ministry of Higher Education and Scientific Research under the TOMendo Project to Dr. Nutan Kaushik and Dr. Naceur Djébali, respectively.

Declarations

Conflict of interest

The authors declare that there is no conflict of interest regarding the publication of this manuscript.

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Research involving human participants and/or animals

This article does not contain any studies with human participants or animals performed by any of the authors.