Abstract:
This study aimed to investigate the effects of Bacillus thuringiensis
on Liriomyza trifolii (Burgess) (Dipetera: Agromyzidae).
L. trifolii is an important pest in vegetable
growing areas in Sanliurfa, Turkey. Field trials were carried out in the experimental
field of Faculty of Agriculture in Harran University through the July to November
in 2005, employing randomized complete block design with three replicates. Adults
of L. trifolii were obtained from laboratory
culture and 5
INTRODUCTION
The serpentine leaf miner Liriomyza trifolii (Burgess) (Diptera: Agromyzidae) is a world-wide pest of ornamental and vegetable crops. L. trifolii arrived in Turkey in 1995 (Uygun et al., 1995). L. trifolii is a polyphagous leaf miner (ca.400 different host plants; Murphy and La Salle, 1999; Parella, 1987) that undergoes larval development in the plant leaf tissue and forms serpentine mines within the leaves (Parella and Bethke, 1988). Damage is caused mostly by larvae that feed their was inside the plant-host mesophyll and by the female feeding behavior, (puncturing the leaf with their ovipositor and feeding from the leaf sap), an action that decreases the plant’s photosynthesis (Parella et al., 1985), provides entry sites for plant pathogens (Broadbent and Matteoni, 1990; Matteoni and Broadbent, 1988) and creates small marks that reduces the aesthetic appearance of leaves. This species has had an impact on many field and flower crops in Turkey, beans being among one of the more economically important crops affected.
There are currently no effective insecticides for use against adults, although growers continue to use whatever is available (chlorfenapeyr, chlorfluazuron, chlopyrifos-ethyl, deltamethrin, diazinon, endosulfan and malathion) and few effective larvicides. Two main larvicides are used to control the leaf miner on beans in Turkey, abamectin and cyromazine both of which are translaminar. Although the Turkish Ministry of Agriculture recommends treating vegetable crops only after 4-5 min are observed per leaf (Anonymous, 1996), growers generally apply insecticides earlier when they see a few min on several leaves and may continue to treat once or twice a month until the end of the season. Exclusive use of these larvicides could quickly generate resistance in the leaf miner.
Environmental and regulatory consideration have led to the development of new classes of insecticides that are less environmentally disruptive and have low mammalian toxicity. A major concern regarding the use of new and existing classes of pesticides is prolonging their utility through resistance management. One resistance management strategy is to rotate among pesticides with different modes of action to delay development of resistance to any one insecticide (Tabashnik, 1989; Georghiou, 1990).
In the search for additional active ingredients to control agromyzid pests, B. thuringiensis is called as a biocide and during sporulation, strains of B. thuringiensis make crystalline cytoplasmic protein inclusions that have been used for over 30 years as highly specific insecticides against certain species of Lepidoptera and Diptera (Van Frankenhuyzen, 1993). Although these toxins have been remarkably successful, their high specificity has precluded their use as a general pesticide. A strain of B. thuringiensis has recently been identified that has demonstrable activity against the larvae of common housefly (Musca domestica).
The present study deals with effect of B. thuringiensis biocide of larvae of L. trifolii in bean field conditions in southern Turkey. In the study, B. thuringiensis treated, non-treated and control plants were compared for the control of serpentine leaf miner larvae as well as parasitoid species abundance and diversity in bean field. The result would be used in developing a sound control strategy of L. trifolii in bean field conditions.
MATERIALS AND METHODS
Experiment area: This study was conducted from July to November in 2005 in the Faculty of Agriculture, University of Harran, experiment areas in Şanlıurfa, Turkey on bean grown under open field conditions.
In this study, bean seeds (cv. Perla) were sown on 29 July in 2005. Experiments were carried out in the field conditions and designed as randomly blocks with 3 treatments and three replicates.
Each plot was 6.3 m2 (30 plants) and consisted of 2 rows (4.5 m long). Inter-row spacing was 30 cm and intra-row plant spacing was 70 cm. No insecticide treatment was applied throughout the production period for the all treatments. In order to prevent contamination from the outside to experiment area all treatments were closed off with fly-nets (hole size: 0.5 mm).
