Diaprepes abbreviatus L. (Coleoptera: Curculionidae) is an occasional root pest of plasticulture strawberry in central Florida, USA. There are few chemical insecticide options for larval D. abbreviatus in strawberry. Therefore, we tested soil-applied aqueous Steinernema riobrave Cabanillas, Raulston and Poinar (Rhabditida: Steinernematidae), which is used for D. abbreviatus control in citrus. When S. riobrave was applied (100 infective juveniles per cm2) to the root zones of plants in a D. abbreviatus-affected area of a commercial strawberry field, less than 12% of plants were severely wilted or dead 17 d after treatment, whereas 23% of plants in control plots were wilted or dead. In research plots, peripheral plants and a central plant in each plot were infested with 4 late-instar D. abbreviatus and treated with 1 or 2 applications of S. riobrave (25 infective juveniles per cm2), 1 application of imidacloprid or water (control). Dead S. riobrave-infected D. abbreviatus larvae were recovered from plots to which S. riobrave was applied, but there was no effect of treatment on numbers of live larvae recovered 1, 2, and 3 wk post-application in the peripheral plants. At the end of the experiment, no live D. abbreviatus larvae were recovered from the central plants or the plants proximal to the central plants in plots treated once or twice with S. riobrave. Plant wilting and death due to larval D. abbreviatus root feeding was minimal (averages of 1–1.4 on a 1–5 rating scale) in plants proximal and distal to the central D. abbreviatus-infested plant. Treatment did not affect plant wilting and death rates in the proximal plants, but death of distal plants occurred only in control plots. Our results showed S. riobrave infected and killed late-instar D. abbreviatus in plasticulture Florida strawberry, and further research should be conducted to optimize S. riobrave applications and develop it into a management strategy for D. abbreviatus.
Diaprepes abbreviatus L. (Coleoptera: Curculionidae) is a polyphagous root weevil and pest of approximately 300 agricultural, ornamental, and wild plants (Simpson et al. 1996; Mannion et al. 2003). In citrus (Citrus spp. L.; Rutaceae), adults may damage trees by scoring young leaves; however, larval root feeding causes significant tree damage, leading to yield reductions and eventually tree death (Woodruff 1985; McCoy 1999; Knapp et al. 2000; Stuart et al. 2006). Adults are active from spring to autumn; an adult female lays an average of 11,000 eggs in her life span (Nigg et al. 2004), and neonates drop from foliage to the soil where they feed on roots and develop over several mo.
Florida, USA, is a major source of winter strawberries (Fragaria × ananassa Duchesne; Rosaceae) in the USA, with 4,000 planted ha and a production value of nearly $300 million USD (USDA/NASS 2018). Development of effective pest management practices will help enable Florida producers to be viable economically in an increasingly competitive market space (Suh et al. 2017). While the primary arthropod pests in Florida strawberry are twospotted spider mite (Tetranychus urticae Koch; Trombidiformes: Tetranychidae), flower thrips (Frankliniella occidentalis [Pergande]; Thysanoptera: Thripidae), and Frankliniella bispinosa Morgan; Thysanoptera: Thripidae), chilli thrips (Scirtothrips dorsalis Hood; Thysanoptera: Thripidae), and fall armyworm (Spodoptera frugiperda [J.E. Smith]; Lepidoptera: Noctuidae), secondary pests such as D. abbreviatus, occasionally require control (Caruso et al. 2019). Root feeding by larval D. abbreviatus causes foliar wilting often followed by plant death. Eggs are laid on young strawberry plants shortly after planting in early autumn by late-flying female D. abbreviatus, and damage typically is concentrated in small patches near field edges. However, plant damage was more widespread in a few fields adjacent to old, neglected, or recently uprooted citrus groves (J. Renkema, personal observation). Insecticides are available for weevils and grubs in Florida strawberry (Renkema et al. 2019), but they either are not labelled for application through drip irrigation to control root-feeding larvae or have long pre-harvest intervals not compatible with in-season harvest schedules. For insecticides that may be chemigated, growers typically are not able to restrict applications to areas of a field where damage occurs, and treating entire fields is not economical or sustainable. A tactic for control of D. abbreviatus in small areas of strawberry fields is needed.
Diaprepes abbreviatus has been managed successfully with the entomopathogenic nematode Steinernema riobrave Cabanillas, Raulston and Poinar (Rhabditida: Steinernematidae) in Florida citrus (Dolinski et al. 2012). Steinernema riobrave, an indeterminate forager, was first found in southern Texas, USA, and is acclimated to warm soils (Kaspi et al. 2010). Applications of commercially produced S. riobrave have caused larval mortality rates of D. abbreviatus in the field of 50% to greater than 90%, depending on application rates and other factors such as soil conditions (Duncan et al. 1996, 2003; Bullock et al. 1999). Over a yr, adult D. abbreviatus emergence was reduced by half by S. riobrave in one study (Duncan et al. 2007). In citrus, S. riobrave is applied to the soil using tractor-mounted sprayers or through drip irrigation (Duncan et al. 1999), but more targeted application methods used in other cropping systems for other entomopathogenic nematode species may be better suited to or adjusted for small applications in strawberry (Shapiro-Ilan & Dolinski 2015). Here we tested aqueous preparations of S. riobrave applied as a soil drench around the base of individual strawberry plants, but formulation of entomopathogenic nematodes may affect efficacy and ease-of-application, with pelletized baits showing promise in some scenarios (e.g., Hiltpold et al. 2012).
Our goal was to provide new knowledge on a potential management strategy for D. abbreviatus in Florida plasticulture strawberry production. Control methods that are effective, sustainable, reduce insecticide use, and are practical for treating only small, affected areas of strawberry fields are needed for mitigating losses due to D. abbreviatus root feeding. Therefore, the objectives of the research were to determine the efficacy of S. riobrave for control of late-instar D. abbreviatus in combination with determining the movement of larvae among strawberry plants.
Materials and Methods
In late Dec 2016, wilting plants were noticed in a small area (about 0.15 ha) of a commercial, raised-bed, plasticulture strawberry (cv. ‘Radiance') field (27.9386111°N, 82.1836111°W) near Dover, Florida, USA. Large, late-instar D. abbreviatus larvae were found in the root zone of wilted plants. To test the efficacy of S. riobrave for control of D. abbreviatus, 7 plots of 32 plants each (plot = 1 raised bed, 6.5 m long) were marked out in the affected field area. Each plot was randomly assigned a treatment of S. riobrave (4 replications) or water (3 replications).
Steinernema riobrave (Nemasys®R, BASF Corp., Florham Park, New Jersey, USA) were prepared in distilled water in the laboratory according to the manufacturer's instructions (rate of 100 IJ per cm2) and transported to the field in 1 L bottles (Thermo Scientific™ Nalgene™, Thermo Fisher Scientific, Waltham, Massachusetts, USA). The bottles were shaken by hand to redistribute the S. riobrave before pouring aliquots into plastic containers (120 mL, Fisherbrand™, Thermo Fisher Scientific, Waltham, Massachusetts, USA). On 3 Jan 2017, the plastic mulch around the base of each plant was pulled back, and S. riobrave or water (60 mL aliquots) was poured evenly and slowly onto an 80 cm2 area of the soil surface around the base of all plants in all plots.
To determine an effect of S. riobrave, plants in each plot were categorized as either: (1) healthy, showing no visual wilting symptoms; (2) wilting, with leaves and stems lacking rigidity as result of low turgor pressure; (3) dead, with plants entirely brown; or (4) missing, where empty planting holes occurred in the plastic mulch. Plants were rated pre-treatment (3 Jan) and 10 and 17 d after treatment (13 and 20 Jan). Plants were not removed to determine D. abbreviatus mortality at the end of the experiment by request of the grower.
An experiment was conducted during the 2017 to 2018 strawberry season in research plots at the Gulf Coast Research and Education Center, Balm, Florida, USA (27.7619444°N, 82.2272222°W). Bare-root strawberry transplants (cv. ‘Radiance') were set on 9 Oct 2017 (38 cm in-row spacing) in 2 rows in a raised, double-pressed, black plastic/ virtually impermeable film (Blockade™, Berry Plastics, Evansville, Indiana, USA) mulched beds (1.2 m spacing) that were fumigated with Telone®C-35 (1,3-dicholoroprepene + chloropicrin) at bed formation (early Aug 2017). Plants were overhead irrigated for 7 to 10 d following transplanting and drip fertigated with 0.27 and 0.34 kg N per d from Nov to mid-Jan and mid-Jan through Apr, respectively. Herbicides Round-up® (glyphosate) and Chateau® (flumioxazin) were applied to row aisles before transplanting. Plants received regular applications of fungicides to control prominent strawberry diseases and early-season applications each yr of DiPel® DF (Bacillus thuringiensis, subsp. kurstaki Berliner; Bacillaceae) to control lepidopteran larvae.
Plots (3 m × 1 raised bed) were arranged in a randomized complete block design with 6 replications in a checkerboard pattern, so that the raised beds adjacent to each plot were unplanted and the total plot area was 12 m × 12 raised beds. In each plot, there were 13 contiguous plants, with 1 infested plant, and 4 peripheral, infested plants, separated from the 13 plants by unplanted space (Fig. 1).
Larval D. abbreviatus were from a laboratory colony maintained at the US Department of Agriculture, Agricultural Research Service facility in Fort Pierce, Florida, USA. The larval cohort used in both experiments were from eggs obtained 9 Sep 2017, with neonate larvae transferred to artificial diet in small cups on 15 Sep and older larvae transferred singly to diet cups on 17 Oct. Larvae were removed from diet and placed singly in empty cups, and the cups (n = 4 per plant) were overturned onto the soil around the base of the plants in the evening of 2 Nov. The following morning, the cups were removed and dead larvae that did not burrow into the soil were replaced with healthy larvae. On 14 Nov, dead leaves and ripe and ripening fruit were removed from plants, and ripe fruit was harvested for the duration of the experiment.
Treatments were S. riobrave applied once, S. riobrave applied twice, imidacloprid applied once and water. On 16 Nov, S. riobrave (Nemasys®R), prepared in distilled water in the laboratory according to the manufacturer's instructions (25 IJ per cm2), imidacloprid (Admire®Pro, Bayer CropScience LP, Research Triangle Park, North Carolina, USA; 767 mL per ha) or water were applied as a drench (50 mL) to 100 cm2 around the base of each plant in a plot using a hand-held CO2-powered backpack sprayer (0.34 atm.) (R&D Sprayers, Opelousas, Louisiana, USA) and a single wand with the nozzle removed. The end of the wand was used to pull up the plastic around the base of the plant as the drench was being made. Steinernema riobrave were prepared and reapplied to plots on 22 Nov.
Healthy compared to wilting, dead, or missing strawberry plants due to Diaprepes abbreviatus root feeding in a commercial field near Dover, Florida, USA, treated 3 Jan 2017 with soil drenches of Steinernema riobrave nematodes (80,000 infective juveniles per plant) (n = 128 plants) or water (n = 96 plants) and evaluated 10 and 17 d after treatment.
Effects of drenches were evaluated on 16, 22, and 30 Nov, and 7 and 14 Dec by categorizing each of the 13 contiguous plants in all plots as either (1) healthy, (2) minor wilting, (3) major wilting, (4) mix of brown and green leaves, or (5) dead with all brown leaves. A randomly selected peripheral plant with a core of soil from its central root zone (11 cm diam × 30 cm depth) was removed with a golf-cup cutter from each plot on each of the first 4 evaluation dates. The soil was hand-sorted in the field for live and dead larvae. Dead or moribund larvae were placed in white traps (reference) in the laboratory to determine infection by S. riobrave. On 14 Dec, the root-zones of all 13 contiguous plants in each plot were removed and assessed for larvae in the same manner.
For the commercial field experiment, the distribution of plant categories was compared between S. riobrave-treated and control plots with a Fisher's exact test for small sample sizes. For the research plot experiment, a mixed-model analysis of variance (ANOVA) was used to compare the total number of larvae recovered from peripheral plants on each sample date among treatments, with the block included as a random effect. Data transformation was not necessary. The same model was used to determine the effect of treatments on numbers of live larvae recovered from the central, proximal, and distal plants (14 Dec). For the 4 proximal and 8 distal plants, the average number of larvae per plant were calculated for analysis. The distribution of plant damage categories was compared among treatments separately for each sample date, and central, proximal, or distal plants in each plot with a Fisher's exact test for small sample sizes. All analyses were conducted using JMP® 15.0.0 (SAS 2019) at α = 0.05.
Categorization of strawberry plant health was not affected by S. riobrave 10 d after treatment but was affected 17 d after treatment (Table 1). There was 16% more healthy plants in plots with S. riobrave compared to those treated with water at 17 d after treatment. Numbers of healthy plants declined by 8% in water-treated plots and increased by 6% in S. riobrave-treated plots from pre-treatment to 17 d after treatment (Table 1).
The number of live larvae recovered from peripheral infested plants was similar among treatments pre-application (F = 0.70; df = 3,15; P = 0.57) and was not affected by treatment at 1 (F = 1.16; df = 3,15; P = 0.36), 2 (F = 2.41; df = 3,15; P = 0.13), or 3 wk (F = 1.00; df = 3,15; P = 0.42) post-application. Dead larvae infected with S. riobrave were recovered only from plants to which S. riobrave was applied (Fig. 2).
The number of live larvae recovered from centrally infested plants was similar among treatments (F = 1.28; df = 3,15; P = 0.32) and similar for the distal plants (F = 1.43; df = 3,15; P = 0.27). For the proximal plants, larval recovery varied by treatment (F = 6.07; df = 3,15; P = 0.007) with fewer larvae found in plots treated once or twice with S. riobrave than in control plots (Table 2).
Categorization of strawberry plant health was not affected by treatments in central, infested plants at pre-application, (P = 0.80), at 1 wk after application (P = 0.31), 2 wk (P = 0.60), 3 wk (P = 0.55), or 4 wk (P = 0.55) (Fig. 3A), or in proximal plants at pre-application (P = 0.90), 1 wk (P = 0.31), 2 wk (P = 0.58), 3 wk (P = 0.75), or 4 wk (P = 0.90) (Fig. 3B). For distal plants, plant health categorization did not differ among treatments at pre-application (P = 0.99), 1 wk (P = 0.26), or 2 wk (P = 0.57), but it did differ at 3 wk (P = 0.01) and 4 wk (P = 0.01) (Fig. 3C).
Based on our results, aqueous applications of S. riobrave provided a modest level of control of D. abbreviatus in plasticulture strawberry production in Florida. In the grower field experiment, there were 17% more healthy and 7% fewer dead plants by 17 d after an application of S. riobrave compared to an application of water. In the research plot experiment, no live D. abbreviatus were recovered from plants proximal to the D. abbreviatus-infested plants when S. riobrave was applied to all plants, and almost all dead larvae that were recovered were infected with S. riobrave. In proximal and distal plants, there were low rates of plant wilting in all treatment plots, up to about 10% in control plots. Wilting was reduced by 1 or 2 applications of S. riobrave in distal plants, but a similar effect was not clear in proximal plants. Applications of S. riobrave may not be able to rescue a wilting plant with significant root feeding by D. abbreviatus, but preventative applications to visually healthy plants minimized further plant wilting and loss.
Mean (± 95% CI) numbers of late-instar Diaprepes abbreviatus recovered from soil in the root-zone of strawberry plants when 1 or 2 applications of infective juvenile Steinernema riobrave (25 per cm2 in 50 mL water), 1 application of imidacloprid (Admire® Pro at 767 mL per ha), or water (UTC: untreated control) were made 16 and 22 Nov 2017 around the base of each plant. Plants were in raised, plasticulture earthen beds in research plots at the Gulf Coast Research and Education Center, Balm, Florida, USA. See Figure 1 for location of central, proximal, and distal plants in plots. Means followed by the same letter are not significantly different (Tukey's HSD test, P < 0.05).
There were no significant differences in the efficacy of 1 or 2 applications of S. riobrave and 1 application of imidacloprid for control of D. abbreviatus. In citrus, the efficacy of imidacloprid and S. riobrave on D. abbreviatus also was similar (Bender et al. 2014). Admire® Pro (42.8% imidacloprid) is labelled for soil and chemigation application in strawberry for aphids and whiteflies. However, the pre-harvest interval for Admire® Pro is 14 d, limiting use in Florida strawberry to the early season before plants produce ripe fruit, because the harvest interval typically is 3 or 4 d once fruit begins to ripen. Similar efficacy between 1 and 2 applications of S. riobrave may be due to most larval infection occurring after the first application, and to high S. riobrave survival rates in soil in irrigated strawberry beds protected by black plastic mulch. Entomopathogenic nematodes are susceptible to desiccation and have reduced virulence and viability in dry soil conditions (Patel et al. 1997). Less than optimal soil temperatures may have reduced the efficacy of S. riobrave, because average hourly Jan nighttime soil temperatures under black plastic in strawberry root zones were less than 21 °C (Deschamps et al. 2019), and applications in citrus are recommended when soil temperatures are consistently above 21 °C (Duncan & Mannion 2021).
In plants infested with D. abbreviatus (central plants), there were similar numbers of dead and wilted plants in plots with 2 applications of S. riobrave as in control plots (mean plant ratings of about 4) by the last sample date, but more healthy plants on average in plots with 1 application of S. riobrave. However, there was already 50% plant death in plots with 2 applications of S. riobrave before the first application (mean plant rating of about 2.5), which was more than in other treatments. Conversely, plots with 1 application of S. riobrave had more dead and wilting proximal plants than other treatments, meaning D. abbreviatus larvae may have moved away from central plants to proximal plants more quickly in plots with 1 application of S. riobrave than other in other plots. From this experiment, the value of a second application of S. riobrave is not clear, but a second application at a longer interval, greater than a wk may be warranted, if wilting progresses to greater levels on plants adjacent to the initially infested and highly wilted plant.
In addition to determining efficacy of S. riobrave in plasticulture strawberry, we designed our research plot experiment to assess larval movement of D. abbreviatus. In untreated control plots, larval recovery after 7 wk (2 Nov to 14 Dec) was similar between central and proximal plants, which were about 40 cm apart. Movement likely was motivated by crowding at the central plant and a resulting search for new food resources, because most of the central plants were severely wilted or dead by the end of the experiment. Vertical movement of D. abbreviatus neonates has been studied (Quintela & McCoy 1998), and larvae typically remain in about 1 m radius of citrus tree trunks (Bates et al. 2015), but little is known about horizontal movement of late-instar D. abbreviatus in soil. A couple D. abbreviatus larvae were recovered from distal plants, suggesting movement of almost 1 m in 7 wk is possible but not common.
Overall, larval D. abbreviatus recovery rates were low at an average of 0.7 out of 4 larvae from central plants in control plots, and no larvae were found in peripheral plants from control plots by 3 wk after treatment applications. We searched only for larvae in plant root zones and missed any larvae moving between plants. In addition, some larvae likely died, because uninfected dead larvae were found in the peripheral untreated control plants. Diaprepes abbreviatus late-instars are capable of moving from 1 strawberry plant to another but caused a relatively low level of injury to plants to which they moved.
In the research plot experiment, S. riobrave were applied to all plants in a plot 2 wk after placing D. abbreviatus on 1 central plant. No live D. abbreviatus were recovered from proximal plants treated with S. riobrave even though minimal plant wilting occurred. It is likely that uninfected larvae moved from the central to proximal plants, began feeding on plant roots, were infected by S. riobrave, but ceased feeding and died before significant plant damage was caused. Since we allowed D. abbreviatus 2 wk to begin feeding and moving before applying S. riobrave, an earlier application to central plants only as soon as wilting was noticed, may have produced similar or even better results. This possibility needs to be tested in a future experiment. From a practical perspective, it will be more economical and thus feasible to selectively apply S. riobrave in large strawberry fields to wilting plants only and not to large numbers of plants adjacent to them.
There is increasing interest in managing D. abbreviatus through entomopathogenic nematode conservation, as abundant and suppressive entomopathogenic nematode communities were found in some Florida citrus orchards (Duncan et al. 2013). Conventional strawberry fields are fumigated annually and will require annual S. riobrave augmentation, but entomopathogenic nematode communities should be evaluated in organic strawberry production where cover crops and physical methods are used in place of chemical fumigation, and may result in conserved and suppressive entomopathogenic nematode communities. Because it appears that D. abbreviatus-damaged plants are localized to certain fields or in field areas, yearly monitoring of larval patches and plant wilting will determine whether or not they are seasonally stable and, if so, then S. riobrave applications can be consistently applied to these fields or field areas. More knowledge about D. abbreviatus flight and oviposition periods in strawberry is needed to guide S. riobrave application timing decisions, and persistence of S. riobrave in plasticulture earthen beds should be determined to guide timing of repeat applications. Currently, S. riobrave applications may not be cost-effective as they are only sold in large quantities for use in citrus; localized applications would require a change in marketing and production of S. riobrave. In conclusion, S. riobrave successfully infected and killed late-instar D. abbreviatus in strawberry and was as effective as imidacloprid in reducing plant wilting. With further research to determine optimal application strategies, S. riobrave can be successfully developed into safe and relatively easy-to-use biological control method to reduce impacts of D. abbreviatus in strawberry.
We are grateful to Shashan Devkota, Rosa Infante, Franklin Dubon, Marc Santos, Ryan Batts, and Braden Evans for technical assistance in the field. We thank Stephen Lapointe and Anna Sara Hill at USDA-ARS Fort Pierce for providing D. abbreviatus larvae, Mark Peacock at BASF for providing S. riobrave, and Larry Duncan at University of Florida, Citrus Research and Development Center for methodological advice. The project was supported by University of Florida start-up funds to JMR.