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RESEARCH PAPER
How maize root volatiles affect the efficacy of entomopathogenic
nematodes in controlling the western corn rootworm?
Ivan Hiltpold •Stefan Toepfer •Ulrich Kuhlmann •
Ted C. J. Turlings
Received: 10 April 2009 / Accepted: 25 November 2009 / Published online: 22 December 2009
ÓBirkha
¨user Verlag, Basel/Switzerland 2009
Abstract Because the ferocious maize pest Diabrotica
virgifera virgifera LeConte can adapt to all currently used
control strategies, focus has turned to the development of
novel, more sustainable control methods, such as biological
control using entomopathogenic nematodes (EPN). A good
understanding of the biology and behaviour of these
potential control agents is essential for their successful
deployment. Root systems of many maize varieties emit
(E)-b-caryophyllene (EbC) in response to feeding by larvae
of the beetle D. v. virgifera. This sesquiterpene has been
shown to attract certain species of EPN, thereby enhancing
their control potential. In this study, we tested the effect
of this root-produced volatile on the field efficacy of the
three EPN Heterorhabditis bacteriophora, Heterorhabditis
megidis and Steinernema feltiae against D. v. virgifera
larvae in southern Hungary. By comparing beetle emer-
gence and root damage for two maize varieties, one that
emits EbC and one that does not, it was found that root
protection by H. megidis and S. feltiae was higher on the
emitting variety, but this was not the case for H. bacte-
riophora. Overall, all three nematode species showed good
control potential. We conclude that, if properly applied and
in combination with the right maize variety, the release of
these nematodes can be as effective as other control
methods.
Keywords Heterorhabditis megidis Heterorhabditis
bacteriophora Steinernema feltiae Diabrotica
virgifera virgifera Zea mays Tritrophic interaction
Crop protection Biological control Root exudates
Nematode attraction
Introduction
Since the domestication of maize, Zea mays (L.), about
5,000–7,000 years ago (Piperno and Flannery 2001; Sluyter
and Dominguez 2006), this crop has been targeted by a
variety of arthropod pests, often causing tremendous yield
losses (Oerke 2006). In nature, plants have evolved various
defence strategies to fend off their herbivorous attackers
either directly (Baldwin and Preston 1999; Agrawal 1998;
Dicke et al. 2003; Karban et al. 1997; Karban and Baldwin
1997; Schoonhoven et al. 1998) or indirectly (Agrawal
1998; Dicke and Sabelis 1998; Dicke et al. 2003; Turlings
and Wa
¨ckers 2004). Direct defence traits of plants com-
prise physical or chemical barriers, whereas indirect
defences consist of the attraction and maintenance of the
herbivore’s natural enemies by providing shelter and/or
food (Janzen 1966; Stapley 1998) and/or the emission of
inducible volatile organic compounds (Dicke et al. 2003;
Turlings and Benrey 1998; Turlings and Wa
¨ckers 2004).
For maize, the attractiveness of such herbivore-induced
plant volatiles to natural enemies of herbivores has been
demonstrated in both laboratory and field experiments
(Turlings et al. 1990; Bernasconi et al. 1998; Hoballah and
Turlings 2005). For instance, green leaf volatiles, as well as
I. Hiltpold T. C. J. Turlings (&)
FARCE Laboratory, University of Neucha
ˆtel, Emile-Argand 11,
cp158, 2009 Neucha
ˆtel, Switzerland
e-mail: ted.turlings@unine.ch
S. Toepfer
CABI Europe, c/o Plant Health Service, Rarosi ut 110,
6800 Hodmezovasarhely, Hungary
U. Kuhlmann
CABI Europe-Switzerland, Rue des Grillons 1,
2800 Dele
´mont, Switzerland
Chemoecology (2010) 20:155–162
DOI 10.1007/s00049-009-0034-6 CHEMOECOLOGY
terpenoids such as monoterpenes, sesquiterpenes and
homoterpenes, have been found to attract parasitoids
aboveground (D’Alessandro and Turlings 2005; Hoballah
and Turlings 2005; Schnee et al. 2006).
