Effects of far-red light on the behaviour and reproduction of the
zoophytophagous predator Macrolophus pygmaeus and its interaction
with a whitefly herbivore
Authors
Davy Meijer1, Syb Hopkoper1, Berhane
Tekle Weldegergis1, Wendy van ’t
Westende2, Joop Johannes Antonius van
Loon1, Marcel Dicke1
1Laboratory of Entomology, Wageningen University, PO
Box 16, NL-6700, AA Wageningen, the Netherlands;2Laboratory of Plant Breeding, Wageningen University,
PO Box 16, NL-6700, AA Wageningen, the Netherlands
Abstract
Plants can detect neighbouring plants through a reduction in the ratio
between red and far-red light (R:FR). This provides a signal of
plant-plant competition and induces rapid plant growth while inhibiting
defence against biotic stress, two interlinked responses designated as
the shade avoidance syndrome (SAS). Consequently, the SAS can influence
plant-herbivore interactions that could cascade to higher trophic
levels. However, little is known on how the expression of the SAS can
influence tritrophic interactions. We investigated whether changes in
R:FR affect the emission of herbivore-induced plant volatiles (HIPVs),
and whether these changes influence the attraction of the
zoophytophagous predator Macrolophus pygmaeus . We also studied
how the expression of the SAS and subsequent inhibition of plant
defences affects the reproduction of M. pygmaeus in both the
presence and absence of the greenhouse whitefly (Trialeurodes
vaporariorum ) as arthropod prey. The results show that changes in R:FR
have little effect on HIPV emissions and predator attraction. However, a
reduction in R:FR leads to increased reproduction of both the predator
and the whiteflies. We conclude that shade avoidance responses can
increase the population development of M. pygmaeus directly by
reducing plant defences, and indirectly by supporting higher herbivore
densities.
Keywords: shade avoidance, volatiles, tritrophic interactions,
predator-prey interactions, arthropod performance, Trialeurodes
vaporariorum
Introduction
Plants possess elaborate mechanisms to sense and respond to changes in
their dynamic environment. These mechanisms help plants to
differentially allocate their resources to many physiological and
developmental processes in order to adequately respond to the risks and
challenges posed by their environment. These allocation patterns often
come down to trade-offs between growth and defence (Zust and Agrawal,
2017). An example of such a trade-off is the shade avoidance syndrome
(SAS). The SAS is aimed at avoiding shade and light competition from
neighbouring plants and consists of morphological and physiological
adaptations that stimulate growth and reproduction (Casal, 2012), but it
is also associated with reduced defensive responses towards biotic
stress (Ballaré, 2014). Over the past decades, there has been increasing
interest in understanding how plants perceive and respond to shading by
neighbouring plants (Ballaré & Pierik, 2017; Fernández-Milmanda &
Ballaré, 2021), and how these responses interact with responses to other
biotic and abiotic stresses (Courbier & Pierik, 2019; Lazzarin et
al. , 2021). These studies have resulted in valuable insights into how
plants balance responses between different stressors, and thereby
provide opportunities to improve plant resilience in agricultural
settings.
Shade avoidance depends on the detection of far-red light, which is
reflected from vegetative plant tissue and therefore provides a signal
of plant density and the degree of competition for light (Ballaréet al. , 1990). Reflection of far-red light lowers the ratio
between red and far-red light (R:FR), which is detected by the
phytochrome B (phyB) photoreceptor. Red light activates phyB, which is
then transported to the nucleus and inhibits SAS responses. With
increasing far-red light, the R:FR drops and phyB is inactivated,
allowing for the expression of SAS (Ballaré and Pierik, 2017). The SAS
is characterised by morphological adaptations such as stem elongation,
upward leaf movement and reduced branching, which serve to increase
light capture in crowded canopies (Casal, 2012). These adaptations are
mediated through far-red induced stimulation of the growth-promoting
phytohormones auxin and gibberellin (Ballaré and Pierik, 2017). At the
same time, inactivation of phyB inhibits the signalling of jasmonic acid
(JA) and salicylic acid (SA), the two principal phytohormones involved
in defence against biotic stress (Ballaré, 2014; Fernández-Milmanda &
Ballaré, 2021). Consequently, plants exposed to far-red light are more
susceptible to biotic stress, leading to the increased performance of
arthropod herbivores (Izzaguirre et al. , 2006; Moreno et
al. , 2009; Cortés et al. , 2016; Meijer et al. , 2022).
