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Interference of ants (Hymenoptera: Formicidae) with biological control of the vine mealybug Planococcus ficus (Signoret) (Hemiptera: Pseudococcidae)
Biological Control 49 (2009) 180–185
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Interference of ants (Hymenoptera: Formicidae) with biological control of the vine mealybug Planococcus ficus (Signoret) (Hemiptera: Pseudococcidae) Nyembezi Mgocheki *, Pia Addison Department of Conservation Ecology and Entomology, Faculty of AgriSciences, Stellenbosch University, Private Bag X1, Matieland, Stellenbosch 7602, South Africa
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Article history: Received 26 August 2008 Accepted 2 February 2009 Available online 12 February 2009 Keywords: Anagyrus sp. Anoplolepis steingroeveri Ant species Coccidoxenoides perminutus Crematogaster peringueyi Linepithema humile Parasitism Mortality Trophobiosis
a b s t r a c t Anagyrus sp. and Coccidoxenoides perminutus are well known parasitoids used for the biological control of the vine mealybug Planococcus ficus, a key pest in vineyards. In South Africa, three ant species, Anoplolepis steingroeveri, Crematogaster peringueyi and Linepithema humile form a trophobiotic relationship with the vine mealybug in vineyards and promote the latter’s infestations to unacceptable levels. In a manipulative laboratory experiment, ants and parasitoids were allowed to forage on vine mealybug-infested butternuts and numbers were recorded for a 1 min period at 10 min intervals for 2 h. Parasitoid mortality and the number of parasitized vine mealybug females were then recorded in the presence and absence of the three ant species. Data were analyzed using a repeated measures generalized linear model (GEEs). The mean number of ants and parasitoids on the mealybug-infested butternuts differed significantly between ant species and time intervals (p < 0.0001 in all cases). Crematogaster peringueyi and L. humile caused significantly higher mortality of both parasitoids than A. steingroeveri during the 24-h exposure period (p < 0.0001). Coccidoxenoides perminutus parasitized significantly more vine mealybugs than Anagyrus sp. for all treatments (p < 0.0001). Ants should therefore be controlled prior to release of parasitoids to suppress populations of ant-tended Hemiptera in vineyards. Crown Copyright Ó 2009 Published by Elsevier Inc. All rights reserved.
1. Introduction Finding new ways of understanding animal behavior is fundamental to the biological control of arthropod pest species. The trophobiotic relationship between ants and their adopted Hemiptera (Gibernau and Dejean, 2001; Jiggins et al., 1993; Hölldobler and Wilson, 1990; Buckley, 1987; Pierce and Mead, 1981; Adenuga, 1975; Bradley, 1973; Bartlett, 1961; Steyn, 1954) needs further investigation to understand various levels of aggression exhibited by ants towards natural enemies of their attended Hemiptera. Field behavioral studies of parasitoids are difficult to carry out due to the complexity of these interactions in the field. Behavior of ants and parasitoids can be broken down into simple components (e.g. ant recruitment and parasitoid oviposition) that can be manipulated under controlled conditions to predict field response that can be considered in planning and implementing a pest management programme. Although many studies have abundantly documented the detrimental impacts of ants on biological control (Bartlett, 1961; Buckley, 1987; Itioka and Inoue, 1996; Martinez-Ferrer et al., 2003), the individual impact of each ant species on individual parasitoids has received limited experimental attention. In agricultural and natural ecosystems, ants obtain carbohydrate-rich honeydew from * Corresponding author. Fax: +27 218084807. E-mail address: [email protected] (N. Mgocheki).
Hemiptera such as mealybugs, scale insects, aphids, among others, while they render protection, sanitation and sometimes transport services to sedentary Hemiptera (Buckley, 1987; Lach, 2003). Carbohydrates serve as a metabolic fuel for most insects and therefore behavioral dominance can be associated with relative availability of and demand for sugar (Grover et al., 2007). Arboreal ants exhibit greater activity and aggression where carbohydrate resources are limited compared to epigeic ants probably due to their numerical superiority in canopies and greater dependency on hemipteran honeydew. Similarly, high levels of activity and aggression exhibited by invasive ants may well be linked to their reliance on carbohydrate-rich honeydew (Way, 1963; Markin, 1970). The Argentine ant Linepithema humile (Mayr) was found to be disruptive to the black scale Saissetia oleae Olivier, parasitoid Coccophagus scutellaris (Dalman) in California (Horton, 1918). In South Africa Metaphycus helvolus (Compere), a parasitoid of black scale was found to be effective in the absence of L. humile ( Flanders, 1943; Compere, 1940). In California, Daane et al. (2007) found that L. humile promoted populations of obscure mealybug Pseudococcus viburni (Signoret) while lowering populations of its parasitoids Pseudaphycus flavidulus and Leptomastix epona. They also noted increased densities of the Argentine ant-tended grape mealybug Pseudococcus maritimus (Erhorn) accompanied by a serious reduction in its parasitoid populations. The cocktail ant Crematogaster peringueyi Emery is disruptive to natural enemies of soft brown scale Coccus hesperidum L., and vine
1049-9644/$ - see front matter Crown Copyright Ó 2009 Published by Elsevier Inc. All rights reserved. doi:10.1016/j.biocontrol.2009.02.001
N. Mgocheki, P. Addison / Biological Control 49 (2009) 180–185
mealybugs Planococcus ficus (Signoret). This ant species provides protection to its hemipteran hosts by constructing carton shelters over the mealybugs (Kriegler and Whitehead, 1962). The common pugnacious ant, Anoplolepis custodiens (Smith), incidentally disturbed the parasitoids of California red scale Aonidiella aurantii (Maskell), while tending soft brown scale in citrus orchards in South Africa (Samways and Tate, 1984; Steyn, 1954). Buckley and Gullan (1991) concluded that the incidence of coccid parasitization was correlated with the relative inoffensiveness of the attendant ant species in a field study in Australia. These authors measured low parasitism rates (at least 15%) of coccids in the presence of Tapinoma and Iridomyrmex spp. and <10% in the presence of the more aggressive Oecophylla and Solenopsis species. In California, L. humile reduced parasitism and host mutilation of the California red scale by the parasitoids Comperiella bifasciata (Howard) (59.1%) and Aphytis melinus De Bach (79.5%) in a laboratory trial, even if there were no honeydew-excreting soft scales (Martinez-Ferrer et al., 2003). Itioka and Inoue (1996), in a comparative field investigation, found a 94% decrease of the mealybug Pseudococcus citriculus Green by natural enemies in the absence of the attendant ant Lasius niger (L.). Previous (unpublished) studies revealed that more than one ant species and parasitoid species were sampled from the same sites and therefore parasitism rates could not reflect the individual impact of each ant species on mealybug biological control. Anagyrus sp. and Coccidoxenoides perminutus (Timberlake) (Hymenoptera: Encyrtidae) are widely distributed primary parasitoids of Planococcus spp. and Pseudococcus spp. (Davies et al., 2004; Triapitsyn et al., 2007), that have been used for classical and augmentative biological control of the vine mealybug and citrus mealybug Planococcus citri (Risso) in some localities (Daane et al., 2004; Walton and Pringle, 2005). This study tests the interference of three ant species on the biological control of the vine mealybug P. ficus using the parasitoids Anagyrus sp. near pseudococci (as identified by Triapitsyn et al., 2007) and C. perminutus. This investigation used the presence and absence of these ant species to quantify their individual impact on the parasitoids under controlled laboratory conditions. For the purposes of this study, Anagyrus sp. near pseudococci shall be referred to as Anagyrus sp. from this point onwards. 2. Materials and methods 2.1. Insect colonies 2.1.1. Vine mealybug colonies Colonies of vine mealybugs were maintained on mature fruit of butternut squash Cucurbita moschata (L.) in the laboratory at 27 ± 1 °C with a 12:12 (L:D) hour photoperiod and 65 ± 5% RH. Butternut squash were obtained fresh from the local grocery store and were more or less of equal size. Before use in experiments, butternuts were washed in 5% bleach solution (to prevent fungal growth), triple rinsed, air dried and then inoculated with vine mealybug crawlers after 1 h. After the first molt, mealybugs were thinned to approximately 100 individuals that were allowed to develop up to a desired stage before use in experiments as hosts to parasitoids. For Anagyrus sp., 3rd instar to preovipositing female vine mealybugs were used while for C. perminutus, 2nd instar mealybugs were used (Islam and Copland, 1997; Joyce et al., 2001). 2.1.2. Ant colonies A single average ant nest (at least 500 workers) for all three ant species, consisting of workers, queens and immatures, was collected from commercial vineyards. For epigeic ants, a spade was used to collect a portion of the nest and maintained in soil from the original nest. A small size carton nest was collected for the