Recently it was found that roots also are able to recruit
belowground enemies of soil dwelling herbivorous insects
by releasing volatile signals. These volatiles can attract
entomopathogenic nematodes (EPN) (van Tol et al. 2001;
Boff et al. 2001; Bertin et al. 2003; Rasmann et al. 2005;
Rasmann and Turlings 2008; Degenhardt et al. 2009),
predatory mites (Aratchige et al. 2004) and even parasit-
oids (Neveu et al. 2002). Maize roots fed upon by larvae of
Diabrotica virgifera virgifera LeConte (western corn
rootworm, WCR, Coleoptera: Chrysomelidae), one of the
most destructive maize pests worldwide (Miller et al. 2005;
Vidal et al. 2005), release the sesquiterpene (E)-b-caryo-
phyllene (EbC). EbC diffuses well in soil (Hiltpold and
Turlings 2008) and plays an important role in the recruit-
ment of the EPN Heterorhabditis megidis Poinar
(Rhabditida: Heterorhabditidae) (Rasmann et al. 2005),
which is highly virulent to WCR larvae (Kurtz et al. 2009).
Two other species of EPNs, Heterorhabditis bacteriophora
Poinar and Steinernema feltiae Filipjev, are also promising
candidates as biological control agents against WCR larvae
(Kurtz et al. 2009; Toepfer et al. 2005), but it is unknown if
their host finding ability is also improved by attraction to
belowground signals.
The aim of the current study was to determine the rel-
ative importance of EbC emission by WCR-damaged
maize roots for the efficacy of H. bacteriophora,H. megidis,
and S. feltiae in controlling WCR larvae under field con-
ditions. For this purpose, the three nematode species were
released at two different time points in separate, WCR-
infested maize plots in Hungary. The results of the
field study prompted us to also conduct additional labora-
tory assays to test the apparent lack of attraction of
H. bacteriophora towards EbC. We discuss the control
potential of the tested nematode species and the importance
of choosing the right maize variety to fully exploit this
potential.
Materials and methods
Field sites and maize varieties
The study was carried out in four maize fields (referred to
as fields A to D) in Csongrad County in southern Hungary
in 2005 and 2006 (Table 1). All fields contained an
experimental section that had been planted with non-host
plants of WCR the year before to ensure the initial absence
of this pest in the experimental plots. Experimental fields
were divided in two plots, the first planted with the variety
Magister (UFA Semences, Bussigny, Switzerland) that
emits EbC after WCR feeding (Hiltpold 2008) and the
second with the variety Pactol (Syngenta, Budapest, Hun-
gary) that does not emit EbC (Rasmann et al. 2005). The
seeds were sown between late April and early May
(Table 1). All maize seeds were sown in rows with plant
spacing of 15 cm and row spacing of 75 cm. The fields
were treated once with 0.16 l of the herbicide Merlin SC
(75% Izoxaflutol, Bayer Crop Science) per hectare when
maize was at the 3–5 leaves stage. No insecticides were
applied.
Table 1 Characteristics of the study fields in southern Hungary and the timing of EPN application
Field A B C D
Location Northwest of Hodmezovasarhely North of Szatymaz North of Szatymaz Hodmezo-vasarhely
