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doi: 10.5958/2249-4677.2018.00005.1
Phytopathogenic Mollicutes
Vol. 8 (1), June 2018, 32-39
Corresponding author e-mail: Simone S. Prado (simone.prado@embrapa.br)
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Research Article
Role of Dalbulus maidis (Hemiptera: Cicadellidae) gender on maize
bushy stunt phytoplasma transmission
Maria Cristina Canale1, João Roberto Spotti Lopes2, Cristiano Nunes Nesi1 and Simone de Souza Prado3
1Agricultural Research and Rural Extension Company of Santa Catarina State (Epagri), Laboratory of Phytopathology and
Entomology, Rua Ferdinando Ricieri Tusseti, S/N, 89803-904, Chapecó, SC, Brazil
2Luiz de Queiroz College of Agriculture (ESALQ), University of São Paulo (USP), Department of Entomology and Acarology,
Avenida Pádua Dias, 11, 13418-900, Piracicaba, SP, Brazil
3Brazilian Agricultural Research Corporation (Embrapa) - Environment, Rodovia SP 340, KM 127,5, S/N, 13820-000,
Jaguariúna, SP, Brazil
Received: May 20, 2018; Accepted: May 30, 2018
Abstract
There is a behavioral difference between male and female of insect vectors of plant pathogens that may reflect on the transmission
of such pathogens.
Dalbulus maidis
is the leafhopper vector of the maize bushy stunt (MBS) phytoplasma. In this work, sex
ratio of
D. maidis
collected directly from the maize whorl or with yellow sticky cards in a field plot in Piracicaba, SP (Brazil) was
evaluated during the first month of the crop development. The natural MBS infection of the captured leafhoppers was assessed
by PCR and the disease incidence was visually evaluated in the maize plots. Female
D. maidis
were more easily captured from
the maize whorl, whereas male leafhoppers were more abundant in the cards. The incidence of MBS diseased plants in the plot
was 21.2%. MBS phytoplasma was detected in 8% of the captured females whilst 2% of males carried the prokaryote. It is possible
that the infected leafhoppers in the early stage of the crop contribute to the final disease incidence. Additionally, the acquisition
and transmission rates to maize seedlings by males and females of
D. maidis
, virgin or mated, was studied. Acquisition rate of
MBS phytoplasma was not significantly different between male and female leaf hoppers, however, female, either virgin or mated,
transmitted the phytoplasma in a higher rate than males. The behavior of females of spending more time on the plant feeding
may explain the higher natural infection of MBS phytoplasma in field condition and the increased transmission rate observed
in the experiment.
Keywords:
Zea mays
, insect-borne bacteria, insect vector, phytoplasma transmission, leafhopper abundance
Introduction
The corn leafhopper,
Dalbulus maidis
(DeLong and
Wilcott, 1923) (Hemiptera: Cicadellidae) is one of the
most important pests to maize crop in the American
continent, where it is broadly distributed from
southeastern and southwestern United States to
Argentina (Triplehorn and Nault, 1985; Virla
et al.
, 2013).
D. maidis
transmits the pathogens associated with the
corn stunt complex due to
Spiroplasma kunkelii
, the
maize bushy stunt
(MBS) phytoplasma, and the
Maize
rayado fino virus
- MRFV (Nault, 1980; Bradfute
et al.
,
1981). The maize bushy stunt disease was reported in
Brazil in the 1970ies. At that time, although this disease
was considered a secondary problem in maize crop, it
was remarkably threatening when it was occuring in
high percentage of plants (Costa
et al.
, 1971; Bedendo
et
al.
, 2000). Since its first report, the seasonality of MBS
disease has been registered in Brazil along the years, with
records of serious outbreaks. From 2016 to the date, high
levels of MBS disease were registered in the Brazilian
states of Mato Grosso, Bahia, Goiás and Minas Gerais,
with total yield reduction of 90%. The disease
symptoms are more evident later in the crop cycle and
are characterized by the reddening of leaves, ear
proliferation and plant stunting (Orlovskis
et al.
