Enhancing Biological Control With Insectary Plantings

Enhancing Biological Control With Insectary Plantings

Summary

This multi-year project examined the use of beneficial insectary plantings to enhance biological control in commercial broccoli fields in western Oregon.
We have conducted various on-farm experiments in a broccoli (Brassica oleracea) production systems to explore the potential of using insectary flowers to enhance the biological control of cabbage aphid (Brevicoryne brassicae) by naturally-occuring predacious hoverflies Our conclusions include:
(1) Insectary plantings were shown to increase abundance of predator eggs on broccoli fields, however no suppression of cabbage aphid was demonstrated.
(2) The use of insectary plants may cause more harm than good in farming systems since pest insects as well as beneficials use nectar and pollen from planted floral resources. This approach to conservation biological (beneficial insectary plantings) must consider multitrophic level impacts on pests, beneficials, and hyperparasitoids across all crops occurring within the system. This research cannot focus solely on a single crop/pest/beneficial relationship because other key interactions in the system may be neglected.
(3) Selected insectary plants exhibit preferential attractiveness among entomophagous and pest species
(4) Biological pest control efforts need to be conducted at the species level of ecological inquiry, with considerable error possible from decisions made at the family level of taxonomic groupings.
(5) Appropriate scale of experimental design needs to incorporate dispersal behaviors of target species and natural enemies.
(6) Oviposition (egg-laying) behavior of adult entomophagous species may be key factor limiting effective biological control

Objectives/Performance Targets

1. To evaluate the relative attractiveness of selected insectary plants to entomophagous arthropods and key insect pests
2. To evaluate the potential of using beneficial insectary plants to enhance biological control of specific insect pests in broccoli production systems, including: (1) the cabbage aphid complex and (2) the worm complex.
3. To develop a multi-faceted educational program for growers and agricultural professionals on integrating beneficial insectary plantings into various kinds of farming operations to enhance biological pest control.

We have previously reported on Project Objectives 1 above. This year’s report focuses on work associated with Objective 2. The following are specific sub-objectives for Objective 2:
1. To quantify the within-field effect that added flowering plants have on the attraction and oviposition activities of hoverflies, and on the abundance of other arthropod natural enemies and herbivores in broccoli.
2. To evaluate the prey-finding and oviposition behaviors of adult hoverflies in commercial broccoli fields.
3. To evaluate the ability of hoverflies to limit populations of aphids on single plants in controlled cage and field experiments.