Coordinates N 46°26.022 N 46°20.945 N 46°20.945 N 46°25.998
E20°20.143 E 20°00.574 E 20°00.574 E 20°20.348
Elevation (m) 83 87 87 83
Size (ha) 0.5 0.2 0.3 0.2
Soil bulk density (g/cm
3
) 1.04 ±0.13 1.4 ±0.13 1.7 ±0.07 1.1 ±0.13
Soil moisture (wt%)* 17.2 ±1.1 11.6 ±0.3 7.1 ±2.5 18.5 ±2.1
Sand content (%) 36 85 85 14
Loam content (%) 34 5 5 44
Clay content (%) 30 10 10 42
pH (H
2
O) 8.3 8.4 8.4 8.3
Maize sown 25 April 2005 8 May 2005 8 May 2006 28 April 2005
EPN applications 25 April 2005 8 May 2005 8 May 2006 28 April 2005
14 June 2005 15 June 2005 7 June 2006 14 June 2005
Significance is indicated by asterisks
156 I. Hiltpold et al.
Entomopathogenic nematodes
Three EPN species were used in this study: (1) a cross of
European and US strains of Heterorhabditis bacteriophora
Poinar provided from liquid culture by e-nema GmbH
(Raisdorf, DE), (2) the NL-HW79 strain of H. megidis
Poinar, Jackson & Klein from The Netherlands, re-isolated
from Swiss soils and provided from a semi-liquid culture
by Andermatt Biocontrol AG (CH), and (3) a cross of
European strains of Steinernema feltiae Filipjev provided
from liquid culture by e-nema GmbH. H. bacteriophora
and S. feltiae were shipped in clay from the producer to the
experimental sites, and H. megidis was shipped in ver-
miculite. All EPNs were stored in their shipping material at
7–9°C in darkness until use. About 2–3 h prior to appli-
cation, EPNs together with the carrier material were diluted
in tap water. Before application, aliquots of EPNs were
taken to determine the quality of the shipment batches. For
this purpose Galleria mellonella L. (Lepidoptera: Pyrali-
dae) larvae were exposed to nematodes in plastic cups
(40 mm diameter, 60 mm height). Each cup was filled with
200 g of 10% moist sterilised sand to which five larvae and
100 infective juvenile nematodes were added. Three rep-
licates per EPN shipment batch were used for this assay.
After 1 week in darkness at 22°C, mortality of 80–100%
was found for all EPN batches, which was considered
sufficient for use.
Diabrotica virgifera virgifera
WCR eggs were obtained from eggs laid by field-collected
beetles from southern Hungary (for procedures see Singh
and Moore 1985). Eggs were kept in diapause in moist sand
at 6–8°C. The diapause of WCR eggs was broken in early
April by transferring them to a climate chamber at 25°C for
3 weeks. The sand was sieved through a 250 lm mesh to
recover the eggs. The eggs were then mixed into a solution
of water and 0.15% agar in order to obtain an egg sus-
pension of 38 eggs/ml. Maize plants of each field were
infested in early May (1–3 leaf stage) with the suspension
of viable and ready-to-hatch eggs. Using a standard pipette
(Eppendorf Company, Hamburg, Germany), 2 ml of the
egg suspension was applied into each of two 12 cm deep
holes at a distance of 5–8 cm from either side of the maize
plant (*150 eggs/plant). The larvae were expected to
hatch by mid-to-late May and to reach the second larval
instar in June (Toepfer and Kuhlmann 2006).
Experimental setup and EPN application
Each of the four fields contained two plots of at least 14
rows of Magister and Pactol plants. Seven groups of six to
seven maize plants were randomly selected from either
the 3rd, 6th, 9th or 12th row of each section, thereby
ensuring buffer rows between experimental groups. The
three different entomopathogenic nematode species were
applied at two different times (early: during sowing in
April/May; late: in June, see Table 1). Thus six groups,
each treated with one nematode species at one particular
date, were distributed over one experimental plot. The
seventh group served as control and was not treated with
nematodes.
The EPN suspensions were poured by hand in a con-
tinuous stream into a 10 cm deep groove dug into the soil
directly along each row. When applied at the earlier date in
April/May, this was done at the same time as maize was
hand-sown. Suspended in 0.2 l of water, 2.1 910
5
±0.07
SD infective juvenile nematodes were applied per metre.
At the later EPN application date in June, they were sus-
pended in 0.2 l of water and 2.6 910
5
±0.07 SD
infective juvenile nematodes were applied per metre. All
applications were carried out in the evening or during
cloudy afternoons to avoid harmful UV radiation.
