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Integrated Management of Pathogens
and Microbes on
Cannabis sativa
L.
(Cannabis) under Greenhouse
Conditions
Liam Buirs and ZAMIR K. PUNJA *
Posted Date: 6 February 2024
doi: 10.20944/preprints202402.0361.v1
Keywords: biological control; bud rot; cultural control; fungal diseases; plant pathogens, root rots
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Review
Integrated Management of Pathogens and Microbes
on Cannabis sativa L. (Cannabis) under
Greenhouse Conditions
Liam Buirs 1 and Zamir K. Punja 2,*
1 Pure Sunfarms Corp., Delta, BC V4K 3N3
2 Department of Biological Sciences, Simon Fraser University, Burnaby, BC V5A 1S6, Canada
* Correspondence: punja@sfu.ca
Abstract: The increased cultivation of high THC-containing Cannabis sativa L. (cannabis), particularly in
greenhouses, has resulted in a greater incidence of diseases and molds that can negatively affect the growth
and quality of the crop. Among them, the most important diseases are root rots (Fusarium and Pythium spp.),
Botrytis bud rot (Botrytis cinerea), powdery mildew (Golovinomyces ambrosiae), cannabis stunt disease (caused
by Hop latent viroid), and a range of microbes that reduce post-harvest quality. An integrated management
approach to reduce the impact of these diseases/microbes requires combining different approaches that target
the reproduction, spread and survival of the associated pathogens, many of which can occur on the same plant
simultaneously. These approaches will be discussed in the context of developing an integrated plan to manage
the important pathogens of greenhouse-grown cannabis at different stages of plant development. These stages
include maintenance of stock plants, propagation through cuttings, vegetative growth of plants, and flowering.
The cultivation of cannabis genotypes with tolerance or resistance to various pathogens is a very important
approach, followed by the maintenance of pathogen-free stock plants. When combined with cultural
approaches (sanitation, management of irrigation, and monitoring for diseases) and environmental approaches
(greenhouse climate modification), a significant reduction in pathogen development and spread can be
achieved. The use of preventive applications of microbial biological control agents and reduced risk biorational
products can also reduce disease development at all stages of production in jurisdictions where they are
registered for use. The combined use of promising strategies for integrated disease management on cannabis
plants during greenhouse production will be reviewed. Future areas for research are identified.
Keywords: biological control; bud rot; cultural control; fungal diseases; plant pathogens; root rots
1. Introduction
Integrated disease management (IDM) incorporates the coordinated use of multiple approaches
to reduce the impact of disease-causing agents (pathogens) on agricultural crops [1]. When applied
in parallel or consecutively, these tactics can achieve control of multiple pathogens using different
and sometimes synergistic suppression tactics. IDM builds upon the concept of Integrated Pest
Management (IPM), which has been widely utilized for decades to target and manage insect pests on
agricultural crops, and requires different strategies to be employed in a coordinated manner, often
with resounding success [2,3]. When IDM approaches are considered for cannabis (Cannabis sativa L.,
high THC-containing genotypes) grown under greenhouse conditions, several aspects need to be
modified from traditional IDM programs. First and foremost is the fact that there are no synthetic
fungicides available for use on cannabis crops, thus eliminating a widely-used disease management
strategy. Instead, only reduced risk “biological” and “biorational” products are permitted. These
products are mostly protective in action i.e. non-fungicidal, so they are best suited for preventative
applications, although some products can also be deployed as sanitizers. While claims of product
efficacy and applications for disease reduction on cannabis are often made, not all are supported by
data from replicated research trials or third-party evaluations. This adds to the difficulty in
identifying the specific IDM approaches that are best suited for each pathogen. The recent expansion
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of hemp cultivation (C. sativa, low THC-containing cultivars) in the USA following federal
government approval should provide useful information on disease and pest management
approaches which could be extended to cannabis [4]. The lack of synthetic fungicides for cannabis
production has prompted the registration of several biological control products which can be used at
different stages of production [5,6]. However, efficacy data for these products are often not available,
and the modes of action of the biocontrol agents are not often fully understood, in the context of
cannabis IDM, highlighting the need for further research in this area [6,7]. Fortunately, efficacy data
may exist for many of these products on other crops e.g., for organic production, and can likely be
extrapolated to cannabis crops [8]. A second challenge for IDM development in cannabis is that
highly-bred cultivars containing specific resistance genes against important pathogens are lacking.
Instead, genetic selections (genotypes) that target higher yields of inflorescences and THC content,
and which display unique morphological traits, have been made a priority [9]. In most instances,
these efforts have excluded the specific incorporation of disease resistance traits. Consequently, some
high-yielding genotypes frequently show high susceptibility to various pathogens, as will be
illustrated in this review. Fortunately, the broad genetic variation that currently exists among
cannabis genotypes has led to the identification of resistance in various genotypes to specific
pathogens, such as powdery mildew [6,10-12]. The mechanisms underlying this resistance are
currently under investigation [13].
A third challenge is that when cannabis is compared to other widely-grown greenhouse crops,
such as tomatoes, cucumbers, and peppers, the optimal cultural and environmental conditions for
cultivation have not yet been fully established. Since different cannabis greenhouse operations can
experience variable growing conditions, standardized research trials are needed to establish these
parameters. Recent research has identified integral aspects of controlled environment cultivation
practices that can be used as a baseline reference [14,15]. The prevalent pathogens affecting cannabis
crops in greenhouses have been recently characterized and described [7], providing diagnostic
information that is required for IDM implementation. Accurate diagnosis of the pathogen(s) involved
in a disease syndrome is an important component of IDM and several diagnostic methods have been
described [4,7,16-19]. In this review article, we describe the most important pathogens of cannabis
crops cultivated under greenhouse conditions and highlight the various growth stages at which IDM
approaches can be implemented during the crop production cycle, which generally occurs over 12-
15 weeks (Figure 1).
Figure 1. The different stages of cannabis production under greenhouse conditions. Each crop
cultivation cycle from propagation to harvest spans ~12-15 weeks. This is followed by a final stage of
post-harvest processing that includes drying, trimming, curing and storage.
The first stage of production of a cannabis crop is stock (mother) plant cultivation (Figure 2a),
which provides a source of vegetative cuttings (Figure 2b). Once rooted, these are transferred to
greenhouse growing conditions for 2-3 weeks (Figure 2c). The developing vegetative plants are then
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transferred to flowering rooms for 8 weeks (Figure 2 d, e), after which time the inflorescences are
harvested (Figure 2f).
Figure 2. The stages of cannabis crop development. (a) Stock plants. (b) Rooting of cuttings. (c)
Vegetative plants. (d, e) Flowering plants. (f) Harvested inflorescences.
During each crop production year, up to 4-5 cropping cycles may take place per greenhouse
compartment. The IDM approaches that can be developed include selection of disease-tolerant
genotypes, implementation of cultural practices, modification of environmental climate settings, and
application of reduced risk products (Figure 3).
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Figure 3. Integrated disease management strategies (left, in brown) are developed according to the
crop development stage (top). The hexagons (in green) illustrate the specific diseases being targeted,
which are discussed in more detail below. HLVd = Hop latent viroid, PM = powdery mildew, Botrytis
= bud rot.
We also discuss aspects of the microbial colonization of cannabis inflorescences by yeasts and
molds and propose IDM strategies to reduce the total microflora present. Monitoring of microbial
colonization of inflorescences is an important quality aspect for cannabis which is under strict
regulatory control and presents a unique and challenging component of crop management that is not
found in most other crops [19,20] This review should aid in the design or refinement of further IDM
programs in greenhouse-cultivated cannabis operations. Detailed descriptions of the symptoms
caused by various pathogens at different stages of cannabis growth during commercial production
and the approaches that can be taken to manage them are described below.
2. Cannabis Pathogens: Symptoms and Management Approaches at Different Stages of Growth
2.1. Stock Cultivation Stage
Stock (mother) plants provide a source of vegetative cuttings which are commonly used in
commercial cannabis production. These plants generally constitute a range of genotypes that are
chosen for their desired phenotypic characteristics and biochemical profiles. They are grown in
designated areas within the greenhouse or in separate indoor rooms. Physical separation of stock
plants from larger-scale commercial production is important to prevent the spread of pathogens. The
ages of these stock plants can vary, and typically range from 3 to 12 months, depending on the facility.