Materials and Methods
Objective 1. To quantify the within-field effect that added flowering plants have on the attraction and oviposition activities of hoverflies, and on the abundance of other arthropod natural enemies and herbivores in broccoli.
A field experiment was set up in a 160 x 400 m broccoli field at Stahlbush Island Farm, 3 km East of Corvallis, OR. Winter season vegetation was killed with Roundup in March 2001, and a strip-tillage machine used to till 4 cm wide strips on 0.9 m centers. and broccoli was transplanted on May 28 along with a soil incorporation of Neemix and Goal (Oxyflourfen). An 18 m (= 20 row) block of broccoli (running the whole 400 m length of the field) was left untreated with the aphicides which were applied to the rest of the field (Provado 43 days after transplant and Pyrin 67 days after transplant). This block was 90 m from one border and 50 m from the border of the other side. Two insecticide applications were made for lepidopteran pests, to the strip and the rest of the field (Confirm 43 days after transplant and Success 67 days after transplant).
Within the 18 x 400 m strip, two “insectary flower” plots (18 x 18 m) were established. Alyssum seedlings (Lobularia maritima var. ‘New Carpet of Snow’) were planted, with a spacing of 45 x 90 cm, 24 days after broccoli transplant. One of the blocks was set 90 m from one end of the strip, and the other 90 m from the other end, with a distance of 180 m between the two flower blocks.
Broccoli crop plants were sampled for arthropod abundance every 10 m from 0 to 80 m in the North and South directions from each flower plot. We visually examined six randomly-selected broccoli plants from evenly spaced rows in the 16 middle rows (rows 3, 6, 9, 12, 15, and 18) in each 10 m section of the 18 x 400-m no-aphicide block. Crop plants were sampled every 4-6 days from 19 days after broccoli transplant (= 5 days pre-alyssum transplant) until broccoli harvest. Each plant chosen in the 10 m section had its 6 largest leaves examined for the following pests and natural enemies: hoverfly eggs and larvae, cabbage aphid (CA), green peach aphid (GPA), potato aphid, imported cabbage worm (ICW), cabbage looper, diamond back moth, hoverfly larvae and eggs, cecidomyiid larvae and eggs, parasitized aphid mummies, spiders, lady beetles and lacewings. Herbivorous pests were further categorized into developmental stage.
The abundance of aphids, parasitized aphid mummies and hoverfly eggs at different distances from the flower plots were also sampled using bait plants of potted broccoli containing cabbage aphid colonies. These plants were produced in the greenhouse by forming aphid colonies of 10-30 apterous individuals within 13 mm diameter clip cages on 3 leaves on 3 week old broccoli seedlings in 4 l pots. These pots were then buried in the ground at distances of 0, 1, 5, 10, 20, 40 and 80 m from the flower plots along transects radiating in all 4 compass directions (there was only room for 40 m transects in each of the eastward transects however). Bait plants were left out in these transects for 3-5 days, then all aphids and hoverfly eggs were counted and recorded. All leaves containing aphids for each plant were then placed in a paper bags, allowed to sit for one week to permit the formation of mummies, and then placed in the freezer to prevent eclosion of parasitic wasps before the mummies were counted. Bait plants were placed out for a total of 8 rounds, starting 15 days after broccoli transplant and ending at harvest. All hoverfly eggs encountered on both crop and bait plants were collected, photographed and placed in Petri dishes with moist cotton wool and aphids to identify the hoverfly species depositing the eggs, once development was complete.
To measure the presence of adult flies, transects of yellow pan traps (18 cm diameter Solo® plastic bowls painted with Rustoleum® ‘safety yellow’) were placed in transects oriented identically to the bait plants. Pan trap height was kept within 15 cm of the upper broccoli canopy surface. Adult hoverflies and ladybird beetles were collected from each trap and stored in 70% alcohol, recording the number of trap-days. Pan traps were placed over a total of 9 rounds, from 15 days after broccoli transplant until harvest. Yellow sticky traps (2-sided, 18 x 40 cm) were staked in the same transects for one round, the week prior to harvest. Sticky trap height went from 20 cm below to 20 cm above the upper canopy surface. To check if the two types of yellow traps were collecting different species of hoverflies in equivalent proportions to those seen in the field, the alyssum flowers were observed on a weekly basis by both stationary and ‘census’ walking methods of observation.
Just before harvest, mature broccoli heads were randomly selected in the same grid described above for the crop plant sampling, for a total of 192 heads. Each head was examined for 5 minutes by destructive sampling of the heads into pieces smaller than 1cm3.
Analysis. The analysis of the crop and bait plant data will be by multiple linear regression, with hoverfly egg number as the response variable and distance from flowers and aphid number as the main explanatory variables. Explanatory variables of block, direction of sampling transect, and date, will also be considered for the analyses. The pan trap data will be analyzed by simple linear regression with distance, block, transect direction and date, as the explanatory variables. To see if the response varies by type of hoverfly species, a multivariate analysis of the different eggs types sampled will also be considered.
Objective 2. To evaluate the prey-finding and oviposition behaviors of adult hoverflies in commercial broccoli fields.