Effects of EPN application, EPN species, and maize
variety on WCR adult emergence
Each of the 14 experimental groups (6–7 plants) of fields A
to C (because of technical problems, emergence was not
assessed in field D) was covered with a fine-mesh screen
cage (1.3 m height 90.75 m width 91.5 m length, maize
plants had been cut to a height of 1 m). WCR adult
emergence within these cages was recorded weekly
between 20 June and 16 August 2005 and between 27 June
and 16 August 2006. Total adult emergence was normalised
to 100 eggs per plant. Nematode efficacy was calculated as
percentage reduction in WCR adults compared to their
untreated controls (corrected efficacy% =(1 -WCR in
treated plots/WCR in the control) 9100) (Abbott 1925).
Effect of application time, nematode species, and maize
variety on root damage by D. v. virgifera
In mid-September, after adult emergence was completed,
field cages were removed and all plants of each group were
dug up. Plants from field D were also used for this part of
the experiment. Soil and other particles were removed from
the roots using a high-pressure water sprayer. Damage was
rated according to Oleson’s Node Injury Scale from 0.00 to
3.00 with 0.00 being no damage and 3.00 being three or
more damaged root nodes (Oleson et al. 2005).
The efficacy of EPNs was calculated as percentage
reduction in root damage compared to the respective con-
trol groups that did not receive any nematodes (corrected
efficacy % =(1 -root damage in treated plots/root
damage in the control) 9100) (Abbott 1925).
Controlling rootworms with nematodes by exploiting root signals 157
Olfactometer assays
Following the methodology developed by Rasmann et al.
(2005), attraction of H. bacteriophora was assessed in six
belowground olfactometers filled with moist sand. EPNs
had to choose between a Pactol maize plant damaged by
four WCR larvae, a healthy Pactol maize plant and four
empty control pots. After 1 day of exposure, the olfac-
tometers were disassembled, the sand from each of the six
connectors was placed in a Baermann extractor (Hass et al.
1999), and the next day, nematodes were counted under a
microscope on a counting plate.
Statistical analyses
The effect of the tested parameter (EPN species, application
periods and maize varieties) on reduction of WCR emer-
gence and root damage was analysed using a three-way
ANOVA. Then EPN species, maize varieties and applica-
tion periods were compared using Tukey’s post hoc tests.
All statistical tests of field data were performed using
SAS 9.1 with a three-way ANOVA (GLM procedure) with
EPN species, application period, maize variety, EPN spe-
cies 9application period, EPN species 9maize variety,
application period 9maize variety and EPN species 9
application period 9maize variety as independent vari-
ables and WCR emergence (relative to control) and node
injury rate (relative to control) as dependent variables.
Differences were analysed using LSMEANS with Tukey–
Kramer adjustments for the Pvalues (SAS 9.1).
The nematodes’ behavioural responses in the six-arm
olfactometer were tested with a log-linear model. The entity
computing a repetition in the statistical analysis corresponds
to the response of a group of 2,000 nematodes released,
which was shown to follow a multinomial distribution. As
the data did not conform to simple variance assumptions
implied in using the multinomial distribution, we used quasi-
likelihood functions to compensate for the over dispersion of
nematodes within the olfactometer (Turlings et al. 2004).
The model was fitted by maximum quasi-likelihood esti-
mation in the software package R (http://www.R-project.org),
and its adequacy was assessed through likelihood ratio
statistics and examination of residuals (Turlings et al. 2004).
Results
Effect of EPN application, EPN species, and maize
variety on WCR adult emergence
All tested EPN species significantly reduced the percentage of
emerging D. v. virgifera, and the time of application had no
major effect on their respective efficacies (Fig. 1;Table2).
Overall, WCR emergence was significantly different
between the two maize varieties (Fig. 1; Table 2) with a
lower emergence from rows with the EbC-emitting Magister
than from rows with the non-emitting Pactol. H. megidis
reduced WCR emergence 2.5-fold more in Magister plots
than in Pactol plots (P\0.001). There was no significant
difference in the efficacy of H. bacteriophora and S. feltiae
between the two maize varieties (Fig. 1,P=0.08 and
P=0.12, respectively).