Reared and inoculation of L. trifolii to experiment plots: Bean leaves which were infested by L. trifolii were collected from bean plants grown area in Şanlıurfa. Collected leaf samples were reared in pots covered with plastic bags in the laboratory at 25±1°C and 65±5% relative humidity to reached the pupae and adults of L. trifolii. The identification of the L. trifolii was made by Dr. E. Çıkman (Harran University, Turkey). Adults of L. trifolii were obtained from laboratory culture and 5 ♀♀ and 5 ♂♂ adults of L. trifolii were inoculated each plots at the first week of the production period. The control plots were non-inoculated by L. trifolii and also, closed off with fly-nets.
Application of Bacillius thrungiensis Berliner: B. thrungiensis Berliner was applied at a concentration of 60x106 mg-1 (Secrotype 3a, 3b, strain SA-11001 C98-1-1) (53000 Spodoptera μ mg-1 (32 000 IU mg-1), 60x106 mg-1 Bacillius thrungiensis spore (Certis Inc., Maryland, USA). B. thrungiensis was applied at the recommended rate of 75 g 100 L water. All treatments were applied with a low-pressure backpack sprayer. It was applied ones a two weeks in the late of afternoon until at the end of the experiment period. Application dates were set when the pest density reached a level of 4-5 larvae/leaf which are economic threshold (E.T) (Table 1), the density at which B. thrungiensis treatment is advocated.
The application of B. thrungiensis was done the 3rd week and application continued once a 15 day throughout the 9th week production periods, because the pest density reduced a level of 4-5 larvae/leaf the last 5 weeks. The application was made from 3rd week of August to 2nd week of October. Total application of B. thrungiensis was 4 times throughout the bean production period.
Sampling: The experiment area was checked weekly during the entire production period starting with the emergency of plants. The production period lasted for 15 weeks. From the very first appearance of L. trifolii larval mines on the leaves, 10 leaves were taken per plot.
| Table 1: | Average number of live larvae of Liriomyza trifolii per leaf in Bacillus thuringiensis treated and non-treated leaf |
| Ns = not significant | |
Because the plants were immature and during the flowering and early fruiting period, leaves are necessary to protect the ripening beans from the sun. Another reason was to keep the study uniform at every stage. All leaves were brought to the laboratory, where the number of live larvae were counted and recorded. The leaves containing the larvae was cut and placed in a small glass vial and then closed with a cotton ball covered with muslin. The vials were kept in plastic culture containers (30x20 cm) at 25°C and 65% relative humidity to allow larvae to develop to adults. They were daily checked for the emerging leaf miners and their parasitoids. They were counted and recorded. The identification of the L. trifolii was made by Dr. E. Çıkman (Harran University, Turkey) and the identification of the chalcidoidea was made by Dr. M. Doğanlar (Mustafa Kemal University, Turkey).
Bean was hand harvested once a 4-5 day throughout the production period and weighed. Trial was terminated at the end of the vegetation period which was lasted 15 weeks in 2005.
Data analysis: The data were analyzed by complete randomized design using TARIST statistic program. Treatment means were separated by LSD test at (p<0.05).
RESULTS
Results of the number of live larvae from B. thuringiensis-treated, non-treated and control leaves are shown in Table 1.
There were approximately 0-3 leaf miner larvae per leaf at the beginning of the trial, the most were in the first or second instar.
In the B. thrungiensis treated plots the number of live larvae were at the E.T. at the 3rd week and application of B. thrungiensis was done. The B. thrungiensis treatments had significantly fewer live larvae than in non-treated leaves. The number of live larvae continued to decrease until 7th week, after which a slight increase was noted until 9th week.
| Table 2: | Average yield of bean for Bacillus thuringiensis treated, non-treated and control |
However, the whole production period which was 15 weeks, from 4th week to 15th week, the number of live larvae was still fewer than the number at which treatment is recommended (4-5 larvae per leaf) (Anonymous, 1996).
In the B. thrungiensis non-treated plots, the number of live larvae were at the E.T. at the 3rd week, after which a slight decrease was noted at the 4th and 5th weeks. The number of live larvae were increased until 11th week, after which decreases were noted at the and of the vegetation period.
As it can be seen from Table 2, yield values for treatments are different from each other. The lowest yield was achieved on B. thrungiensis non-treated plots, while the highest yield was recorded with the B. thrungiensis treated plots.
Results of the number of L. trifolii adults, parasitoid species and percentage of parasitization from B. thrungiensis-treated and non-treated treatments in bean leaves are shown Table 3. Seven parasitoids species invaded the bean grown areas: Aprostocetus sp., Crytogaster vulgaris Walker, Cirrospilus vittatus Walker, Diglyphus isaea Walker, D. pachyneurus Graham, Hemiptarsenus zilahisebessi Erdös, Neochrysocharis formosa (Westwood), Pachyneuron groenladicum Holm.
