Development of a fungal based-product for the control of Western flower thrips Frankliniella occidentalis (Pergande)

Abstract

The Western Flower Thrips (WFT) Frankliniella occidentalis (Pergande) is a major constraint to many cultivated crops in the world, causing important economical yield losses. They cause direct damage on plants and contribute exclusively in the propagation of tospovirus to plant. Thrips control is essentially based on the use of synthetic chemical pesticides; however most thrips species including Foccidentalis are resistant to a wide group of chemical pesticides. Hence there is the need to develop alternative strategies that are environmentally friendly. Entomopathogenic fungi are among the options being considered as a promising alternative for F. occidentalis management. The aim of this study was to develop a fungal based-product for the control of F. occidentalis. Larval stages of F. occidentalis are often considered to be refractory to fungal infection as compared to the adult stage; hence, screening of 10 isolates of Metarhizium anisopliae (Metschnikoff) Sorokin and 8 of Beauveria bassiana (Balsamo) Vuillemin was carried out in order to select virulent entomopathogenic fungal isolate(s) for their control. The most virulent isolates were compared in terms of conidial production and genetic polymorphism. All the fungal isolates tested were pathogenic to the second-instar larvae of WFT, causing mortalities ranging between 24 and 56%. M. anisopliae isolates ICIPE 7, ICIPE 20, ICIPE 69 and ICIPE 665 had the shortest LT50 values, causing mortalities within 8.0-8.9 days. M. anisopliae isolates ICIPE 69, ICIPE 7 and ICIPE 20 had the lowest LC50 values of 1.1 × 107, 2.0 × 107 and 3.0 × 107 conidia ml-1, respectively. In terms of conidia production, M. anisopliae isolate ICIPE 69 produced significantly more conidia than the other two isolates. ITS sequence comparison indicated that ICIPE 69 differed from the other isolates for 2bp corresponding to a restriction site SfoI which could be due to the difference in geographical origin. These results coupled with previous studies on its virulence and field efficacy to other thrips species make M. anisopliae isolate ICIPE 69 a suitable candidate for development as fungal biopesticide for thrips management. In order to understand the underlying genetic mechanism behind virulence of the M. anisopliae ICIPE 69, 8 isolates of M. anisopliae (ICIPE 7, ICIPE 20, ICIPE 30, ICIPE 41, ICIPE 62, ICIPE 63, ICIPE 69 and ICIPE 78), which had previously shown to be pathogenic to F. occidentalis, were characterized using chitinase genes (chi1, chi2, BbTrch and chi4). Results suggest that although chitin digestion is critical in fungal infection, the use of chitinase genes for genotyping might not be appropriate for virulence characterisation. The compatibility of M. anisopliae isolate ICIPE 69, with 12 agrochemicals in an integrated pest management perspective was assessed, under laboratory conditions. The agrochemicals included 5 insecticides, one botanical insecticide, 2 acaricides, and 3 fungicides, commonly used in French bean production in terms of conidia production, vegetative growth and mycelial mass. The insecticides abamectin and imidacloprid were highly compatible with M. anisopliae; thiamethoxam was compatible whereas azadirachtin and L-cyhalothrin were toxic to the fungus. The acaricide spiromesifen was moderately toxic while the fungicides carbendazim, probineb and copper-hydroxide were very toxic to the fungus. The association of the M. anisopliae ICIPE 69 with imidacloprid or thiamethoxam did not result in any synergistic or antagonistic effects on larvae of F. occidentalis. However, the combination of lower concentrations of both fungus and thiamethoxam resulted in shorter LT50 values compared to individual treatments. Our results suggest that application of the fungus along with agrochemicals like imidacloprid and thiamethoxam can be an efficient IPM strategy in pest control. The intra and inter specific effects of host-plant on the virulence of M. anisopliae on F. occidentalis was evaluated in the laboratory. Three varieties of French bean, Phaseolus vulgaris L. var. Alexandria, Julia and Samantha and Snow pea Pisum sativum L. were selected for