Research Questions and hypotheses. The oviposition behavior of hoverflies in relation to the aphid distribution on each sampled broccoli plant in the field was assessed according to the following observational research questions: 1) How many aphids per leaf, or per plant, are needed before hoverfly eggs are seen? 2) At what aphid densities are the peak numbers of hoverfly eggs observed? 3) If these thresholds and peaks exist, do they vary by hoverfly species, by aphid species, by aphid colony type, by field site, or over time? 4) Does this oviposition response vary on different parts of the broccoli plant?, and 5) How are the different aphid species distributed on different parts of the broccoli plant, especially at the end of the season?
Methods. Data for these questions were obtained from the same two field experimental sites at Stahlbush Island Farm described in the 2000 annual report to SARE. Data were collected in two 15 x 66 m rectangles of broccoli, untreated with aphicides, at each of the two fields. Sampling began 25 days after broccoli transplant, and was repeated every 4 days until harvest. In each rectangle, every leaf on 50, randomly selected, broccoli plants was examined for the presence of all aphid species, hoverfly eggs, and the other key arthropods mentioned above in the methods for ‘Objective 1’. The numerical rank position of each leaf (starting from the center and radiating out to the lower leaves) was also recorded. At the 10-leaf stage of broccoli plant development, data from each leaf of each plant were recorded in separate categories of ‘inner’ (leaves closest to head under 6cm2), ‘middle’ (leaves next closest to head over 6 cm2), and ‘lower’ (leaves closest to ground which had begun to senesce).
Analysis. The analyses of these data will also be through multiple linear regression, with number of hoverfly eggs as the response variable and numbers of aphids per leaf and per plant as the main explanatory variables. Aphid species, aphid colony type, field site and leaf location on plant will also be considered as explanatory variables in additional analyses.
Objective 3. Method development for single plant behavioral studies: To evaluate the ability of local hoverfly species to limit populations of aphids on single plants in controlled cage and field experiments.
Preliminary work for this objective has taken the form of a greenhouse trial varying hoverfly larva/aphid ratios on individual broccoli plants in a replicated glasshouse bench trial. The hoverfly species that has shown the highest rate of oviposition at aphid colonies in several commercial broccoli field experiments, Eupeodes fumipennis, was chosen for this experiment
Second instar larvae belonging to the same cohort were taken from a colony of this species (a description of this colony is given below) and placed on broccoli plants with cabbage aphids in the following ratios: 1/10, 1/50 and 1/100. These ratios were matched with positive control treatments of: 0/10, 0/50 and 0/100, and all six treatments were replicated four times. Broccoli plants in gallon pots, 40 days post transplant, which had at least 6 leaves over 15 cm2 were stripped down to these 6 leaves. The aphids were applied to one of the central leaves and allowed to equilibrate on the leaf in clip cages for a period of 24 hours in individual ‘colony’ batches no larger than 25 aphid individuals. Resulting aphid numbers were recorded every 12 hours for a period of 5 days.
To objectively assess the predatory activity of local hoverfly species in experiments on individual plants, a continuous supply of uniformly aged and naïve hoverflies are needed. At present we have a recycling colony of the species that has demonstrated the highest potential in the field, Eupeodes fumipennis.
Hoverfly culturing. Attempts at hoverfly culturing began in early Spring 2001, following the preparatory work necessary for this operation, including greenhouse setups with cages, broccoli plants, aphid colonies, floral resources and artificial food resources. Initial phases involved the collection of any commonly available hoverflies from the field. They were then placed in 1m3 no-see-um mesh cages containing various combinations of: aphid species (cabbage aphid, green peach aphid and pea aphid), aphid plant hosts (broccoli, collards, brussels sprouts, kale, radish, bell beans, bush beans and peas), floral resources (alyssum, coriander, phacelia, buckwheat and baby’s breath), artificial food resources (sugar cubes, honey, hazel pollen, oak pollen, grass pollen, bee-collected pollen, commercial protein extract and sugar water), as well as temperature.
Certain fly species survived cage conditions and oviposited on aphid-bearing plants. These plants were then placed in covered buckets where subsequent larval and puparial development was tracked, while adding additional aphid food as needed. Those flies that eclosed from puparia in the buckets were then placed in cages with other eclosed flies of the same species to see if mating and another round of oviposition would occur.