, 2017).
Phytoplasmas are obligate biothrophic plant
pathogenic bacteria that do not have a cell wall and
belong to the class
Mollicutes
. These pathogens reside
in the sieve tube elements and spread systemically
throughout the plant (Christensen
et al.
, 2004; Marcone,
2014). Phytoplasmas require that the vector feed over
periods of hours in infected plants to be acquired and
successively transmitted to another plant (Hogenhout
et al.
, 2008). MBS is a ‘
Candidatus
Phytoplasma asteris’-
related phytoplasma, classified in the subgroup 16SrI-B
(Lee
et al.
, 2004). It is very efficiently transmitted by the
maize specialist corn leafhopper in a persistent
propagative manner. The pathogen circulates and
multiplies within the vector and when it has reached
the salivary glands, the leafhopper become competent
to inoculate the bacteria into the phloem of a healthy
plant (Moya-Raygosa and Nault 1998; Sugio
et al.
, 2011;
Orlovskis
et al.
, 2017).
MBS disease occurrence in maize fields can be better
understood by the knowledge of the complex
pathosystem enclosing the MBS phytoplasma, its
leafhopper vector and the plant host (Power, 1987). There
is a known behavioral difference between male and
female of leafhoppers vectors of plant pathogens.
Females usually spend more time on the host plant
feeding or ovipositing, whereas males exhibit a more
active flight behavior in searching for females to
copulate (Heady and Nault 1985; Hunt and Nault 1991;
Larsen and Nault 1994). Moreover, it has already been
reported that vector gender can influence disease
transmission. Antolínez
et al.
, (2016) verified a higher
transmission efficiency of ‘
Candidatus
Liberibacter
solanacearum’ by the females of the carrot psyllid
Bactericera trigonica
(Hemiptera: Psylloidea), due to
higher frequency of phloem contacts during feeding
activity. Females of the aster leafhopper,
Macrosteles
quadrilineatus
(Hemiptera: Cicadellidae), infected with
the aster yellows phytoplasma, were more likely to
transmit the pathogen to lettuce plants (Beanland
et al.
,
1999).
In spite of already published research on behavior of
males and females of
D. maidis
, it has not been
approached yet if such sex-based behavioral difference
could influence MBS phytoplasma transmission. The
objective of this work was to clarify the role of
D. maidis
gender on transmission of MBS phytoplasma. The
population of
D. maidis
and the sex ratio of this insect
was assessed in a maize field by both directly counting
the leafhoppers from the maize whorl and by using
yellow sticky cards. The MBS phytoplasma natural
infection of the captured male and female of
D. maidis
was evaluated, and the disease incidence was estimated
in the field. Furthermore, the acquisition and
transmission rates by gender of
D. maidis
was
experimentally evaluated. The contribution of male
and female
D. maidis
in MBS disease epidemiology is
discussed.
Materials and methods
Dalbulus maidis population
Population of adult
D. maidis
was assessed weekly in a
plot of maize established in January 2000, in Piracicaba,
Brazil (-22.713287, -47.625723).
D. maidis
was sampled
by using two methods. The first one, carried out at 8, 15,
22, 29 and 36 d.a.e. (days after emergence), consisted in
the insect collection from 40 whole maize plants
randomly chosen in the plot by covering the plants with
a transparent plastic bag. Plants where shacked and the
leafhoppers present at the whorl flew into the bag. The
plastic bag was removed with the insects inside and
closed with a rubber band, taken to the lab where the
captured insects were anesthetized with CO2 fo r
counting and gender separation (sexing). Then, the
leafhoppers were transferred to 1.5 ml microtubes
containing 70% alcohol and placed into the freezer for
further evaluation of phytoplasma presence by PCR
tests. For the other sampling method five yellow sticky
cards measuring 10 x 24.5 cm (Biocontrole®, Indaiatuba,
SP), were installed 8 d.a.e. 0.50 m above the plants. As
the plants were getting taller, the yellow cards were
installed at higher points, at 1 and then 1.5 m above soil,
in order to follow the crop growth. Yellow cards were
replaced weekly during 5 weeks. Fifty
D. maidis
were
removed from each yellow cards, sexed and stored in
freezer for PCR analysis.