Both JA and SA are known to inhibit the signalling of growth-promoting
phytohormones (Wasternack and Feussner, 2018; van Butselaar and van den
Ackerveken, 2020). Downregulation of defensive signalling is therefore
required for the full and rapid expression of SAS, and serves to
prioritize SAS responses over defensive responses when experiencing
competition for light (Ballaré and Austin, 2019). There are also
indications that plants can compensate for the reduced direct defences
against arthropod herbivores by stimulating interactions with the
natural enemies of the herbivores (Cortés et al. , 2016). Plants
that are attacked by herbivores emit herbivore-induced plant volatiles
(HIPVs), a blend of volatile organic compounds (VOCs) that attract the
natural enemies of the attacking herbivores (Dicke and Baldwin, 2010).
These volatile blends are highly specific and can contain information on
the herbivore species, density and on the abiotic conditions of the
plant (Aartsma et al. , 2017; Cabedo-López et al. , 2019).
Exposure to far-red light can influence VOC emissions, with consequences
for plant-arthropod interactions (Kegge et al. , 2013; Cortéset al. , 2016). Plants treated with methyl jasmonate (MeJA), a
volatile form of the stress hormone JA commonly used to induce
anti-herbivore defences, became more attractive to the predatory bugMacrolophus pygmaeus after exposure to far-red light (Cortéset al. , 2016). These results indicate that plants might balance
their direct and indirect defences in response to signals of
competition.
Macrolophus pygmaeus is a generalist predator that can feed on a
broad range of herbivore species (Messelink et al. , 2011; Bouaggaet al. , 2018; Leman et al. , 2020; Kenway et al. ,
2022). However, M. pygmaeus is a zoophytophagous predator,
meaning that it is also able to sustain itself on plant material in the
absence of arthropod prey. At higher densities, M. pygmaeus can
itself cause plant damage (Castañé et al. , 2011; Sanchez et
al. , 2018). The extent of plant-feeding by M. pygmaeus is
dependent on the availability of arthropod prey and on the defensive
status of the plants. In the absence of prey, induction of plant
defences by a non-pathogenic strain of Fusarium oxysporumreduced M. pygmaeus reproduction (Eschweiler et al. ,
2019). In the presence of arthropod prey, inoculation with F.
oxysporum did not affect M. pygmaeus reproduction, but did lead
to decreased prey abundance compared to non-inoculated plants, while
prey abundance was not influenced by F. oxysporum inoculation
alone (Eschweiler et al. , 2019). Together, these results indicate
a shift toward more prey consumption after induction of plant defences.
Reversely, reduced plant defences in low R:FR conditions might make the
plant a more readily available food source for M. pygmaeus and
might cause a shift toward more plant-feeding.
In this study, we further investigated the effects of changes in R:FR on
the attraction and feeding behaviour of M. pygmaeus . Cortéset al. (2016) tested the effects of far-red light on the VOC
emission and attractiveness of plants treated with MeJA. Although MeJA
application can induce anti-herbivore responses, there are no actual
herbivores present. The results reported by Eschweiler et al.(2019) highlight the importance of prey availability for the interaction
between plants and M. pygmaeus . We therefore studied the effect
of R:FR on the HIPV emission of plants infested with the greenhouse
whitefly (Trialeurodes vaporariorum ), and how this influences the
attraction of M. pygmaeus . We tested both an increase and
decrease in R:FR compared to sunlight levels, as previous work indicated
that both changes in R:FR could influence plant-whitefly interactions
(Shibuya et al. , 2010; Meijer et al. , 2022), with possible
consequences for plant-predator interactions. We also investigated how
supplemental far-red light influenced the reproduction of M.