On average, the reduction in WCR emergence was
higher for plots treated with H. bacteriophora than for plots
treated with either H. megidis or S. feltiae (Fig. 1; Table 2).
This was reflected in an average WCR emergence per
plant, which was 0.65 and 0.75 adult WCR per 100 eggs for
the Magister rows treated with H. megidis and S. feltiae,
respectively, versus 0.28 adults for rows treated with H.
bacteriophora (1.13 WCR adults emerged per plant from
Magister control rows). In Pactol rows, 0.3 WCR adults
emerged when treated with H. bacteriophora, whereas on
average 0.8 WCR adults emerged from Pactol rows when
treated with either H. megidis or S. feltiae (1.60 WCR
adults emerged per plant from Pactol control rows).
Effect of EPN application, EPN species, and maize
variety on root damage by D. v. virgifera
All tested EPN species significantly reduced root damage
caused by WCR larvae, and the time of application had no
0
10
20
30
40
50
60
70
80
90
100
magist er pactol magis ter pactol magister pactol
H. bacteriophora H. megidis S. feltiae
% of reduction of WCR adults emergence
BBA
a
a
a
b
a
a
Maize varieties and EPN species
Fig. 1 Comparison of the reduction of WCR emergence relative to
the untreated controls in maize fields with the EbC-emitting Magister
variety and the non-emitting Pactol variety (pooled data for the two
release dates). Uppercase letters indicate statistical differences
between the three EPN species (Tukey post hoc test, H. bacteriophora
vs. H. megidis P \0.001, H. bacteriophora vs. S. feltiae P \0.001
and H. megidis vs. S. feltiae P =0.70). Lowercase letters above bars
indicate statistical differences between maize varieties within each
EPN species (Tukey post hoc test, H. bacteriophora Magister vs.
Pactol P=0.08, H. megidis Magister vs. Pactol P\0.001, S. feltiae
Magister vs. Pactol P=0.12). Error bars represent the standard error
of mean
158 I. Hiltpold et al.
effect on their efficacies (Fig. 2; Table 3). However, the
efficacy of the nematodes was different for the two maize
varieties (Fig. 2; Table 3). This difference was due to H.
megidis and S. feltiae, which reduced root damage to a
higher degree in rows with the EbC-emitting Magister than
in rows with the non-emitting Pactol (Fig. 2; Table 3).
Olfactometer assays
When offered a choice between volatiles emitted by a
WCR-damaged Pactol maize plant or a healthy Pactol
plant, H. bacteriophora preferred the arm with the pest
feeding on the roots (Fig. 3, ANOVA, F
2,33
=6.6,
P\0.001). Surprisingly, the healthy plants were found to
be repellent, evidenced by the fact that fewer nematodes
were collected from arms connected to pots with healthy
plants than from arms connected to the control pots with
sand only.
Discussion
The results from the field experiment confirm that the
choice of maize variety and/or nematode species can sig-
nificantly affect the control efficacy of EPNs. Kurtz et al.
(2009) had already compared the efficacy of the three
nematode species against WCR in a laboratory study and
also reported that WCR was most susceptible to H. bac-
teriophora. EPN persistence in the soil has been shown to
rapidly decrease with time (Kurtz et al. 2007). It was,
therefore, surprising to find that there was no difference in
the efficacy of EPN between the two application periods
(during sowing in April/May or later in June) (Tables 1,2).
Apparently some early applied nematodes persisted,
probably by producing a new generation on alternative
hosts that resulted in sufficiently high abundance to reduce
the later hatching WCR population. For inundative bio-
logical control strategies, it remains essential to find the
optimal dose and release timing (Fenton et al. 2002).