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arboreal C. peringueyi. Nests of all three species were placed into five liter square plastic containers. The set up was designed to mimic a natural situation as far as possible, where ants are regarded as pests and as such ants were collected from vineyards where their infestation levels were above the action threshold. Anoplolepis steingroeveri and C. peringueyi were collected from Ashton ( 33.85°S, 20.08°E, 186 m) in the Breede River Valley (BRV) while L. humile were collected from Simondium ( 33.83°S, 18.83°E, 175.2 m) in the Stellenbosch area, South Africa. The three ant species were maintained in plastic containers (18 18 16 cm) in the laboratory containing soil or material from the original nests. Each ant nest was connected to a clear Perspex container (25 25 20 cm) with clear plastic tubing (20 cm long and 6 mm in diameter). A butternut infested with about 100 mealybugs was placed into each Perspex container and ants were allowed to forage freely on this butternut for honeydew until 48 h prior to the experiment. All ant colonies were kept at 27 ± 0.5 °C, 65 ± 5% RH and a 12:12 (L:D) photoperiod. 2.1.3. Parasitoid colonies 2.1.3.1. Anagyrus sp.. Field collected vine mealybugs were incubated individually in gelatin capsules at room temperature. Incubated mealybugs were monitored daily, and emerged Anagyrus sp. were selected for experimental use through stereo microscope identification. Only Anagyrus sp. were selected, based on antennal coloration according to Triapitsyn et al. (2007). The parasitoids were placed in a cage (66 66 37 cm) containing butternuts infested with vine mealybugs. Parasitoids were offered a 50:50 honey:water solution and kept at 27 °C, 65 ± 5% RH with a 12:12 (L:D) photoperiod. After seven days, parasitized mealybugs were moved into another cage for parasitoid emergence. Anagyrus sp. colonies were maintained in the laboratory on 3rd instar to adult vine mealybugs feeding on butternut squash (Islam et al., 1997). Testing was done when a total of 40 newly emerged female parasitoids (total per trial) were available and each individual was used only once. Newly emerged parasitoids were allowed to feed and mate before use in trials. One male parasitoid was given access to five newly emerged females for 24 h (Tingle and Copland 1988, 1989). Only mated two-day old females were used in the experiment. After every three generations, Anagyrus sp. from the field were added to laboratory colonies to prevent inbreeding of the laboratory colony. 2.1.3.2. Coccidoxenoides perminutus. Coccidoxenoides perminutus were obtained from DuRoi Integrated Pest Management (Letsitele, South Africa) as mature pupae. Newly emerged individuals were allowed to feed for 24 h after which they were used in the experiments. No mating was necessary as C. perminutus are thelytokous (Davies et al., 2004). Both parasitoid species were deprived of hosts 24 h prior to the experiment. 2.2. Quantitative observations Ants foraged on a butternut infested with 100 vine mealybugs in each of the six experimental cages of 21.5 21.5 16 cm (one for each ant species per parasitoid species). An ant-free cage was included as a control for each parasitoid species. Each experimental cage had a hole (6 mm in diameter) in the side which was plugged with cotton wool. This hole was only unplugged during introduction of parasitoids into the cages. A single tube aspirator was used to collect parasitoids from their rearing cages and release the parasitoids into the experimental cage through the side hole by gently blowing through the tube. The ants were allowed to forage for 3 h before 20 two-day old fertilized Anagyrus sp. females and 20 one-day old C. perminutus were introduced. Observations were made 10 min after the release of parasitoids for each treatment
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whereby the number of ants and parasitoids on the whole butternut was recorded during a 1 min period at 10 min intervals for 2 h. The aspirator tube was used to ensure the accuracy of the counted parasitoids before and after release. This also allowed the parasitoids to be in the experimental cage at approximately the same time and therefore equal exposure to ants however minimizing vulnerability of parasitoids to ants. Parasitoids were then left in the experimental cages for 24 h after which they were removed and the numbers of surviving, dead and/or missing parasitoids, if any, were recorded. Mortality of the parasitoids was defined as the number of dead + missing parasitoids/total number of parasitoids and expressed as a percentage. Magnifying lenses were used to search for missing parasitoids, which could also have been dismembered or carried back to the nest by ants. After 24 h, all mealybugs were removed and incubated individually in gelatin capsules at 27 °C, 65 ± 5% RH with a 12:12 (L:D) photoperiod, for two weeks after which they were examined for parasitism under a stereo microscope. Percentage parasitism of the vine mealybug was defined as the number of parasitized mealybugs/total number of mealybugs. The tests were performed on five different dates for each parasitoid species with five replicates per ant colony and their controls (ant-free treatment).
workers (32.30 ± 2.21), A. steingroeveri from 5 to 25 workers (16.17 ± 2.11) and C. peringueyi from 315 to 401 (354.98 ± 4.42) (Fig. 2). In C. perminutus-released cages, L. humile ranged from 19 to 36 workers (28.45 ± 2.18), A. steingroeveri from 0 to 15 (5.65 ± 1.93) and C. peringueyi from 290 to 335 (313 ± 1.66) during 1 min observation periods (n = 5 in all cases). Significantly more ants foraged when Anagyrus sp. were released than when C. perminutus were released (Fig. 3). The number of foraging ants for both parasitoid species generally decreased with time. The mean number of ants on the mealybug-infested butternuts was influenced by the ant and parasitoid species and the time intervals so that the interaction; ant species parasitoid species time was significant (v2 = 5.491E8; df = 33; p < 0.0001). 3.2. Parasitoid behavior in the absence and presence of ants During 1 min observation periods, the number of Anagyrus sp. increased from 0 to 1 (0.07 ± 0.06) for C. peringueyi, 0 to 3
370 360
Data were analyzed using Generalized Estimating Equations (GEEs) (Liang and Zeger, 1986) in GENMOD procedure in SAS (SAS Enterprise Guide 3, 2004) using Poisson distribution and an identity link function. Abbott’s correction formula (Abbott, 1925) was used to correct for control mortality.
350 No. of ants
2.3. Data analysis
340 330
3. Results
320
3.1. Ant behavior in the presence of parasitoids
310
Where Anagyrus sp. were released, the number of L. humile on butternuts during 1 min periods (Fig. 1) ranged from 22 to 42
300 10
20 30
40
50 60 70 80 Time (minutes)
C. peringueyi/C. perm.
90 100 110 120
C. peringueyi/Anagyrus sp.
40 Fig. 2. The number of Crematogaster peringueyi observed during 1 min intervals on a vine mealybug-infested butternut during a 2 h observation period.
