EFFECT OF WATER EXTRACTS FROM GERANIACEAE (GERANIACEAE) PLANTS ON FEEDING AND DEVELOPMENT OF COLORADO POTATO BEETLE (LEPTINOTARSA DECEMLINEATA SAY)

Robert Lamparski, Maria Wawrzyniak

ABSTRACT

There were tested water extracts from selected Geraniaceae plants. It was observed that the plant extracts investigated inhibit feeding and development of Colorado potato beetle. The greatest antifeedant activity towards Colorado potato beetles and their larvae was noted for Pelargonium x hortorum Bailey and Geranium sanquineum L. extracts. The Pelargonium x hortorum Bailey extract added to food showed unfavourable effect on the development of female reproductive organs and significantly inhibited the number of eggs laid, however it showed effective neither towards the period of winter diapause nor towards spring emergence of beetles. The highest effectiveness under field conditions was recorded for extract from Erodium cicutarium L.

Key words: Colorado potato beetle, Geraniaceae, plant extracts.

INTRODUCTION

Colorado potato beetle is one of the most dangerous potato pests. It is controlled mostly with insecticides which inhibit the abundance of pest below the commercial harmfulness threshold, but they can also show a negative effect on the environment and cause resistance in insects to some chemicals [5]. For that reason more and more frequently alternatives to chemical control are searched for to find methods of inhibiting the abundance of this pest. One of the concepts involves biologically active substances of plant origin as pesticides or compounds affecting the behaviour of insects. Bioactivity of some plants depends on the presence of various chemical compounds in their tissues which would inhibit insect feeding [3,8]. Great hopes are related to secondary plant metabolites to include triterpene – azadirachtin, isolated from Meliaceae [14,24], limonoids – obtained from citruses [22,27], diterpenes isolated from plants of Rhododendron spp. and Delphinium spp. genera [6, 12]. The effect of secondary plant metabolites contained in Geraniaceae was researched towards various organisms: wireworm (Melanotus communis) beetle larvae [35], mosquitoes [29] and Aspergillus niger van Tieghen fungus [21].

So far not many researches have been carried out into the use of Geraniaceae plants in inhibiting the abundance of different Colorado potato beetle development stages. Such researches were originated by Wyrostkiewicz [39] who tested water and acetone extracts obtaining results which suggested a need for further research.

The present working hypothesis assumes that secondary metabolites contained in Geraniaceae plants show active towards insects and that they can be used in the form of natural extracts to inhibit the harmfulness of Colorado potato beetle. The aim of the present research was to define the effect of water extracts obtained from selected Geraniaceae plants on feeding of Colorado potato beetles and larvae and the development of this pest.

MATERIAL AND METHODS

Research carried out over 1998-1999 covered laboratory experiments carried out in the Department of Applied Entomology as well as field observations on experimental plots located at Mochełek in the vicinity of Bydgoszcz, of the Experiment Station of the Bydgoszcz University of Technology and Agriculture.

There was analyzed the effect of water extracts obtained from six species of Geraniaceae:

• Pelargonium x hortorum Bailey (Photo 1),

• Erodium cicutarium L. (Photo 2),

• Geranium pusillum L. (Photo 3),

• Geranium sanquineum L. (Photo 4),

• Geranium pratense L. (Photo 5),

• Geranium robertianum L. (Photo 6).

The plants were collected by cutting overground parts (herbage) over flowering on barren land. Geraniaceae plants were obtained from horticultural plantation. Plants crushed with pruning shears were dried at room temperature and then ground with lab WŻ – 1 lab grinder obtaining dried plant material (ground-coffee-like) which over the experiments (a few weeks) was stored in hermetic glass containers. The material constituted the raw material for water extracts obtained following the method given by Wyrostkiewicz [39]. The ground dried plant material was poured for 24 hours with cold water (municipal water intake); 100 ml of water per 10 g of dried material. Having filtered it, the extract was obtained (defined as 10%) used in the experiments. For each experimental series a fresh plant extract was prepared.