The choice of the right maize variety seems particularly
important for two of the three EPN species investigated,
H. megidis and S. feltiae (Figs. 1,2). H. megidis was more
effective near the EbC-emitting Magister variety was
expected from the results of previous studies (Rasmann
et al. 2005; Rasmann and Turlings 2007). However, that
this was also the case for S. feltiae was surprising,as
S. feltiae is considered to mainly use the so-called
ambusher (nictating) foraging strategy. Although S. feltiae
is known to actively move through soil (Grewal et al. 1994;
Lewis 2002),it never responded to any cues in olfactom-
eter experiments (Rasmann and Turlings 2008; personal
observations), suggesting that they were not very mobile or
not responding to the compounds tested. S. feltiae has been
shown to be effective against WCR (Kurtz et al. 2009).
Yet, the trend of reduced of adult emergence and a sig-
nificant reduction of root damage for the Magister plants
Table 2 Effects of EPN species, application period and maize variety on WCR adult emergence (% efficacy relative to control) according to the
three-way ANOVA
Factor Sum of squares df Mean of squares FP
EPN species 2.10 2 1.05 14.41 \0.001***
Application period 0.02 1 0.02 0.32 0.567
Maize variety 0.55 1 0.55 7.57 0.007**
EPN species 9application period 0.05 2 0.02 0.30 0.735
EPN species 9maize variety 0.03 2 0.02 0.23 0.795
Application period 9maize variety 0.00 1 0.00 0.02 0.863
EPN species 9application period 9maize variety 0.04 2 0.02 0.23 0.788
Significance is indicated by asterisks
0
10
20
30
40
50
60
70
80
90
100
magist er pactol m agister pact ol magist er pactol
H.bact eriophora H. megi dis S. felt iae
% of reduction in root damage
ABBA
aa
a
b
a
b
Maize varieties and EPN species
Fig. 2 Comparison of the reduction of root damage relative to the
untreated controls in maize fields with the EbC-emitting Magister
variety and with a non-emitting Pactol variety (pooled data for the
two release dates). Uppercase letters indicate statistical differences
between the three EPN species (Tukey post hoc test, H. bacteriophora
vs. H. megidis P =0.012, H. bacteriophora vs. S. feltiae P=0.226
and H. megidis vs. S. feltiae P =0.480). indicates statistical
differences between maize varieties within each EPN species (Tukey
post hoc test, H. bacteriophora Magister vs. Pactol P=1.00, H.
megidis Magister vs. Pactol P=0.042, S. feltiae Magister vs. Pactol
P=0.024). Error bars represent the standard error of mean
Controlling rootworms with nematodes by exploiting root signals 159
after S. feltiae application suggests that this nematode also
responds to this root signal under field conditions. How-
ever, S. feltiae foraging efficiency might also have been
affected by other aspects, such as WCR larval behaviour,
affected by Magister plants. The combined results from
these studies suggest that S. feltiae foraging behaviour is
strongly determined by the media in which it has to ambush
or cruise.
The effectiveness of H. bacteriophora was not affected
by EbC emission from WCR-damaged maize roots
(Figs. 1,2) suggesting a random and uniformed spread of
the nematode in the field. However, when offered a Pactol
plant infested with WCR larvae (no emission of EbC) or a
healthy Pactol plant, H. bacteriophora significantly
migrated more toward the damaged plant (Fig. 3), imply-
ing that it uses other chemical cues for host location. These
cues might come from the plants, but also from the hosts
themselves. These and other (Rasmann and Turlings 2008)
olfactometer assays show that healthy maize roots are
repellent to H. bacteriophora. This could imply that it uses
a unique and potentially highly efficient host location
strategy. It remains unknown what signals allow H. bac-
teriophora to make the distinction, but it has been shown
that it is sensitive to long-chain alcohols and possibly
other, more insect specific, volatiles (O’Halloran and
Burnell 2003).
Current WCR management strategies involve crop
rotation and the use of insecticides (Levine and Oloumi-
Sadeghi 1991), but WCR has shown the ability to evolve
resistance to both these methods (Ball and Weekman 1962;
Meinke et al. 1998; Zhou et al. 2002; Levine et al. 2002;
O’Neal et al. 2001). Moreover, soil insecticides that are
still effective pose environmental and human health risks.