In the context of disease development, older plants often exhibit signs of declining growth, such as
reduced shoot growth, leaf yellowing, and poor root development (Figure 4a). These symptoms may
be indicative of sub-lethal infections by Fusarium and Pythium spp. or Hop latent viroid (Figures 4
and 5).
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Figure 4. Symptoms of infection by a range of pathogens commonly observed on cannabis stock
plants. (a) Declining growth with reduced vigour. (b, c) Internal stem discoloration due to F.
oxysporum infection. (d) Isolation of colonies of F. oxysporum from diseased tissues. (e) Browning of
roots due to Pythium infection. (f) Isolation of Pythium colonies from diseased roots. (g) Powdery
mildew infection on leaves. (h, i,) Infection by Hop latent viroid may cause reduced vigor and curling
of young leaves.
A closer inspection of the stems of diseased plants will often reveal internal discoloration in the
pith and xylem tissues (Figure 4 b, c), a symptom of Fusarium infection, and/or root browning that
can be caused by Fusarium or Pythium species [21-24]. Excessive waterlogging may also cause root
browning on cannabis plants. Accurate pathogen diagnosis at this stage is critical to determine the
most effective IDM strategies to implement. Stock plants are also susceptible to powdery mildew,
visible as white colonies on the upper surfaces of leaves (Figure 4 g). A significant challenge in
maintaining healthy stock plants is the recent emergence of hop latent viroid (HLVd) [25-27], which
is mostly asymptomatic on stock plants but may cause occasional curling or mottling on the youngest
leaves (Figure 4 h,i). The impact of HLVd infection on stock plants is seen when rooting frequency
and vigor of cuttings derived from them are examined (Figure 5). HLVd infection leads to poor root
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growth (Figure 5 b) that continues to impact plant growth at the vegetative stage (Figure 5 c, d) and
can also impact flowering (Figure 5 e). HLVd-infected flowering plants derived from infected stock
plants display reduced inflorescence growth as well as lower levels of cannabinoid production [26].
This emphasizes the importance of maintaining pathogen-free stock plants during commercial
production. Routine scouting for the presence of disease symptoms and testing of stock plants for the
presence of HLVd, Fusarium and Pythium species is recommended.
Figure 5. Symptoms of Hop latent viroid infection during propagation, vegetative growth and
flowering stages of the cannabis crop cycle. (a) Infected stock plants may show unthrifty growth and
smaller leaves. (b) Comparison of root development on cuttings derived from an HLVd-infected stock
plant (left) and a healthy plant (right). (c) Vegetative plants may show curling and distortion of the
youngest leaves. (d) Lateral branching may be seen on HLVd-infected vegetative plants. (e) Stunted
growth of HLVd-infected flowering plant (left) compared to a healthy plant (right). (f, g) HLVd-
infected inflorescence with yellowing compared to a healthy one, respectively. (h, i, j) Reduced
inflorescence development in three different genotypes of cannabis resulting from HLVd infection. In
all photos, the infected plant is shown on the left. (k) Dried inflorescences from an HLVd-infected
plant (left) compared to a healthy plant (right).
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2.2. IDM Approaches at Stock Cultivation Stage
During the stock plant cultivation stage, various IDM strategies can be implemented to minimise
the development of plant pathogens. The following are examples of some commonly used practices.
2.2.1. Biosecurity and Quarantine Inspection
Biosecurity practices which include foot baths, wearing protective clothing, and removing
pruned leaves and diseased plants are standard in most horticultural greenhouse operations [28];
these practices should be implemented for cannabis growing operations. In addition, it is important
to establish a quarantine protocol in cases where plant materials, such as unrooted cuttings or whole
plants, are brought in from an external source [2]. Such precautionary measures can prevent pathogen
introduction and are standard biosecurity protocols in commercial crop production [29]. When
applied to cannabis, this necessitates an isolation period for 3-4 weeks during which plants are
monitored for disease symptoms and tested for the presence of potential viruses and other pathogens
[6,7]. After the plants are confirmed to be free of detectable pathogens, they can be used for
commercial propagation.
2.2.2. Cultural and Environmental Management
Environmental management is a component of IDM across all stages of cannabis growth since
climatic conditions can influence both plant growth as well as pathogen growth. Standard cannabis
cultivation environmental setpoints, which are established for baseline pathogen management
during low disease pressure periods, have been described [15,30]. Conditions that are unfavorable
for disease development while at the same time supporting optimal plant growth are required. This
often necessitates maintaining lower humidity levels by enhancing venting, heating, and air
circulation. Seasonal adjustments may also need to be made, as warmer temperatures with higher
humidity in the summer months may increase the incidence of root-infecting pathogens, such as
Fusarium and Pythium species. Similarly, cooler and more humid conditions during winter seasons
may enhance the development of powdery mildew infections.
2.2.3. Testing for Pathogen Presence and Eradication
Early detection of disease symptoms on stock plants is important to prevent pathogens from
spreading within the growing environment. There are several diagnostic approaches that have been
described to detect cannabis pathogens and a number of commercial laboratories provide testing
services for a range of pathogens [4,7,19,26]. The practice of culling and replenishing stock plants is
a standard component of IDM programs when diseased plants are detected [31]. Stock plants should
be replaced after several months of production with new, pathogen-free plants, which is key to
maintaining a healthy and vigorous stock plant population. Plants infected with Fusarium, Pythium
or HLVd should be promptly removed from a facility. Eradication of diseased plants, particularly
those infected with HLVd, is essential. When regular (weekly) pathogen testing is followed by
destruction of those plants infected by HLVd, a gradual decline in the occurrence of diseased stock
plants can be achieved (Figure 6). After many rounds of testing performed over a 6-month period,
this strategy was shown to reduce HLVd frequency on stock plants from 22% to 1% (Figure 6). Peaks
of infection can still be seen which are attributed to the re-introduction of diseased plant material that
went undetected initially and were inadvertently used as a source of cuttings. Removing them upon
detection resulted in the continued downward trend of infection.
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Figure 6. The impact of eradication of HLVd-infected stock plants on the frequency of positively
infected plants over a 6-month duration. The blue line shows the actual incidence of infected plants,
which fluctuates over time. The solid green line is the general trend that shows a decline in numbers
of infected plants.
2.2.4. Sanitary Practices
Thorough sanitation of the growing environment before planting a new crop is important to
reduce residual pathogen inoculum, which can be spread by water, air, on tools, and potentially on
clothing, gloves, or shoes. This is a common practice used on most greenhouse crops, especially
where viruses are of concern [6,28]. To reduce pathogen transmission, all surfaces and equipment, as
well as gutters, tables, floors, drip emitters, and pots should be cleaned by using reduced risk sanitary
products. These products include hydrogen peroxide with peracetic acid (Sanidate
®
or Zerotol
®
),
dodecyl dimethyl ammonium chloride (Chemprocide
®
or KleenGrow
®
), isopropyl alcohol, and
bleach [6]. The efficacy of these products in inhibiting pathogen growth can vary, depending on the
pathogen, product and concentrations used. A comparison of two products used at four
concentrations against growth of two pathogens is shown in Figure 7. At increasing concentrations,
both Zerotol
®
and hypochlorous acid reduced pathogen growth but Pythium showed a greater
sensitvity compared to Fusarium (Figure 7c). These products can potentially also negatively affect the
growth of beneficial Trichoderma species when applied as biocontrol agents (Figure 7d). Therefore,
care must be taken to consider the potential impact of applying reduced risk products in conjunction
with biocontrol products. These types of evaluations are important to conduct for any reduced risk
product targeted for the cannabis market to demonstrate efficacy and possible non-target effects.
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Figure 7. The effect of reduced risk products on pathogen growth can be evaluated under laboratory
conditions by testing a range of concentrations in liquid culture medium. (a) Growth in potato
dextrose broth containing a range of concentrations of individual products is measured by obtaining
mycelium dry weights after a 7-day exposure. (b) The effect of Zerotol
®
and hypochlorous acid on
growth of two pathogens at increasing concentrations is shown. Both Fusarium and Pythium are
reduced at higher concentrations but growth of Pythium shows greater sensitivity compared to
Fusarium. (c) Growth of Trichoderma can also be reduced by the presence of specific compounds.