35 30
c
350 d
20 No. OF ANTS
No. of ants
400 25
15 10 5
Anagyrus sp. C. perminutus
300 250 200 150 100
0 10
20 30
40 50 60 70 80 90 100 110 120 Time (minutes) A. steingroeveri/Anagyrus sp. A. steingroeveri/C.perm. L. humile/Anagyrus sp. L. humile/C.perm.
Fig. 1. The number of Anoplolepis steingroveri and Linepithema humile observed during 1 min intervals on a vine mealybug-infested butternut during a 2 h observation period.
50
a ab
0 L.humile
b
e
C. peringueyi A. steingroeveri ANT SPECIES
Fig. 3. The mean number of ants, Linepithema humile, Crematogaster peringueyi and Anoplolepis steingroeveri on a mealybug-infested butternut for different parasitoid species during 1 min observation periods over 2 h. Means followed by different letters are significantly different at p 6 0.05.
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(0.57 ± 0.22) for L. humile, 0 to 9 (3.33 ± 3.42) for A. steingroeveri and 7 to 16 (12.92 ± 1.93) in the ant-free cage. The number of C. perminutus ranged from 0 to 2 (0.1 ± 0.15) for C. peringueyi, 0 to 7 (1.12 ± 1.53) for L. humile, 0 to 18 (10.57 ± 4.25) and 9 to 18 (13.65 ± 1.03) in the ant-free cage (n = 5 in all cases). Fig. 4 illustrates the trends in the number of parasitoids on butternuts in the presence or absence of ants. The mean number of parasitoids foraging on the mealybug-infested butternut differed significantly
18
3.3. Effects of ants on parasitoid mortality and mealybug parasitism
16
Parasitoid mortality was significantly different between parasitoid species (v2 = 13.47; df = 1; p < 0.001) and ant species (v2 = 2168.53; df = 3; p < 0.0001) (Fig. 5). Anoplolepis steingroeveri caused the least parasitoid mortality during the 24-h exposure period, while C. peringueyi caused the most. Percentage parasitism differed significantly (p < 0.0001) between parasitoid species (v2 = 38.18; df = 1) and ant species (v2 = 10351.8; df = 3). Coccidoxenoides perminutus caused significantly more parasitism than Anagyrus sp. while percentage parasitism was lowest in the presence of C. peringueyi (Fig. 6).
14 No. of parasitoids
over the 2 h period and was influenced by ant species parasitoid species time interval (v2 = 3.883E7; df = 66; p < 0.0001). Coccidoxenoides perminutus searched for mealybugs in higher numbers than Anagyrus sp. The highest number of parasitoids (both species) searched for vine mealybug on A. steingroeveri-infested butternuts and the lowest on C. peringueyi-infested butternuts. These data are an indication that all three ant species are capable of preventing parasitoids from accessing the vine mealybug, but to varying degrees.
12 10 8 6 4 2
4. Discussion
0 10
20
30
40 50 60 70 80 90 100 110 120 Time after release (minutes)
Anagyrus sp. (A. steingroveri)
C. perm (A.steingroeveri)
Anagyrus sp.(L. humile)
C. perm (L. humile)
Anagyrus sp.(C. peringueyi)
C. perm (C. peringueyi)
Anagyrus sp.(Ant free)
C. perm (Ant free)
Fig. 4. The number of Anagyrus species near pseudococci and Coccidoxenoides perminutus observed on a vine mealybug-infested butternut during 1 min intervals over a 2 h observation period in the presence and absence of one of three ant species.
Different ant species have different strategies for defending Hemiptera against parasitoids (Buckley and Gullan, 1991; Kaneko, 2003). Attendance by all three ant species significantly reduced the number and percentage mortality of parasitoids on P. ficus colonies and resultant parasitism in the present study. However, certain ant species were found to be more aggressive, while certain parasitoids were found to be more ant-resistant. Parasitism occurred less frequently in A. steingroeveri attended P. ficus probably due to incomplete protection. Martinez-Ferrer et al. (2003) noted that larger ants do not easily recognize small natural enemies. Anoplolepis
% mortality
100 80
Anagyrus sp. C.perminutus
60 40 20 0 A.steingroeveri
L. humile C.peringueyi Ant treatment
Control
Fig. 5. The mean (±standard error) percentage mortality of Anagyrus species near pseudococci and Coccidoxenoides perminutus after a 24-h exposure to different ant treatments. Means with different letters are significantly different at p 6 0.05.
% parasitism
100
C.perminutus Anagyrus sp.
80 60 40 20 0
A.steingroeveri
L. humile C.peringueyi Ant treatment
Control
Fig. 6. Parasitism rates (% mortality ± standard error) of P. ficus by Anagyrus species near pseudococci and Coccidoxenoides perminutus after a 24 h exposure period to various ant species. Means with different letters are significantly different at p 6 0.05.
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steingroeveri are mainly predatory, epigeic ants which tend to attack other insects within their foraging territory, promoting mealybug infestations due to incidental protection from natural enemies (Way, 1963; Henschel, 1998). However, their impact on mealybug biological control is limited to drier regions as they appear to be negatively affected by flooding (e.g. flood and micro jet-irrigated vineyards) (Addison and Samways, 2000). As this study indicated furthermore, their impact on parasitism was not as marked as the other ant species. Crematogaster peringueyi recruited in large numbers to the food source and therefore negatively affected P. ficus parasitism rates the most for both parasitoids species. Almost complete protection is afforded by creation of a biological barrier that covered the mealybug-infested butternut and mealybugs were immune to parasitoid attack. Schatz and Hossaert-McKey (2003) and Varon et al. (2007) described arboreal ants like Crematogaster spp. as predacious on other insects in plant canopies. This allows such ant species to obtain protein to complement their carbohydrate-rich diet. Although very aggressive, C. peringueyi are not widely distributed in Western Cape vineyards (Addison and Samways, 2000). They are confined to old neglected vineyards where they utilize old or diseased canes for nest building, presumably providing better protection to mealybugs than would epigeic ant species. Their impact is significant when their infestations are high but at low infestations, this species often confines itself to their nests with small colonies of mealybugs and therefore parasitoid activity in the vine foliage may not be disturbed much by the ants (Kriegler and Whitehead, 1962). Given their invasive nature and wide distribution, possibly perpetrated by global warming (Roura-Pascual et al., 2004), L. humile present a serious threat to the biological control of hemipteran pests like the vine mealybug, as they are capable of affecting parasitoids in vineyards over large areas of the Western Cape, South Africa (Addison and Samways, 2000). In vineyards, they make numerous nests in the floor and construct temporary nests on vine stems, leaves and bunches (Horton, 1918; personal observation), thereby posing a serious threat to mealybug parasitoids. Nixon (1951) noted that the behavior of a parasitoid in the presence of ants largely determines its own effectiveness as a biological control agent. Anagyrus sp. and C. perminutus evoked different responses in the ants. From personal observation, Anagyrus sp. often got entangled with the host while ovipositing and were then seized more readily by ants than C. perminutus, while struggling to retract their ovipositor. Parasitoids abandoned oviposition and kept away from mealybugs to avoid ants, limiting the number of eggs that could be laid into the host. While some parasitoids have developed escape strategies from ants to improve their efficacy, others are so ant sensitive that after an encounter with ants, they are deterred not only by ants, but by any moving object including other parasitoids or the host itself, thereby greatly reducing their potential as biological control agents (Martinez-Ferrer et al., 2003). It is apparent that ants not only interfere with percentage parasitism of their adopted Hemiptera, but also reduce parasitoid abundance by causing direct mortality and low reproductive success. Daane et al. (2007) noticed an almost complete absence of parasitoids in vineyards infested with P. maritimus attended by L. humile. Results from this investigation showed that C. perminutus were more tolerant towards ants than Anagyrus sp., as indicated by their higher parasitism rates which they achieved in the presence of ants. 5. Conclusion These results are important to growers who should be aware of the species of pest ants foraging in their vineyards. Since the responses of parasitoids used in this investigation differed between