pygmaeus in both the presence and absence of T. vaporariorum as
prey. We expect that a reduction in R:FR influences the emission of
HIPVs and increases the attraction of M. pygmaeus to
whitefly-infested plants. An increase in R:FR is expected to have little
or no effect, as an increase in R:FR also did not influence the
performance of T. vaporariorum (Meijer et al. , 2022). We
further expect that supplemental far-red light increases M.
pygmaeus reproduction in both the absence and presence of prey, related
to reduced plant defences in low R:FR conditions.
Materials & Methods
Plants and arthropods
Tomato plants (Solanum lycopersicum cv. Moneymaker) were used in
all experiments. Greenhouse whiteflies, Trialeurodes vaporariorumWestwood (Hemiptera: Aleyrodide), were obtained from the stock colony of
the Laboratory of Plant Breeding (Wageningen University, the
Netherlands). The T. vaporariorum colony is maintained on tomato
(Moneymaker) at 21/17˚C, 60% RH and 16h/8h L/D. The predatory bugs,Macrolophus pygmaeus (Hemiptera: Miridae), were obtained from a
commercial strain of Biobest Biological Systems (Westerlo, Belgium).
Experimental design
Laboratory experiments were performed to investigate the effects of R:FR
on tomato HIPV emissions and the attraction of the predatory bugM. pygmaeus . Tomato seeds were sown in rockwool blocks (7.5 x 7.5
cm) soaked in Tomato 2.0 nutrient solution (Unifarm, Wageningen, the
Netherlands). Two weeks after germination, the plants were transferred
to a climate-controlled growth chamber (25/18˚C day/night, 70±3% RH and
16h/8h L/D) and divided over three separate compartments with different
light treatments. Each treatment consisted of 150
μmol/m2/s white light (5700K) supplemented with 60
(+FR), 30 (CL) or 0 (-FR) μmol/m2/s of far-red light
(735 nm). The control light treatment (CL) was set at a R:FR of 1.2,
which is equivalent to sunlight, and was reduced (+FR) or increased
(-FR) to 0.6 and 8.2, respectively. Both white and far-red light were
provided by Dyna LED-modules (Heliospectra, Gothenburg, Sweden). The
different compartments were separated with white reflective plastic to
prevent light contamination between treatments. Plants were exposed to
the light treatments for one week before being exposed to T.
vaporariorum whiteflies (WF). Plants were individually covered with
mesh bags, tightly fastened at the base of the rockwool cube. One
hundred adult whiteflies were released inside the mesh bags and left to
feed freely for five days before plants were used in further
experiments. Other plants remained uninfested (U) for five days under
the same conditions, resulting in six treatment combinations
(+FRWF, +FRU, CLWF,
CLU, -FRWF and -FRU).
Y-tube olfactometer
The response of M. pygmaeus to plant volatiles was observed in a
two-choice Y-tube olfactometer as described previously (Lins et
al. , 2014). A Y-shaped Pyrex tube, formed by an entry arm (20 cm) and
two side arms (13 cm, 80° angle), was positioned vertically. The two
side arms were each connected to 15 L glass jars containing tomato
plants as odour sources. Compressed air was passed through the jars at a
regulated flow of 2.5 L/min to carry the plant volatiles into the arms
of the Y-tube. Before reaching the jars, the air was filtered by passing
it through active charcoal. The glass jars were isolated in light-tight
cabinets, equipped with a Dyna LED module, to provide plants with their
respective light treatment throughout the choice assays while preventing
visual detection of the plants by the predators. A single plant was
introduced in each glass jar. The rockwool base of the plants was
wrapped in aluminium foil before placement into the jar and the plants
were left to acclimate for 15 min before starting choice assays.
