Optimizing the spray application of entomopathogenic nematodes in vegetables: from spray tank to nematode deposition

Abstract

Several insect pests of economic importance are encountered on vegetable crops. Being high value crops, the introduction of biological control agents such as entomopathogenic nematodes (EPN) has stimulated great interest for above and below-ground pests. However, despite promising laboratory and field trials, success with EPN in vegetables has rarely been achieved in field applications under practical conditions. Improvements in application technology for EPN that aim at minimizing losses during the transfer from the mixing tank to the target insect, increasing the competitiveness of EPN, are badly needed. Twenty years ago, the effect of application technology on EPN was underestimated. Laboratory experiments investigated the possible detrimental effects of application equipment and resulted in some rules of thumb, e.g., considering spray liquid temperature, nozzle size and pressure. Numerous succeeding studies, however, indicated that those restrictions do not prevent nematode damage. The general objective of this thesis was, therefore, to systematically screen all steps of the spray application process, from spray tank to nematode deposition, for possible detrimental or efficacy reducing effects on EPN applied in vegetables. Research started with a literature review on the effects of application technique on EPN, and on the characteristics of three important vegetable pests (cabbage maggot, onion thrips and cabbage moth) and their control with EPN. Experiments were designed to investigate two important processes of the spray system, i.e., agitation and spray formation. More practical oriented experiments were performed to study the effect of the spray application technique on nematode deposition. Finally, the effect of spray volume on nematode deposition, viability and infectivity was studied. For the application of EPN, a good agitation system is indispensable as the nematodes tend to sediment fast in a spray tank without agitation. Three agitation systems, viz. mechanical, pneumatic and hydraulic agitation were tested for their ability to keep the nematodes suspended. Hydraulic agitation was tested using a centrifugal and a diaphragm pump. Nematode damage was quantified based on viability and infectivity of the EPN. Mechanical agitation at a speed of approximately 696 revolutions min-1 did not influence the viability nor the infectivity of the nematodes. Also pneumatic agitation during 120 min was not detrimental to the EPN. Viability, nor infectivity were affected. However, the effect of hydraulic agitation differed depending on the pump type used. Hydraulic agitation using the diaphragm pump did not harm the nematodes, whereas the use of the centrifugal pump clearly affected viability. After 120 minutes of recirculation, only 19.3% of the nematodes survived; infectivity was even reduced to 0%. An additional experiment revealed that the temperature increase, from 21.7°C to 45.4°C, was responsible for the observed nematode damage. The ability of the agitation system to keep the nematodes in suspension was examined by comparing the nematode concentration observed in the samples taken at different agitation times. These measurements showed that the pneumatic agitation was unstable. Agitation during 120 minutes using the other agitation systems resulted in a significant loss of nematodes at 15 cm above the spray tank bottom. In general, the experiments prove that the agitation systems developed to agitate a chemical solution are not always suited to agitate a nematode suspension and can attribute to reduced efficacy of EPN. One of the major considerations related to the selection and use of an application system is the application distribution pattern. A completely uniform distribution of the nematodes in soil applications is not essential because the nematodes can move over short distances; however, uniform distribution is important in foliar EPN applications. The volumetric distribution pattern of EPN was tested beneath four nozzle types, i.e., a standard flat fan nozzle (TeeJet XR 110 08), an air induction flat fan nozzle (TeeJet AI 110 08), a drift reducing deflector type nozzle (TeeJet TT 110 08) and a TwinJet spray nozzle (TeeJet TJ60 110 08). A comparison with the distribution of a chemical tracer (Brilliant Sulfo Flavine, BSF) was made to reveal possible distribution problems. A significant effect of nozzle type on the distribution of EPN beneath a spray nozzle was found. The differences between the nematode distribution and the distribution of the chemical tracer were negligible for the flat fan and the TwinJet nozzle. Small differences were measured for the air induction nozzle, while a remarkable difference in EPN-BSF distribution was found for the deflector nozzle. The nematode concentration shows a sharp peak in the center of the spray fan and declines much faster toward the edges as compared with the BSF concentration. Ideally, the distribution of EPN beneath the spray boom should be uniform. A theoretical calculation of the coefficient of variation of the nematode distribution beneath a spray boom was performed and showed that the spray overlap from a spray boom decreases differences in nematode distribution. An acceptable value for the coefficient of variation was found for all nozzles, except for the TwinJet nozzle where the coefficient of variation was slightly above 7%. Effective control of insect pests using EPN requires more than a judicious choice of the nematode species. The biological agent must also be delivered in a way that enables the nematodes to infect the host. The optimization process needs to account for the particular requirements of the EPN species used, the target pest, and the crop. The effect of spray application equipment was studied on the deposition of EPN in five pest control applications, viz. cabbage maggot (Delia