In the laboratory experiments the following were evaluated:

1. The effect of the plant extracts tested on feeding of Colorado potato beetles and larvae

The potato leaves weighed were soaked for about 3 seconds in the extracts tested and then dried at room temperature and put on Petri dishes, a couple of beetles or 10 weighed L₂ larvae into each. After 24 hours larvae were weighed again as well as food which had not been consumed yet was removed and the dishes were filled with fresh leaves prepared the same way. The dish with a couple of beetles or 10 larvae constituted a single replication. Each experimental combination and control in which extract-untreated potato leaves was tested in 4 replications. The weight differences helped to calculate:

1. the weight of food consumed by a beetle couple or a larva,

2. the larva body weight gain,

3. the absolute deterrence index (for beetles and larvae – bwd, calculated based on the amount of feeding on control food and treated with extract for both feed sources placed separately [15].

2. Effect of extracts on the development of Colorado potato beetle, including:

1. oviposition by beetles fed with potato leaves with extracts added – winter generation Colorado potato beetles were fed with potato leaves treated with plant extracts. The total number of eggs laid by 1 couple of beetles in respective test combinations is given as a percentage of the number of eggs laid in the control combination;

2. course of larvae hatch from eggs treated with extracts; from eggs obtained in respective experimental combinations four very numerous deposits were selected (as replications), onto which adequate extracts were placed with a brush. After 5-7 days the hatch of larvae was observed. At the same time larvae hatch from eggs untreated with extracts was being observed.

3. pupation of larvae fed with leaves with extracts; larvae from respective experimental combinations were transferred into sand-filled containers covered with a net. Until the larvae came down for pupation, they were fed with potato leaves treated with adequate water extracts. The control leaves were untreated. Summer generation Colorado potato beetles which emerged after pupation were weighed and counted. The results are given as percentage of beetles of the initial number of larvae and of the number of beetles obtained in the control combination;

4. gonads development; ovarioles were prepared (following the method developed by Błażejewska [2]) from summer generation female beetles from larvae fed with leaves treated with extract from Pelargonium x hortorum (stored in Ringer’s body fluid). Then ovarial tubes were counted and in each 10 randomly sampled tubes were measured with stereoscopic microscope equipped with measuring eyepiece.

Field experiments

Field experiments were carried out over 1998-1999 on experimental plots located at the Mochełek Experiment Station of the Bydgoszcz University of Technology and Agriculture. The following were evaluated:

1. Effectiveness of extracts towards Colorado potato beetles and larvae. The potato field of 1 ha was divided into 5 x 5 m plots (8 rows 20 plants each) which accounted for respective test and control combinations. From each plot four potato rows (20 plants each) were randomly selected on which prior to treatment and after 2 and 6 days after treatment, the abundance of insects was observed. Always whole plots were sprayed with 1 dm³ of extract per plot (400 dm³ˇha^-1). 10% water extracts were used. The effectiveness of extracts under field experiments (in %) was calculated with the Henderson-Tilton formula [31]:

where:
K₁ – number of insects on control plot prior to treatment,
K₂ – number of insects on control plot after treatment,
T₁ – number of insects on test plot prior to treatment,
T₂ – number of insects on test plot after treatment.

2. Effect of extracts on oviposition; number of eggs deposits was observed on plants treated with extracts 2 and 6 days after treatment and compared to the number of eggs deposits prior to treatment (%).

3. Effect of Pelargonium x hortorum extract on wintering of imagines; place of wintering: plastic bags with potato plants dug 80 cm deep, covered with large insulators (metal framework covered with mill gauze). At least 10 beetles were put onto each plant. The experiment and control combination consisted of 4 replications (one insulator with a single plant constituted a single replication). All the beetles were placed into insulators prior to the end of the first decade of August, after which potato plants were sprayed with geranium extract. In spring emerging beetles were counted and weighed.

The lab and field experiments results were analyzed with single variance analysis method in completely randomized design. The significance of the differences between means for respective factor levels (plant species or extract type) was defined with Tukey test at α = 0.05.