Individual predators were released at the downwind arm of the Y-tube,
and their choice for either odour source was recorded when they passed
at least 10 cm into one of the side arms, or no choice was recorded if
they did not pass this mark within 10 min after their release. Pair-wise
comparisons were made between whitefly-infested plants from the three
light treatments (+FRWF, CLWF and
-FRWF). Because M. pygmaeus is known to prefer
whitefly-infested plants over undamaged plants (Ingegno et al. ,
2011; Leman et al. , 2020), choice assays between uninfested and
infested plants of the control-light treatment (CLU and
CLWF) were added as a positive control. For each
pair-wise comparison, the response of 10 female M. pygmaeuspredators of 7-10 days old was tested daily for a total of 13 days (130
predators per comparison). After each set of five predators, the
position of the odour sources was switched between the left and right
side arms to prevent positional bias. After every 10 predators, the
plants were removed and the Y-tube system was flushed with clean air for
10 min before introducing new plants.
Collection of headspace VOCs
Volatiles were collected from plants of all six treatment combinations.
The rockwool base of plants was wrapped in aluminium foil before plants
were placed in 15 L glass jars. They were left to acclimate for 15 min
before starting the headspace collection. Air was filtered through
active charcoal before reaching the jars, and volatiles were collected
by drawing air with a suction pump through a stainless steel cartridge
containing 200 mg of Tenax TA (20/35 mesh, CAMSCO, Houston, Texas, USA)
at 150 mL/min for two hours. During volatile collection, plants remained
exposed to their respective light treatments. Samples were collected
from at least 10 plants for each of the six treatments. Volatile samples
of empty rockwool blocks soaked with Tomato 2.0 nutrient solution and
wrapped in aluminium foil were also collected to correct for background
odours. The Tenax cartridges with VOC samples were dry purged under a
stream of helium (50 mL/min) for 15 min to remove excess moisture.
The collected volatiles were thermally released from the Tenax TA
adsorbent using an Ultra 50:50 thermal desorption unit (Markes,
Llantrisant, Glamorgan, UK) at 250 °C for 10 min under a 20 mL/min
helium flow, while the volatiles were simultaneously re-collected in a
thermally cooled universal solvent trap: Unity (Markes) at 0 °C. When
desorption was completed, the volatile compounds were released from the
cold trap by ballistic heating at 40 °C/s to 280 °C, which was then kept
for 10 min, while all the volatiles were transferred to a ZB-5 MS
analytical column (30 m x 0.25 mm ID x 1 mm F.T. with 10 m built-in
guard column (Phenomenex, Torrance, CA, USA), placed inside the oven of
a Thermo Trace GC Ultra (Thermo Fisher Scientific, Waltham, MA, USA) for
further separation of the plant volatiles. The gas chromatograph (GC)
oven temperature was initially held at 40 °C for 2 min and was then
raised at 6 °C/min to a final temperature of 280 °C, where it was kept
for 4 min under a constant helium flow of 1 mL/min. A Thermo Trace DSQ
quadrupole mass spectrometer (Thermo Fisher Scientific) coupled to the
GC was operated in an electron impact ionization (EI) mode at 70eV in a
full scan with a mass range of 35–400 amu at 4.70 scans/s. The mass
spectrometer (MS) transfer line and ion source were set at 275 °C and
250 °C, respectively. Automated baseline correction, peak selection (S/N
> 3) and alignments of all extracted mass signals of the
raw data were processed following an untargeted metabolomic workflow
using MetAlign software, producing detailed information on the relative
abundance of mass signals representing the available metabolites
(Lommen, 2009). This is followed by reconstruction of the extracted mass
features into potential compounds using the MSClust software through
data reduction by means of unsupervised clustering and extraction of
putative metabolite mass spectra (Tikunov et al. , 2012).
Tentative identification of volatile metabolites was based on comparison
of the reconstructed mass spectra with those in the NIST 2008 and
Wageningen Mass Spectral Database of Natural Products MS libraries, as
well as experimentally obtained linear retention indices (LRIs).