Recently, genetically modified maize expressing Cry3
proteins, a Bacillus thuringiensis toxin against WCR lar-
vae, has become available on US market (Moellenbeck
et al. 2001). Bt maize appears to be effective against WCR,
reducing populations of by 80–96% in the field/lab (Sieg-
fried et al. 2005; Storer et al. 2006; Vaughn et al. 2005).
This high but incomplete efficacy can be expected to lead
to rapid resistance to Bt-maize in WCR populations. While
some models estimate that resistance will not occur until at
least 20 years after farmers start growing Bt maize with
5–10% refuge (Storer et al. 2006), others have shown that
resistance developed within three generations under
greenhouse conditions (Meihls et al. 2008).
In the current study, we show that the synergetic effect
of using the appropriate EPN species combined with
attractive maize varieties can result in a control of WCR
that is almost as effective as the use of pesticides or Bt
maize (Figs. 1,2). WCR populations are unlikely to be able
to develop resistances against EPNs. Moreover, EPNs are
able to infect and kill all the larval instars of WCR
(Jackson and Brooks 1995, Kurtz et al. 2009; Toepfer et al.
2005), whereas transgenic maize seems to be efficient only
against the first instar (Oyediran et al. 2005). Neonate
WCR larvae may survive on neighbouring weed roots and
as second instar larvae could move back to the Bt maize
Table 3 Effects of EPN species, application period and maize variety on WCR’s root damage (% efficacy relative to control) according to the
three-way ANOVA
Factor Sum of squares df Mean of squares FP
EPN species 16.13 2 8.07 4.07 0.017*
Application period 0.00 1 0.00 0.00 0.957
Maize varieties 15.25 1 15.25 7.81 0.005**
EPN species 9application period 10.04 2 5.02 0.41 0.663
EPN species 9maize variety 1.56 2 1.56 2.53 0.080
Application period 9maize variety 1.63 1 0.82 0.78 0.377
EPN species 9application period 9maize variety 0.44 2 0.22 0.11 0.894
Significance is indicated by asterisks
0
100
200
300
400
500
600
WCR infested healthy control pots
Pactol plant
a
b
c
EPN per arm
Fig. 3 H. bacteriophora is attracted by EbC non-emitting plants.
When offered choice between a healthy, a WCR damaged Pactol
plant or sand, this nematode species is significantly attracted towards
the damaged plant even if no EbC is emitted (ANOVA, F
2,33
=6.6,
P\0.001). The healthy plant appears to repel H. bacteriophora
compared to the control pots filled with sand only. Letters indicate
statistical differences. Error bars represent the standard error of mean
160 I. Hiltpold et al.
roots on which they can survive (Moeser and Vidal 2004,
Oyediran et al. 2005). In contrast, EPN will also be
effective against WCR larvae on roots of other plants
(Christen et al. 2007; Gaugler and Campbell 1991; Rae
et al. 2006; Ramos-Rodriguez et al. 2007).
Conclusion
In conclusion, the efficacy of the tested EPN species in
controlling WCR populations is promising. Based on our
findings, it should be possible for farmers to match their
crops with the most effective nematode. Further studies are
needed to take optimal advantage of the biology and
behavioural plasticity of EPN to maximise their persistence
and their responses to plant-provided signals in the soil.
Acknowledgments This work was possible thanks to the hospitality
of the Plant Health Service in Hodmezovasarhely in Hungary, offered
by Ibolya Zseller, Jozsef Gavallier, Kataline Buzas, Erzsebet
Dormannsne, Piroska Szabo, Andras Varga and others. We would
also like to thank the summer students Bobe Kovacs, Benedikt Kurtz,
and Ferenc Koncz for their help in field work, as well as Arne Peters
(e-nema GmbH, Germany) and Erich Frank (Andermatt Biocontrol
AG, Switzerland) for providing nematodes. This study was funded by
the CTI Innovation and Technology Fund of Switzerlandin collaboration
with Landi REBA (Basel, Switzerland) and e-nema GmbH (Raisdorf,
Germany).
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