Dalbulus maidis sex ratio and MBS phytoplasma natural
infectivity and disease incidence in the field
All the leafhoppers captured in the maize whorl with a
bag at 8, 15 and 22 d.a.e. were anesthetized with CO2 fo r
counting and sexing. Those insects were further assessed
for MBS phytoplasma natural infection. After counting
the captured insects by the yellow sticky traps which
Phytopathogenic Mollicutes, Vol. 8 (1), June 2018 33
Role of Dalbulus maidis (Hemiptera: Cicadellidae) gender on maize bushy stunt phytoplasma transmission
Maria Cristina Canale et al.
Phytopathogenic Mollicutes, Vol. 8 (1), June 201834
remained in the field in the 2nd, 3rd and 4th weeks after
emergence, 50 leafhoppers (25 males and 25 females)
were removed from the cards for sexing and MBS
phytoplasma detection. Leafhoppers were sexed using
a stereomicroscope, based on the presence of the
ovipositor in the female genital area. Sex ratio was
calculated dividing the number of females by the
number of total individuals found in the maize whorl
or removed from each trap. After sexing, leafhoppers
were placed in 1.5 ml microtubes and kept in the freezer
for evaluation of MBS phytoplasma presence. The
number of MBS phytoplasma carrying leafhoppers was
determined for both capture methods and the
percentage of phytoplasma presence was calculated.
MBS disease incidence was evaluated in the field plot 3
weeks after tasseling assessing 938 plants by visual
observation of typical symptoms such as leaf reddening,
ear proliferation and stunting.
Obtention of D. maidis colony and MBS phytoplasma
strain
A colony of
D. maidis
was established in the laboratory
using insects collected in a maize field in Piracicaba.
Around 100 insects were placed onto maize plants (3 – 4
true leaves) in cages aluminum-framed with an acrylic
door, measuring 32 x 32 x 50 cm, covered with insect
proof net. The eggs laid by these leafhoppers were
collected from the leaves under a stereomicroscope and
placed onto healthy fresh maize seedlings in another
cage. The progeny hatched from the eggs, therefore, never
had contact with field or diseased plants. Some insects
from the colony were regularly tested for MBS
phytoplasma presence in order to ensure that the insects
were not infected. In addition a MBS phytoplasma
strains was obtained by collecting a symptomatic plant
from a field in Piracicaba. The MBS-infected plant was
brought to the laboratory and the MBS phytoplasma
recovered from this plant was maintained
in vivo
in
maize plants via insect transmission according to
Orlovskis
et al.
(2017).
MBS phytoplasma acquisition and transmission rate by
gender
A group composed of 100 nymphs of 2nd and 3rd instar
were confined onto a MBS infected plant for a 24 h
acquisition access period (AAP). After the AAP, the
leafhoppers were maintained on maize plants for a
latent period of 2 weeks, when the nymphs were in the
4th instar and before they became adults, the insects
were separated by gender, as described previously. The
separated insects were confined in 3 healthy maize
plants, composing three different groups: virgin males,
virgin females and same amount of nymphs of both
genders. This mixed group was set with the objective
for the insects to copulate right after they become adults
since adult insects are able to mate within 1 or 2 days
after emergence (Ramirez-Romero
et al.