Macrolophus pygmaeus reproduction
A greenhouse experiment was conducted to study the effects of
supplemental FR light on the reproduction of M. pygmaeus , in both
the absence and presence of T. vaporariorum as arthropod food
source. Tomato seeds were sown in rockwool blocks as described above and
transported to a greenhouse compartment (22/18˚C, 70% RH and 16h/8h
L/D) three weeks after germination. Plants were individually placed in
mesh cages (60 x 40 x 40 cm; BugDorm, Taiwan) and divided over two
tables, both illuminated with 150 μmol/m2/s broad
spectrum white light provided by VYPR series LEDs (model VR-3X-BW4,
Fluence, Rotterdam, the Netherlands) as supplement to natural daylight.
One table was further supplemented with 60 μmol/m2/s
far-red light (Philips Greenpower LEDs, Eindhoven, the Netherlands),
creating a R:FR of 0.8 (+FR) compared to 1.6 in the control (CL). Ten
plants were placed on either side of the greenhouse and were exposed to
the light conditions for one week. After four weeks, 100 adult
whiteflies of mixed age and sex were introduced to half of the cages,
five in either light treatment, creating four treatments based on the
supplementation of FR light and the presence of whiteflies (CL/-WF,
CL/+WF, +FR/-WF and +FR/+WF). The introduction of 100 adult whiteflies
occurred weekly to maintain a viable population and food source forM. pygmaeus throughout the experiment. At six weeks, 12 adultM. pygmaeus of approximately 5-10 days old (six males and six
females) were introduced to each cage. After 25 days, all M.
pygmaeus were collected and counted per cage. The experiment was
repeated three times between April and September 2022, resulting in 15
replicates per treatment. During the second and third run, leaflets from
the 6th and 9th leaf in the C/+WF
and +FR/+WF treatments (10 plants per treatment) were collected to
provide estimates of the whitefly population within the cage. Leaf discs
of 2.5 cm2 were excised from the three most terminal
leaflets per leaf (30 leaf discs per leaf per treatment) and the number
of whitefly eggs and nymphs was counted. The 9th leaf
was the youngest fully developed leaf.
Statistical analysis
Choice responses of M. pygmaeus in the two-choice Y-tube
experiment were analysed with a two-sided binominal test with α = 0.05,
for each pair-wise comparison separately. Data are presented as the
total number of M. pygmaeus choosing either odour source in each
pair-wise comparison.
The volatile emission data, expressed as peak heights, were imported
into SIMCA-P 17 statistical software (Umetrics, Umea, Sweden), followed
by log-transformation, mean-centering and unit-variance scaling before
being subjected to multivariate data analysis. Unsupervised principal
component analysis (PCA) and/or supervised partial least
squares-discriminant analysis (PLS-DA), and its extension orthogonal
partial least squares-discriminant analysis (OPLS-DA) were used as tools
to compare and correlate treatment groups. The results of the analysis
are visualized in score plots, which reveal the sample structure
according to model components, and in loading plots, which display the
contribution of the variables (individual VOCs) to these components as
well as the relationships among the variables. R2 and
Q2 metrics are provided for PLS-DA or OPLS-DA
analysis. These metrics describe the explained variation within the data
set and the predictability of the model, respectively, and were
calculated based on the averages of the sevenfold cross-validation.
R2 and Q2 values range between 0–1,
and the closer these metrics are to 1, the higher the variance explained
by the model and the more reliable the predictive power of the model.
Significant differences in the total emission of individual VOCs between
light treatments of either infested or uninfested plants were analysed
using Kruskal-Wallis analysis. Pair-wise comparisons, as performed in
the Y-tube choice assays, were analysed using Mann-Whitney U analyses
for comparisons showing significant separation in multivariate data
analysis.
Macrolophus pygmaeus reproduction, expressed as the total number
of nymphs produced per six females, were analysed using a univariate
general linear model (GLM) with light treatment and whitefly presence as
fixed factors, and the experimental replicates as random factor.