RESULTS

Laboratory experiments

Effect of extracts from Geraniaceae plants on feeding of Colorado potato beetle

Feeding of Colorado potato beetles
There was observed a considerable effect of the extracts tested on inhibiting feeding of beetles (Table 1). Considerably less food was taken up by insects fed with potato leaves with extracts from Geranium sanquineum and Pelargonium x hortorum added, respectively 13.0 and 17.3% of the food consumed by a couple of beetles in the control combination. A less powerful effect on inhibiting feeding was observed for extract obtained from Geranium robertianum (31.3%). The differences between the weight of food consumed by beetles in combinations with all the water plant extracts tested and the control were significant. Absolute deterrence index (bwd) calculated based on the differences between the weight of food consumed by beetles in respective experimental tests and the control ranged from 51 to 76 (Fig. 1). Significantly greatest antifeedant effect was shown for extract from Geranium sanquineum (bwd – 76.1).

Larvae feeding
It was recorded that in all the experiment tests larvae consumed less leaves than in the control combination (from 28.7 to 55.3% of the weight of food consumed in the control). The least feeding of larvae was noted in tests which involved Pelargonium x hortorum and Geranium sanquineum extracts, while the most intensive in the combination with Erodium cicutarium (Table 1). The 48-hour larvae body weight gain ranged from 23 mg (Pelargonium x hortorum) to 41 mg (Erodium cicutarium). In control, body weight of a single larva increased by 46 mg. The calculated absolute deterrence index was relatively high (Fig. 1). Its highest value was recorded in the test in which larvae were fed with potato leaves with Pelargonium x hortorum extract (56), while the lowest after the application of Erodium cicutarium extract (30). Statistical analysis of the results did not show significant differences of bwd for respective extracts .

The effect of Geraniaceae plants extracts on the development of Colorado potato beetle

Oviposition
In all the combinations tested there were observed fewer eggs laid than in the control (Table 2). The lowest number of eggs was laid by beetles which were fed with leaves treated with Geranium pratense and Pelargonium x hortorum extracts (respectively 6.5 and 14.7% as compared with the number of eggs in the control), however the biggest number in the experiment which involved extract from Geranium robertianum (65.6%). There was observed a significance of differences in the number of eggs laid by beetles fed with leaves with Pelargonium x hortorum, Geranium sanquineum and Geranium pratense extracts and by the control beetles.

Larvae hatching
The observations made showed that the greatest effect on inhibiting Colorado potato beetle larvae hatch was attributed to Pelargonium x hortorum extract (Table 2). After its application out of 32% of eggs larvae hatched. In the other tests more larvae were obtained; the greatest number in the test which involved Geranium robertianum extract (larvae hatched from 85.6% eggs).

Pupation
A significant effect on the process of Colorado potato beetle pupation was noted for Pelargonium x hortorum extract. After its application 47% of beetles were obtained, as compared with the number of beetles observed in the control. Their body weight was lower, however differed significantly neither from the control combination beetle body weight nor from the other experiment combinations (Table 2).

Gonad development
Based on the calculations made, it was observed that the number of ovarioles and their length in the Colorado potato beetle females tested, obtained from larvae fed with leaves treated with water extract from geranium, was significantly lower than in the control combination (Table 3).

Field experiments

Effectiveness of extracts from Geraniaceae plants towards Colorado potato beetle

The results obtained over two research years showed that extract from Geranium robertianum discouraged the pest from infesting potato plants most significantly, as compared with the other extracts applied (37.9% effectiveness) (Fig. 2). A lower effectiveness was observed for extracts from Geranium pusillum and Pelargonium x hortorum. The greatest effectiveness towards larvae was shown by Geranium pusillum extract (39.1%). In the other cases the calculated extracts effectiveness ranged from 26.1% (Geranium sanquineum) to 37.6% (Geranium pratense) (Fig. 2).

Effect of extracts tested on oviposition

With the number of Colorado potato beetle eggs laid on plants treated with the extracts investigated, one can state that the most effective inhibiting effect on oviposition was noted for Erodium cicutarium extract. Having treated potato leaves with this extract, no new eggs deposits were recorded. As for the other plant extracts and in the control, the calculated percentage increase in the number of eggs deposits laid ranged from 38 to 120% (Table 4).