ant species, this affects the choice of biological control agent. Currently C. perminutus is commercially available to growers for augmentative releases. The commercial availability of Anagyrus sp. is being investigated, but due to their sensitivity to ants, augmentative releases of Anagyrus sp. near pseudococci in ant infested vineyards may not be as effective. Producers should engage in vineyard management practices that improve and conserve natural and commercially released populations of this parasitoid species. Of these, ant control should be considered a priority when parasitoids are to be used as biological control agents of hemipteran pests as ant presence will not only affect parasitoid abundance but also reproductive success and possibly oviposition strategy of female parasitoids. In vineyards, exclusion of ants, particularly the epigeic L. humile and Anoplolepis sp., is recommended through the use of chemical stem barriers ( Addison, 2002). These barriers prevent ants from gaining access into the vineyard canopy and therefore allow effective biological control of the vine mealybug. Crematogaster peringueyi could also be controlled by removing old and diseased canes that provide nesting sites for this ant species. Acknowledgments Special thanks to Deciduous Fruit Producers Trust (DFPT), THRIP and Winetech for funding this investigation. Many thanks to Dr. K.A. Achiano (Agricultural Research Council, Infruitec-Nietvoorbij, Stellenbosch) for supplying mealybug colonies, and DuRoi IPM, Letsitele, South Africa, for supplying C. perminutus. We also thank Dr. J.S. Terblanche (Stellenbosch University) for statistical assistance. References Abbott, W.S., 1925. A method of computing the effectiveness of an insecticide. Journal of Economic Entomology 18, 265–267. Addison, P., 2002. Chemical stem barriers for the control of ants (Hymenoptera: Formicidae) in vineyards. South African Journal of Enology and Viticulture 23, 1–8. Addison, P., Samways, M.J., 2000. A survey of ants (Hymenoptera: Formicidae) that forage in the vineyards of the Western Cape Province, South Africa. African Entomology 8, 251–260. Adenuga, A.O., 1975. Mutualistic association between ants and some Homoptera – its significance in cacao production. Psyche 82, 24–28. Bartlett, B.R., 1961. The influence of ants upon parasites, predators and scale insects. Annals of Entomological Society of America 54, 543–551. Bradley, G.A., 1973. Effect of Formica obsuripes (Hymenoptera: Formicidae) on the predator–prey relationship between Hyperaspis congressis (Coleoptera: Coccinelidae) and Toumeyella numismaticum (Homoptera: Pseudococcidae). Canadian Entomology 105, 1113–1118. Buckley, R.C., 1987. Interactions involving plants, homoptera and ants. Annual Review of Ecology and Systematics 18, 111–135. Buckley, R., Gullan, P., 1991. More aggressive ant species (Hymenoptera: Formicidae) provide better protection for soft scale and mealybugs (Homoptera: Coccidae; Pseudococcidae). Biotropica 23, 282–286. Compere, H., 1940. Parasites of the black scale, Saissettia oleae, in Africa. Hilgardia 13, 387–425. Daane, K.M., Bentley, W.J., Weber, E.A., 2004. Vine mealybug: a formidable pest spreads throughout California vineyards. Practical Winery Vineyard Management 3, 35–40. Daane, K.M., Sime, K.R., Fallon, J., Cooper, M.L., 2007. Impacts of Argentine ants on mealybugs and their natural enemies in California’s coastal vineyards. Ecological Entomology 32, 583–596. Davies, A.P., Ceballo, F.A., Walter, G.H., 2004. Is the potential of Coccidoxenoides perminutus, a mealybug parasitoid, limited by climatic or nutritional factors? Biological Control 31, 181–188. Flanders, S.E., 1943. The Argentine ant versus the parasites of the black scale. The California Citrograph, March. pp. 117–137. Gibernau, M., Dejean, A., 2001. Ant protection of a Heteroptera trophobiont against a parasitoid wasp. Oecologia 126, 53–57. Grover, C.D., Kay, A.D., Monson, J.A., Marsh, T.C., Holway, D.A., 2007. Linking nutrition and behavioural dominance, carbohydrate scarcity limits aggression and activity in Argentine ants. Proceedings of the Royal Society Bulletin 274, 2951–2957. Henschel, J.R., 1998. Predation on social and solitary individuals of the spider Stegodyphus dumicola (Araneae, Eresidae). Journal of Arachnology 26, 61–69. Hölldobler, B., Wilson, E.O., 1990. The Ants. Belknap Press, Harvard University, Cambridge, Massachusetts. 732pp.
N. Mgocheki, P. Addison / Biological Control 49 (2009) 180–185 Horton, J.R., 1918. The Argentine Ant in Relation to Citrus Groves. Bulletin No. 647. Government Printing Office, Washington, USA. Itioka, T., Inoue, T., 1996. The role of predators and attendant ants in the regulation and persistence of a population of citrus mealybug, Pseudococcus citriculus, in a Satsuma orange orchard. Applied Entomology and Zoology 31, 195–202. Islam, K.S., Copland, M.J.W., 1997. Host preference and progeny sex ratio in a solitary koinobiont mealybug endoparasitoids, Anagyrus pseudococci (Girault) in response to its host stage. Biocontrol Science and Technology 7, 449–456. Islam, K.S., Perera, H.A.S., Copland, M.J.W., 1997. The effects of parasitism by an encyrtid parasitoid, Anagyrus pseudococci, on the survival, reproduction and physiological changes of the mealybug, Planococcus citri. Entomologia Experimentalis et Applicata 34, 77–83. Jiggins, C., Majerus, M.E.N., Gough, U., 1993. Ant defence of colonies of Aphis fabae Scolopi (Hemiptera: Aphididae), against predation by lady birds. British Journal of Natural History 6, 129–137. Joyce, A.L., Hoddle, M.S., Bellows, T.S., Gonzalez, D., 2001. Oviposition behaviour of Coccidoxenoides peregrinus, a parasitoid of Planococcus ficus. Entomologia Experimentalis et Applicata 98, 49–57. Kaneko, S., 2003. Impacts of two ants, Lasius niger and Pristomyrmex pungens (Hymenoptera: Formicidae), attending the brown citrus aphid, Toxoptera citricidus (Homoptera: Aphidiidae), on the parasitism of the aphid by a primary parasitoid, Lysiphlebus japonicus (Hymenoptera: Aphidiidae), and its larval survival. Applied Entomology and Zoology 38, 347–357. Kriegler, P.J., Whitehead, V.B., 1962. Notes on the biology and control of Crematogaster peringueyi var. angustior, Arnold on grape vines (Hymenoptera: Formicidae). Journal of Entomological Society of Southern Africa 25, 287–290. Lach, L., 2003. Invasive ants: unwanted partners in ant–plant interactions? Annals of the Missouri Botanical Garden 90, 91–108. Liang, K.Y., Zeger, S.L., 1986. Longitudinal data analysis using general linear models. Biometrika 73, 13–22. Markin, G.P., 1970. Foraging behavior of the Argentine ant in California citrus grove. Journal of Economic Entomology 63, 740–744. Martinez-Ferrer, M.T., Grafton-Cardwell, E.E., Shorey, H.H., 2003. Disruption of parasitism of the Californian red scale (Homoptera: Diaspididae) by three ant species (Hymenoptera: Formicidae). Biological Control 26, 279–286. Nixon, G.E.J., 1951. The Association of Ants with Coccids. Commonwealth Institute of Entomology, London. 36pp.