Whitefly numbers are expressed as the total number of eggs, crawlers
(1st instar nymphs) and nymphs per leaf disc. A
separation between 1st instar nymphs and older stages
is made because crawlers are more easily distinguished from other
nymphal stages. Data were analysed using multivariate GLM with light
treatment and leaf number as fixed factors, and plant ID and the
experimental replicates as random factors.
Results
Y-tube choice assays
In the positive control, M. pygmaeus significantly preferred
whitefly-infested plants over uninfested plants (Fig. 1). No preference
was observed when M. pygmaeus were offered the choice between
whitefly-infested plants exposed to different R:FR light ratios in
pair-wise comparisons. This indicates that changes in R:FR did not
influence the attraction of M. pygmaeus to whitefly-infested
plants.
Headspace VOC emissions
We identified 70 VOCs across the six different treatment combinations
(Table S1). These compounds were detected in at least 50% of the
samples from at least one of the treatment combinations. Multivariate
analysis (OPLS-DA) comparing the volatile blends from uninfested plants
of all three light treatments (+FRU, CLUand -FRU) did not result in a significant separation
between volatile blends (permutation test: R2 = 0.287,
Q2 = 0.024; pcv-ANOVA = 1.000),
indicating that changes in R:FR did not influence the composition of
constitutive volatile blends (Fig. 2A and B). Similarly, OPLS-DA
including the whitefly-infested plants of all three light treatments
(+FRWF, CLWF and -FRWF)
also did not result in significant separation (permutation test:
R2 = 0.320, Q2 = 0.056;
pcv-ANOVA = 0.998), indicating that HIPV emission is
also not affected by changes in R:FR (Fig. 2C and D). In uninfested
plants, nine VOCs were emitted in significantly different quantities by
plants exposed to different R:FR ratios, all showing a gradient from
highest emission in +FR to lowest in -FR, with the exception of
(Z )-3-hexen-1-ol (Table S1). In whitefly-infested plants, only
four VOCs showed significant differences, following the same gradient
from +FR to -FR, with the exception of 2-methylbutanal-O-methyloxime
(Table S1).
We also compared volatile blends pair-wise following the pair-wise
comparisons performed in the Y-tube choice essays. Surprisingly,
multivariate analysis (PCA) did not show a separation between the
volatile blends of CLU and CLWF (Fig.
S1). Similarly, no separation was observed between the volatile blends
of CLWF and +FRWF, or between VOC blends
of CLWF and -FRWF (Fig. S2). We did
observe significant separation between the blends of
+FRWF and -FRWF using PLS-DA analysis
(permutation test: R2 = 0.967, Q2 =
0.731; pcv-ANOVA = 0.011) (Fig. S3). Twenty-three
compounds contributed most to the separation, with five VOCs showing
significantly higher emission in +FRWF and one compound
showing significantly lower emission in +FRWF (Table
S2).
Macrolophus pygmaeus reproduction
The total number of M. pygmaeus nymphs collected after the 25-day
period was significantly higher in the presence of whitefly prey
compared to when no prey was available (Fig. 3). Furthermore, the number
of nymphs was significantly higher on plants exposed to +FR compared to
CL treatments. No significant interaction between the presence of prey
and the light treatments was found, indicating that also in the absence
of prey, supplemental FR light increased M. pygmaeusreproduction.
Trialeurodes vaporariorum density estimate
Both light treatment and leaf number had a significant effect on the
number of whiteflies of all three life stages (Fig. 4). +FR
significantly increased whitefly numbers compared to CL. The effect of
leaf number was related to the leaf age; younger leaves
(9th leaf) contained more eggs and crawlers while
older leaves (6th leaf) had more nymphs. A significant
interaction between light treatment and leaf number was only observed
for nymphs (Fig. 4).