Ten species of hoverflies were collected, demonstrating differential abilities to survive, oviposit on aphid-bearing plants, develop into adults, mate, and repeat the cycle. Those species that have made it through a complete life cycle under artificial conditions to date are Eupeodes fumipennis and Sphaerophoria sulphuripes.
Results and Discussion
Objective 1. To quantify the within-field effect that added flowering plants have on the attraction and oviposition activities of hoverflies, and on the abundance of other arthropod natural enemies and herbivores in broccoli.
Crop plants. Hoverfly eggs first appeared in low numbers, 2-3 weeks before harvest, but did not appear on broccoli crop plant leaves to any considerable extent until the last few sampling dates, within a week of harvest. There is a trend of greater oviposition at distances closer to the flowers on the final sampling date. Through the rearing of collected eggs, the majority of eggs on all sampling dates were found to be either Eupeodes fumipennis or Sphaerophoria sulphuripes (data not shown).
Bait plants. Hoverfly eggs appeared about the same time on the bait plants as on the leaves of the broccoli crop. However, the trend of greater oviposition on leaves at sampling points closer to the flowers was not observed on the latest dates for this sampling method. This was true for both blocks, and all transect directions (graphs not shown).
Pan traps. Hoverfly adults appeared in the pan traps in the broccoli crop only after the alyssum flowers were planted. Large numbers were found in the traps by the 6th sampling date, two weeks later. At this time in the broccoli crop season, capture rate peaked at distances 5 m from the flowers, and gradually decreased out to 80 m. Traps at distances of 0 m and 1 m from the flowers also trapped fewer flies on this date.
More than 90% of the flies captured in the traps on this date were Toxomerus marginatus (data not shown). The data from the final three pan trapping dates has not been processed yet. Yellow sticky card traps did not capture local hoverfly species (sticky trap data not shown). Several species of ladybird beetles were captured in large numbers, but no distance relationship to flowers was observed. Green lacewings were trapped at distances close to the flower plots, but in lesser numbers than those seen for the ladybird beetles.
Insect Contamination in Broccoli Heads. Out of 192 broccoli heads sampled in the no-aphicide strip, only 7% contained aphids. Of these, 14 heads, none had more than 10 aphids. When these data are broken down by distance class, no effect with distance to flowers was seen.
Objective 2. To evaluate the prey-finding and oviposition behaviors of adult hoverflies.
Processing of data collected for this objective in the 2 field trials in 2000 is continuing and certain trends are already evident. The dataset is large, consisting of aphid and hoverfly egg counts on each individual leaf on each sampled plant. The analysis should be complete in the spring of 2002. The information in Figure 6 is included here to show one trend that has been observed in many of the sampling dates looked at so far, that is, plants with less than 50 aphids did not have hoverfly eggs. No such threshold has yet been seen for individual leaves, many leaves with no aphids had hoverfly eggs. No peaks in oviposition response have been observed per leaf or per plant, rather, what has been seen is a sustained increase in number of eggs with an increasing numbers of aphids. Remaining data manipulations and analyses to address this objective are outlined in the ‘Future Work’ section below.
Objective 3. To evaluate the ability of local hoverflies to limit populations of aphids on single plants in controlled cage and field experiments.
Greenhouse studies were conducted in 2002 to evaluate relative voracity of key hoverfly species to consume aphids. Data clearly showed that Eupeodes fumipennis consumed the greatest number of aphids in the shortest time period, however Syrphus opinator and Sphaerophoria sulphuripes were also shown to have high levels of consumption of aphid prey. These data clearly support the importance of the predators in aphid biological control. (Data to be presented in the final report).
Discussion
The first objective of quantifying the effect that added flowering plants have on the activity of predatory hoverflies was addressed by measuring the amount of: oviposition on crop plants, oviposition on bait plants, aphids on crop plants, aphids in broccoli heads, and trapped adult hoverflies at different distances from the flowering plants over most of the crop’s development.