2.2.5. Utilizing Disease Tolerant Genotypes
The utility of disease-tolerant genotypes that may have been developed through selective
breeding and genotype screening is an important aspect of IDM for stock plants. Disease-tolerant
genotypes of cannabis to a number of pathogens have been identified, including to Fusarium
oxysporum [23], powdery mildew [10,11,12,32], Alternaria leaf blight [33] and Botrytis bud rot (34,35)
(Figure 8). Recent research suggests that specific defense genes may play a role in certain host-
pathogen interactions, leading to a resistant phenotype [11,12,13,36]. The impact of cannabis
genotype on disease development at the flowering stage will be discussed later in this review.
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Figure 8. Examples of cannabis genotypes that exhibit a level of disease tolerance to different
pathogens. (a) Fusarium damping-off, with susceptible genotype on the left and tolerant genotype on
the right. (b) Powdery mildew, with susceptible genotype on the left and tolerant one on the right.
(c,d) Alternaria leaf blight, with tolerant genotype on the left and susceptible one on the right. (d)
Botrytis bud rot, with tolerant genotype on the left and susceptible one on the right.
.
2.3. Propagation Stage
Cannabis plants can be initiated from seeds or from vegetative cuttings which originate from
stock plants, but the latter is more commonly used in commercial production, since large-scale
propagation from seeds is less common. Routine testing for pathogens that may be present on seeds
is not currently a standard practice in the cannabis industry, which can result in the spread of seed-
borne pathogens. Cannabis and hemp seeds are known to harbor species of Alternaria, Fusarium, and
several post-harvest molds [33,37], as well as HLVd [26,27]. Testing for pathogen presence and
sanitation are important IDM approaches during plant propagation in greenhouse crops, including
cannabis [38,39]. These steps can minimize the subsequent spread of fungal, bacterial, and
viral/viroid pathogens. Vegetative cuttings used for propagation are required to be rooted under
high-humidity conditions over a two-week period. This environment is conducive for the spread of
pathogens such as Fusarium spp. and Botrytis cinerea (Figure 9), as well as a number of bacteria that
may be spread by water or in the air. Testing conducted in the rooting environment by swabbing of
surfaces, sampling of water and air, or plating of surface sterilised plant material can be used to assess
total microbes that may be present [21-23]. Cuttings may unknowingly harbor inoculum of Fusarium
spp., and infection by powdery mildew or HLVd are likely to be present if the original stock plants
were infected [6,7,23,26]. Cuttings taken from stock plants infected internally by Fusarium species can
result in spread of the pathogen, resulting in damping-off symptoms, particularly in susceptible
genotypes (Figure 9). The infection causes the pith and xylem tissues to collapse, resulting in death
of the cuttings. Powdery mildew symptoms may also appear on cuttings, from inoculum either
carried over from the stock plants or introduced at the propagation stage. The most significant
pathogen affecting root development and growth of cuttings is HLVd that originates from infected
stock plants [26]. Additionally, under high humidity conditions, vegetative cuttings may be affected
by gray mold (B. cinerea) and common saprophytic fungi, including Penicillium spp., which can
potentially reduce the appearance and quality of the cuttings [7,23]. Many of these fungal pathogens
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that affect cannabis cuttings as well as other stages of plant development produce large numbers of
spores, which can be spread by water, air and on tools throughout the growing facility (Figure 10).
These spores can serve as sources of initial inoculum and can be challenging to manage. The inclusion
of HEPA filters and HVAC systems is advisable to reduce the total counts of air-borne fungal
propagules.
Figure 9. Propagation of cannabis from vegetative cuttings and development of Fusarium damping-
off. (a) A tray of healthy cuttings. (b) A tray of cuttings infected with Fusarium oxysporum. (c, d, e)
Close-up views of damped-off cuttings. (f) A cross-sectional view of the stem a healthy cutting (left)
compared to a diseased one (right) in which tissue browning can be seen. (g) A scanning electron
microscopic view of a section through the stem of a healthy cutting. The central pith can be seen. (h)
A collapsed stem of a diseased cutting viewed through the scanning electron microscope. The central
pith has collapsed as well as surrounding cells.
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Figure 10. Spores of a range of pathogens that can affect cannabis plants at various stages of crop
growth. (a) Fusarium oxysporum micro-conidia. (b) Botrytis cinerea spores. (c) Large cluster of spores of
Aspergillus sp. (d, e) Chains of spores of Penicillium sp. (f) Powdery mildew spores. .
2.4. Propagation Stage IDM Approaches
2.4.1. Cultural and Environmental Management
During the propagation stage, ensuring that cuttings are obtained from healthy stock plants
reduces the probability of pathogens being transferred through these cuttings. In particular, the
incidence of F. oxysporum is reported to be greater in cuttings taken from the base of the plant
compared to locations higher up the plant [7,23]. Therefore, cuttings from the uppermost part of stock
plants may limit transmission of this pathogen and possibly of HLVd, although sufficient data is
lacking at the present time for this pathogen. Ensuring that cuttings are acclimatized in a transitional
environment prior to resuming vegetative growth reduces stress on the rooted plants [38,39].
2.4.2. Application of Biological Control Agents
Several biological control products containing Trichoderma spp. or Gliocladium cantenulatum are
registered for use on cannabis in Canada [5]. These products are classified as “reduced risk” and
provide an alternative disease management option in the absence of registered synthetic fungicides.
They can be used at all stages of cannabis crop growth but are particularly useful for managing
damping-off caused by Fusarium spp. on cuttings. When applied at the vegetative stage of plant
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growth, they can reduce mortality due to Fusarium and Pythium species [40]. Several weeks after
application, the biocontrol agents can be recovered from cannabis tissues, indicating they are able to
survive for a period of time (Figure 11). Their effectiveness is based on protection of tissues and
therefore they should be applied before pathogen infection occurs, ideally as a drench or as a dip
when cuttings are being rooted, or as a drench at later stages of crop growth. The biocontrol agents
protect susceptible root tissues from infection by these pathogens and can colonize cuttings
internally, possibly functioning as endophytes; they can potentially enhance root and shoot growth
in addition to providing protection against pathogens [40]. Trichoderma sp. also exhibit direct
antagonism to F. oxysporum in dual culture (Figure 12). Therefore, several registered biocontrol agents
have been demonstrated to be effective against root-infecting pathogens when applied preventatively
to cannabis plants.
Figure 11. Application of biological control agents provides protection to cannabis cuttings against
Fusarium damping-off. (a) Rootshield-treated cuttings (left) show greater survival compared to
pathogen-only (right). (b) Growth of Trichoderma harzianum from Rootshield-treated cuttings. (c)
Asperello-treated cuttings (right) show greater survival compared to pathogen-only (left). (d) Growth
of Trichoderma asperellum from Asperello-treated cuttings. (e) Prestop-treated cuttings (left) show
greater survival compared to pathogen-only. (f) Growth of Gliocladium catenulatum from Prestop-
treated cuttings.
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Figure 12. Growth of T. asperellum (top) is observed to stop the growth of Fusarium oxysporum (bottom)
when both are placed on a Petri dish. After a few days, the biocontrol agent continues to grow over
and inhibit further growth of the pathogen..
2.5. Vegetative Growth Stage
Following the establishment of rooted plants from cuttings, the plants are allowed to continue
vegetative growth for an additional 2-3 weeks before being transferred to flowering rooms. During
this growth stage, root-infecting pathogens, including Fusarium and Pythium species, as well as
HLVd, may continue to develop and spread. Development of powdery mildew may also become
more severe at this stage of production. Internal stem infections by Fusarium spp. in rooted cuttings
can significantly reduce the growth and development of vegetative plants. Symptoms such as
yellowing, stunted growth, browning of roots, and plant death are often linked to infection by
Fusarium and Pythium species (Figure 13). The development of these pathogens can be exacerbated
by root damage and excessive watering or flooding, which can also spread pathogen inoculum.
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Figure 13. Pythium and Fusarium infection on vegetative plants of cannabis. (a) Symptoms of
yellowing of the foliage are indicative of root infection by these pathogens. (b) Death of rooted
cuttings due to Fusarium infection. (c) Root development on healthy plant (left) compared to one
infected by Fusarium (right). (d) Internal stem discoloration is indicative of infection by Fusarium. (e,
f) Infection by Pythium can cause significant stunting of plant growth and death (right) compared to
healthy plants (left).