Effect of Pelargonium x hortorum extract on wintering of imagines

An analysis of the number of beetles emerging in spring against their number prior to coming down to soil for the period of winter diapause, did not show the effect of Pelargonium x hortorum extract on Colorado potato beetle wintering (no significance of differences compared with the control). The survival over period of winter diapause was noted in 32.5% of the control beetles, while after the application of water extract from pelargonium - 37.9%.

DISCUSSION

The present research showed that the water extracts tested obtained from selected Geraniaceae plants affected feeding and development of Colorado potato beetle. Under the control conditions the greatest effect on inhibiting Colorado potato beetle and larvae was attributed to extracts from Pelargonium x hortorum and Geranium sanquineum. The herbage of these plants contains geranium oil [20,28], flavonoids (quercetin) and considerable amounts of tannins [10,25]. A clear inhibition of the pest feeding following the application of Geraniaceae plants extracts was also observed by Wyrostkiewicz [39]. A similar reaction of Colorado potato beetle under lab conditions was also seen by Schearer [30] and Hough-Goldstein [11] following the application of water extracts from common tansy, Klingauf and Weil [16] – extracts from western thuja and sessile oak, Murray et al. [26] – limonoids from citrus plants, Zahnder and Warthen [42] – extracts from Azadirachta se eds and saponins obtained from alfalfa – Waligóra [36,37], Waligóra and Krzymańska [38].

Geraniaceae plants as well as other plants which inhibit feeding of Colorado potato beetle include numerous antifeedant substances, including flavonoids, polyphenols and tannins, which they represent, considered crucial in the way insects behave while food intake [4,7,17]. Numerous researchers claim that those produced over millions of years, the so-called secondary plant metabolites, affect insects neither as single substances nor their specific groups, however as a mixture of all these chemical compounds [1,19,40]. It is therefore difficult to define clearly the importance of respective compounds of secondary metabolism for the process of inhibiting the pest feeding. For that reason it would be recommended to carry out detailed chemical analysis of the plant raw material (from Geraniaceae plants) and its extracts. The return on investment into analysis and other pilot research could be possible quite soon as commercial production of natural insecticides can be very beneficia l, proving e.g. a better protection for beneficial insects. Bioinsecticides can be also used in the control of pests which developed strains resistant to synthetic insecticides [13].

In the present research the development of Colorado potato beetle under laboratory conditions was most inhibited by water extract from Pelargonium x hortorum. Similar results were obtained by Wyrostkiewicz [39] as well as Wyrostkiewicz and Wawrzyniak [41] for extracts from field bindweed, royal marigold and white deadnettle. Numerous authors report on insects fertility being related to the content of secondary metabolites in plants [9,23,34], especially phenols [19] and flavonoids [32,33]. Under field conditions the greatest effectiveness towards Colorado potato beetles, larvae and eggs was recorded for Erodium cicutarium extract. Its herbage contains tannins, organic acids (malic acid, citric acid, gallic acid), caffeine, choline, histamine, mineral salts (mainly potassium salts), vitamin C [18,25]. Similar results for Erodium cicutarium (at a lower concentration) were reported by Wyrostkiewicz [39].

CONCLUSIONS

1. Water extracts from selected Geraniaceae plants tested under laboratory and field conditions showed various effects on Colorado potato beetle.

2. A high antifeedant activity towards Colorado potato beetles and larvae was recorded for extracts obtained from Pelargonium x hortorum and Geranium sanquineum.

3. Potato leaves covered with Pelargonium x hortorum extract showed an unfavourable effect on the development of reproductive organs in females, significantly inhibited the number of eggs laid by them, however they showed effective neither towards the period of winter diapause nor spring beetle emergence.

4. Water extracts obtained from Geraniaceae plants applied under field conditions cannot replace chemical insecticides in Colorado potato beetle control now, however their application can be an element of integrated control of this pest.

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