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Pierce, N.E., Mead, P.S., 1981. Parasitoids as selective agents in the symbiosis between Lycaenid butterfly larvae and ants. Science 211, 1185–1187. Roura-Pascual, N., Suarez, A.V., Gomez, C., Pons, P., Touyama, Y., Wild, A.L., Peterson, A.T., 2004. Geographical potential of Argentine ants (Linepithema humile Mayr) in the face of global climate change. Proceedings of the Royal Society of London Series B-Biological Sciences 271, 2527–2534. Samways, M.J., Tate, B.A., 1984. Evaluation of several trunk barriers used to prevent the movement of the pugnacious ant (Anoplolepis custodiens (Smith)) into citrus trees. Citrus and Subtropical Fruit Journal 608, 9–12. Schatz, B., Hossaert-McKey, M., 2003. Interactions of the ant Crematogaster scutellaris with fig/fig wasp mutualism. Ecological Entomology 28, 359–368. Steyn, J.J., 1954. The pugnacious ant Anoplolepis custodiens (Smith) and its relation to the control of the citrus scale at Letaba. Memoirs of the Entomological Society of Southern Africa No. 3, Pretoria. Tingle, C.C.D., Copland, M.J.W., 1988. Predicting development of the mealybug parasitoids Anagyrus pseudococci, Leptomastix dactylopii and Leptomastidea abnormis under glasshouse conditions. Entomologia experimentalis et Applicata 46, 19–28. Tingle, C.C.D., Copland, M.J.W., 1989. Progeny production and adult longevity of the mealybug parasitoids Anagyrus pseudococci, Leptomastix dactylopii and Leptomastidea abnormis (Hymenoptera: Encyrtidae) in relation to temperature. Entomophaga 34, 111–120. Triapitsyn, S.V., Gonzalez, D., Vickerman, D.B., Noyes, J.S., White, E.B., 2007. Morphological, biological and molecular comparisons among the different geographical populations of Anagyrus pseudococci (Hymenoptera: Encyrtidae), parasitoids of Planococcus spp. (Hemiptera: Pseudococcidae), with notes on Anagyrus dactylopii. Biological Control 41, 14–24. Varon, E.H., Hanson, P., Longino, J.T., Borbon, O., Carballo, M., Hilje, L., 2007. Temporal and spatial distribution of ants in a light gradient in a coffee agroforestry system, Turrialba, Costa Rica. Revista de Biologia Tropical 55, 943– 956. Walton, V.M., Pringle, K.L., 2005. Developmental biology of vine mealybug, Planococcus ficus (Signoret) (Homoptera: Pseudococcidae), and its parasitoid Coccidoxenoides perminutus (Timberlake) (Hymenoptera: Encyrtidae). African Entomology 13, 143–147. Way, M.J., 1963. Mutualism between ants and honeydew producing homoptera. Annual Review of Entomology 8, 307–344.
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Interference of ants (Hymenoptera: Formicidae) with biological control of the vine mealybug Planococcus ficus (Signoret) (Hemiptera: Pseudococcidae) Nyembezi Mgocheki *, Pia Addison Department of Conservation Ecology and Entomology, Faculty of AgriSciences, Stellenbosch University, Private Bag X1, Matieland, Stellenbosch 7602, South Africa
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Article history: Received 26 August 2008 Accepted 2 February 2009 Available online 12 February 2009 Keywords: Anagyrus sp. Anoplolepis steingroeveri Ant species Coccidoxenoides perminutus Crematogaster peringueyi Linepithema humile Parasitism Mortality Trophobiosis
a b s t r a c t Anagyrus sp. and Coccidoxenoides perminutus are well known parasitoids used for the biological control of the vine mealybug Planococcus ficus, a key pest in vineyards. In South Africa, three ant species, Anoplolepis steingroeveri, Crematogaster peringueyi and Linepithema humile form a trophobiotic relationship with the vine mealybug in vineyards and promote the latter’s infestations to unacceptable levels. In a manipulative laboratory experiment, ants and parasitoids were allowed to forage on vine mealybug-infested butternuts and numbers were recorded for a 1 min period at 10 min intervals for 2 h. Parasitoid mortality and the number of parasitized vine mealybug females were then recorded in the presence and absence of the three ant species. Data were analyzed using a repeated measures generalized linear model (GEEs). The mean number of ants and parasitoids on the mealybug-infested butternuts differed significantly between ant species and time intervals (p < 0.0001 in all cases). Crematogaster peringueyi and L. humile caused significantly higher mortality of both parasitoids than A. steingroeveri during the 24-h exposure period (p < 0.0001). Coccidoxenoides perminutus parasitized significantly more vine mealybugs than Anagyrus sp. for all treatments (p < 0.0001). Ants should therefore be controlled prior to release of parasitoids to suppress populations of ant-tended Hemiptera in vineyards. Crown Copyright Ó 2009 Published by Elsevier Inc. All rights reserved.
1. Introduction Finding new ways of understanding animal behavior is fundamental to the biological control of arthropod pest species. The trophobiotic relationship between ants and their adopted Hemiptera (Gibernau and Dejean, 2001; Jiggins et al., 1993; Hölldobler and Wilson, 1990; Buckley, 1987; Pierce and Mead, 1981; Adenuga, 1975; Bradley, 1973; Bartlett, 1961; Steyn, 1954) needs further investigation to understand various levels of aggression exhibited by ants towards natural enemies of their attended Hemiptera. Field behavioral studies of parasitoids are difficult to carry out due to the complexity of these interactions in the field. Behavior of ants and parasitoids can be broken down into simple components (e.g. ant recruitment and parasitoid oviposition) that can be manipulated under controlled conditions to predict field response that can be considered in planning and implementing a pest management programme. Although many studies have abundantly documented the detrimental impacts of ants on biological control (Bartlett, 1961; Buckley, 1987; Itioka and Inoue, 1996; Martinez-Ferrer et al., 2003), the individual impact of each ant species on individual parasitoids has received limited experimental attention. In agricultural and natural ecosystems, ants obtain carbohydrate-rich honeydew from * Corresponding author. Fax: +27 218084807. E-mail address: [email protected] (N. Mgocheki).
