Microbial Solutions to Combat Cannabis sativa Fungal Pathogens

Abstract

Cannabis (Cannabis sativa L.) is one of the earliest cultivated crops and is valued for its medicinal compounds, food, fibre, and bioactive secondary metabolites. The rapid expansion of the cannabis industry has surpassed the development of production system knowledge. The scientific community currently focuses on optimising agronomic and environmental factors to enhance cannabis yield and quality. However, cultivators face significant challenges from severe pathogens, with limited effective control options. The principal diseases include root rot, wilt, bud rot, powdery mildew, cannabis stunt disease, and microorganisms that reduce post-harvest quality. Sustainable management strategies involve utilising clean planting stocks, modifying environmental conditions, implementing sanitation, applying fungal and bacterial biological control agents, and drawing on decades of research on other crops. Plant–microbe interactions can promote growth and regulate secondary metabolite production. This review examines the recent literature on pathogen management in indoor cannabis production using biocontrol agents. Specific morphological, biochemical, and agronomic characteristics hinder the implementation of biological control strategies for cannabis. Subsequent investigations should focus on elucidating the plant–microbe interactions essential for optimising the effectiveness of biological control methodologies in cannabis cultivation systems.

Introduction

Cannabis sativa L. (Hemp), an annual and dioecious plant belonging to the Cannabaceae family, encompasses the following two significant crop species cultivated globally: cannabis (marijuana), valued for its medicinal and psychotropic properties, and hemp, utilised for fibre and oil seed production [1,2,3,4]. Globally, hemp cultivation serves both agricultural and industrial purposes [5]. This versatile crop finds applications in numerous sectors, including the production of medicinal substances, fabric goods, renewable fuels, building supplies, paper products, and materials for thermal insulation [5,6]. Hemp and cannabis contain a broad range of other cannabinoids, terpenes, and phenolic compounds [7]. The plant is rich in substances of pharmaceutical interest, notably phytocannabinoids such as tetrahydrocannabinol (THC) and cannabidiol (CBD), which exhibit various psychotropic effects [2,5,7,8,9,10].

In comparison to other domesticated plant species, scientific knowledge regarding cannabis production remains limited, primarily due to the illicit status of marijuana, which has directed researchers’ focus towards distinguishing marijuana from hemp for law enforcement purposes. Over the past two decades, there has been a significant increase in scientific publications on cannabis, coinciding with legislative relaxation in several countries, attributable to its substantial economic value [8,11,12]. Medicinal and recreational marijuana are cultivated both outdoors and indoors, whereas hemp is predominantly grown outdoors [11,13]. The cultivation of cannabis and its cannabinoid yield are subject to various environmental and biological influences. Abiotic factors, including photoperiod, ambient temperature, soil pH levels, and nutrient availability, significantly impact plant development [14]. Additionally, biotic elements such as pathogenic microorganisms, insect infestations, and predator interactions play crucial roles in shaping cannabis growth and cannabinoid synthesis [15]. The expansion of hemp cultivation faces challenges including seed quality and pressure from weeds, insects, and diseases [7]. Both cropping systems require improvements in cultivation practices and fungal disease management [11].

The expansion of C. sativa cultivation has been linked to a rise in the occurrence and intensity of fungal infections, including new pathologies such as vascular wilt, root and crown rot, damping-off, powdery mildew, head blight, and bud rot [1,2,3,4,10]. These are caused by various fungi, including Fusarium, Botrytis, Diaporthe, Pythium, Sclerotinia, and Sclerotium [1,2,3,4,10]. These fungal diseases present considerable challenges to worldwide agriculture and food security [1,2,3,5,15]. Synthetic fungicides are the primary control method in most crops, targeting various cellular processes such as respiration, cytoskeleton assembly, osmoregulation, sterols, and amino acid biosynthesis [12]. Nevertheless, the application of fungicides has led to the development of resistant fungal strains due to metabolic detoxification, increased activity of efflux membrane transporters, and alterations in target sites [12]. Furthermore, fungicide application to cannabis is heavily regulated in numerous countries owing to health and environmental concerns [12]. Although chemical fungicides can effectively control these diseases, they also present risks including environmental pollution, fungal resistance, and reduced populations of beneficial soil microbes [2,11,16].

The endophytic/epiphytic microbiome influences the plant phenotype, determines its phytochemical profile, and mediates functional trait expression, irrespective of the colonisation mode [5,9,15,17,18]. These microbes are vital in the production of plant growth promoters (PGPs), which facilitate plant and root expansion whilst bolstering plant resistance mechanisms [2,9,11,15]. The defensive strategy known as antagonism manifests in two forms—direct and indirect. Direct antagonism employs protective substances such as antibiotics, antibacterial toxins (including peptides and proteinaceous compounds), bacteriocins, hydrolytic enzymes, volatile organic compounds (VOCs), siderophores, and cyclic lipopeptides. Indirect antagonism, on the other hand, activates the plant host’s defensive mechanisms, specifically induced systemic resistance (ISR) and systemic acquired resistance (SAR), to combat rival organisms [9,15]. Considering these attributes, biological control agents are regarded as a sustainable substitute for chemical interventions in cannabis production. Nevertheless, it is crucial to recognise that the capacity of these agents to suppress diseases through direct or indirect antagonistic interactions within the cannabis plant has not been specifically investigated [2,11,15].