, 2014). The
groups remained confined for 7 days. After this period,
individuals from each group were individually
confined on a maize test-seedling 2 weeks old, planted
in a 300 ml pot containing sterilized soil. The plants
were inside cages made of transparent plastic cylinders,
10 x 25 cm (diameter x height), with lateral openings
covered with insect proof net. Four treatments were
considered: virgin male, virgin female, mated male and
mated female. Insects remained confined for a 24 h
inoculation access period (IAP). The experiment setting
is schematized in Figure 1. After IAP, the leafhoppers
were removed from the test-plants, stored in freezer and
then individually tested by PCR to assess the presence
of MBS phytoplasma. Plants were kept in an insect-
proof greenhouse examined for typical MBS symptoms,
such as reddening of leaves, about one month after IAP.
Leaves of asymptomatic plants were sampled and
submitted to PCR for MBS phytoplasma detection. The
experiment was repeated four times.
DNA extraction from insect and plant tissues
In order to analyze the MBS phytoplasma presence in
the leafhoppers captured in the field and the presence
of MBS phytoplasma in insects and plants used in the
experiment of acquisition and transmission rate by
gender, plant samples and insect specimens were
submitted to PCR. Briefly, total DNA from leafhoppers
was extracted by macerating each insect individually
in 30 µl of extraction buffer (1 M Tris-HCl, 0.1% SDS, 20
mM EDTA) in a 1.5 ml microtube by using an autoclaved
pestle. The mixture was maintained at 65°C for 15
minutes, then centrifuged at 12,000 rpm for 10 minutes.
The supernatant was removed and after the DNA pellet
was dried, it was eluted in 30 µl of TE buffer. PCR was
performed using the primer pair R16F4 and R16R1 (Davis
and Lee, 1993).
The total DNA from maize leaves was extracted
using the CTAB method as described in Canale
et al.
(2017). Nested PCR assay was performed using the two
Phytopathogenic Mollicutes, Vol. 8 (1), June 2018 35
Role of Dalbulus maidis (Hemiptera: Cicadellidae) gender on maize bushy stunt phytoplasma transmission
universal primer pairs R16mF2/R1, followed by R16F2n/
R2, as described by Gundersen and Lee (1996). The
reaction mix contained PCR buffer (1 X), 0.2 mM of each
dNTP, 2.5 mM MgCl2, 0.25 U of Taq DNA polymerase,
0.5 µM of each primer, 1 µl of undiluted sample, in a
final volume of 10 µl adjusted with water. PCR were
performed in a PTC-100 thermocycler (M.J. Research, Inc.,
USA), where the temperature and length of each cycle
was adjusted accordingly to Davis and Lee (1993) for
insect samples, or to Gundersen and Lee (1996) for plant
samples. Amplicons were visualized in a 1% agarose gel
stained with ethidium bromide using a UV
transilluminator.
Data analysis
MBS phytoplasma acquisition and transmission by
gender was evaluated by calculating the acquisition
rate, which is the proportion of insects that acquired
MBS phytoplasma (PCR-positive) out of the total.
Transmission rate was calculated considering the
number of successful inoculation events out of the total
number of leafhoppers tested (Canale
et al.
, 2017). Since
acquisition and transmission rates were found not to be
normally distributed, a multiple comparison of
treatments was performed with Kruskal-Wallis test,
which is a nonparametric method. Statistical analysis
was carried using Agricolae package (version 1.2-4, 2016)
of the R software (R Core Team, 2015).
Results
D. maidis
population in maize field was assessed using
two different methods (Table 1). While the amount of
insects collected in the maize whorl increased over time,
the insects captured by yellow cards decreased over time.
Larger number of insects were collected by the cards
that remained in the field 15 to 28 days after emergence,
and after that, the number of captured leafhoppers
decreased drastically. The amount of female was always
higher in the maize whorl than in the yellow cards
(Table 1).