In addition, HLVd infection of rooted cuttings can adversely affect root development and plant
growth at the vegetative stage, leading to reduced plant size, particularly in susceptible genotypes
(Figure 5). Molecular diagnostic methods should be used to ensure that vegetative plants are not
infected by this viroid [4,26]. In greenhouse environments where recycling of nutrient solution is
practiced, monitoring for the presence of Fusarium and Pythium inoculum is necessary since both are
known to be present in hydroponic nutrient solutions [21]. Regular testing of electrical conductivity
(EC) and potential hydrogen (pH), coupled with testing of drip and drain nutrient ratios, will ensure
that the nutrient profiles remain within the optimal range for crop development, preventing nutrient
deficiencies that could lead to predisposition to pathogen infection [41,42]. In addition, monitoring
of water temperature and oxygen levels can reduce extremes which can enhance root infection by
pathogens [42]. Treatment of recirculated water with reduced risk products, such as those indicated
in Section 2.2.4, can reduce the incidence of root pathogens (Figure 7).
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2.6. Vegetative Growth Stage IDM Approaches
2.6.1. Cultural and Environmental Management
Root pathogen development on vegetative plants can be minimized by increasing the interval
between watering events, leading to fewer and shorter irrigation events as long as adequate moisture
is provided for optimal root development. This strategy has been used to reduce root pathogen
development on various crops [2]. On the foliage, exposure of plants to ultraviolet radiation,
especially UV-C light, can suppress powdery mildew development [43]. Night-time exposure
enhances pathogen susceptibility by limiting light-activated DNA repair mechanisms [44]. UV-C may
also enhance plant defense responses including accumulation of reactive oxygen species [45]. To
avoid phytotoxicity, exposure of plants to UV-C should be made gradually over several weeks,
according to manufacturer’s guidelines. Treated plants may show a reduction in plant height and
increased lateral branching as has been observed in some ornamental plant species [46, 47].
2.6.2. Application of Biological Control Agents
Biological control agents can also be applied as drenches to vegetative plants to reduce the
severity of root pathogens similar to treatments made at the propagation stage [40]. The extent to
which these agents can survive following application at this stage has not been determined.
2.7. Flowering Stage
After vegetative plants have been transferred to greenhouse compartments where the
photoperiod is reduced from 18:6 hr light:dark to 12:12 hr or other iterations of light:dark [48,49]; the
onset of inflorescence development is triggered within 1-2 weeks. At this stage of crop development,
symptoms of root infection by Fusarium or Pythium spp. originating from the propagation/vegetative
stage may rapidly become apparent. These symptoms include leaf yellowing, plant wilting, crown
and root rot, and stunted growth (Figure 14). There is no evidence that new infections from residual
inoculum is occurring on flowering plants if all sanitary practices have been followed and
recirculated water is not being used. Symptoms attributed to HLVd infection, which may have been
previously undetected on vegetative plants, will typically manifest within 1-3 weeks after transfer to
the flowering room. These symptoms are distinct, appearing as reduced inflorescence size, yellowing
of the bract leaves, and stunted plant growth [26] (Figure 5). The environmental conditions during
inflorescence development, which includes higher humidity due to increased plant biomass, may
also promote the development of powdery mildew, particularly in more susceptible genotypes.
Closer towards the harvest period when inflorescences begin to mature, bud rot caused by B. cinerea
is likely to become visible, depending on environmental conditions and the genotype. This can lead
to significant reductions in inflorescence quality and yield (Figure 14).
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Figure 14. Symptoms due to pathogen infection on flowering cannabis plants. (a) Yellowing of the
foliage and stunted growth due to infection by Fusarium. (b) Wilting of plants and yellowing of foliage
due to infection by Pythium. (c) Powdery mildew development on inflorescences and surrounding
leaves. (d,e) Bud rot caused by B. cinerea destroys the inflorescence.
In addition to the above pathogens that infect the crop during plant growth, colonization of
inflorescences by yeasts and molds prior to harvest is common and generally remains undetected
until after harvest when quality tests are performed. On the inflorescence tissues, the most common
fungal genera recovered include Penicillium, Alternaria, Cladosporium, and Fusarium (Figure 15). These
microbes can be detected by conducting bud swab tests as described in recent studies [19,20,50]. This
buildup of yeasts and molds can lead to the final dried product failing to meet quality standards
[20,50]. Various factors influence the levels of yeast and mold contamination, which are discussed in
the following sections.
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Figure 15. The most common fungi recovered from inflorescences of cannabis plants. The Petri dishes
show the results from swabbing of samples and plating onto an agar medium that allows growth of
yeasts and molds to occur. On top row – (left to right) Penicillium, Cladosporium, Aspergillus. (On
bottom row) Botrytis, Penicillium and Fusarium. Photos were taken after 7 days.
2.8. Flowering stage IDM approaches
2.8.1. Cultural and Environmental Management
The increased plant biomass resulting from plant development during the flowering stage
creates challenges for maintenance of consistent environmental conditions, particularly with regard
to ambient humidity. Reducing plant densities can significantly lower humidity levels in the
greenhouse and also allows for better light penetration and ease of application of disease control
products. However, lower plant densities can decrease overall yield per unit area of production
[35,51]. A lower ambient relative humidity can also be achieved by increasing air circulation with
circulating fans placed near the plants in the weeks leading up to harvest. Maintaining air movement
at 0.5-1.0 m/s appears to be an optimal target for microbial suppression in cannabis [51]. Under
experimental conditions, enhanced air flow around maturing inflorescences was demonstrated to
significantly reduce the populations of various microbes within the tissues of genotype ‘PH’ (Figure
16). This reduction in humidity, combined with appropriate climate control settings, can mitigate the
severity of diseases such as bud rot (B. cinerea) and powdery mildew during high-risk periods.
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Figure 16. (a) Effect of enhanced air flow around cannabis plants using circulating fans on total
colony-forming units of microbes in these tissues. Vertical bars show total colony-forming units of
total aerobic count (TAMC), bile-tolerant Gram-negative count (BTGN) and total yeast and mold
count (TYMC) with and without air circulation. (b) Fans were positioned 35 cm above the crop to
circulate air continuously at ~7 m/s over ~40 plants, beginning in week 2 of the flowering period until
harvest. The trial was replicated three times in different greenhouse compartments. Inflorescences
were dried prior to microbial analysis.
In relation to seasonal effects on disease development in the greenhouse, bud rot development
was shown to be influenced by external vapour pressure deficits that impacted moisture levels in the
air and hence ambient humidity [35]. To avoid periods of high disease pressure brought on by
external environmental conditions, one IDM strategy is to alter the time of seasonal plantings. By
scheduling planting and harvest times to avoid periods of high disease pressure, particularly with
desirable but susceptible cannabis genotypes, producers can reduce the impact of seasonal pathogens
such as B. cinerea [35], as well as reduce the build-up of total inflorescence microbes that are also
impacted by seasonal fluctuations (Figure 17).
Figure 17. Influence of cannabis genotype and time of year (season) on total microbes present in dried
cannabis inflorescences. Vertical bars denote total aerobic microbial count (TAMC), bile-tolerant
Gram-negative count (BTGN) and total yeast and mold count (TYMC). Samples were taken from three
genotypes during three harvests in each season (fall, winter, summer season) of the same year.
Highest microbial counts were observed in the September harvest period corresponding to late-
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summer production. The failure thresholds for each microbial group are shown by the horizontal
lines. Genotype ‘PD’ contained the highest microbial levels.
An alternate approach is to harvest inflorescences after a shorter crop development period to
avoid prolonged exposure to environmental conditions that favour disease development at the
maturation stage. For example, harvesting at 6 weeks of inflorescence development instead of 8
weeks can reduce bud rot incidence but could result in compromised yield and potency in certain
genotypes unless they are close to maturity [35,52]. Areas within a greenhouse that have localized
disease or “hot spots” should be identified followed by the eradication of the affected plants to
minimize pathogen spread. The diseased plants should be recorded, and if the causal pathogen is
unclear, diagnostic testing should be performed, typically through submissions of samples to a
diagnostic laboratory [4,7]. In addition to visualization of these areas with the naked eye, the utility
of infra-red (IR) and artificial intelligence (AI) powered scouting technologies could be of value as
they have been used in a range of other crops [53-57] but further evaluation of how these technologies
could be modified for application to cannabis is needed.
2.8.2. Utility of Disease Tolerant Genotypes
The cannabis genotype being grown can have a profound impact on the development of certain
pathogens especially under disease-conducive conditions. The impact of genotypes on disease
development at the stock cultivation and propagation stages were described previously. A similar
effect of genotypes on pathogen infection can also be demonstrated at the flowering stage. A
comparison of the response of six genotypes to four pathogens is shown in Figure 18. The genotype
‘LO’ showed high susceptibility to powdery mildew but low susceptibility to HLVd, Botrytis bud rot
and root pathogens. A second genotype ‘LB’ showed high tolerance to all four pathogens while the
remaining genotypes varied in their response to these specific diseases. These data were collected
from observation trails under natural infection and not from replicated trials. They demonstrate,
however, that cannabis producers have an option to select those genotypes that show tolerance to
pathogens under the specific cultivation conditions of greenhouse production. While the genetic basis
for this level of tolerance has not been determined, it indicates there is a basis on which to establish
breeding programs that can lead to the development of disease-tolerant cannabis cultivars.
Figure 18. Comparison of disease incidence on six cannabis genotypes to four pathogens,
demonstrating variation in susceptibility to Botrytis bud rot, powdery mildew, hop latent viroid and
Pythium or Fusarium root diseases. Incidence data were obtained from scouting reports made during
the cultivation of batches of genotypes in comparable greenhouse compartments over three
production cycles in the summer season.
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2.8.3. Application of Biological Control Agents.
As described for cuttings during propagation and during vegetative growth of plants, biological
control agents also show promise in reducing specific diseases at the flowering stage of cannabis
plants. The diseases of importance that can be targeted are Botrytis bud rot and powdery mildew.
Application of several biological control products and reduced risk chemicals at weekly intervals as
a fine spray, at full label rates, onto developing inflorescences of the genotype ‘PH’, was observed to
reduce the development of Botrytis bud rot under both low and high disease pressure resulting from
natural infection during the Fall growing season (Figure 19).
Figure 19. Comparative efficacy of six biological control products and reduced risk chemicals on
Botrytisbud rot development on flowering cannabis plants. Three applications were made at weeks 2,
3 and 4 of the flowering period at maximum label rates. The sprays were applied to ca. 216 plants
using a robotic pipe rail sprayer that delivered ~60 mL of product to each plant. Disease assessments
were made at harvest (week 8) in a greenhouse compartment with low and high Botrytis bud rot
pressure from natural inoculum. (a) Low disease pressure flower room; (b) High disease pressure
flower room.
The most effective product was Rootshield HC
®
(containing Trichoderma harzianum) followed by
Regalia
®
(Reynoutria sachalinensis), Double Nickel
®
(Bacillus amyloliquefaciens), Lifegard
®
(Bacillus
mycoides) and Prestop
®
(Gliocladium catenulatum). Zerotol
®
(hydrogen peroxide) did not show an effect
(Figure 19). The efficacy of the various biocontrol agents likely stems from their ability to pre-
emptively colonize the inflorescence tissues and exert competition against the pathogen, a mode of
action also reported on other crops [58,59]. The application of T. harzianum was also found to suppress
the development of other microbes naturally present within the inflorescences, including Penicillium
spp., and this was reflected by a reduction in all three categories of microbial counts (Figure 20).
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Figure 20. Effect of Rootshield HC
®
(T. harzianum) applications made at weeks 2, 3, and 4 of the
flowering period on final microbial levels in harvested cannabis inflorescences. (a) Total counts of all
microbes in both untreated and sprayed plants are shown. Total microbes were reduced following
Rootshield applications. (b) The growthof microbial colonies after blending of the treated
inflorescences in distilled water and subsequent plating onto agar medium. A comparison is shown
of samples following applications of Rootshield made at weeks 2, 3 and 4 of the flowering period.
Samples treated atweek 4 show maximum suppression of Penicillium growth compared to week 2
where there is no suppression and no colonies of Trichoderma were recovered.
Following this spray trial, a second trail demonstrated that applying T. harzianum (Rootshield
HC
®
) thrice to the foliage of flowering cannabis plants also reduced the development of powdery
mildew compared to untreated plants, as shown in Figure 21. These results indicate that a single
biological control agent may target two important diseases affecting cannabis, namely Botrytis bud
rot and powdery mildew. Trichoderma applications have been shown to suppress powdery mildew
in several crops [60-62].
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Figure 21. Effect of Rootshield HC
®
applications on development of powdery mildew. Three weekly
applications were made to the foliage of flowering plants as preventative treatments and compared
to an untreated control and a water control. (a) Untreated control leaves. (b) Rootshield HC
®
treated
leaves. (c) Water treated leaves.
2.8.4. Application of Reduced-Risk Products
A number of reduced risk products are available for use on cannabis plants at the flowering
stage. During this phase of crop development, care must be taken to avoid damage to inflorescence
tissues and to avoid visual quality changes. Products including Agrotek vaporized sulfur
®
, Regalia
Maxx
®
, Suffoil-X
®
and Milstop
®
are registered to reduce powdery mildew development [43]. Sulfur
is applied via vaporizing pots, a method that ensures uniform dispersal and is commonly used on
many other greenhouse crops [31] while the remainder are applied as sprays. In a comparative study
to evaluate these and other products for powdery mildew control on flowering cannabis plants, nine
products were applied thrice at days 0, 7 and 14 of the flowering period (~ 60 mL per plant) during
the spring season on ‘MP’, a susceptible cannabis genotype, prior to disease appearance.
Subsequently, disease severity was rated visually using a leaf infection coverage scale, as follows: 0
= 0%, 1 = 1-33% = 2 = 34-66%, 3 =67-100% (Figure 22a). Results showed that Suffoil-X
®
applied at a rate
of 10 mL/L and Regalia Maxx
®
applied at a rate of 2.5 mL/L were the best preventative products
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(Figure 22b). In a subsequent trial with same genotype, flowering plants visibly infected with
powdery mildew (disease rating of 1) received one application of seven products at their maximum
label rates made at day 42 of the flowering period to evaluate their curative potential. The findings
showed that Milstop
®
applied at a rate of 3 g/L and Cyclone
®
applied at a rate of 12 mL/L were the
best for curative treatments (Figure 22c). The remaining products provided varying levels of disease
reduction. No phytotoxicity was observed in any of the treatments. The active ingredients in Milstop
®
(potassium bicarbonate), Cyclone
®
(citric and lactic acid) and Suffoil-X
®
(mineral oil) are all
considered to be ‘physical’ in their mode of action, altering leaf surface pH and osmotic pressure or
desiccating/coating mycelium and spores [6]. The active ingredient in Regalia Maxx
®
is an extract
from the giant knotweed Reynoutria sachalinensis and was shown to be effective against pathogens
such as B. cinerea and powdery mildew on cannabis as well as on various other crops [63-66]. This
product enhances plant defense responses through the salicylic acid (SA)-dependent pathway by
inducing the accumulation of plant defence chemicals such as hydrogen peroxide and formation of
mechanical plant defences such as callose papillae [66,67]. Additional research is needed to explore
the breadth to which Regalia Maxx
®
can control other pathogens and the duration of the protection
offered following application.
Figure 22. Comparative efficacy of reduced risk products at managing powdery mildew development
on cannabis genotype ‘MP’. (a-d) Disease was rated according to the scale shown, from 0 (a) to 3 (d).
(e) Products were applied as preventative treatments at days 0, 7, and 14 of the flowering period. (f)
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Products were applied as a curative treatment, once at day 42 of the flowering period, after the onset
of disease development. The trials were conducted during the spring growing season.
A summary of the IDM approaches that can be used against four important pathogens of
cannabis is provided in Table 1.
Table 1. Summary of IDM strategies for four important pathogens affecting cannabis plants.
HLVd Stunting
Disease
Fusarium/Pythium Root
& Crown Rot Botrytis Bud Rot Powdery Mildew
Prevention
Test propagative
materials and stock
plants; utilize
pathogen-free planting
materials.
Test propagative
materials and stock
plants; utilize pathogen-
free planting materials.
Reduce canopy humidity
by adjusting planting
density and enhancing air
circulation.
Maintain an even climate,
above 21°C, and vaporize
sulfur nightly.
Sanitation
Clean equipment and
bench surfaces; destroy
diseased plants.
Clean equipment and
bench surfaces; actively
remove dead or diseased
tissues.
Fog growing environment
with reduced risk products
prior to planting.
Fog growing
environment with
reduced risk products
prior to planting.
Protection
Isolate propagative
materials and stock
plants in controlled
access areas.
Apply Trichoderma
harzianum and
Gliocladium cantenulatum
as a drench to rooted
cuttings and plants.
Apply Rootshield HC® on
developing inflorescences,
from day 14 to day 28 of
flowering.
Preventatively spray
reduced risk products
such as Suffoil-X, Regalia
Maxx, on susceptible
genotypes.
Monitoring
Scout regularly for
symptoms; routinely
sample water and
suspect plants.
Scout regularly for
symptoms; routinely
sample water and
suspect plants.
Conduct daily scouting for
bud rot from the sixth
week of flowering
onwards.
Conduct weekly scouting
at all plant development
stages.
Eradication
Immediately remove
and safely dispose of
diseased plants at all
stages of growth.
Immediately remove and
safely dispose of
diseased plants at all
stages of growth.
Remove and dispose
infected inflorescences;
perform post-drying bud
rot severity checks.
Remove infected leaves
and dispose; spot spray
reduced risk products.
Genotype
Selection
Avoid highly
susceptible genotypes;
evaluate tolerant
genotypes.
Avoid highly susceptible
genotypes; evaluate
tolerant genotypes.
Avoid planting highly
susceptible genotypes
during Botrytis-prone
periods; evaluate tolerant
genotypes.
Avoid highly susceptible
genotypes; evaluate
tolerant genotypes.
2.9. Post-Harvest IDM Approaches
Following harvest of cannabis inflorescences, they undergo a phase of drying to reduce moisture
content to levels that would minimize the development of microbes [19,20,52], following which they
are trimmed and prepared for packaging and stored prior to shipment. During each of the post-
harvest processing stages, there is the potential for microbial contamination to be increased, primarily
consisting of total yeasts and molds (TYM), total aerobic microbial count (TAMC), and bile-tolerant
Gram-negative count (BTGN). Some of these microbes likely originated from the original fresh
harvested inflorescences in the greenhouse or otherwise may have been picked up through
contamination during harvesting and post-harvest processing stages. Detailed studies are lacking
regarding at which specific stages the levels of microbes may build-up to cause the final product to
potentially fail to meet regulatory standards. However, pre-harvest, it has been shown that cannabis
genotype and growing conditions can significantly influence TYM build-up; in addition, post-harvest
drying methods and handling practices can affect TYM levels [19,20,52]. A number of commonly
encountered fungi have been identified on dried cannabis products pre- and post-harvest (Figure 15)
and they contribute to TYM levels [19].
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The implementation of Integrated Disease Management (IDM) approaches to reduce total yeast
and mold (TYM) is complicated by several pre-harvest variables. For example, TYM levels tend to be
higher in the summer season than in winter, certain cannabis genotypes accumulate much higher
TYM than others, and post-harvest handling practices influence TYM levels (hang-dried
inflorescences have lower TYM than those that are rack-dried) [19]. Managing these factors to
minimize microbial build-up depends on the appropriate application of IDM strategies that were
previously outlined for stock plants, and during propagation, vegetative growth, and flowering.
Post-harvest processing practices, such as reducing moisture by hang-drying plants at a high vapor
pressure deficit (VPD) and trimming only after inflorescences are dry, along with thorough cleaning
of post-harvest processing equipment using sanitizing agents, can significantly reduce microbial load
on inflorescences. Additionally, conducting detailed inspections at each stage of post-harvest
processing to detect presence of molds is critical. This may involve various standard practices,
including predefined in-process Acceptable Quality Level (AQL) checks, to ensure that any quality
issues are identified and addressed prior to shipment, as is commonly done in many food processing
plants [68].
Irradiation of cannabis products with gamma and electron beam irradiation has been shown to
be an effective option for producers; they can be used to sterilize commercial batches of inflorescences
without major changes in quality, but they are costly [69-71]. Irradiation is typically used in cases
where microbial levels have exceeded regulatory limits or where a zero tolerance is recommended
i.e. for medical patients with immunocompromised immune systems that rely on cannabis [20]. Other
approaches have been described that require more in-depth studies to demonstrate their commercial
utility [72,73]A summary of the various approaches that can be implemented as a part of an IDM
program for greenhouse-cultivated cannabis is presented in Figure 23. These are organized according
to growth stages of the cannabis crop. These approaches can be readily implemented, and examples
of their successful use have been included in this review. Additional potential IDM approaches for
cannabis that require further research, but which have shown potential in other crops, are described
below.
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2.10. Future Potential Areas for IDM Development for Cannabis
2.10.1. Evaluation of Endophytes and Microbial Antagonists in Cannabis
Endophytes, consisting primarily of fungal and bacterial species, are present within various
tissues and organs of cannabis and hemp plants and vary in species composition, depending on the
tissue source, such as roots, stems, petioles, leaves, flowers and seeds [37,40,74-76]. For example,
endophytes which are consistently present in stems of cannabis plants include species of Penicillium
and Chaetomium, as well as others (Figure 24).
Figure 24. Examples of endophytic fungal and bacterial species recovered from cannabis stem
segments following sterilization. On the left Petri dish are Penicillium species and on the right are
Chaetomium and bacterial species. .
Various plant growth-promoting rhizobacteria, including species of Bacillus and Pseudomonas,
have also been reported to be present in roots and can inhibit the growth of root pathogens [40,77].
These endophytes can potentially improve plant growth and development [78,79] although research
evaluating their growth benefits to cannabis and hemp plants are currently lacking. Dumigan and
Deyholos [37] reported that seed-borne bacterial endophytes, including Bacillus subtilis and B.
inaquosorum, showed inhibitory activity in dual culture assays against fungal pathogens, including
Alternaria and Fusarium species. These endophytes were also present in hemp seeds and included
Bacillus velezensis and Paenibacillus polymyxa, which were also inhibitory to growth of Alternaria,
Aspergillus, Fusarium and Penicillium species. Pseudomonas species have also shown growth inhibition
of Fusarium species in vitro [80]. In previous research, antagonism to B. cinerea in dual culture assays
was demonstrated for several cannabis-derived endophytes (Paecilomyces lilacinus and
Penicillium spp.) [75] and for several hemp-derived endophytes (Pseudomonas fulva and P. orientalis )
[80]. In a study conducted by Gabriele et al. [81] on investigating the endophytes present in the seeds
and young plants of a cannabis cultivar, a unique resistance to the plant's own antimicrobial
compounds was discovered, along with an enhancement of nutraceutical aspects such as polyphenol
content and antioxidant activity in the plants. This finding suggests the potential for introducing
these endophytes as natural biostimulants and biological control agents against pathogenic microbes,
unhindered by the plant’s inherent antimicrobial properties. Such symbiotic relationships underscore
the significant potential of endophytes in cannabis cultivation but further research is needed to
establish their potential applications. The antagonistic properties of endophytic bacteria have been
attributed to antibiotic production, host defense response induction, growth promotion, competition,
parasitism and quorum signal interference [82-85]. Despite these promising studies, however, whole
plant assays demonstrating the benefits of these bacteria and other fungal endophytes are presently
lacking for cannabis. It should be noted that fungal endophytes can also be present in stem tissues of
mother plants, including those shown in Figure 24, and they could negatively impact the health of
these plants over time and complicate attempts to initiate tissue cultures using explants from these
plants [86].
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Another aspect of potential microbial antagonism against fungal pathogens infecting cannabis
that requires research is the diverse microflora that can be present in organic soils compared to
conventional hydroponic cultivation. Punja and Scott [87] reported that a diverse range of microbes
were recovered from cannabis inflorescences grown in organic soil compared to coco-fibre medium
commonly used in hydroponic cannabis production. These communities were comprised of
pathogenic, saprophytic, and beneficial microbes. Of the beneficial microbes detected, Trichoderma
harzianum and Metharzium anisopliae are currently used as biological control agents for root disease
suppression and for insect suppression, respectively. M. anisopliae may hold some potential for
cannabis pathogen suppression as well [86]. In the context of disease management, similar microbes
which originate from organic soils that exhibit general antagonistic properties such as
mycoparasitism, host defense response induction, competition, and antibiotic production are worthy
of evaluation [85,89,90]. Cannabis plants grown in ‘living soil’ or growing media amended with
‘compost teas’ may foster greater colonization of roots by these beneficial endophytes although more
research is needed to demonstrate their utility in an IDM program. Caution should be exercised in
ensuring these microbes do not colonize the inflorescences internally or externally, potentially
leading to a failure of the product due to excessive levels of TYM.