the study. F. occidentalis cohorts (Second-instar larvae cohorts of WFT were reared for three generations before the bioassays. They were then exposed to pods of host plants previously sprayed with three concentrations of M. anisopliae (1 × 106; 1 × 107; 1 ×108 conidia ml-1). There were no significant differences in mortality between host-plants. However, the comparison of LT50 values showed that thrips were more susceptible when reared on Alexandria. Inter specific comparison showed that there were no significant differences in LT50 between snow pea and the French bean variety Julia. Results suggest that intra-specific and inter-specific host-plant effects need to be taken into consideration during pest management programmes. The performance of an autoinoculation device for M. anisopliae was evaluated in field cage experiments for the control of F. occidentalis in French bean. Treatments consisted of an autoinoculation device with semiochemical (LUREM-TR a commercial thrips attractant), a M. anisopliae treated device without semiochemical and a fungus-free device as control. Parameters assessed included conidial acquisition by thrips, thrips mortality, and conidial persistence in the device. The overall mean of conidia acquired by single insect was higher (5.0 ± 0.6 × 104 conidia/insect) in the fungus-treated semiochemical-baited device than in the device without semiochemical (2.2 ± 0.4 × 104 conidia/insect). The overall thrips mortality was also higher in the fungus-treated semiochemical-baited device (59.3 ± 3.9%) as compared to the device without the semiochemical (41.7 ± 3.5%). Viability of M. anisopliae conidial was not affected in autoinoculation device without semiochemical until 7 days after treatment. However, it was considerably affected in the autoinoculation device with semiochemical, where the conidial viability decreased from 81.0 ± 1.3%, 2 days postinoculation to 6.5 ± 1.1%, 7 days post-inoculation. There was a positive correlation between mortality and conidial persistence and a negative correlation between conidial persistence and conidial acquisition, suggesting that the semiochemical volatiles significantly affected the conidial viability which in turn resulted in significant reduction in thrips mortality despite the highest conidial acquisition in the fungus-treated semiochemical-baited device. Thrips density per plant was significantly reduced in both the autoinoculation device with semiochemical (autoinoculation adults/plant) autoinoculation device without semiochemical (8.7 ± 1.7 adults/plant) and autoinoculation (6.6 ± 1.4 adults/plant) as compared to the control (19.8 ± 2.6 adults/plant). These results demonstrate the prospects of autoinoculation device strategy for dissemination of M. anisopliae in the control of thrips, particularly in screenhouses. The effects of the entomopathogenic fungus M. anisopliae (ICIPE 69) infection on T. tabaci feeding and its competence to vector IYSV in onion plant were investigated. Newly-emerged T. tabaci larvae were allowed to feed on IYSV-infected onion leaves in order to get infection and to become adults. Newly-emerged adult T. tabaci were treated with three sublethal concentrations of M. anisopliae (105; 106 and 107 conidia ml-1) and placed in a sterile 9-cm Petri dish on a clean onion leaf-disc and allowed to feed for 5 days. The numbers of feeding punctures were recorded daily for five days. The viral titer in adult insects and onion leaves was measured using DAS ELISA technique. Infection by M. anisopliae significantly reduced (ANOVA: F3, 480 = 50.4; P < 0.0001) the feeding punctures by adult T. tabaci. Although there was no significant effect of M. anisopliae infection on IYSV titer in adults T. tabaci, virus transmission was significantly reduced (ANOVA: F3,240 = 14.4, P = 0.0001) in M. anisopliaeinfected insects compared to the control. There was a significant positive correlation between feeding punctures and IYSV transmission in onion leaf-disc, suggesting that M. anisopliae application in addition to the effective control of the vector, Thrips tabaci can also influence IYSV propagation by reducing T. tabaci feeding punctures. However, further investigations are needed to determine effect of M. anisopliae on IYSV acquisition by T. tabaci.