D. maidis
-MBS phytoplasma presence by gender was
assessed from insects collected both from maize whorl
and yellow sticky cards and 10 insects were positive to
phytoplasma presence out of the 345 PCR tested which
gives a natural infection rate of 2.90%. Higher number
of infected leafhoppers were captured from the plant
whorl than by the yellow sticky traps. Eight females
and two males were positive (Table 2). MBS disease
Figure 1. Schematic illustration of
the experiment to evaluate
acquisition and transmission of
MBS phytoplasma by Dalbulus
maidis. After acquisition access
period (AAP) on a MBS
phytoplasma source plant, nymphs
were transferred to healthy plants
for a 2 week latent period. During
latency, insects were separated by
gender and a group was
composed with insects to mate
after emergency of the adults. After
latent period, leafhoppers were
individually transferred to new
healthy maize seedlings, and
separated in four groups: mated
females, mated males, virgin
females, and virgin males.
Maria Cristina Canale et al.
Phytopathogenic Mollicutes, Vol. 8 (1), June 201836
Table 1. Mean per plant and total number of Dalbulus maidis sampled on maize plants and on yellow stick cards followed by sex ratio in each
evaluation period.
Maize whorl Sticky cards
D.a.e. Mean/plant Total Sex ratio D.a.e. Mean/card Total Sex ratio
8 1.55 62 0.52 8 - 14 32.0 160 0.27
15 1.67 67 0.57 15 - 21 62.8 314 0.35
22 1.65 66 0.56 22 - 28 40.8 204 0.21
29 2.70 108 0.55 29 - 35 22.4 112 0.42
36 4.37 175 0.64 36 - 42 27.4 137 0.30
Higher number of male D. maidis are captured by yellow sticky cards, which means that male leafhoppers exhibit greater flight activity. Leafhoppers
captured in 40 plants and 5 yellow stick traps at different days after emergence (d.a.e.) periods. Sexual ratio calculated dividing the number of females
by the total.
incidence in the field calculated on 938 plants was 21.2%
since 199 plants showed MBS symptoms.
MBS phytoplasma acquisition and transmission rates
by mated male and female, and virgin male and female
of
D. madis
was experimentally assessed, using insects
derived from a colony of insects and a MBS phytoplasma
strain maintained in laboratory. Leafhoppers were
allowed to acquire the phytoplasma as nymphs while
feeding from an infected source-plant. Then, they were
sexed and a group of virgin and mated males and
females was formed. After the groups were formed, the
mated or virgin insects were individually confined on
healthy maize seedlings for IAP. Acquisition rate of MBS
phytoplasma was not significantly different among
mated male and female and virgin male and female of
D. maidis
(Table 3). However, there was a significant
difference for transmission rate of MBS phytoplasma
by mated and virgin female of
D. maidis
(Table 3).
Female leafhoppers, either the virgin or the ones allowed
to mating before the IAP, resulted in a transmission rate
of 0.37, higher than the males. Every PCR-positive plant
Table 2. Natural phytoplasma presence in male and female Dalbulus
maidis weekly evaluated on maize field, sampled directly from the plants
whorl and by yellow sticky cards.
Sampling Gender Proportion and percentage of infectivity1
method 8 d.a.e. 15 d.a.e. 22 d.a.e.
Plant whorlaMale 1/301 (3.3)21/30 (3.3) 0/33 (0.0)
Female 2/32 (6.2) 2/37 (5.4) 1/33 (3.0)
2nd wk 3rd wk 4th wk
Sticky cardbMale 0/25 (0.0) 0/25 (0.0) 0/25 (0.0)
Female 0/25 (0.0) 0/25 (0.0) 3/25 (12.0)
Leafhoppers captured in a40 plants at different days after emergence
(d.a.e.) and b5 yellow sticky traps at different week (wk) periods after
emergency. Fifty insects (25 male and 25 female) were removed from
sticky traps to be assessed. 1Number of MBS phytoplasma positive insects
out of the total insects sampled and the 2correspondent percentage of
the infectivity between brackets.
sample for MBS phytoplasma showed typical symptoms,
whereas the PCR-negative samples did not show
phenotypical alterations.