Researchers have employed diverse experimental techniques to investigate and elucidate beneficial relationships between plants and soil bacteria [19]. These methodologies encompass various approaches, with some examining plant alterations, whereas others focus on analysing beneficial plant growth-promoting microbes’ physiological and biochemical aspects (PGPM). In this context, we have presented a synthesis of several contemporary studies that have explored the interactions between cannabis pathogens and PGPM to identify efficacious biocontrol agents.

Biocontrol by PGPM: A Mechanistic Point of View

Various biological (bacterial and fungal) strategies have been implemented to mitigate plant fungal diseases, thereby reducing dependence on synthetic agrochemicals for vegetable production [20,21]. PGPMs are advocated as an ecological and economical disease management approach and a sustainable biological alternative to agrochemicals thus conserving natural resources [22]. PGPMs are extensively utilised in sustainable agriculture to enhance biodiversity, improve crop yields, and restrict pathogen infection [20,21]. These free-living soil microorganisms effectively colonise plant roots and form endophytic bacterial populations, demonstrating their adaptability to specific ecological niches [20,21]. They are recognized for optimising plant development and performance through both direct and indirect processes [20,21,22].

Microorganisms produce a diverse array of low-molecular-weight compounds known as secondary metabolites, which are not crucial for survival under typical conditions. These substances encompass a wide range of compounds, including “antibiotics, toxins, and various peptides such as ribosomal peptides (RPs) and non-ribosomal peptides (NRPs). Additionally, they comprise chitinase, hydrogen cyanide (HCN), polyketides (PKs), enzymes that detoxify reactive oxygen species (ROS), antioxidant enzymes, and volatile organic compounds (VOCs)”. The list includes alkaloids, polyenes, phenazines, lipopeptides, amino sugars, macrolactone, aminoglycosides, indole-3-acetic acid (IAA), and iron-chelating siderophores [18,23,24,25]. These secondary metabolites exhibit remarkable structural diversity and serve various functions beyond the essential survival needs. These compounds compete for ecological niches or nutrients (carbon/energy sources) and secrete organic acids, phosphatases, and other enzymes to break down complex phosphorus compounds [20,21,26,27,28,29]. These mechanisms are widely recognized as direct biocontrol strategies (Figure 1).

Figure 1. The diagram illustrates both direct and indirect pathways of microbe-associated biocontrol for Cannabis sativa pathogens. In the indirect route, pattern recognition receptors (PRRs) detect microbe-, pathogen-, or damage-associated molecular patterns (MAMPs, PAMPs, DAMPs) from PGPR/PGPF/Pathogens, whilst R-proteins recognise intracellular effectors. This recognition triggers PAMP-triggered immunity (PTI) or effector-triggered immunity (ETI). These immune responses initiate calcium influx (Ca²⁺) via G-protein-coupled membrane receptors and Ca²⁺ channels, generate reactive oxygen species (ROS) and reactive nitrogen species (RNS), and produce salicylic acid (SA) to activate SAR, as well as jasmonic acid (JA) or ethylene (ETH) to activate ISR. Additionally, they stimulate mitogen-activated protein kinase (MAPK) cascades, calcium- and calmodulin-dependent protein kinase (CCaMK), and NPR1. These processes lead to the transcriptional regulation of defence genes, including pathogenesis-related (PR) proteins, lignin and callose deposition, suberin synthesis, and the production of secondary metabolites (terpenoids, oxylipins, flavonoids, phenolics, cannabinoids). In the direct mechanism, PGPR/PGPF function as bioinoculants, enhancing plant growth and modulating gene expression in secondary metabolite biosynthesis pathways. This includes the production of hormones, siderophores, AMP, VOC, PKS, NRPS, PTS, hybrid PKS–NRPS, PTS, enzymes for nutrient solubilisation, and ethylene dissociation enzymes (ACC deaminase). MAMP, microbe-associated molecular pattern; PAMP, pathogen-associated molecular pattern; DAMP, damage-associated molecular pattern; PGPR, plant growth-promoting rhizobacteria; PGPF, plant growth-promoting fungi; PKS, polyketide synthases; AMP, antimicrobial peptides; NRPS, non-ribosomal peptide synthetases; TPS, terpene synthases; PTS, prenyltransferases; NPR1, nonexpressor of pathogenesis-related genes 1; SAR, systematic acquired resistance; ISR, induced systemic resistance.

Immunity is triggered when PRRs or R-proteins detect microbe-, pathogen-, or damage-associated molecular patterns (MAMPs, PAMPs, DAMPs) or intracellular effectors, respectively, leading to PTI/ETI [30,31,32,33,34,35]. PRRs, which include receptor-like kinases (RLKs) and receptor-like proteins (RLPs), initiate the production of signalling molecules such as ROS, Ca²⁺, RNS (NO), G-protein, and SA. These molecules contribute to SAR in biotrophic pathogen–plant interactions. Alternatively, JA or ET are involved in ISR during beneficial plant–microbe interactions [30,31]. This process activates NPR1, modifies transcription factors (TFs) and MAPK cascades and regulates defence gene expression. The latter includes the production of antimicrobial compounds, PR, and secondary metabolites [26,31,32,36]. Additionally, it promotes callose deposition, suberin and lignin formation [20,21,31], programmed cell death (PCD), and hypersensitive response (HR), ultimately enhancing the plant’s resistance to pathogens [20,21,31].