LABORATORY TEST

A laboratory test was conducted to evaluate the effect of 2 imidacloprid formulations used for hemipteran control on N. fulva. The test used a completely randomized design with 4 replications. Each replicate consisted of 3 plastic trays (27.2 × 19.6 × 9.4 cm) with sides coated with Fluon® and filled to a depth of 2.5 cm with untreated soil (50% autoclaved sand and 50% potting mix; Fafard®, Agawam, Massachusetts, USA). Two 0.47 L pots filled with the same soil were placed at the center of each tray. The soil in 1 pot was treated with either CoreTect® or Merit® 2F (20% and 21.4% imidacloprid, respectively) (Bayer Environmental Science, Research Triangle Park, North Carolina, USA), whereas the other pot remained untreated. One CoreTect® tablet was placed 5.1 cm below the soil surface to follow the label rate of 1 tablet per 3.75 L or smaller pot. Merit® 2F was applied as a soil drench with a pipette at the rate of 146.2 μL per m². After treatment, all pots, including controls, received 25 mL of water.

A N. fulva colony fragment consisting of 500 workers and 2 queens, originating from source colonies collected in Gainesville, Alachua County, Florida, USA, was placed on the soil in each tray. The colony was provided water and a 10% sucrose solution through cotton-plugged 20 mL vials, and frozen house crickets were provided as a protein source. Ant mortality was assessed after 2 wk. The soil from each pot and tray was spread separately in larger tubs (45.5 × 34.3 × 7.6 cm) and allowed to dry. Because ants would move from dry soil to moist areas, artificial nests were established to entice living ants out of the soil, which facilitated the determination of ant survivorship. Placed in the middle of each tub, the artificial nest consisted of an inverted black plastic container (6 × 17 × 12 cm; Sterling King Products, Lyons, Illinois, USA), with holes on each side that covered 2 nest tubes. A nest tube was a 20 mL glass test tube, half filled with water and plugged with cotton and dental plaster (Castone®, Dentsply International, York, Pennsylvania, USA) (Banks et al. 1981).

The number of live N. fulva in the nest tubes were counted after 48 h and the percentage of surviving ants relative to the initial number of ants was calculated. Analysis was conducted on percent survival after square-root transformation. Differences in N. fulva survival among the 2 imidacloprid treatments and untreated control were determined by a 1-way analysis of variance (R Core Team 2012).

FIELD TEST

Twenty-four pittosporum plants (< 1 m tall) in 11.34 L pots were kept in a greenhouse together with pittosporum naturally infested with I. purchasi until all the plants were equally infested (approximately 1 mo). The plants were then transferred to a location in Gainesville, Florida, that was infested with N. fulva. In Aug 2012, pots were placed on the soil surface 2 m apart between a hedge and a road. Nylanderia fulva naturally moved into the potted plants. One wk after the pots were placed on site, the mean number of I. purchasi from 3 randomly selected branches per plant was determined to characterize the scale infestation. Nylanderia fulva activity was indicated by the following: (a) the mean number of N. fulva on 2 sausage slices (22 mm diam × 33 mm wide) placed at the base of the main stem, or trunk, of each plant for approximately 30 min; (b) trailing intensity reflected by the mean number of trailing ants crossing a single spot in 20 s; and (c) the mean number of ant trails per main plant stem (Sharma et al. 2013). The initial counts for each of the variables above were not significantly different (P > 0.05) among the plants.

The treatments were assigned to pots in a completely randomized design, with 8 replications per treatment. The scale and antinfested pittosporum pots were randomly assigned 1 of the 2 imidacloprid treatments used in the laboratory test, or designated as a control. For the CoreTect® treatment, we inserted 3 tablets, spaced equally apart, about 5 cm deep into each pot (about 15 cm from the base of the plant), and watered (28.8 mL per pot). Merit® 2F was applied as a soil drench following the label rate of 6 mL of Merit® 2F in 28.8 mL water per 30.5 cm of shrub height. The untreated control pots received 28.8 mL of water. All pots were drip-irrigated daily for 10 min.

Icerya purchasi and N. fulva survival and activity in the infested plants were monitored weekly for 2 mo when ambient temperatures were > 21 °C and favorable for ant activity. Because plants were small (≤ 1 m tall), destructive sampling was not conducted; instead, 3 branches were randomly flagged on each plant before the study began and the mean number of I. purchasi per 3 branch sample was determined weekly.

Two mo after treatment, the plants were removed from their pots, placed in fluon-coated tubs (45.5 × 34.3 × 7.6 cm), the roots were removed, and the soil was air-dried in the laboratory. The ants were supplied with water, a 10% sucrose solution, and frozen house crickets. All live ants that moved from the soil into the nest tubes were counted after 48 h, and the mean number of N. fulva per pot was determined for each treatment.

Repeated measures analysis of variance was conducted to assess differences in the numbers of I. purchasi on the pittosporum, and the number of N. fulva counted, on the sausage slices, on trailing intensity (20 s counts), and on the number of trails among treatments for the pre-treatment and weekly post-treatment data. If significant, weekly means were separated with the Tukey HSD test (R Core Team 2012). Pearson's correlation was used to assess the association between the number of I. purchasi and N. fulva trailing intensity for each treatment.

Results

LABORATORY TEST

Nylanderia fulva survival when exposed to soil treated with either imidacloprid formulation did not differ significantly from the untreated control. Mean percent survivorship ranged from 71 to 86% of the 500 ants initially exposed to the treatments (Table 1). Because a majority of N. fulva survived both treatments, these treatments were used in the field test to examine N. fulva activity relative to the treatment of I. purchasi.

Table 1.

Mean (± SE) numbers of live Nylanderia fulva workers and percent survival of workers given access to treated and untreated soil in the laboratory after 2 wk of exposure (n = 4 pots per treatment).

FIELD TEST

Numbers of I. purchasi Scales on Branches

The numbers of I. purchasi were significantly higher on the control, compared to both imidacloprid treatments on all sampling dates (F = 8.9; df = 2, 21; P < 0.001). However, statistical differences were not observed for the numbers of I. purchasi between the 2 insecticide treatments. The numbers of scales on treated plants began to decrease 1 wk post-treatment, and it was almost zero at 7 wk post-treatment. Scale density decreased by 67% to an average of 20 scales per plant in Sep and Oct in the control pots (Fig. 1), possibly due to declining temperatures. In Sep and Oct, the mean average daily temperatures were 24 and 20 °C, respectively, compared to 26 °C in Aug (Florida Automated Weather Network).

Activity of N. fulva on Potted Plants

The numbers of N. fulva workers foraging on the sausage slices placed at the base of the main stem of each potted plant were significantly lower in the 2 imidacloprid treatments than the control over all sampling dates (F = 11.1; df = 2, 21; P < 0.0005). The mean numbers of ants in the pots treated with either CoreTect® or Merit® 2F did not differ significantly (Fig. 2). The trailing intensities were significantly lower (2 ants per 20 s) in the treated plants than in the control plants (15 ants per 20 s) (F = 20.6; df = 2, 21; P < 0.0001). However, the trailing intensities did not differ significantly between the imidacloprid treatments (Fig. 3). Similarly, significantly fewer ant trails occurred on the main stems of treated plants than on the stems of control plants (F = 26.0; df = 2, 21; P < 0.0001). The average number of trails between the 2 insecticide treatments did not differ significantly (Fig. 4). The mean number of I. purchasi and the N. fulva trailing intensity counts were positively correlated in the pots treated with CoreTect® (r = 0.88) and Merit® 2F (r = 0.88), and to a lesser extent, the control (r = 0.60) (Fig. 5).

Fig. 1.

Mean (n = 8) numbers of Icerya purchasi per pittosporum plant counted on 3 branches of each plant. Different letters indicate significantly different (P < 0.05) means by repeated measures analysis of variance and Tukey HSD test (R Development Core Team 2012).

Fig. 2.

Mean (n = 8) numbers of Nylanderia fulva counted from sausage samples placed near the base of the trunk of pittosporum plants. Different letters indicate significantly different (P < 0.05) means by repeated measures analysis of variance and Tukey HSD test (R Core Team 2012).

Nesting by N. fulva in Potted Plants

After 2 mo in the N. fulva infested area, all pots were examined for the presence of N. fulva nests. All untreated control pots either had nests or ants present in them. Half (n = 4) of the untreated pots had tunnels and brood present, and contained an average of 545 ± 222 (± SE) N. fulva workers per pot. The remaining control pots contained 250 ± 86 adults but lacked brood. Three (19%) of the treated pots (2 with Merit® 2F, and 1 with CoreTect®) contained a small number of worker ants averaging 100 ± 50 workers per pot. The rest of the treated pots did not harbor any N. fulva.

Discussion

In the laboratory test, ant survivorship averaged 72% and 78% in the imidacloprid treatments. Some of the observed mortality could be due to initial contact with treated soil. However, the low concentrations of imidacloprid present in CoreTect® and Merit® 2F formulations (20% and 21.6%, respectively) may have allowed some survivorship of N. fulva because substantial mortality of N. fulva was observed in a preliminary test where potted soil was treated with higher concentrations of imidacloprid (Merit® 75 WP; S. Sharma, unpublished data). Costa and Rust (1999) noted that even though Argentine ants showed avoidance toward the insecticide-treated soil in pots, ants would initially excavate the treated soil, thereby contacting the insecticide, before displaying avoidance. We also observed avoidance behavior by N. fulva of the imidacloprid-treated soil inside the pots. Although N. fulva nested within the potting soil of the untreated pots, they nested beneath the pots treated with imidacloprid. This suggested that the treated soil deterred N. fulva from nesting in the potted soil, even though imidacloprid is considered a non-repellent insecticide for ants (Rust et al. 2004). Oi and Williams (1996) reported red imported fire ants, Solenopsis invicta Buren, clinging to the exterior of pots containing pyrethroid-treated soil, apparently due to the repellent effects of the insecticide.

Fig. 3.

Mean (n = 8) numbers of Nylanderia fulva crossing a specific point on the main stems of pittosporum plants in 20 s (trailing intensity). Different letters indicate significantly different (P < 0.05) means by repeated measures analysis of variance and Tukey HSD test (R Core Team 2012).

Fig. 4.

Mean numbers of Nylanderia fulva trails on the trunk of pittosporum plant. Different letters indicate significantly different (P < 0.05) means by repeated measures analysis of variance and Tukey HSD test (R Core Team 2012).

Fig. 5.

Correlations between the mean numbers of I. purchasi and N. fulva in pots treated with (A) CoreTect® (r = 0.88; n = 8), (B) Merit® 2F (r = 0.88; n = 8), and (C) control (untreated) (r = 0.60; n = 8).

In the field test, it was observed that most of the N. fulva on the sausages at the base of the control plants emerged from the potting soil within min of sausage placement. In contrast, N. fulva found on sausages at the base of treated plants appeared to originate in the surrounding area beyond the pots. Because these ants were not nesting in the pots, searching and recruitment times to the sausages would take longer, thus resulting in the significantly lower counts for the treated soil. These results are consistent with the laboratory test where N. fulva tended to nest under the pots, rather than in the treated soil.

The reduced trailing intensity and the lower number of ant trails on the treated plants may be attributed to the lower abundance of I. purchasi, resulting in less availability of honeydew as a food resource. This is supported by the positive correlation between I. purchasi counts and the trailing intensity of N. fulva (Fig. 5). This also is consistent with the results of Sharma et al. (2013), where ants and hemipteran numbers were positively correlated throughout a sampling season, presumably due to the availability of honeydew. As seen with other ant species, when the number of hemipterans decreased, honeydew became scarce, which may have triggered the ants to search for food elsewhere (Bach 1991; Blüthgen et al. 2000). We also documented fewer N. fulva nests in treated pots, which also could be due to the reduced availability of honeydew. Establishment of nests near food sources has been reported for Argentine ants (Holway & Case 2000).

Removal of hemipterans to reduce honeydew resources for invasive ant control also is being attempted for yellow crazy ants, Anoplolepis gracilipes (Smith) (Green et al. 2011, 2013). In addition, Brightwell et al. (2010) recorded reductions in ground foraging, a decline in nest density, and canopy foraging by Argentine ants when honeydewproducing scale insects were reduced with the use of trunk-injected insecticide. Our study showed that N. fulva activity was reduced by managing I. purchasi on pittosporum. These results suggested that eliminating honeydew-producing insects in landscapes may contribute to the control of N. fulva. However, because soil applications of imidacloprid were used to systemically reduce I. purchasi, we cannot conclude that the sole removal of honeydew producing hemipterans can significantly reduce N. fulva activity. Indeed, the partial avoidance of imidacloprid-treated soil by N. fulva may be an important component of the observed decline in N. fulva activity, but it was not a predominant factor because foraging on sausage was not completely excluded in treated pots. Nylanderia fulva avoidance of insecticide residues has been documented for several insecticides used to control N. fulva (Calibeo et al. 2017). Sustained reductions of overwhelming numbers of invasive ants such as N. fulva most likely will require an integrated pest management program consisting of several control tactics (Oi et al. 2016). Eliminating natural food sources, such as honeydew producing hemipterans, either directly with insecticides, or by preventing food access via repellents, could be part of an integrated pest management program for N. fulva.