Discussion
By using two sampling methods, i.e. collecting insects
from the maize whorl and with the yellow sticky cards,
it was recorded that females
D. maidis
were more easily
captured from the maize whorl, whereas male
leafhoppers were more abundant in the cards, indicating
a different behavior of
D. maidis
accordingly with the
insect gender. While the female tends to stay feeding
and ovipositing in the plant whorl, male leafhoppers
fly in search for female to copulate (Heady and Nault
1985; Todd
et al.
, 1991; Larsen and Nault 1994). Regardless
of the possibility of capturing
D. madis
directly from
the maize whorl, the sticky cards are the most
appropriate method for leafhoppers monitoring (Larsen
et al.
, 1992; Meneses
et al.
, 2016). In this study, higher
number of insects was collected in the yellow traps
between 15 and 28 days after emergence probably due
to the plants height in the plot, recording approximately
0.50 m.
D. maidis
distributes vertically at different
heights according to the development stage of maize
plants. For insect monitoring, yellow traps shall be
installed at 0.50 and 1.50 m according to the crop
development (Meneses
et al.
, 2016).
The incidence of MBS diseased plants in the plot was
21.2%, and it is possible that the verified 2.90% of a MBS
natural infection rate of leafhoppers collected in the
field in the early weeks of the crop conduction was
linked to the disease incidence representing the average
of that 6.2 to 49.9% previously reported by Oliveira
et
al.
(2003). However, serious outbreaks of MBS disease
have been recently registered in Brazil, with nearly total
losses of maize crop. MBS disease outbreak is related to
Phytopathogenic Mollicutes, Vol. 8 (1), June 2018 37
Role of Dalbulus maidis (Hemiptera: Cicadellidae) gender on maize bushy stunt phytoplasma transmission
population levels of the corn leafhopper, which also
exhibits a cyclic dynamics yet not well understood
(Summers
et al.
, 2004). It has been observed that, in fact,
the population levels of this leafhopper is high in maize
plantations throughout Brazil. Therefore, maize growers
must be aware of the population levels of leafhoppers
for the disease management. Additionally, the amount
and extent of maize crops in a region and the maize
variety used may influence the disease incidence.
In field collection, it was verified that there was more
MBS phytoplasma-infected female than male of
D.
maidis
. This might also be related to the female behavior
of spending more time feeding from maize plant, which
might increase the chances of bacterial acquisition from
infected plants by female leafhopper under natural
circumstances. However, there was no difference on the
acquisition rate by gender of leafhoppers during nymph
stage in the present laboratory experiment suggesting
that
D. maidis
nymphs had the same chances of feeding
and acquire MBS phytoplasmas after hatching.
A significant difference was detected in the
transmission rate between males and females. Female
leafhoppers, whether copulated or not, presented a
higher transmission rate compared to male
D. maidis
,
which shall be explained by the female behavior of
spending longer periods feeding (Heady and Nault 1985;
Hunt and Nault, 1991; Oliveira
et al.
, 2004). Increased
phloem ingestion observed in females of hemipteran
vectors (Narayana and Muniyappa 1996; Ning
et al.
, 2015;
Antolínez
et al.
, 2016) can be explained by the high
Table 3. Maize bushy stunt phytoplasma (MBS) acquisition and transmission rates by mated and virgin male and female Dalbulus maidis.