Hemiptera such as mealybugs, scale insects, aphids, among others, while they render protection, sanitation and sometimes transport services to sedentary Hemiptera (Buckley, 1987; Lach, 2003). Carbohydrates serve as a metabolic fuel for most insects and therefore behavioral dominance can be associated with relative availability of and demand for sugar (Grover et al., 2007). Arboreal ants exhibit greater activity and aggression where carbohydrate resources are limited compared to epigeic ants probably due to their numerical superiority in canopies and greater dependency on hemipteran honeydew. Similarly, high levels of activity and aggression exhibited by invasive ants may well be linked to their reliance on carbohydrate-rich honeydew (Way, 1963; Markin, 1970). The Argentine ant Linepithema humile (Mayr) was found to be disruptive to the black scale Saissetia oleae Olivier, parasitoid Coccophagus scutellaris (Dalman) in California (Horton, 1918). In South Africa Metaphycus helvolus (Compere), a parasitoid of black scale was found to be effective in the absence of L. humile ( Flanders, 1943; Compere, 1940). In California, Daane et al. (2007) found that L. humile promoted populations of obscure mealybug Pseudococcus viburni (Signoret) while lowering populations of its parasitoids Pseudaphycus flavidulus and Leptomastix epona. They also noted increased densities of the Argentine ant-tended grape mealybug Pseudococcus maritimus (Erhorn) accompanied by a serious reduction in its parasitoid populations. The cocktail ant Crematogaster peringueyi Emery is disruptive to natural enemies of soft brown scale Coccus hesperidum L., and vine
1049-9644/$ - see front matter Crown Copyright Ó 2009 Published by Elsevier Inc. All rights reserved. doi:10.1016/j.biocontrol.2009.02.001
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mealybugs Planococcus ficus (Signoret). This ant species provides protection to its hemipteran hosts by constructing carton shelters over the mealybugs (Kriegler and Whitehead, 1962). The common pugnacious ant, Anoplolepis custodiens (Smith), incidentally disturbed the parasitoids of California red scale Aonidiella aurantii (Maskell), while tending soft brown scale in citrus orchards in South Africa (Samways and Tate, 1984; Steyn, 1954). Buckley and Gullan (1991) concluded that the incidence of coccid parasitization was correlated with the relative inoffensiveness of the attendant ant species in a field study in Australia. These authors measured low parasitism rates (at least 15%) of coccids in the presence of Tapinoma and Iridomyrmex spp. and <10% in the presence of the more aggressive Oecophylla and Solenopsis species. In California, L. humile reduced parasitism and host mutilation of the California red scale by the parasitoids Comperiella bifasciata (Howard) (59.1%) and Aphytis melinus De Bach (79.5%) in a laboratory trial, even if there were no honeydew-excreting soft scales (Martinez-Ferrer et al., 2003). Itioka and Inoue (1996), in a comparative field investigation, found a 94% decrease of the mealybug Pseudococcus citriculus Green by natural enemies in the absence of the attendant ant Lasius niger (L.). Previous (unpublished) studies revealed that more than one ant species and parasitoid species were sampled from the same sites and therefore parasitism rates could not reflect the individual impact of each ant species on mealybug biological control. Anagyrus sp. and Coccidoxenoides perminutus (Timberlake) (Hymenoptera: Encyrtidae) are widely distributed primary parasitoids of Planococcus spp. and Pseudococcus spp. (Davies et al., 2004; Triapitsyn et al., 2007), that have been used for classical and augmentative biological control of the vine mealybug and citrus mealybug Planococcus citri (Risso) in some localities (Daane et al., 2004; Walton and Pringle, 2005). This study tests the interference of three ant species on the biological control of the vine mealybug P. ficus using the parasitoids Anagyrus sp. near pseudococci (as identified by Triapitsyn et al., 2007) and C. perminutus. This investigation used the presence and absence of these ant species to quantify their individual impact on the parasitoids under controlled laboratory conditions. For the purposes of this study, Anagyrus sp. near pseudococci shall be referred to as Anagyrus sp. from this point onwards. 2. Materials and methods 2.1. Insect colonies 2.1.1. Vine mealybug colonies Colonies of vine mealybugs were maintained on mature fruit of butternut squash Cucurbita moschata (L.) in the laboratory at 27 ± 1 °C with a 12:12 (L:D) hour photoperiod and 65 ± 5% RH. Butternut squash were obtained fresh from the local grocery store and were more or less of equal size. Before use in experiments, butternuts were washed in 5% bleach solution (to prevent fungal growth), triple rinsed, air dried and then inoculated with vine mealybug crawlers after 1 h. After the first molt, mealybugs were thinned to approximately 100 individuals that were allowed to develop up to a desired stage before use in experiments as hosts to parasitoids. For Anagyrus sp., 3rd instar to preovipositing female vine mealybugs were used while for C. perminutus, 2nd instar mealybugs were used (Islam and Copland, 1997; Joyce et al., 2001). 2.1.2. Ant colonies A single average ant nest (at least 500 workers) for all three ant species, consisting of workers, queens and immatures, was collected from commercial vineyards. For epigeic ants, a spade was used to collect a portion of the nest and maintained in soil from the original nest. A small size carton nest was collected for the
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arboreal C. peringueyi. Nests of all three species were placed into five liter square plastic containers. The set up was designed to mimic a natural situation as far as possible, where ants are regarded as pests and as such ants were collected from vineyards where their infestation levels were above the action threshold. Anoplolepis steingroeveri and C. peringueyi were collected from Ashton ( 33.85°S, 20.08°E, 186 m) in the Breede River Valley (BRV) while L. humile were collected from Simondium ( 33.83°S, 18.83°E, 175.2 m) in the Stellenbosch area, South Africa. The three ant species were maintained in plastic containers (18 18 16 cm) in the laboratory containing soil or material from the original nests. Each ant nest was connected to a clear Perspex container (25 25 20 cm) with clear plastic tubing (20 cm long and 6 mm in diameter). A butternut infested with about 100 mealybugs was placed into each Perspex container and ants were allowed to forage freely on this butternut for honeydew until 48 h prior to the experiment. All ant colonies were kept at 27 ± 0.5 °C, 65 ± 5% RH and a 12:12 (L:D) photoperiod. 2.1.3. Parasitoid colonies 2.1.3.1. Anagyrus sp.. Field collected vine mealybugs were incubated individually in gelatin capsules at room temperature. Incubated mealybugs were monitored daily, and emerged Anagyrus sp. were selected for experimental use through stereo microscope identification. Only Anagyrus sp. were selected, based on antennal coloration according to Triapitsyn et al. (2007). The parasitoids were placed in a cage (66 66 37 cm) containing butternuts infested with vine mealybugs. Parasitoids were offered a 50:50 honey:water solution and kept at 27 °C, 65 ± 5% RH with a 12:12 (L:D) photoperiod. After seven days, parasitized mealybugs were moved into another cage for parasitoid emergence. Anagyrus sp. colonies were maintained in the laboratory on 3rd instar to adult vine mealybugs feeding on butternut squash (Islam et al., 1997). Testing was done when a total of 40 newly emerged female parasitoids (total per trial) were available and each individual was used only once. Newly emerged parasitoids were allowed to feed and mate before use in trials. One male parasitoid was given access to five newly emerged females for 24 h (Tingle and Copland 1988, 1989). Only mated two-day old females were used in the experiment. After every three generations, Anagyrus sp. from the field were added to laboratory colonies to prevent inbreeding of the laboratory colony. 2.1.3.2. Coccidoxenoides perminutus. Coccidoxenoides perminutus were obtained from DuRoi Integrated Pest Management (Letsitele, South Africa) as mature pupae. Newly emerged individuals were allowed to feed for 24 h after which they were used in the experiments. No mating was necessary as C. perminutus are thelytokous (Davies et al., 2004). Both parasitoid species were deprived of hosts 24 h prior to the experiment. 2.2. Quantitative observations Ants foraged on a butternut infested with 100 vine mealybugs in each of the six experimental cages of 21.5 21.5 16 cm (one for each ant species per parasitoid species). An ant-free cage was included as a control for each parasitoid species. Each experimental cage had a hole (6 mm in diameter) in the side which was plugged with cotton wool. This hole was only unplugged during introduction of parasitoids into the cages. A single tube aspirator was used to collect parasitoids from their rearing cages and release the parasitoids into the experimental cage through the side hole by gently blowing through the tube. The ants were allowed to forage for 3 h before 20 two-day old fertilized Anagyrus sp. females and 20 one-day old C. perminutus were introduced. Observations were made 10 min after the release of parasitoids for each treatment
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whereby the number of ants and parasitoids on the whole butternut was recorded during a 1 min period at 10 min intervals for 2 h. The aspirator tube was used to ensure the accuracy of the counted parasitoids before and after release. This also allowed the parasitoids to be in the experimental cage at approximately the same time and therefore equal exposure to ants however minimizing vulnerability of parasitoids to ants. Parasitoids were then left in the experimental cages for 24 h after which they were removed and the numbers of surviving, dead and/or missing parasitoids, if any, were recorded. Mortality of the parasitoids was defined as the number of dead + missing parasitoids/total number of parasitoids and expressed as a percentage. Magnifying lenses were used to search for missing parasitoids, which could also have been dismembered or carried back to the nest by ants. After 24 h, all mealybugs were removed and incubated individually in gelatin capsules at 27 °C, 65 ± 5% RH with a 12:12 (L:D) photoperiod, for two weeks after which they were examined for parasitism under a stereo microscope. Percentage parasitism of the vine mealybug was defined as the number of parasitized mealybugs/total number of mealybugs. The tests were performed on five different dates for each parasitoid species with five replicates per ant colony and their controls (ant-free treatment).