radicum) and cabbage moth (Mamestra brassicae) in both cauliflower (Brassica oleracea var. botrytis) and savoy cabbage (Brassica oleracea var. sabauda) and onion thrips (Thrips tabaci) in leek (Allium porrum). The nematode deposition (number of nematodes) and spray pattern (distribution of EPN in the droplets) were compared after applying a nematode suspension using a 5-nozzle spray boom equipped with flat fan (TeeJet XR 110 08), air induction flat fan (TeeJet AI 110 08) or TwinJet spray nozzles (TeeJet TJ60 110 08). Two additional spray application systems, viz. air support and row application, were evaluated on their effectiveness to deliver nematodes to their target site. Deposition of Steinernema feltiae was observed in the leek shaft (habitat of onion thrips) and at the cauliflower and savoy cabbage foot (habitat of cabbage maggot), while the deposition of S. carpocapsae was measured at the underside of cauliflower and savoy cabbage leaves (habitat of cabbage moth). Control measurements were performed using Petri dishes filled with water. Empty Petri dishes were used after spraying to measure the spray pattern. The cabbage plants were young but their foliage already sheltered the nematode spray significantly; approximately 40% of the applied nematodes did not reach the foot of the plants. Changing nozzle type did not affect the deposition results, except for the TwinJet nozzle. A spray boom equipped with these dual fan nozzles delivered significantly fewer EPN at the foot of the cauliflower as compared with the flat fan nozzle. The use of an air support system or a row application system improved nematode deposition at the foot of the savoy cabbage. These systems, however, did not significantly increase deposition at the cauliflower foot. With the standard boom spray application technique, relative nematode deposition on the bottom side of the savoy cabbage leaves was 27.2%, while only 2.6% of the applied nematodes reached the bottom side of the cauliflower leaves. Neither nozzle type nor application technique affected nematode deposition at the bottom side of the savoy cabbage or cauliflower leaves. After spraying leek with a standard boom, a low nematode deposition was measured in the leek shaft. With this technique, only 7.3 EPN cm-2 reached the transition zone in the leek shaft. Changing nozzle type or using the air support system did not significantly increase nematode deposition in the leek shaft. Overall, no differences in nematode deposition were found on Petri dishes at different sampling positions along the spray boom, thus an even nematode distribution was obtained underneath the 5-nozzle spray boom. Relative deposition on the horizontal Petri dishes, calculated based on the theoretical maximum deposition, was significantly higher for the air induction nozzle as compared to the flat fan nozzle for both nematode species. Nozzle type also affected the spray pattern of nematodes. The Petri dish surface covered with nematode containing droplets was very low and varied from 13.6% (flat fan nozzle) to 15.8% (air induction nozzle). The experiments provide evidence that EPN frequently do not reach their target sites using standard application techniques. Moreover, the nematodes reaching their target are applied in droplets that cover only a small part of the treated surface. The use of adapted spray application techniques that direct the spray to the target site are indispensable to increase chances for contact of EPN with their target and to result in a cost-effective and reliable application. Spray volume can influence the amount of free water on the leaf surface and subsequently the ability for EPN to move. Therefore, the effect of spray volume, viz. 548, 730 and 1095 L ha-1, was investigated on the deposition, viability and infectivity of EPN against Galleria mellonella on savoy cabbage, cauliflower and leek. Leaf disks and filter paper disks, placed at different angles to the spray nozzle, were exposed to a nematode spray. Increasing spray volume decreased nematode deposition on top of 7.1 cm² leek leaf disks in a 15° angle with the spray nozzle. Although the number of living nematodes observed after 240 min of incubation (24°C and 60% RH) was not significantly different between the low and the high volume application, a greater infectivity was obtained in the latter application. No significant effect of spray volume was observed on the relative deposition of S. carpocapsae on the bottom side of cauliflower and savoy cabbage leaf disks. Despite the low S. carpocapsae deposition on the bottom side of the savoy cabbage disks, high infectivity was obtained against G. mellonella. Using the lowest spray volume, infectivity decreased with increasing exposure time, while infectivity was not affected by exposure time when a spray volume of 730 L ha-1 or more was used. Based on these experiments, spray volume can be considered as an important application parameter since it can affect nematode infectivity. The results of this research confirm that the technique used for the spray application can have severe efficacy reducing effects on EPN. These effects can be observed in the different steps of the application process and vary from nematode death, loss of nematodes and problems to keep the nematodes suspended in the spray tank to difficulties to reach the target pest and covering the target site evenly. The application of chemical insecticides is known to be very inefficient since only small amounts of the applied product reaches the insect, but eventually results in acceptable pest control due to the high persistence of the chemicals. Entomopathogenic nematodes, however, have low persistence on exposed surfaces and are currently too expensive to be applied in excessive amounts. More research is therefore badly needed to develop new or adjusted spray equipment or other application techniques which can deliver nematodes more efficiently at their target site in large field applications