The number of parasitoids and the percentage of parasitization in the B. thrungiensis non-treated plots were higher than B. thrungiensis treated plots and the percentage of parasitization were 59.14 and 50.69% respectively. In other words, in the B. thrungiensis treated plots the percentage of parasitization were relatively low because B. thrungiensis also reduced the number of parasitoids, most probably due to host death.
| Table 3: | The number of Liriomyza trifolii, parasitoid species and percentage of parasitism in Bacillius thrungiensis treated and non-treated plots |
DISCUSSION
L. trifolii has been a serious pest in Turkey sine 1995 and is found from autumn to spring. Growers following the advice of pesticide advisors use translaminar insecticides against the larval stages once populations reach 4-5 larvae/leaf, however many growers do not treat according to recommendations and will apply a wide range of conventional insecticides against both larval and adult stages. Although studies have shown that a few leaf miners (1-50/plant) actually cause an increase in fruit production (Kotze and Denill, 1996), growers will make multiple applications of insecticides in an attempt to completely eliminate the leaf miners. This non-judicious use of insecticides raises serious concern about the possibility of resistance developing in leaf miners as well as other species.
B. thrungiensis has been used for over 30 years as highly specific insecticides against certain species of Lepidoptera and Diptera (Van Frankenhuyzen, 1993), although was also recommended for control of Liriomyza sativae Blanchard (Diptera: Agromyzidae) (Sengonca and Lıu, 2003).
There have been few trials with B. thrungiensis for the control of agromyzid species. Sengonca and Liu (2003) There have been few trials wit B. thrungiensis for the control of agromyzid species. Sengonca and Liu (2003) studied effect of GCSC-BtA (Germany-China Scientific Cooperation-B. thrungiensis-Abamectin) biocide on abundance and diversity of some important cabbage pests, i.e., Plutella xylostella (L.) (Lep., Plutellidae), Pieris rapae (L.) (Lep., Pieridae), Brevicoryne brassicae (L.) (Hom., Aphididae), Liriomyza sativae Blanch. (Dip., Agromyzidae), Phyllotreta vittata Fabric. (Col., Chrysomelidae), as well as their naturel enemies, i.e., Apanteles plutellae Kurdj. (hym., Braconidae), Erigonidium graminicola (Sundv.) (Araneida, Linyphiidae), Coccinella septempunctata L. (Col., Coccinellidae), were determined in comparison to that of methomyl insecticide in cabbage fields in Fuzhou region of the southern China. The result of their study, they found that GCSC-BtA biocide showed high efficacy in reducing abundance of all the tested pest species while very low harmfulness to their natural enemies. GCSC-BtA treatment resulted in significantly lower abundance of all the pests with 10.32% of P. xylostella, 6.10% of P. rapae, 2.11% of B. brassicae, 8.68% of L. sativae as well as 1.02% of P. vittata as compared to methomyl treatment with 26.68, 26.33, 25.18, 33.89, 32.08% or control with 62.99, 67.55, 71.74, 57.42, 46.41%, respectively. The biocide gave higher species evenness of the natural enemies than the methomyl or control treatments. They recorded that after application, diversity index was higher with a value of 2.4111 with GCSC-BtA treatment than 1.1859 with methomyl or 1.2166 control treatment and concluded GCSC-BtA has high efficacy against cabbage pests and low harmfulness to their naturel enemies as compared methomyl. Khyami-Horani (2002) reported that in Jordan Valley, B. thrungiensis and B. thrungiensis local isolated, in addition to the reference strains B. thrungiensis kurstaki and B. thrungiensis israelensis, high toxicity towards 3rd instar larvae of Drosophila melanogaster. The toxic concentration ranged between 2.0x106 and 4.4x107 viable spores mL-1. Also, Jorge et al. (1999) described toxic effect of the B. thuringiensis β-exotoxin toward 3rd instars of 3 fruit fly species: Anastrephe ludens (Loew), A. obliqua (Macquart) and A. serpentine (Wiedemann). And they found that β-exotoxin was highly toxic to all 3 species tested.
This study shows that B. thrungiensis is effective in controlling the larvae of L. trifolii. Application of B. thrungiensis significantly improved efficacy; as a result, B. thrungiensis should be treated only once every 2-3 weeks for effective control.