Treatment Trial Total of MBS-positive Acquisition Diseased Transmission
individualsxleafhoppersyrate plantszrate
Mated male I 21 5 0.24 3 0.14
II 21 10 0.48 9 0.43
II I 20 8 0.40 3 0.15
IV 20 12 0.60 9 0.45
mean 0.43 0.29
rank 8.5 a 8.12 ab
Mated female I 21 11 0.52 9 0.43
II 22 11 0.50 10 0.45
II I 20 11 0.55 5 0.25
IV 20 9 0.45 7 0.35
mean 0.50 0.37
rank 12.12 a 11.00 a
Virgin male I 20 2 0.10 2 0.10
II 16 4 0.25 2 0.13
II I 20 9 0.45 5 0.25
IV 6 3 0.50 1 0.17
mean 0.32 0.16
rank 5.87 a 4.12 b
Virgin female I 21 7 0.33 5 0.24
II 22 8 0.36 4 0.18
II I 20 10 0.50 9 0.45
IV 13 6 0.46 8 0.62
mean 0.41 0.37
rank 7.50 a 10.75 a
χ
23.73 5.44
p0.2910 0.1418
As the leafhoppers were individually confined on healthy maize seedlings, total number of individualsx tested correspond to the number of leafhoppers
evaluated and the number of plant-test used in the experiment. Leafhoppers were tested by PCR for MBS phytoplasma detectiony. Acquisition rate was
determined considering the number of insects which acquired MBS phytoplasmas out of the total. Plants were evaluated for MBS symptoms, and
asymptomatic plants were tested by PCR for MBS phytoplasma detectionz. Transmission rate was calculated considering the number of successful
inoculation (diseased plants) out of the total number of leafhoppers tested. Data obtained in 4 independent trials. Letters show statistical comparison
among treatments using Kruskal-Wallis test.
Maria Cristina Canale et al.
Phytopathogenic Mollicutes, Vol. 8 (1), June 201838
requirement of nutrients to support eggs production.
Furthermore, this increased phloem ingestion would
result in a higher efficiency in acquisition and
transmission of a plant pathogen.
Given the long persistence of MBS phytoplasmas
within the
D. maidis
, it is suggested that it might be
transmitted to healthy plants anytime during the vector
lifespan. However, the differential behavior between
male and female
D. maidis
may suggest different roles
in MBS disease epidemiology. As female are less active,
they may be ready to transmit MBS phytoplasma to
the plants immediately adjacent to those where the
phytoplasma was acquired, increasing the MBS disease
incidence within the field. Spatial analysis of aster
yellows incidence in field-grown and in greenhouse
lettuce indicates a clustering of diseased plants, a pattern
suggesting that field spread of aster yellows
phytoplasma in lettuce is more likely due to female
transmission (Beanland
et al.
, 1999). Furthermore, female
D. maidis
is proven to live longer than males and survive
longer periods without access to food, with ability to
overwinter and migrate (Larsen
et al.
, 1992; Taylor
et al.
,
1993; Moya-Raygosa
et al.
, 2007; Carloni
et al.
, 2013;
Oliveira
et al.
, 2013). This characteristic shall increase
the chance for MBS phytoplasma to be spread farther
away from where it was acquired. In spite the tendency
of the female leafhopper on MBS phytoplasma
acquisition and transmission verified in this study, the
role of infected male leafhopper may not be disregarded
since male
D. maidis
tend to exhibit a more intense
flight activity, helping to increase MBS disease incidence
or even taking the inoculum to neighbor maize crops.
In addition to
D. maidis
monitoring using yellow
cards, it would be of great importance the development
of a fast detection tool such as the molecular analysis in
order to check if the leafhoppers are infected and
consequently pose a risk for maize crop (Beanland
et al.
,
1999; Demeuse
et al.
, 2016). The leafhopper population
must be tested specially in the initial phase of maize
crop thus MBS disease management measures for the
vector must be taken as earlier as possible. It has been
shown in this study that males and females of
D. maidis
have a differential role on MBS disease epidemiology
and this might be due to insect behavior. Therefore, as it
occurs for
Macrosteles quadrilineatus
and the aster
yellows phytoplasma, MBS disease outbreaks may be
also influenced by the MBS phytoplasma infected female
and male population of
D. maidis
.
Acknowledgements
We would like to thank Maria Tereza Vieira de Camargo
Lopes and Luciana Bianchini for technical assistance
and CNPq (Brazilian National Council for Scientific and
Technological Development).
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