workers (32.30 ± 2.21), A. steingroeveri from 5 to 25 workers (16.17 ± 2.11) and C. peringueyi from 315 to 401 (354.98 ± 4.42) (Fig. 2). In C. perminutus-released cages, L. humile ranged from 19 to 36 workers (28.45 ± 2.18), A. steingroeveri from 0 to 15 (5.65 ± 1.93) and C. peringueyi from 290 to 335 (313 ± 1.66) during 1 min observation periods (n = 5 in all cases). Significantly more ants foraged when Anagyrus sp. were released than when C. perminutus were released (Fig. 3). The number of foraging ants for both parasitoid species generally decreased with time. The mean number of ants on the mealybug-infested butternuts was influenced by the ant and parasitoid species and the time intervals so that the interaction; ant species parasitoid species time was significant (v2 = 5.491E8; df = 33; p < 0.0001). 3.2. Parasitoid behavior in the absence and presence of ants During 1 min observation periods, the number of Anagyrus sp. increased from 0 to 1 (0.07 ± 0.06) for C. peringueyi, 0 to 3
370 360
Data were analyzed using Generalized Estimating Equations (GEEs) (Liang and Zeger, 1986) in GENMOD procedure in SAS (SAS Enterprise Guide 3, 2004) using Poisson distribution and an identity link function. Abbott’s correction formula (Abbott, 1925) was used to correct for control mortality.
350 No. of ants
2.3. Data analysis
340 330
3. Results
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3.1. Ant behavior in the presence of parasitoids
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Where Anagyrus sp. were released, the number of L. humile on butternuts during 1 min periods (Fig. 1) ranged from 22 to 42
300 10
20 30
40
50 60 70 80 Time (minutes)
C. peringueyi/C. perm.
90 100 110 120
C. peringueyi/Anagyrus sp.
40 Fig. 2. The number of Crematogaster peringueyi observed during 1 min intervals on a vine mealybug-infested butternut during a 2 h observation period.
35 30
c
350 d
20 No. OF ANTS
No. of ants
400 25
15 10 5
Anagyrus sp. C. perminutus
300 250 200 150 100
0 10
20 30
40 50 60 70 80 90 100 110 120 Time (minutes) A. steingroeveri/Anagyrus sp. A. steingroeveri/C.perm. L. humile/Anagyrus sp. L. humile/C.perm.
Fig. 1. The number of Anoplolepis steingroveri and Linepithema humile observed during 1 min intervals on a vine mealybug-infested butternut during a 2 h observation period.
50
a ab
0 L.humile
b
e
C. peringueyi A. steingroeveri ANT SPECIES
Fig. 3. The mean number of ants, Linepithema humile, Crematogaster peringueyi and Anoplolepis steingroeveri on a mealybug-infested butternut for different parasitoid species during 1 min observation periods over 2 h. Means followed by different letters are significantly different at p 6 0.05.
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(0.57 ± 0.22) for L. humile, 0 to 9 (3.33 ± 3.42) for A. steingroeveri and 7 to 16 (12.92 ± 1.93) in the ant-free cage. The number of C. perminutus ranged from 0 to 2 (0.1 ± 0.15) for C. peringueyi, 0 to 7 (1.12 ± 1.53) for L. humile, 0 to 18 (10.57 ± 4.25) and 9 to 18 (13.65 ± 1.03) in the ant-free cage (n = 5 in all cases). Fig. 4 illustrates the trends in the number of parasitoids on butternuts in the presence or absence of ants. The mean number of parasitoids foraging on the mealybug-infested butternut differed significantly
18
3.3. Effects of ants on parasitoid mortality and mealybug parasitism
16
Parasitoid mortality was significantly different between parasitoid species (v2 = 13.47; df = 1; p < 0.001) and ant species (v2 = 2168.53; df = 3; p < 0.0001) (Fig. 5). Anoplolepis steingroeveri caused the least parasitoid mortality during the 24-h exposure period, while C. peringueyi caused the most. Percentage parasitism differed significantly (p < 0.0001) between parasitoid species (v2 = 38.18; df = 1) and ant species (v2 = 10351.8; df = 3). Coccidoxenoides perminutus caused significantly more parasitism than Anagyrus sp. while percentage parasitism was lowest in the presence of C. peringueyi (Fig. 6).
14 No. of parasitoids
over the 2 h period and was influenced by ant species parasitoid species time interval (v2 = 3.883E7; df = 66; p < 0.0001). Coccidoxenoides perminutus searched for mealybugs in higher numbers than Anagyrus sp. The highest number of parasitoids (both species) searched for vine mealybug on A. steingroeveri-infested butternuts and the lowest on C. peringueyi-infested butternuts. These data are an indication that all three ant species are capable of preventing parasitoids from accessing the vine mealybug, but to varying degrees.
12 10 8 6 4 2
4. Discussion
0 10
20
30
40 50 60 70 80 90 100 110 120 Time after release (minutes)
Anagyrus sp. (A. steingroveri)
C. perm (A.steingroeveri)
Anagyrus sp.(L. humile)
C. perm (L. humile)
Anagyrus sp.(C. peringueyi)
C. perm (C. peringueyi)
Anagyrus sp.(Ant free)
C. perm (Ant free)
Fig. 4. The number of Anagyrus species near pseudococci and Coccidoxenoides perminutus observed on a vine mealybug-infested butternut during 1 min intervals over a 2 h observation period in the presence and absence of one of three ant species.
Different ant species have different strategies for defending Hemiptera against parasitoids (Buckley and Gullan, 1991; Kaneko, 2003). Attendance by all three ant species significantly reduced the number and percentage mortality of parasitoids on P. ficus colonies and resultant parasitism in the present study. However, certain ant species were found to be more aggressive, while certain parasitoids were found to be more ant-resistant. Parasitism occurred less frequently in A. steingroeveri attended P. ficus probably due to incomplete protection. Martinez-Ferrer et al. (2003) noted that larger ants do not easily recognize small natural enemies. Anoplolepis
% mortality
100 80
Anagyrus sp. C.perminutus
60 40 20 0 A.steingroeveri
L. humile C.peringueyi Ant treatment
Control
Fig. 5. The mean (±standard error) percentage mortality of Anagyrus species near pseudococci and Coccidoxenoides perminutus after a 24-h exposure to different ant treatments. Means with different letters are significantly different at p 6 0.05.
% parasitism
100
C.perminutus Anagyrus sp.
80 60 40 20 0
A.steingroeveri
L. humile C.peringueyi Ant treatment
Control
Fig. 6. Parasitism rates (% mortality ± standard error) of P. ficus by Anagyrus species near pseudococci and Coccidoxenoides perminutus after a 24 h exposure period to various ant species. Means with different letters are significantly different at p 6 0.05.
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steingroeveri are mainly predatory, epigeic ants which tend to attack other insects within their foraging territory, promoting mealybug infestations due to incidental protection from natural enemies (Way, 1963; Henschel, 1998). However, their impact on mealybug biological control is limited to drier regions as they appear to be negatively affected by flooding (e.g. flood and micro jet-irrigated vineyards) (Addison and Samways, 2000). As this study indicated furthermore, their impact on parasitism was not as marked as the other ant species. Crematogaster peringueyi recruited in large numbers to the food source and therefore negatively affected P. ficus parasitism rates the most for both parasitoids species. Almost complete protection is afforded by creation of a biological barrier that covered the mealybug-infested butternut and mealybugs were immune to parasitoid attack. Schatz and Hossaert-McKey (2003) and Varon et al. (2007) described arboreal ants like Crematogaster spp. as predacious on other insects in plant canopies. This allows such ant species to obtain protein to complement their carbohydrate-rich diet. Although very aggressive, C. peringueyi are not widely distributed in Western Cape vineyards (Addison and Samways, 2000). They are confined to old neglected vineyards where they utilize old or diseased canes for nest building, presumably providing better protection to mealybugs than would epigeic ant species. Their impact is significant when their infestations are high but at low infestations, this species often confines itself to their nests with small colonies of mealybugs and therefore parasitoid activity in the vine foliage may not be disturbed much by the ants (Kriegler and Whitehead, 1962). Given their invasive nature and wide distribution, possibly perpetrated by global warming (Roura-Pascual et al., 2004), L. humile present a serious threat to the biological control of hemipteran pests like the vine mealybug, as they are capable of affecting parasitoids in vineyards over large areas of the Western Cape, South Africa (Addison and Samways, 2000). In vineyards, they make numerous nests in the floor and construct temporary nests on vine stems, leaves and bunches (Horton, 1918; personal observation), thereby posing a serious threat to mealybug parasitoids. Nixon (1951) noted that the behavior of a parasitoid in the presence of ants largely determines its own effectiveness as a biological control agent. Anagyrus sp. and C. perminutus evoked different responses in the ants. From personal observation, Anagyrus sp. often got entangled with the host while ovipositing and were then seized more readily by ants than C. perminutus, while struggling to retract their ovipositor. Parasitoids abandoned oviposition and kept away from mealybugs to avoid ants, limiting the number of eggs that could be laid into the host. While some parasitoids have developed escape strategies from ants to improve their efficacy, others are so ant sensitive that after an encounter with ants, they are deterred not only by ants, but by any moving object including other parasitoids or the host itself, thereby greatly reducing their potential as biological control agents (Martinez-Ferrer et al., 2003). It is apparent that ants not only interfere with percentage parasitism of their adopted Hemiptera, but also reduce parasitoid abundance by causing direct mortality and low reproductive success. Daane et al. (2007) noticed an almost complete absence of parasitoids in vineyards infested with P. maritimus attended by L. humile. Results from this investigation showed that C. perminutus were more tolerant towards ants than Anagyrus sp., as indicated by their higher parasitism rates which they achieved in the presence of ants. 5. Conclusion These results are important to growers who should be aware of the species of pest ants foraging in their vineyards. Since the responses of parasitoids used in this investigation differed between
ant species, this affects the choice of biological control agent. Currently C. perminutus is commercially available to growers for augmentative releases. The commercial availability of Anagyrus sp. is being investigated, but due to their sensitivity to ants, augmentative releases of Anagyrus sp. near pseudococci in ant infested vineyards may not be as effective. Producers should engage in vineyard management practices that improve and conserve natural and commercially released populations of this parasitoid species. Of these, ant control should be considered a priority when parasitoids are to be used as biological control agents of hemipteran pests as ant presence will not only affect parasitoid abundance but also reproductive success and possibly oviposition strategy of female parasitoids. In vineyards, exclusion of ants, particularly the epigeic L. humile and Anoplolepis sp., is recommended through the use of chemical stem barriers ( Addison, 2002). These barriers prevent ants from gaining access into the vineyard canopy and therefore allow effective biological control of the vine mealybug. Crematogaster peringueyi could also be controlled by removing old and diseased canes that provide nesting sites for this ant species. Acknowledgments Special thanks to Deciduous Fruit Producers Trust (DFPT), THRIP and Winetech for funding this investigation. Many thanks to Dr. K.A. Achiano (Agricultural Research Council, Infruitec-Nietvoorbij, Stellenbosch) for supplying mealybug colonies, and DuRoi IPM, Letsitele, South Africa, for supplying C. perminutus. We also thank Dr. J.S. Terblanche (Stellenbosch University) for statistical assistance. References Abbott, W.S., 1925. A method of computing the effectiveness of an insecticide. Journal of Economic Entomology 18, 265–267. Addison, P., 2002. Chemical stem barriers for the control of ants (Hymenoptera: Formicidae) in vineyards. South African Journal of Enology and Viticulture 23, 1–8. Addison, P., Samways, M.J., 2000. A survey of ants (Hymenoptera: Formicidae) that forage in the vineyards of the Western Cape Province, South Africa. African Entomology 8, 251–260. Adenuga, A.O., 1975. Mutualistic association between ants and some Homoptera – its significance in cacao production. Psyche 82, 24–28. Bartlett, B.R., 1961. The influence of ants upon parasites, predators and scale insects. Annals of Entomological Society of America 54, 543–551. Bradley, G.A., 1973. Effect of Formica obsuripes (Hymenoptera: Formicidae) on the predator–prey relationship between Hyperaspis congressis (Coleoptera: Coccinelidae) and Toumeyella numismaticum (Homoptera: Pseudococcidae). Canadian Entomology 105, 1113–1118. Buckley, R.C., 1987. Interactions involving plants, homoptera and ants. Annual Review of Ecology and Systematics 18, 111–135. Buckley, R., Gullan, P., 1991. More aggressive ant species (Hymenoptera: Formicidae) provide better protection for soft scale and mealybugs (Homoptera: Coccidae; Pseudococcidae). Biotropica 23, 282–286. Compere, H., 1940. Parasites of the black scale, Saissettia oleae, in Africa. Hilgardia 13, 387–425. Daane, K.M., Bentley, W.J., Weber, E.A., 2004. Vine mealybug: a formidable pest spreads throughout California vineyards. Practical Winery Vineyard Management 3, 35–40. Daane, K.M., Sime, K.R., Fallon, J., Cooper, M.L., 2007. Impacts of Argentine ants on mealybugs and their natural enemies in California’s coastal vineyards. Ecological Entomology 32, 583–596. Davies, A.P., Ceballo, F.A., Walter, G.H., 2004. Is the potential of Coccidoxenoides perminutus, a mealybug parasitoid, limited by climatic or nutritional factors? Biological Control 31, 181–188. Flanders, S.E., 1943. The Argentine ant versus the parasites of the black scale. The California Citrograph, March. pp. 117–137. Gibernau, M., Dejean, A., 2001. Ant protection of a Heteroptera trophobiont against a parasitoid wasp. Oecologia 126, 53–57. Grover, C.D., Kay, A.D., Monson, J.A., Marsh, T.C., Holway, D.A., 2007. Linking nutrition and behavioural dominance, carbohydrate scarcity limits aggression and activity in Argentine ants. Proceedings of the Royal Society Bulletin 274, 2951–2957. Henschel, J.R., 1998. Predation on social and solitary individuals of the spider Stegodyphus dumicola (Araneae, Eresidae). Journal of Arachnology 26, 61–69. Hölldobler, B., Wilson, E.O., 1990. The Ants. Belknap Press, Harvard University, Cambridge, Massachusetts. 732pp.
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