Discussion
Shade avoidance responses are part of a complex network of ecological
interactions within the canopy that together shape the growth-defence
dynamics in plants (de Vries et al. , 2017). The prioritization of
shade avoidance responses over defensive responses leads to changes in
plant-arthropod interactions that span across trophic levels (Lazzarinet al. , 2021). Previous work indicated that plants experiencing
competition might trade direct defences for indirect defences by showing
that plants expressing the SAS are more attractive to the predatory bugM. pygmaeus (Cortés et al. , 2016). In this paper, we
further examined the effects of shade avoidance on the interaction
between M. pygmaeus and tomato plants, thereby taking into
account the zoophytophagous nature of this predator. We show that the
volatile-mediated attraction of this predator to whitefly-infested
plants is not affected by changes in R:FR, but that far-red light does
influence the reproduction of M. pygmaeus and whiteflies.
Our results indicate that changes in R:FR do not strongly affect the
emission of tomato volatiles and also do not influence the attraction ofM. pygmaeus to whitefly-infested plants. Cortés et al.(2016) previously reported that exposure to far-red light increased the
attraction of this predator to MeJA-treated tomato plants. Whiteflies
are known to induce SA-dependent defences in plants (Elzinga et
al. , 2014; Xu et al. , 2019). SA- and MeJA-induced volatiles can
be differentially affected by changes in R:FR, resulting in differences
in predator behaviour in response to these volatiles. However, previous
work showed that the attraction of predatory mites to plants infested
with JA-inducing spider mites (Tetranychus urticae ) was also not
affected by changes in R:FR (Meijer et al. , 2023), indicating
that changes in R:FR do not influence the emission of both SA- and
JA-induced HIPVs. The observed results by Cortés et al. (2016)
might therefore result from the use of MeJA instead of actual herbivory.
Although application of MeJA is an effective method to induce
anti-herbivore responses in plants, there are differences in the
responses induced by MeJA application and true herbivory (Dicke et
al. 1999; Kappers et al. , 2010; Lortzing et al. , 2017;
Papazian et al. , 2019), which can affect predator preference
behaviour. For example, volatiles emitted by spider mite-infested plants
were more attractive to a predatory mite than volatiles from JA-treated
plants (Dicke et al. , 1999). VOC blends emitted by
hormone-induced plants are missing the specific information pertaining
to herbivore identity and density and might therefore provide a more
general stress signal (Kappers et al. , 2010). Generalist
predators are proposed to be more sensitive to general stress indicators
compared to specific HIPVs (Silva et al. , 2021). Due to the
zoophytophagous nature of M. pygmaeus , it is also possible that
the presence or absence of actual herbivores can change the
informational value of the volatile blends. Far-red light might alter
the information provided by MeJA-induced volatile blends as it provides
information on the defensive status of the plant and the accessibility
of plant material, while changes in R:FR might not change the
attractiveness of herbivore-infested plants as the presence of prey is
the overriding element. It would be interesting to perform comparative
studies between MeJA-treated and herbivore-infested plants to determine
whether the presence of prey is a determining factor for the attraction
of predators in plants experiencing competition.
The lack of preference between whitefly-infested plants exposed to
different R:FR light ratios largely corresponds with the lack of
differences in volatile blend composition, with the exception of the
pair-wise comparison between +FRWF and
-FRWF. A significant separation between the volatile
blends of these treatments was observed, although M. pygmaeus did
not show a preference for plants exposed to either +FRFWor -FRWF. Reversely, we did not find a significant
separation between volatile blends of whitefly-infested or uninfested
plants grown under control light conditions (CLWF and
CLU), while M. pygmaeus showed a clear preference
for CLWF plants. A possible explanation is that
compounds emitted by whitefly-infested tomato plants that are relevant
for attracting M. pygmaeus were emitted at levels below the
detection ability of the analytical instruments used. The volatiles
detected in the headspace of tomato plants in this study primarily
consisted of terpenoids (60 out of 70 compounds), which play a minor
role in the attraction of mirid predators (Silva et al. , 2021).
Feeding by T. vaporariorum is also known to induce the emission
of the ester methyl salicylate (MeSA) by tomato plants (López et
al. , 2012; Conboy et al. , 2020), which was not detected in the
headspace analysis of the current study. MeSA is an important compound
for the attraction of mirids (Silva et al. , 2021), and its
emission can be modulated after exposure to far-red light (Cortéset al. , 2016).
The results of this study also show that exposure to supplemental
far-red light increases the reproduction of M. pygmaeus in both
the presence and absence of prey. These results support the hypothesis
that exposure to low R:FR and the subsequent downregulation of defences
increases plant quality for M. pygmaeus . Although no significant
interaction between light treatment and whitefly presence was found, the
effect of far-red light on predator reproduction appears to be stronger
in the presence of whitefly prey, which might be related to increased
prey availability. Along with the increased reproduction of M.
pygmaeus , we also found increased numbers of T. vaporariorum in
the +FR treatment, which corresponds with previous findings (Meijeret al. , 2022). This indicates that supplemental far-red light can
stimulate M. pygmaeus population development both directly
through changes in plant quality and indirectly by supporting higher
herbivore densities. In previous studies, supplemental far-red light
increased the population growth of spider mites (T. urticae ),
allowing a more rapid population growth of the predatory mitePhytoseiulus persimilis and leading to a stronger suppression of
the herbivore (Meijer et al. , 2023). Unfortunately, the time
frame of the current study was too short to study population development
in M. pygmaeus and to make a proper assessment on the
effectiveness of whitefly control by M. pygmaeus . However, the
observed results provide promising opportunities for the use of far-red
light for biological control in greenhouses.
The use of far-red LEDs is an application in horticulture to manipulate
plant morphology and reproductive development and thereby increase crop
yield (Demotes-Mainard et al. , 2016), but at the same time, it
can lead to an increased pest pressure (e.g. Meijer et al. ,
2022). Cortés et al. (2016) suggested that far-red light might
have a role in biological control by increasing the attraction of
natural enemies. Although our current results indicate that changes in
R:FR neither enhance nor interfere with the attraction of the predatory
bug M. pygmaeus , supplemental far-red light might still stimulate
biological control by allowing a more rapid build-up of the predator
population, both in the presence and absence of pests. Macrolophus
pygmaeus is often used in preventative release strategies by
maintaining predator populations on banker plants, but its successful
establishment depends on the species of banker plant, the crop and the
availability of supplemental food (Bresch et al. , 2014; Messelinket al. , 2015; Sanchez et al. , 2021). Supplemental far-red
light might contribute to increased establishment of M. pygmaeusduring preventative release.
We conclude that exposure to low R:FR conditions could indirectly lead
to increased plant protection against arthropod herbivores through
changes in plant defences and predator-prey dynamics, with possible
applications to stimulate biological control in horticulture. However,
the tritrophic interactions between plants, arthropod herbivores andM. pygmaeus are complex, and more research is required to
understand how far-red light shapes the ecological interactions between
trophic levels. For example, plant feeding by mirid predators can induce
plant defences and increase resistance to herbivores (Pappas et
al. , 2015; Bouagga et al. , 2019; Pérez-Hedo et al. , 2022;
Silva et al. , 2022). Herbivores also avoid plants that have been
exposed to M. pygmaeus (Zhang et al. , 2019). On the other
hand, too high densities of M. pygmaeus can cause plant damage
through direct feeding or the transmission of viruses (Castañé et
al. , 2011; Moerkens et al. , 2016; Moerkens et al. , 2017;
Sanchez et al. , 2018). As far-red light exerts strong effects on
plant defensive signalling, it has the potential to influence both
direct and indirect interactions between all three trophic levels. A
proper understanding of how these interactions are affected by changes
in R:FR is required to adequately assess the efficiency of supplemental
far-red light to stimulate biological control.
Acknowledgements
This research was funded by the Dutch Research Council (NWO), project
number ALWGR.2017.004 with financial contributions by Biobest Biological
Systems, Enza Seeds, Florensis, Schoneveld Breeding, WPK Vegetable
Plants, and Heliospectra through the NWO.
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