Oviposition did not occur to any significant extent until just before (less than one week) broccoli harvest. Much lower levels of oviposition were recorded on both crop and bait plant leaves for the two weeks prior to this period. This same result was seen in both field trials reported on in the 2000 annual report to SARE. It was hypothesized in the discussion of the 2000 report that providing floral resources sooner, and in greater amounts might induce earlier, and greater amounts of, oviposition. The 2001 trial used over 6 times as many flowers provided approximately one month earlier in the broccoli field. Although the onset of hoverfly oviposition occurred at the same time as that seen in the 2000 trials (i.e. 3 weeks prior to harvest), the relative proportion of plants with hoverflies was higher at the end of the season. Also, unlike the 2000 trials, oviposition on crop plants was generally greater at sampling points closer to the flowers.
This same trend, of reduced oviposition at distances greater from the flowers, was not seen with the bait plant sampling method however. These differences may not be surprising when the differences in plant quality, size and number between these two sampling methods are considered. Additionally, since the oviposition activities of predatory hoverflies throughout a crop field are also influenced by the relative amounts of aphid hosts at each sampling point, the most appropriate analysis of oviposition response as a function of distance from floral resources will also take into account the number of aphids present at each sampling point. Once these aphid data are included in the analysis, the spatial patterns of oviposition response on both crop and bait plant sampling points at different distances from the flowers may be different from that reported here.
These aphid data, once processed, will also be used to assess the indirect effect that the floral blocks may have on aphid infestations at varying distances from the blocks on the last few sampling dates. Indirect effects on aphid infestation in the broccoli heads were not seen. The very low incidence of aphid-infested broccoli heads at all distances from the flower blocks sampled in the field (in terms of both relative percentage of infested heads and number of aphids in heads) probably indicates that the overall infestation of the heads in this no-aphicide strip was insufficient to detect any distance relationship which may have occurred. Levels of other key arthropod herbivore pests and natural enemies on both crop and bait plants were also too low in the sampling area to identify any spatial or temporal relationship with the added flower blocks.
More than 90% of the flies trapped in the first 6 pan trap sampling dates were the species Toxomerus marginatus, a species which has not been observed to oviposit on broccoli plants with aphids in local broccoli fields. The data from the last three trapping dates remain to be processed, but certain trends are already evident at this point.
The pan trap data from the 6th sampling date, midway through the broccoli season, showed the total number of adult hoverflies trapped (mostly represented by T. marginatus) to taper off from a peak at 5 meters, showing a greater number of hoverflies flying in zones closer to the flower blocks. Very low trap numbers at the 0 meter distance, and moderately low numbers at the 1 meter distance, indicate that flower-mimicking yellow pan traps are less attractive when a dense block of real flowers is in the immediate vicinity.
Although eggs of E. fumipennis, S. sulphuripes and Platycheirus spp. did not appear until 3-4 weeks after the alyssum flowers were transplanted, adults of at least some individuals of these species were trapped within days of flower transplant. The reason for this lag could be either insufficient egg maturation in hoverflies present on these early sampling dates, insufficient numbers and arrangements of aphids to elicit oviposition responses, or a combination of both of these. This will be investigated further by dissecting the ovaries of the preserved specimens trapped on each date to assess egg development over time. These data will then be compared to the information derived from the analysis of oviposition response to varying levels of aphids on crop and bait plant leaves over time, to assess the relative importance of each.
We are continuing to develop appropriate experimental methodology to examine the ability of locally-occuring hoverfly species to limit aphid populations on broccoli. Our success with hoverfly culturing, suggests that this experimental approach can be developed further. The early results demonstrated that a combination of glasshouse conditions and host plant quality (particularly the removal of most leaves) affect hoverfly larval survival. Attempts will be made to enhance microclimate, and to improve the colonization and survival rate of larvae, by using clip cages for an initial period of acclimation.
This technique is could provide valuable, quantitative insights into the rate and patterns of hoverfly predation, interactions with other predators, and the ability of hoverflies to limit aphid population growth and the colonization of broccoli heads. Studies of this type are expected to provide more accurate information than simple voracity studies in Petri dishes, which can only provide minimum and maximum feeding rates.