2.10.2. Tissue Culture Applications for Cannabis
Tissue culture of cannabis has received considerable recent interest in efforts to obtain a source
of clean plant materials that can be free of pathogenic microbes, including fungi, viruses and viroids.
Detailed methods have been described from several laboratories and various techniques have been
developed [38, 86,91,92]. The interest among cannabis producers in utilizing tissue culture methods
is to obtain pathogen-free plants and minimize pathogen re-introduction into commercial production
facilities. This is particularly relevant in the context of Hop latent viroid, which is known to be spread
through vegetative cuttings taken from infected mother plants [26]. Meristem tip culture technology
has been used for many decades to eliminate the potential for virus introduction in other vegetatively
propagated crops, such as potatoes, bananas, and strawberries [93-95]. Meristem and shoot tip culture
techniques have been utilized not only for virus elimination but also for rapid clonal multiplication
and germplasm preservation of many vegetatively propagated crops [96,97]. In some cases, these
methods are augmented with cryotherapy (cold treatment), thermotherapy (heat treatment),
chemotherapy (anti-viral chemical treatment), electrotherapy (electrical current treatment) and
shoot-tip grafting (micrografting technique) to enhance the chances of obtaining pathogen-free
planting materials [98-102]. Research to evaluate the applicability of these methods to obtain
pathogen-free planting materials of cannabis, particularly for HLVd, are still in the early stages of
evaluation and development. Tissue culture-derived plants can be obtained from meristems and
nodal explants of cannabis, resulting in shoot growth of a number of genotypes in vitro (Figure 25).
However, confirmation of the eradication of pathogens of importance requires additional research.
Hence, while tissue culture approaches hold promise for potential inclusion in an IDM program for
cannabis, more effort to generate high frequencies of plants confirmed to be pathogen-free on an
economically feasible scale is needed. The confirmation of pathogen-free planting materials could be
utilized for certification programs for cannabis, similar to many agriculturally important crops.
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Figure 25. Tissue-culture derived plants of cannabis can be obtained from meristem tips (a) and nodal
explants (b) resulting in growth of a number of genotypes (c). The feasibility to generate large-scale
production of pathogen-free planting materials awaits further research and development.
2.10.3. Pathogen Control Product Registration for Cannabis
To evaluate new products aimed at managing fungal pathogens in agricultural crops, screening
for pathogen growth inhibition is an initial step. For example, effective concentration (EC
50
) values
are determined for the product to identify the fungicide levels needed to inhibit 50% of the pathogen's
growth in vitro. However, such studies are less commonly reported for products intended for
cannabis. EC
50
studies, which are relatively straight-forward to conduct as demonstrated in Figure 7,
are informative about the potential of new products to inhibit the growth of specific pathogens
affecting cannabis, especially when followed by whole plant assays. These studies can also determine
if there are any secondary effects on biocontrol fungi, such as Trichoderma spp. (Figure 7). Recent
evaluations of products for powdery mildew control in organic hemp production [103] serve to
identify products that may be acceptable for registration in cannabis. Such products could be utilized
for pathogen control at the stock plant and propagation stages, which are critical to ensure that the
subsequent vegetative and flowering stages do not carry over pathogen inoculum and to reduce
concerns about product residues on the finished flower. Currently, the majority of products
registered for use on cannabis can be applied up until harvest, as outlined by Scott et al. [5]. Several
commercial products have since been added to the registered list that are not included in [5]. For soil
fumigation: Pic Plus Fumigant
®
(Chloropicrin – 85.1%), Chloropicrin 100 Liquid Soil Fumigant
®
(Chloropicrin – 85.1%), and Mustgrow Crop Biofumigant
®
(Oriental Mustard Seed Meal – 100%) can
be used pre-plant. For powdery mildew suppression: Vegol Crop Oil
®
(Canola Oil – 96%), Suffoil-X
®
(Mineral Oil – 80%), Purespray FX (Mineral Oil - 80%), and General Hydroponics Suffocoat (Canola
Oil – 96%) can be applied as foliar sprays. For suppression of Botrytis cinerea, powdery mildew, and
Sclerotinia sclerotiorum: Timorex Gold
®
(Tea Tree Oil – 23.8%) can be used. For Phytophthora spp. and
Verticillium dahlia suppression: Foretryx
®
(Trichoderma asperellum strain ICC 012 and Trichoderma
gamsii strain ICC 080) can be used. While many of these products are different formulations of the
same active ingredient, unique products have been added each year since the legalization of cannabis
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31
production in Canada. A current list of registered products can be found at Health Canada - Pesticide
Label Search (hc-sc.gc.ca).
2.10.4. Nutrient Supplements for Cannabis Disease Suppression
Nutrient amendments have been shown to impact plant susceptibility to infection by a range of
pathogens, often reducing disease development through various mechanisms. These changes involve
a wide range of macronutrients and micronutrients. In hydroponic greenhouse cultivation, nutrient
levels are carefully monitored to prevent deficiencies; thus, additional nutrient supplements must be
approached cautiously to avoid phytotoxicity. Formulation and rates are critical factors when
considering these nutrients for disease management [104,105]. The use of nutrient supplements
containing copper, silicon, and calcium show particular promise for cannabis and can be applied via
the roots or foliage.
Copper has a long history of use as a bactericide and fungicide on various crops against
numerous pathogens since the discovery of 'Bordeaux mixture' in 1885. It disrupts fungal cell
membrane integrity and interferes with key enzyme activities, thereby inhibiting pathogen growth
and survival [106,107]. Copper can be applied to cannabis as rootzone drenches, foliar sprays, or seed
treatments. For instance, Mayton et al. [108] assessed different seed treatments to manage damping-
off caused by Pythium and Fusarium species on industrial hemp. Seeds treated with a copper-
containing product, Ultim® at 0.05 mg Cu/seed, showed efficacy comparable to fungicide treatments.
Moreover, copper nanoparticles have been successfully applied as dips and foliar treatments on
tomatoes and watermelons to reduce Fusarium infection [109,110]. A copper formulation, Copper
CropTM, reduced powdery mildew on melons [111]. This suppression aligns with the conventional
use of copper sulfate pentahydrate as a foliar fungicide on plant species such as roses and dogwood
[112,113]. On grapevines, copper citrate effectively reduced Botrytis cinerea infections [114]. The
diverse range of pathogens suppressed by copper formulations suggests its potential for use on
cannabis; however, copper is not currently registered for this purpose. Silicon is effective against
various bacterial, fungal, and viral pathogens and can strengthen cell walls via silicon deposits and
also induce plant defense responses [115,116]. Scott and Punja [43] reported that multiple weekly
sprays of potassium silicate, Silamol®, on vegetative cannabis plants significantly reduced powdery
mildew development. In contrast, a single application showed no effect in the current study.
Akinrinlola et al. [103] reported that Sil-Matrix®, a fungicide with potassium silicate, significantly
reduced hemp powdery mildew by 88%. Dixon et al. [117] demonstrated that root-applied silicon at
a rate of 600 kg/ha significantly reduced powdery mildew severity in hemp. Similar benefits of silicon
supplementation have been observed in crops such as cucumbers, roses, and strawberries [118-
123].Calcium application has been shown to reduce pathogen infection by strengthening plant cell
walls, thereby providing greater structural integrity against fungal and bacterial infections [124].
However, its effectiveness in reducing pathogens affecting cannabis has not been studied. There are
no reports of a direct toxic effect of calcium-containing compounds on fungal pathogens affecting
cannabis, suggesting that its activity may stem from reducing host susceptibility or through other
mechanisms. In some crops, root-zone supplementation of calcium nitrate was reported to reduce B.
cinerea severity on beans and tomatoes, although higher doses increased disease on beans [125].
Supplementing roses with calcium nitrate and adding calcium chloride or calcium sulfate to solutions
for harvested flowers reduced B. cinerea incidence under conducive disease conditions [126].
Similarly, increasing calcium and reducing nitrogen levels in the irrigation water for sweet basil
plants reduced both sporulation and infection severity of B. cinerea [127]. Whether enhanced calcium
supplementation on cannabis plants can influence development of B. cinerea remains to be
determined.
2.10.5. Alternative Technologies for Cannabis Disease Detection
The use of recently developed robotic and imaging technologies for scouting for diseases has
garnered interest from cannabis producers. Various options are available with pros and cons,
depending on greenhouse scale, layout and operations. For small-scale greenhouses, fixed crop
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32
monitoring cameras or AI-powered phone scouting apps are often utilized. Several cannabis-focused
scouting apps include Koppert’s Natutec Scout app (https://www.koppert.com/natutec-scout/),
BioBest’s Crop-scannerTM app (https://www.crop-scanner.com/), GrowDoc AI’s app
(https://growdoc.ai/), and the IPM ScoutekTM app (https://ipmscoutek.com/). In contrast, larger
greenhouse operations, with a more consistent layout, have trialed autonomous robotic scouting carts
or booms with cameras attached to crop carts, such as I UNU’s LUNA A I scouti ng abo ve-cr op cam eras
or scouting carts (https://iunu.com/luna-ai), Ecoation’s OKO or ROYA scouting carts
(https://www.ecoation.com/integrated-pest-management), and Budscout AI’s Budscout above-crop
cameras (https://budscout.ai/budscout/). The challenge for AI and imaging solutions currently is that
they may not reliably distinguish symptoms caused by various pathogens from nutrient deficiencies
and other environmental stressors. Supplemental and tailored training are likely needed to achieve
accurate results. In the broader agricultural sector, significant progress is being made in the robot AI-
assisted vision space [56]. An example of the training process and customizability of AI scouting
technology is demonstrated in the following two studies. Anagnostis et al. [55] aimed to build a fast
and accurate object detection system to identify anthracnose-infected leaves (by Colletotrichum spp.)
in a commercial walnut orchard. The study involved segmenting high-resolution images into smaller
sub-images and training an object detector to recognize disease-specific features. The deep learning
approach achieved high accuracy under real-field conditions. Similarly, Mahmud et al. [54] focused
on developing an innovative machine vision system to accurately detect powdery mildew in
strawberry fields. This system utilized real-time image processing and artificial neural networks
(ANNs) to distinguish diseased leaves from healthy ones. The study demonstrated the system's
adaptability to field conditions and showed high accuracy in detecting powdery mildew. These
examples point to the potential for applications in integrated disease management and early disease
intervention in different agricultural settings.
Infrared imaging (IR), a spectrum used in some remote sensing technologies, identifies
variations in crop or leaf temperature to reflect reduced transportational activity or metabolic
functions, signalling the potential presence of stressors including disease [53,57,128]. In cannabis,
detection of leaf surface temperature changes due to root diseases or poor root development when
tested at different developmental stages can be captured with a handheld device such as a FLIR E8
ProTM infrared camera (https://www.flir.ca/products/e8-
pro/?vertical=condition+monitoring&segment=solutions) (Figure 26). This method showed
definitively that poorly developed root systems on affected plants was correlated directly with a
reduced rate of transpiration and hence a build-up of leaf surface temperature that was detectable
with the IR camera. However, when powdery mildew-infected plants or cannabis plants affected by
Hop latent viroid were similarly compared to healthy plants, these plants did not show a
corresponding reduced transpirational activity pattern, suggesting that the IR camera was unable to
detect physiological changes in these diseased plants. It is unknown whether infrared or other
spectrums could be used to effectively detect hop latent viroid; limited research has been done on
virus detection with infrared but there may be potential applications [129,130]. Vagelas et al. [131]
utilized a low-cost infrared camera and a standard RGB web camera to analyze vine,
chrysanthemum, and rose leaves that had been infected with various fungi. Results showed that
infected leaves exhibited temperature deviations from uninfected ones, which occurred before visible
symptoms developed. Specifically, infected vine and rose leaves showed a decrease in temperature,
while chrysanthemum and another set of rose leaves demonstrated an increase, compared to healthy
tissue. Lindenthal et al. [132] used infrared thermography to detect downy mildew infection on
cucumbers. In controlled environments, the study showed that the maximum temperature difference
in a leaf could be used to distinguish between healthy and infected tissues. Under natural
environments, while leaf temperatures and transpiration rates were similar in both healthy and
infected plants, diseased leaves showed more varied transpiration rates depending on the severity of
the symptoms. Liaghat et al. [133] utilized Fourier Transform Infrared (FT-IR) spectroscopy to detect
Ganoderma infections in oil palm trees. This method involved analyzing leaf samples from both
healthy and infected trees, examining the infrared spectra of these samples, and using a statistical
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33
model for classification. The researchers successfully identified differences linked to the disease,
accurately identifying infected trees at early, symptomless stages. Therefore, there is a growing body
of evidence to demonstrate that IR approaches could be applied to cannabis for early detection of
infection by foliar pathogens but additional studies are required to validate this approach.
Figure 26. Infrared and digital image comparisons to illustrate changes in plant surface temperatures
at different stages of cannabis propagation. (a, b) A stock plant exhibiting a low transpiration rate
(and high temperature, in yellow) compared to an adjacent plant with high transpiration (and lower
temperature, in purple) shows a difference in surface temperatures that was attributed to infection by
a root pathogen. (c, d) A cutting in the centre of a tray (arrow) with low transpiration (in yellow)
surrounded by cuttings with higher transpiration rates. While the former cutting showed no obvious
visual symptoms (d), early signs of pathogen infection and reduced rooting were observed. (e, f) A
vegetative plant (arrow) with low transpiration (seen in yellow) among other plants with higher
transpiration rates, shows notable differences in root health (g).
2.10.6. Induction of Plant Defence Responses in Cannabis
The potential for inducing plant defense responses before pathogen infection has not yet been
developed to a practical level for cannabis. However, as discussed earlier, Regalia Maxx (Reynoutria
sachalinensis) when applied to cannabis plants can reduce pathogens such as Botrytis cinerea and
powdery mildew, confirming reports in the literature of the efficacy through presumed induction of
defence responses and pathogen reduction [63-67]. Weekly applications are recommended to ensure
on-going protection on cannabis plants. The role of endophytic microbes in the growing medium in
promoting plant health and reducing pathogen infection on cannabis awaits confirmation. Previous
reports demonstrate the defence-boosting properties of endophytic organisms on various plant
species [82-85,134]. The utility of a biological control product containing Trichoderma spp. appears to
be promising when applied preventatively to the rootzone or to inflorescences; however, additional
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34
research is necessary to establish whether induction of defence responses in cannabis can be
confirmed as it has in previous reports on numerous plants [135,136].
3. Conclusions
This comprehensive review of integrated disease management (IDM) approaches for
greenhouse-cultivated cannabis underscores the significance of developing a multifaceted approach
to control the various pathogens of economic concern. The review highlights the importance of pre-
emptive measures, including selection of disease-tolerant genotypes and use of stringent sanitation
practices, in minimizing pathogen incidence. The utilisation of biological control agents and reduced-
risk products, and the modification of cultural and environmental conditions, have shown promising
results in suppressing Botrytis bud rot, powdery mildew, Pythium and Fusarium root diseases and
hop latent viroid causing stunt disease. Moreover, the exploration into alternative strategies
including utility of endophytes, tissue culture, nutrient supplementation and technology-aided
scouting, offers potential new avenues for enhancing plant health. This review underscores the
dynamic nature of IDM in cannabis cultivation, and emphasizes the continuing need for research and
adoption of sustainable strategies to meet the evolving challenges in disease management within the
greenhouse cannabis industry. Such strategies should receive support from governmental regulatory
agencies to ensure they meet the criteria set forth by the appropriate jurisdictions.
Author Contributions: Conceptualization, L.B, Z.K.P.; methodology, L.B.; formal analysis, L.B., Z.K.P.;
investigation, L.B.; resources, L.B., Z.K.P.; writing—original draft preparation, L.B.; writing—review and editing,
L.B., Z.K.P.; visualization, L.B., Z.K.P.; supervision, Z.K.P.; project administration, Z.K.P.; funding acquisition,
Z.K.P. All authors have read and agreed to the published version of the manuscript.
Funding: This research was funded by an Alliance Grant (ALLRP 571270-21) from the Natural Sciences and
Engineering Research Council of Canada (NSERC) with matching funding from Pure Sunfarms Corp.
Data Availability Statement: There was no new data created in this study.
Acknowledgments: We appreciate the assistance and insights provided by several licenced cannabis producers
who have shared their knowledge of disease managements practices and approaches that were included in this
review. We acknowledge the resources, staff and cultivation expertise provided by Pure Sunfarms Corp., a
licensed cannabis facility in Delta, BC, during the experimental trials. Technical help was provided by S. Lung
and C. Sco.
Conflicts of Interest: The authors declare no conflict of interest.
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