Impacts and Contributions/Outcomes

Impact and Results
Late season predation of cabbage aphids by predacious hoverfly larvae may have an important impact on the suppression of this pest and preventing excessive contamination of broccoli heads by these pests. We have documented high numbers of predacious hoverfly eggs in broccoli prior to harvest and we have developed methods to identify the key predacious hoverfly species. In our field experiments to date, however, we have been unable to demonstrate an enhancement of biological control of aphids by planting blocks of insectary flowers either on the margin of the field or in within the fields. We have made significant steps in understanding the key aspects of oviposition behavior of Eupeodes fumipennis, the major predacious hoverfly species occurring in broccoli in Oregon. There seems to be an “oviposition threshold” of approximately 50 aphids per plant that is required before the adult female hoverflies will begin to lay eggs on broccoli plants.
Economic Analysis
This research has contributed significant findings to the science and practice of biological control. However, because we have not been able to demonstrate actual pest reduction resulting from our efforts to enhance biological control through the provisioning of floral resources, we have not pursued an economic analysis of this approach compared to other pest control practices. We do not feel that an economic analysis would have any relevancy here. Further applied research is required to document efficacy in enhancing biological control before implementation strategies can be addressed.
Publications and Outreach
Invited Presentations:
Luna, J. M. 2002. Applying theory to practice in conservation biological control: Lessons from a model system using broccoli, cabbage aphids, and predacious hoverflies. National Meeting, Entomological Society of America. Fort Lauderdale, FL.
2001. Sustainable agriculture as part of working landscapes. Oregon Sustainability Conference, Portland, OR.
2000. Habitat management strategies to enhance conservation biological control. National Meeting, Amer. Soc. of Agronomy., Minneapolis, MN.
We are currently working on an OSU Experiment Station Bulletin on the identification of predacious hoverflies in Oregon farm land. We are also currently preparing three manuscripts to be submitted to refereed scientific journals within the next year.
Farmer Adoption
Stahlbush Island Farms, near Corvallis, has planted beneficial insectary flowers in more than 200 acres of broccoli in 2001. However, because of our inability to document beneficial impacts of this practice, we have not pursued an educational program that would encourage grower adoption.
Areas Needing More Study
Objective 1. To quantify the within-field effect that added flowering plants have on the attraction and oviposition activities of hoverflies, and on the abundance of other arthropod natural enemies and herbivores in broccoli.
Three field experiments have now been completed, adding to a number of other investigations over the last five years. A meta-analysis of data from all of these investigations will be undertaken to determine patterns of hoverfly oviposition relative to plant development and aphid abundance. This analysis will be used to build hypotheses for the development of further field experiments. It will also be used to provide a summary of findings to be communicated to growers and to be incorporated into extension publications. Aphid outbreaks have consistently occurred at a late stage in the season, and hoverfly population build up has consequently, also been at a late stage in crop development. Glasshouse investigations will provide information on the contribution that this late phase of predation may make to aphid colonization of broccoli heads. Further field investigations, or field monitoring exercises, which may be the subject of further grants, will determine the probability that hoverflies could limit aphid population growth in the early season outbreaks that are the most damaging to the crop. An outline of these hypotheses and proposed approaches will be provided in the final report.
Objective 2. To evaluate the prey-finding and oviposition behaviors of adult hoverflies in commercial broccoli fields.
Analysis of this large data set will be completed in the next threes. The analysis should provide a basis for predicting the threshold densities of aphids that elicit egg laying. These predictions will be tested in the meta-analysis of independent data sets referred to above. It will also be possible to test these predictions in small, replicated plot experiments where artificial aphid outbreaks will be created in screen houses, and in the open field, at different times in the season. This will also assist with resolution of the question of seasonal timing of hoverfly oviposition, relative to aphid arrival and population build up.
Objective 3. To evaluate the ability of local hoverflies to limit populations of aphids on single plants in controlled cage and field experiments.
This experimental approach will be the main approach undertaken in 2002. Initial work will involve the resolution of experimental design problems mentioned above. Usable experimental designs resulting from this work will then be used to first investigate and quantify the ability of E. fumipennis larvae to limit aphid populations on individual broccoli plants. Potential follow-up studies after this could include investigations of the aphid-limiting abilities of other local hoverfly species, host range studies of the most common local species as well as the role of inter- and intra-specific interactions between larvae on plants with aphids.

Collaborators: