Journal of Chemical Ecology, Vol. 12, No. 1, 1986
CHEMICAL ATTRACTANTS TESTED AGAINST THE AUSTRALIAN BUSH FLY Musca vetustissima (DIPTERA: MUSCIDAE)
MIR
S. M U L L A
1 and JAMES
T.
RIDSDILL-SMITH
2
IDepartment of Entomology University of California, Riverside Riverside, California 92521 2Division of Entomology, CSIRO Private Bag P. O. Wembley, Western Australia, 6014 (Received March 21, 1985; accepted June 21, 1985)
A b s t r a c t - - A number of chemical compositions known to attract several synanthropic and pest fly species were tested against the Australian bush fly Musca vetustissima, a severe and persistent pest of man and domesticated and wild mammals. A standard composition containing small quantities of trimethylamine and indole, blended with large amounts of ammonium sulfate and anchovy meal, showed good attractancy against this fly. Deletion of ammonium sulfate did not significantly alter attractancy. Similarly, incorporation of n-butyric acid and linoleic acid into the formulation did not alter attmctancy, nor did addition of several carbonates and bicarbonates. Skatole, a closely related compound to indole, decreased attractancy significantly. In all the tests, the attmctant compositions lured greater number of females than males. The attractancy of the standard composition was studied and compared with equal amount of fresh cattle dung. Chemical attractants lured bush flies and not dung beetles, while dung attracted both flies and beetles. The dung resource was 4.6 • more attractive to flies than the standard attractant composition. This suggests that there are other chemical attractants emanating from dung that warrant isolation and identification work. Nevertheless, the chemical attractants studied here provide a starting point for further investigations. The ratio of female to male flies attracted to humans was greater than the ratio of flies attracted to chemicals. The sex ratio of flies netted from human host was 3.0, while the sex ratio of flies attracted to chemical attractants was 1.8. The preponderance of females attracted to both sources was probably due to physiological requirements of the sexes. Key W o r d s - - B u s h fly, Australian bush fly, Musca vetustissima, Diptera, Muscidae, chemical attractants.
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262
MULLA AND RIDSDILL-SMITH INTRODUCTION
Early explorers in Australia noted the pestiferous nature of the Australian bush fly Musca vetustissima Walker, some three centuries ago (Cleland, 1913). One of the explorers, John Lort Stokes, in 1838 related that "the flies are at you all day, crawling into your eyes, up your nostrils and down your throat" (Hooper, 1982). The nature and scope of annoyance brought about by these flies remains the same today. This pest is widely spread over most of the continent (Hughes et al., 1972) and is especially abundant in cattle-raising areas. They breed in dung, preferring freshly deposited dung of cattle (Hughes, 1981; Hughes et al., 1972; Norris, 1966). Adult bush flies are attracted to large animals, including man, females outnumbering the males attacking or landing on hosts (Hughes, 1977, 1981; Hughes et al., 1972; Norris, 1966). We have noted that the flies can find the host in a few seconds in the field and that flies at some distance orient and move upwind toward the hosts. Chemical odors emanating from hosts or breeding medium (dung) are some of the factors providing cues for directed response by the flies. However, no definitive studies on the role of chemical attractants influencing bush fly behavior have been accomplished on this important synanthropic and synzootic fly. The hovering and feeding behavior of bush flies approaching and attacking hosts is quite similar to that of the tiny flies known as eye gnats (Hippelates species, family Chloropidae) prevailing in the Westem hemisphere (Mulla and Stains, 1977). Chemical attractants inducing positive responses in female H. collusor (Townsend) were isolated and identified from putrified chicken eggs (Hwang and Mulla, 1973; Hwang et al., 1975, 1976) and field tested by Mulla and Axelrod (1974) and Mulla et al. (1974, 1976). Some of the principal chemicals attracting H. collusor were also found to manifest a high level of attractancy against the common house fly Musca domestica (Mulla et al., 1977), and the coastal lesser house fly Fannia femoralis on poultry ranches (Mulla et al., 1984). It is well known that many species of synanthropic and synzootic flies are attracted by odors emanating from putrefying proteins. Musca sorbens, a closely related species to M. vetustissima, was found to be attracted to putrefying egg and fish meat (Legner et al., 1974), and baiting of an insular population with these baits reduced populations of M. sorbens markedly. M. vetustissima has been found to be attracted to liver baits with sodium sulfide (Hughes, 1977; Norris, 1966; Vogt et al., 1981). Most recently Vogt et al. (1985) have used a mixture of putrefying liver and dung with live blow fly maggots to stir the bait for the purpose of trapping bush flies. In view of these findings and the extensive research on the isolation and identification of chemicals showing field attractancy to a variety of pest flies, the present studies were initiated. Compositions of chemical attractants with
AUSTRALIAN BUSH FLY ATTRACTANTS
263
proven attractancy to Hippelates, Musca, and Fannia species were evaluated against the Australian bush fly. A simple method for field evaluation of attractant compositions was developed, where attractancy of various chemical compositions and natural attractants emanating from fresh dung was tested against field populations of M. vetustissima.
METHODS
AND MATERIALS
Compositions of chemical attractants found attractive to the eye gnat Hippelates collusor (Hwang et al., 1975, 1976; Mulla et al., 1976), Musca domestica (Mulla et al., 1977), and Fannia femoralis (Mulla et al., 1984) consisted of mixtures of a carder (anchovy meal) with one or more of the following chemicals: trimethylamine (from trimethylamine hydrochloride), ammonia (from ammonium sulfate), indole, linoleic acid, and a few lower aliphatic acids. Against the bush fly, a composition containing anchovy meal cartier (57.25 %), trimethylamine hydrochloride (2.5 %), ammonium sulfate (40 %), and indole (0.25 %) by weight was used as a standard mix with which other compositions with fewer or more chemicals were compared for luring the flies. When additional chemicals were included or deleted, the content of carrier was varied in the composition accordingly. The composition was mixed in 1:1 ratio with Improved Golden Malrin (IGM) (Zoecon Corporation, Dallas, Texas) toxic bait. The ingredients of this bait are: 1% methomyl, 0.025 % (Z)-9-tricosene, and 98.97 % inert ingredients. Fine sand (780 g) forming a 2-cm layer was placed in a plastic bowl (18 cm diam. and 8 cm deep) and wetted with 140 ml of tapwater, bringing the moisture content initially to its field capacity level. For testing, a 5-g quantity of each mix was placed on the damp sand. In some tests, especially in testing attractancy of dung, the IGM was not mixed with the attractants, but rather it was spread on the top and around the dung or other attractant compositions. The bowls (replicated five to six times in each experiment) containing various test attractant compositions were placed on the ground 5 m apart in paddocks infested with bush flies. They were distributed in randomized-block design in a line at right angles to the wind direction or in a circular pattern. The tests were run in two areas near Perth, Western Australia. The Serpentine area, with extensive livestock pastures, is located in the coastal plain 100 km south of Perth, while Cunderdin, a predominantly wheat growing and pasturing area, is located on the Darling plateau 150 km east of Perth. Flies attracted to the compositions oriented toward the attractant source and crawled or flew into the bowls, walking and milling around the attractanttoxicant compositions. Because of the action of the quick-acting toxicant methomyl, the flies on contact with the bait died in the bowl within a few seconds after arrival. All flies killed in the bowls were counted with no loss due to wind
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MULLA AND RIDSDILL-SMITH
or transportation into the laboratory. The compositions were exposed in the field for a period of 4-6 hr for each experiment, depending on the activity of bush flies. After termination of each experiment, the bowls were covered with lids and brought into the laboratory where the flies were identified, sexed, and counted under a dissecting microscope. Analysis of data was carried out using standard computer program design for randomized-block analysis, The data were transformed to log (n + 1) when fly numbers were high and variable. Transformed means were compared for significant differences using Duncans's multiple-range test, letter designations for denoting significant differences are shown on untransformed means for ease of interpretation. Where there were two treatments in a given test, the transformed data were analyzed by means of t test. Significant differences between or among means are expressed in the tables. RESULTS AND DISCUSSION
Since the attractant composition utilized fish meal as carrier and Improved Golden Malrin (IGM) as a toxicant, it was necessary to test the attractancy of these two components. Fish meal and IGM (1:1) and IGM alone were compared in the first test. The fish meal showed a low level of attractancy which was significantly higher than that of IGM alone (Table 1). When the standard composition plus IGM was compared with fish meal plus IGM, the standard composition caught significantly higher numbers than the carrier (Table 1, test II). In a third test, this comparison was repeated, and again the standard attracted significantly higher numbers than the carrier (Table 1, test II-A). On the basis of these data, it was concluded that a composition containing three chemicals, carrier, and IGM was more attractive than the carrier plus IGM to field popuTABLE 1. ATTRACTION OF AUSTRALIAN BUSH FLY Musca vetustissima TO CHEMICAL ATTRACTANTS AND TOXICANT BAIT, IMPROVED GOLDEN MALRIN (IGM)
Ingredients and % composition Experiment and treatment I
A B II A B (std.) II-A A B (std.)
TMA 9 HC1
Indole
NH4SO 4
Anchovy meal
Mean No. flies killed/unit~
0 0 0 2.5 0 2.5
0 0 0 0.25 0 0.25
0 0 0 40.00 0 40.00
100 0c 100 57.25 100 57.25
5.6 *b 1:0 45.5 78.4*** 18.4 39.6**
aN = 5. Significant differences by t-test for each pair in each test. b'*significant at 0.10 level;**, significant at 0.05 level;***, significant at 0.01 level. c 100% IGM toxic bait.
AUSTRALIAN BUSH FLY ATTRACTANTS
265
TABLE 2. ATTRACTANCY OF VARIOUS COMPOSITIONS OF CHEMICAL AGENTS TO
AUSTRALIANBUSH FLY M. vetustissima (EXPERIMENTIV) Ingredients and % composition Treatment A B C D (std.)
TMA 9 HC1
Indole
2.5 2.5
0.25
2.5
0.25 0.25
NHaSO 4
Anchovym e a l
Mean No. flies killed/unita
40.0 40.0 40.0
97.25 57.50 59.75 57.25
26.0 ab 15.3 bc 14.5 c 30.0 a
aN = 6. Means followed by same letters not significantlydifferentfrom each other at 0.05 level. lations of the Australian bush fly, but the carrier itself did show some attractancy also. Compositions were prepared in which one of the three chemicals in the standard was deleted to see if deletion of each chemical would result in increased or decreased attractancy. The standard composition attracted significantly higher numbers than the compositions lacking trimethylamine hydrochloride or indole (Table 2). The composition lacking a m m o n i u m sulfate was not significantly different from the standard. In studies on Hippelates eye gnats and house flies, n-butyric acid was found to increase the attractancy of chemical compositions (Hwang et al., 1975; Mulla et al., 1977). W e deemed it desirable to study attractancy of this chemical, a m m o n i u m carbonate, and sodium carbonate in the standard composition (Table 3). n-Butyric acid and a m m o n i u m carbonate did not enhance attractancy. Addition of sodium carbonate, however, increased attractancy of the composition. Additionally, calcium carbonate and sodium bicarbonate were studied for enhancing attractancy, but neither one significantly increased attractancy (data not presented). TABLE 3. ATTRACTANCY OF VARIOUS COMPOSITIONS OF ATTRACTANT CHEMICALS TO AUSTRALIAN BUSH FLY M. vetustissima (EXPERIMENT V)
Ingredients and % composition Treatment E (n-butyricacid 5%) F (ammoniumcarbonate 10%) G (sodium carbonate 10%) D (standard)
TMA 9 HC1 Indole NH4SO4 Anchovymeal 2.5 2.5 2.5 2.5
0.25 0.25 0.25 0.25
40.0 40.0 40.0 40.0
52.25 47.25 47.25 57.25
Mean No. flies killed/unita 21,6 b 36.0 ab 54.7 a 27.2 b
aN = 6. Means followed by same letters are not significantlydifferent from each other at 0.05 level.
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MULLA AND RIDSDILL-SMITH
Skatole, a chemical closely related to indole, has been reported to be attractive to certain fly species and to H. collusor in particular (Hwang et al., 1976). This material at 0.25% concentration was added to the standard formulation. The standard composition lacking skatole attracted a mean number of 66.5 _+ 18.5 flies per unit compared to 9.3 + 2.0 flies per unit in the composition containing skatole. The means were significantly different from each other at 0.05 level. It is possible that the combined concentration of indole and skatole was too high or that skatole is acting as a repellent against the flies. One further test was designed to evaluate the addition of n-butyric acid and linoleic acid to the standard composition and to delete ammonium sulfate, replacing it with n-butyric acid (Table 4). Addition of both linoleic and n-butyric acid did not significantly alter response of flies from that of the standard. However, deleting ammonium sulfate and replacing it with n-butyric acid significantly decreased attractancy as compared to that of the standard. Ammonium sulfate provides bulk and improves flowability and handling of the formulation. Therefore its inclusion aside from attractancy is desirable. The IGM alone attracted very few flies. Attraction of Bush Flies and Dung Beetles to Chemicals and Dung. Bush flies and dung beetles are attracted to cattle dung. Dung beetles are employed as biological control agents to disperse cattle dung for improving pastures and to reduce bush fly production in dung pads (Ridsdill-Smith, 1981; RidsdillSmith and Matthiessen, 1984). Bush flies are highly attracted to freshly dropped cattle dung pads or older pads when the surface is scraped (Figure 1). During the course of our studies, we noted that large numbers of bush flies were attracted to fresh dung pads for a short period and dung attractancy declined in time as the pads crusted. To test and compare the attractancy of dung and chemical compositions tested, fresh dung and toxicant were tested in the experimental system designed. TABLE 4. ATTRACTION OF BUSH FLIES TO VARIOUS CHEMICAL ATTRACTANT COMPOSITIONS WITH AND WITHOUT CARBOXYLIC ACIDS (EXPERIMENT VI)
Ingredients and % composition
Treatment a A (std) B C Check (IGM)
TMA - HC1 Indole 2.5 2.5 2.5
0.25 0.25 0.25
NH4SO4 40 40
n-Butyric Linoleic Anchovy Mean No. flies acid acid meal killed/unit b
2 2
2
57.25 53.25 95.5
285.4 209.2 63.0 3.8
a a b c
aOne teaspoon of each composition and one teaspoon of IGM put in each treatment except in check when IGM alone was used. bN = 5. Means followed by same letter are not significantly different from each other at 0.01 level.
AUSTRALIAN BUSH FLY ATTRACTANTS
267
FIG. 1. Attraction of M. vetustissima to 1-day-old dung pad when its surface was scraped off. Flies are not attracted to such old pads; attraction ensues when the surface crust is removed. This indicates release of trapped volatiles in the pad covered with a hard crust. Almost five times more flies were attracted to equal amounts of the dung than the chemical attractants (Table 5). The magnitude of attractancy of dung was s!gnificantly different from the chemical attractants. Dung beetles, Onthophagus binodis Thunberg and Euoniticellus pallipes (Fabricius), prevalent during the test, were not attracted to the chemical attractants but were lured to the dung in large numbers. It thus seems that bush flies and dung beetles utilize, in part, different cues for oviposition and feeding on the dung pads. Formulations of toxic chemicals and attractant compositions, if used in bush fly control programs, would not attract and harm the beneficial beetle fauna inhabiting this resource. Relative Attraction to Chemicals and Humans and Fly Sex Ratios. Activity of bush flies is influenced by many factors, among which temperature and wind velocity play an important part. Due to variation in environmental conditions, activity patterns of bush flies are quite variable. The extent of population activity can be roughly measured by sweeping in a standard manner from human hosts (Hughes, 1970) and the numbers sampled corrected for influence of temperature. On each day where chemical attractant compositions were exposed in the field, flies coming to a human host were netted in a 20-min sample, sexed, and counted. The data in Table 6 clearly indicate a great deal of variation in the numbers of flies either attracted to chemicals or human hosts in the different
268
MULLA AND RIDSDILL-SMITH
TABLE 5. COMPARISON OF ATTRACTANCY OF STANDARD CHEMICAL COMPOSITION AND FRESH CATTLE DUNG TO AUSTRALIAN BUSH FLY M. vetustissima AND DUNG BEETLES (EXPERIMENT VIII) ~
Mean No. of insects killed/unit b Treatment A (std.) B (dung)
Bush flies
Dung beetles c
41.3 191.3
0 21
aA teaspoonful of the attractant and toxicant mixture, and a teaspoonful of dung plus toxicant were placed in each unit. bMeans significantly different for each insects group at 0.01 level CDung beetles (Scarabaeinae): Onthophagus binodis 55 %, Euoniticellus pallipes 45 %.
experiments. In general, as the extent o f attractancy to humans increased, so did attractancy to compositions o f chemical attractants. However, there were instances where trends o f attractions to the two sources did not follow the same trend. The sex ratio of bush flies coming to human hosts was more in favor o f females. The sex ratio o f female to male was 3.0 for flies attracted to human TABLE 6. SEX RATIO OF BUSH FLIES ATTRACTED TO STANDARD CHEMICAL ATTRACTANT COMPOSITIONS AND HUMAN HOSTS (TAKEN BY NETTING) MEASURED ON SAME DAY
Mean No. of flies collected Chemical compositions" Experiment I
IA II II-A III IV V VI VII VIII Total Ratio ~4-6 hr trapping. b20-min sample. CAt dung, 756:392.
Human net samplesb
Female
Male
Female
Male
23 5 508 32 293 531 239 234 1949 101 c 3914
10 5 111 23 231 306 209 221 858 147 2121
105 315 105 315 655 655 232 263 385 263 3293
16 117 16 117 204 204 77 90 168 90 1099
1.8
3.0
AUSTRALIAN BUSH FLY ATTRACTANTS
269
hosts, while the same ratio in flies attracted to the standard chemical attractant composition was 1.8. Since the sex ratio of flies emerging from pads is usually 1:1 (Hughes et al., 1972), the bias in sex ratio of flies attracted to hosts or attractant compositions is probably due to the physiological status of females requiring food for oogenic development and may vary with the age of flies, presence of other sources of protein, time of year, and composition of the bait (Hughes, 1981). Measurement of quantitative responses of bush flies to chemical attractant compositions and to natural attractants emanating from fresh dung offers some interesting possibilities for further study and development of chemical attractants in bush fly control programs. It is apparent that the attractant compositions studied here are lacking one or more chemical attractants present in the dung, and isolation, identification, and evaluation of chemical attractants from dung sources warrant further studies. Nevertheless, this is the first attempt to test and evaluate chemical attractants against the widely distributed pest fly M. vetustissima.
It should be pointed out that the behavior of M. vetustissima is quite complex and variable, and is influenced by many environmental and physiological factors. Chemicals showing very low levels of attractancy will not provide a distinct separation in the behavioral response of this fly under different conditions. Further studies are warranted to work out concentration-attractancy relationships for these chemicals under various field conditions. It is equally evident (Figure 1) that, due to the very high potency of chemical attractants emanating from dung pads, the dung pads can be used for attracting natural populations of flies and killing them on the pads with the application of suitable toxicant formulations. Addition of toxic bait to dung pads resulted in the kill of at least 1000 flies on one treated dung pad in 15-30min. In view of the low absolute density of bush flies, found to be 9000 flies/acre in one study (Norris, 1966) and less than 100 flies per acre during March in another study (Vogt et al., 1981), marked local reduction of flies could be achieved by treating dung pads in infested areas. Before such a program can be implemented, detailed studies on population dynamics and behavioral aspects of the pest are in order. Acknowledgments--The authors would like to acknowledgethe assistanceof Harold Axelrod of the Department of Entomology, University of California, Riverside, California, in the preparation of attractant compositions, and of John Matthiessen of CSIRO Division of Entomology, Perth, Westem Australia, for advice on bush fly samplingmethods.
REFERENCES CLELAND,
J.B. 1913. Insects and their relationship to disease in man in Australia. Australas. Med.
Congr. 9:548-570.
HOOPER,M. 1982. Doctor Hunger and Captain Thirst--Stories of Australian Explorers. Methuen, Perth, Australia, 144p.
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HUGHES, R.D. 1970. The seasonal distribution of bush fly (Musca vetustissima Walker) in southeast Australia. J. Anita. Ecol. 39:691-706. HuGuEs, R.D. 1977. Some problems associated with quantitative sampling of bush fly (Musca vetustissima Walker). J. Aust. Entomol. Soc. 16:117-122. HUGHES, R.D. 1981. The Australian bush fly: A climate-dominated nuisance pest of man, pp. 177191, in R.L. Kitching and R.E. Jones (eds.). The Ecology of Pests. CSIRO, Canberrra, Australia. HUGHES, R.D., GREENHAM,P. M., TYNDALE-BIscoE,M., and WALKER,J.M. 1972. A synopsis of observations on the biology of the Australian bush fly (Musca vetustissima Walker). J. Aust. Entomol. Soc. 11:311-331. HWANG, Y.-S., and MULLA,M.S. 1973. Attractants for synanthropic flies: 5. Isolation and attractangy of Hippelates eye gnat attractants. J. Econ. Entomol. 66:1339-1340. HWANG, Y.-S., MULLA, M.S., and AXELROD,H. 1975. Attmctants for synanthropic flies: Evaluation of chemical attractants and coattractants against the eye gnat Hippelates collusor (Diptera: Chloropidae). Environ. Entomol. 4:769-773. HWANG, Y.-S., MULLA, M.S., and AXELROD,H. 1976. Attractants for synanthropic flies: Identification of attractants and coattmctants for Hippelates eye gnats (Diptera: Chloropidae). J. Agric. Food Chem. 24:164-169. LEGNER,E.F., SUGARMAN,B.B., Yu, H.-S., and LUM, H. 1974. Biological and integrated control of the bush fly, Musca sorbens Wiedemann and other filth-breeding Diptera in Kwajalein attol, Marshall Islands. Bull Soc. Vector Ecol. 1:1-14. MULLA, M.S., and AXELROD,H. 1974. Attractants for synanthropic flies: Longevity of attractant and toxicant formulations evaluated against Hippelates coUusor. J. Econ. Entomol. 67:641643. MULLA, M.S., and STAINS, G.S. 1977. Eye gnats, pest and plague of mankind, "The friendly Coachella Valley salute". Proc. Papers Annu. Conf. Calif. Mosq. Vector Control Assoc. 45:205-209. MULLA,M.S., AXELROD,H., and IKnSHOJt,T. 1974. Attractants for synanthropic flies: Area-wide control of Hippelates collusor with attractive baits. J. Econ. EntomoI. 67:631-638. MULLA,M.S., HWANG,Y.-S., and AXELROD,H. 1976. Chemical attractants and their formulations against the eye gnat Hippelates collusor. Bull. Soc. Vector Ecol. 3:47-51. MULLA, M.S., HWANG, Y.-S., and AXEL~OD, H. 1977. Attractants for synanthropic flies: Chemical attractants for domestic flies. J. Econ. Entomol. 70:644-648. MULLA, M.S., AXELROD,H., and HWANG, Y.-S. 1984. Field evaluation of chemical attractants against the fly Fanniafemoralis (Diptera: Muscidae). J. Chem. Ecol. 10:349-360. NORPaS, K.R. 1966. Notes on the ecology of the bush fly, Musca vetustissima Walker (Diptera: Muscidae), in the Canberra district. Aut. J. Zool. 14:1139-1156. PdOSDILL-SMITH,T.J. 1981. Some effects of three species of dung beetles (Coleoptera: Scarabaeidae) in southwestern Australia on the survival of the bush fly Musca vetustissima Walker (Diptera: Muscidae), in dung pads. Bull. Entomol. Res. 71:425-433. RIDSDILL-SMITH,T.J., and MATTIESSEN,J.N. 1984. Field assessments of the impact of night-flying dung beetles (Coleoptera: Scarabaeidae) on the bush fly, Musea vetustissima Walker (Diptera: Muscidae) in south-western Australia. Bull. Ent. Res. 74:191-195. VOt3T, W.G., RUNKOE, S. and STAPaCK, N.T. 1985. A wind-oriented fly trap with a bait that maintains a constant level of attractiveness for quantitative sampling of adult bush fly, Musca vetustissima Walker. J. Aust. Ent. Soc. : In press. VOGT, W.G., WOODBURN,T.L., and COMMON, G.W. 1981. Estimating absolute densities of the bush fly, Musca vetustissima Walker (Diptera: Muscidae) using West Australian blowfly traps. Bull. Entomol. Res. 71:329-337.
CHEMICAL ATTRACTANTS TESTED AGAINST THE AUSTRALIAN BUSH FLY Musca vetustissima (DIPTERA: MUSCIDAE)
MIR
S. M U L L A
1 and JAMES
T.
RIDSDILL-SMITH
2
IDepartment of Entomology University of California, Riverside Riverside, California 92521 2Division of Entomology, CSIRO Private Bag P. O. Wembley, Western Australia, 6014 (Received March 21, 1985; accepted June 21, 1985)
A b s t r a c t - - A number of chemical compositions known to attract several synanthropic and pest fly species were tested against the Australian bush fly Musca vetustissima, a severe and persistent pest of man and domesticated and wild mammals. A standard composition containing small quantities of trimethylamine and indole, blended with large amounts of ammonium sulfate and anchovy meal, showed good attractancy against this fly. Deletion of ammonium sulfate did not significantly alter attractancy. Similarly, incorporation of n-butyric acid and linoleic acid into the formulation did not alter attmctancy, nor did addition of several carbonates and bicarbonates. Skatole, a closely related compound to indole, decreased attractancy significantly. In all the tests, the attmctant compositions lured greater number of females than males. The attractancy of the standard composition was studied and compared with equal amount of fresh cattle dung. Chemical attractants lured bush flies and not dung beetles, while dung attracted both flies and beetles. The dung resource was 4.6 • more attractive to flies than the standard attractant composition. This suggests that there are other chemical attractants emanating from dung that warrant isolation and identification work. Nevertheless, the chemical attractants studied here provide a starting point for further investigations. The ratio of female to male flies attracted to humans was greater than the ratio of flies attracted to chemicals. The sex ratio of flies netted from human host was 3.0, while the sex ratio of flies attracted to chemical attractants was 1.8. The preponderance of females attracted to both sources was probably due to physiological requirements of the sexes. Key W o r d s - - B u s h fly, Australian bush fly, Musca vetustissima, Diptera, Muscidae, chemical attractants.
261
0098-0331/86/0100-0261$05.00/0 9 1986PlenumPublishingCorporation
262
MULLA AND RIDSDILL-SMITH INTRODUCTION
Early explorers in Australia noted the pestiferous nature of the Australian bush fly Musca vetustissima Walker, some three centuries ago (Cleland, 1913). One of the explorers, John Lort Stokes, in 1838 related that "the flies are at you all day, crawling into your eyes, up your nostrils and down your throat" (Hooper, 1982). The nature and scope of annoyance brought about by these flies remains the same today. This pest is widely spread over most of the continent (Hughes et al., 1972) and is especially abundant in cattle-raising areas. They breed in dung, preferring freshly deposited dung of cattle (Hughes, 1981; Hughes et al., 1972; Norris, 1966). Adult bush flies are attracted to large animals, including man, females outnumbering the males attacking or landing on hosts (Hughes, 1977, 1981; Hughes et al., 1972; Norris, 1966). We have noted that the flies can find the host in a few seconds in the field and that flies at some distance orient and move upwind toward the hosts. Chemical odors emanating from hosts or breeding medium (dung) are some of the factors providing cues for directed response by the flies. However, no definitive studies on the role of chemical attractants influencing bush fly behavior have been accomplished on this important synanthropic and synzootic fly. The hovering and feeding behavior of bush flies approaching and attacking hosts is quite similar to that of the tiny flies known as eye gnats (Hippelates species, family Chloropidae) prevailing in the Westem hemisphere (Mulla and Stains, 1977). Chemical attractants inducing positive responses in female H. collusor (Townsend) were isolated and identified from putrified chicken eggs (Hwang and Mulla, 1973; Hwang et al., 1975, 1976) and field tested by Mulla and Axelrod (1974) and Mulla et al. (1974, 1976). Some of the principal chemicals attracting H. collusor were also found to manifest a high level of attractancy against the common house fly Musca domestica (Mulla et al., 1977), and the coastal lesser house fly Fannia femoralis on poultry ranches (Mulla et al., 1984). It is well known that many species of synanthropic and synzootic flies are attracted by odors emanating from putrefying proteins. Musca sorbens, a closely related species to M. vetustissima, was found to be attracted to putrefying egg and fish meat (Legner et al., 1974), and baiting of an insular population with these baits reduced populations of M. sorbens markedly. M. vetustissima has been found to be attracted to liver baits with sodium sulfide (Hughes, 1977; Norris, 1966; Vogt et al., 1981). Most recently Vogt et al. (1985) have used a mixture of putrefying liver and dung with live blow fly maggots to stir the bait for the purpose of trapping bush flies. In view of these findings and the extensive research on the isolation and identification of chemicals showing field attractancy to a variety of pest flies, the present studies were initiated. Compositions of chemical attractants with
AUSTRALIAN BUSH FLY ATTRACTANTS
263
proven attractancy to Hippelates, Musca, and Fannia species were evaluated against the Australian bush fly. A simple method for field evaluation of attractant compositions was developed, where attractancy of various chemical compositions and natural attractants emanating from fresh dung was tested against field populations of M. vetustissima.
METHODS
AND MATERIALS
Compositions of chemical attractants found attractive to the eye gnat Hippelates collusor (Hwang et al., 1975, 1976; Mulla et al., 1976), Musca domestica (Mulla et al., 1977), and Fannia femoralis (Mulla et al., 1984) consisted of mixtures of a carder (anchovy meal) with one or more of the following chemicals: trimethylamine (from trimethylamine hydrochloride), ammonia (from ammonium sulfate), indole, linoleic acid, and a few lower aliphatic acids. Against the bush fly, a composition containing anchovy meal cartier (57.25 %), trimethylamine hydrochloride (2.5 %), ammonium sulfate (40 %), and indole (0.25 %) by weight was used as a standard mix with which other compositions with fewer or more chemicals were compared for luring the flies. When additional chemicals were included or deleted, the content of carrier was varied in the composition accordingly. The composition was mixed in 1:1 ratio with Improved Golden Malrin (IGM) (Zoecon Corporation, Dallas, Texas) toxic bait. The ingredients of this bait are: 1% methomyl, 0.025 % (Z)-9-tricosene, and 98.97 % inert ingredients. Fine sand (780 g) forming a 2-cm layer was placed in a plastic bowl (18 cm diam. and 8 cm deep) and wetted with 140 ml of tapwater, bringing the moisture content initially to its field capacity level. For testing, a 5-g quantity of each mix was placed on the damp sand. In some tests, especially in testing attractancy of dung, the IGM was not mixed with the attractants, but rather it was spread on the top and around the dung or other attractant compositions. The bowls (replicated five to six times in each experiment) containing various test attractant compositions were placed on the ground 5 m apart in paddocks infested with bush flies. They were distributed in randomized-block design in a line at right angles to the wind direction or in a circular pattern. The tests were run in two areas near Perth, Western Australia. The Serpentine area, with extensive livestock pastures, is located in the coastal plain 100 km south of Perth, while Cunderdin, a predominantly wheat growing and pasturing area, is located on the Darling plateau 150 km east of Perth. Flies attracted to the compositions oriented toward the attractant source and crawled or flew into the bowls, walking and milling around the attractanttoxicant compositions. Because of the action of the quick-acting toxicant methomyl, the flies on contact with the bait died in the bowl within a few seconds after arrival. All flies killed in the bowls were counted with no loss due to wind
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or transportation into the laboratory. The compositions were exposed in the field for a period of 4-6 hr for each experiment, depending on the activity of bush flies. After termination of each experiment, the bowls were covered with lids and brought into the laboratory where the flies were identified, sexed, and counted under a dissecting microscope. Analysis of data was carried out using standard computer program design for randomized-block analysis, The data were transformed to log (n + 1) when fly numbers were high and variable. Transformed means were compared for significant differences using Duncans's multiple-range test, letter designations for denoting significant differences are shown on untransformed means for ease of interpretation. Where there were two treatments in a given test, the transformed data were analyzed by means of t test. Significant differences between or among means are expressed in the tables. RESULTS AND DISCUSSION
Since the attractant composition utilized fish meal as carrier and Improved Golden Malrin (IGM) as a toxicant, it was necessary to test the attractancy of these two components. Fish meal and IGM (1:1) and IGM alone were compared in the first test. The fish meal showed a low level of attractancy which was significantly higher than that of IGM alone (Table 1). When the standard composition plus IGM was compared with fish meal plus IGM, the standard composition caught significantly higher numbers than the carrier (Table 1, test II). In a third test, this comparison was repeated, and again the standard attracted significantly higher numbers than the carrier (Table 1, test II-A). On the basis of these data, it was concluded that a composition containing three chemicals, carrier, and IGM was more attractive than the carrier plus IGM to field popuTABLE 1. ATTRACTION OF AUSTRALIAN BUSH FLY Musca vetustissima TO CHEMICAL ATTRACTANTS AND TOXICANT BAIT, IMPROVED GOLDEN MALRIN (IGM)
Ingredients and % composition Experiment and treatment I
A B II A B (std.) II-A A B (std.)
TMA 9 HC1
Indole
NH4SO 4
Anchovy meal
Mean No. flies killed/unit~
0 0 0 2.5 0 2.5
0 0 0 0.25 0 0.25
0 0 0 40.00 0 40.00
100 0c 100 57.25 100 57.25
5.6 *b 1:0 45.5 78.4*** 18.4 39.6**
aN = 5. Significant differences by t-test for each pair in each test. b'*significant at 0.10 level;**, significant at 0.05 level;***, significant at 0.01 level. c 100% IGM toxic bait.
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265
TABLE 2. ATTRACTANCY OF VARIOUS COMPOSITIONS OF CHEMICAL AGENTS TO
AUSTRALIANBUSH FLY M. vetustissima (EXPERIMENTIV) Ingredients and % composition Treatment A B C D (std.)
TMA 9 HC1
Indole
2.5 2.5
0.25
2.5
0.25 0.25
NHaSO 4
Anchovym e a l
Mean No. flies killed/unita
40.0 40.0 40.0
97.25 57.50 59.75 57.25
26.0 ab 15.3 bc 14.5 c 30.0 a
aN = 6. Means followed by same letters not significantlydifferentfrom each other at 0.05 level. lations of the Australian bush fly, but the carrier itself did show some attractancy also. Compositions were prepared in which one of the three chemicals in the standard was deleted to see if deletion of each chemical would result in increased or decreased attractancy. The standard composition attracted significantly higher numbers than the compositions lacking trimethylamine hydrochloride or indole (Table 2). The composition lacking a m m o n i u m sulfate was not significantly different from the standard. In studies on Hippelates eye gnats and house flies, n-butyric acid was found to increase the attractancy of chemical compositions (Hwang et al., 1975; Mulla et al., 1977). W e deemed it desirable to study attractancy of this chemical, a m m o n i u m carbonate, and sodium carbonate in the standard composition (Table 3). n-Butyric acid and a m m o n i u m carbonate did not enhance attractancy. Addition of sodium carbonate, however, increased attractancy of the composition. Additionally, calcium carbonate and sodium bicarbonate were studied for enhancing attractancy, but neither one significantly increased attractancy (data not presented). TABLE 3. ATTRACTANCY OF VARIOUS COMPOSITIONS OF ATTRACTANT CHEMICALS TO AUSTRALIAN BUSH FLY M. vetustissima (EXPERIMENT V)
Ingredients and % composition Treatment E (n-butyricacid 5%) F (ammoniumcarbonate 10%) G (sodium carbonate 10%) D (standard)
TMA 9 HC1 Indole NH4SO4 Anchovymeal 2.5 2.5 2.5 2.5
0.25 0.25 0.25 0.25
40.0 40.0 40.0 40.0
52.25 47.25 47.25 57.25
Mean No. flies killed/unita 21,6 b 36.0 ab 54.7 a 27.2 b
aN = 6. Means followed by same letters are not significantlydifferent from each other at 0.05 level.
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Skatole, a chemical closely related to indole, has been reported to be attractive to certain fly species and to H. collusor in particular (Hwang et al., 1976). This material at 0.25% concentration was added to the standard formulation. The standard composition lacking skatole attracted a mean number of 66.5 _+ 18.5 flies per unit compared to 9.3 + 2.0 flies per unit in the composition containing skatole. The means were significantly different from each other at 0.05 level. It is possible that the combined concentration of indole and skatole was too high or that skatole is acting as a repellent against the flies. One further test was designed to evaluate the addition of n-butyric acid and linoleic acid to the standard composition and to delete ammonium sulfate, replacing it with n-butyric acid (Table 4). Addition of both linoleic and n-butyric acid did not significantly alter response of flies from that of the standard. However, deleting ammonium sulfate and replacing it with n-butyric acid significantly decreased attractancy as compared to that of the standard. Ammonium sulfate provides bulk and improves flowability and handling of the formulation. Therefore its inclusion aside from attractancy is desirable. The IGM alone attracted very few flies. Attraction of Bush Flies and Dung Beetles to Chemicals and Dung. Bush flies and dung beetles are attracted to cattle dung. Dung beetles are employed as biological control agents to disperse cattle dung for improving pastures and to reduce bush fly production in dung pads (Ridsdill-Smith, 1981; RidsdillSmith and Matthiessen, 1984). Bush flies are highly attracted to freshly dropped cattle dung pads or older pads when the surface is scraped (Figure 1). During the course of our studies, we noted that large numbers of bush flies were attracted to fresh dung pads for a short period and dung attractancy declined in time as the pads crusted. To test and compare the attractancy of dung and chemical compositions tested, fresh dung and toxicant were tested in the experimental system designed. TABLE 4. ATTRACTION OF BUSH FLIES TO VARIOUS CHEMICAL ATTRACTANT COMPOSITIONS WITH AND WITHOUT CARBOXYLIC ACIDS (EXPERIMENT VI)
Ingredients and % composition
Treatment a A (std) B C Check (IGM)
TMA - HC1 Indole 2.5 2.5 2.5
0.25 0.25 0.25
NH4SO4 40 40
n-Butyric Linoleic Anchovy Mean No. flies acid acid meal killed/unit b
2 2
2
57.25 53.25 95.5
285.4 209.2 63.0 3.8
a a b c
aOne teaspoon of each composition and one teaspoon of IGM put in each treatment except in check when IGM alone was used. bN = 5. Means followed by same letter are not significantly different from each other at 0.01 level.
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267
FIG. 1. Attraction of M. vetustissima to 1-day-old dung pad when its surface was scraped off. Flies are not attracted to such old pads; attraction ensues when the surface crust is removed. This indicates release of trapped volatiles in the pad covered with a hard crust. Almost five times more flies were attracted to equal amounts of the dung than the chemical attractants (Table 5). The magnitude of attractancy of dung was s!gnificantly different from the chemical attractants. Dung beetles, Onthophagus binodis Thunberg and Euoniticellus pallipes (Fabricius), prevalent during the test, were not attracted to the chemical attractants but were lured to the dung in large numbers. It thus seems that bush flies and dung beetles utilize, in part, different cues for oviposition and feeding on the dung pads. Formulations of toxic chemicals and attractant compositions, if used in bush fly control programs, would not attract and harm the beneficial beetle fauna inhabiting this resource. Relative Attraction to Chemicals and Humans and Fly Sex Ratios. Activity of bush flies is influenced by many factors, among which temperature and wind velocity play an important part. Due to variation in environmental conditions, activity patterns of bush flies are quite variable. The extent of population activity can be roughly measured by sweeping in a standard manner from human hosts (Hughes, 1970) and the numbers sampled corrected for influence of temperature. On each day where chemical attractant compositions were exposed in the field, flies coming to a human host were netted in a 20-min sample, sexed, and counted. The data in Table 6 clearly indicate a great deal of variation in the numbers of flies either attracted to chemicals or human hosts in the different
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MULLA AND RIDSDILL-SMITH
TABLE 5. COMPARISON OF ATTRACTANCY OF STANDARD CHEMICAL COMPOSITION AND FRESH CATTLE DUNG TO AUSTRALIAN BUSH FLY M. vetustissima AND DUNG BEETLES (EXPERIMENT VIII) ~
Mean No. of insects killed/unit b Treatment A (std.) B (dung)
Bush flies
Dung beetles c
41.3 191.3
0 21
aA teaspoonful of the attractant and toxicant mixture, and a teaspoonful of dung plus toxicant were placed in each unit. bMeans significantly different for each insects group at 0.01 level CDung beetles (Scarabaeinae): Onthophagus binodis 55 %, Euoniticellus pallipes 45 %.
experiments. In general, as the extent o f attractancy to humans increased, so did attractancy to compositions o f chemical attractants. However, there were instances where trends o f attractions to the two sources did not follow the same trend. The sex ratio of bush flies coming to human hosts was more in favor o f females. The sex ratio o f female to male was 3.0 for flies attracted to human TABLE 6. SEX RATIO OF BUSH FLIES ATTRACTED TO STANDARD CHEMICAL ATTRACTANT COMPOSITIONS AND HUMAN HOSTS (TAKEN BY NETTING) MEASURED ON SAME DAY
Mean No. of flies collected Chemical compositions" Experiment I
IA II II-A III IV V VI VII VIII Total Ratio ~4-6 hr trapping. b20-min sample. CAt dung, 756:392.
Human net samplesb
Female
Male
Female
Male
23 5 508 32 293 531 239 234 1949 101 c 3914
10 5 111 23 231 306 209 221 858 147 2121
105 315 105 315 655 655 232 263 385 263 3293
16 117 16 117 204 204 77 90 168 90 1099
1.8
3.0
AUSTRALIAN BUSH FLY ATTRACTANTS
269
hosts, while the same ratio in flies attracted to the standard chemical attractant composition was 1.8. Since the sex ratio of flies emerging from pads is usually 1:1 (Hughes et al., 1972), the bias in sex ratio of flies attracted to hosts or attractant compositions is probably due to the physiological status of females requiring food for oogenic development and may vary with the age of flies, presence of other sources of protein, time of year, and composition of the bait (Hughes, 1981). Measurement of quantitative responses of bush flies to chemical attractant compositions and to natural attractants emanating from fresh dung offers some interesting possibilities for further study and development of chemical attractants in bush fly control programs. It is apparent that the attractant compositions studied here are lacking one or more chemical attractants present in the dung, and isolation, identification, and evaluation of chemical attractants from dung sources warrant further studies. Nevertheless, this is the first attempt to test and evaluate chemical attractants against the widely distributed pest fly M. vetustissima.
It should be pointed out that the behavior of M. vetustissima is quite complex and variable, and is influenced by many environmental and physiological factors. Chemicals showing very low levels of attractancy will not provide a distinct separation in the behavioral response of this fly under different conditions. Further studies are warranted to work out concentration-attractancy relationships for these chemicals under various field conditions. It is equally evident (Figure 1) that, due to the very high potency of chemical attractants emanating from dung pads, the dung pads can be used for attracting natural populations of flies and killing them on the pads with the application of suitable toxicant formulations. Addition of toxic bait to dung pads resulted in the kill of at least 1000 flies on one treated dung pad in 15-30min. In view of the low absolute density of bush flies, found to be 9000 flies/acre in one study (Norris, 1966) and less than 100 flies per acre during March in another study (Vogt et al., 1981), marked local reduction of flies could be achieved by treating dung pads in infested areas. Before such a program can be implemented, detailed studies on population dynamics and behavioral aspects of the pest are in order. Acknowledgments--The authors would like to acknowledgethe assistanceof Harold Axelrod of the Department of Entomology, University of California, Riverside, California, in the preparation of attractant compositions, and of John Matthiessen of CSIRO Division of Entomology, Perth, Westem Australia, for advice on bush fly samplingmethods.
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J.B. 1913. Insects and their relationship to disease in man in Australia. Australas. Med.
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HOOPER,M. 1982. Doctor Hunger and Captain Thirst--Stories of Australian Explorers. Methuen, Perth, Australia, 144p.
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HUGHES, R.D. 1970. The seasonal distribution of bush fly (Musca vetustissima Walker) in southeast Australia. J. Anita. Ecol. 39:691-706. HuGuEs, R.D. 1977. Some problems associated with quantitative sampling of bush fly (Musca vetustissima Walker). J. Aust. Entomol. Soc. 16:117-122. HUGHES, R.D. 1981. The Australian bush fly: A climate-dominated nuisance pest of man, pp. 177191, in R.L. Kitching and R.E. Jones (eds.). The Ecology of Pests. CSIRO, Canberrra, Australia. HUGHES, R.D., GREENHAM,P. M., TYNDALE-BIscoE,M., and WALKER,J.M. 1972. A synopsis of observations on the biology of the Australian bush fly (Musca vetustissima Walker). J. Aust. Entomol. Soc. 11:311-331. HWANG, Y.-S., and MULLA,M.S. 1973. Attractants for synanthropic flies: 5. Isolation and attractangy of Hippelates eye gnat attractants. J. Econ. Entomol. 66:1339-1340. HWANG, Y.-S., MULLA, M.S., and AXELROD,H. 1975. Attmctants for synanthropic flies: Evaluation of chemical attractants and coattractants against the eye gnat Hippelates collusor (Diptera: Chloropidae). Environ. Entomol. 4:769-773. HWANG, Y.-S., MULLA, M.S., and AXELROD,H. 1976. Attractants for synanthropic flies: Identification of attractants and coattmctants for Hippelates eye gnats (Diptera: Chloropidae). J. Agric. Food Chem. 24:164-169. LEGNER,E.F., SUGARMAN,B.B., Yu, H.-S., and LUM, H. 1974. Biological and integrated control of the bush fly, Musca sorbens Wiedemann and other filth-breeding Diptera in Kwajalein attol, Marshall Islands. Bull Soc. Vector Ecol. 1:1-14. MULLA, M.S., and AXELROD,H. 1974. Attractants for synanthropic flies: Longevity of attractant and toxicant formulations evaluated against Hippelates coUusor. J. Econ. Entomol. 67:641643. MULLA, M.S., and STAINS, G.S. 1977. Eye gnats, pest and plague of mankind, "The friendly Coachella Valley salute". Proc. Papers Annu. Conf. Calif. Mosq. Vector Control Assoc. 45:205-209. MULLA,M.S., AXELROD,H., and IKnSHOJt,T. 1974. Attractants for synanthropic flies: Area-wide control of Hippelates collusor with attractive baits. J. Econ. EntomoI. 67:631-638. MULLA,M.S., HWANG,Y.-S., and AXELROD,H. 1976. Chemical attractants and their formulations against the eye gnat Hippelates collusor. Bull. Soc. Vector Ecol. 3:47-51. MULLA, M.S., HWANG, Y.-S., and AXEL~OD, H. 1977. Attractants for synanthropic flies: Chemical attractants for domestic flies. J. Econ. Entomol. 70:644-648. MULLA, M.S., AXELROD,H., and HWANG, Y.-S. 1984. Field evaluation of chemical attractants against the fly Fanniafemoralis (Diptera: Muscidae). J. Chem. Ecol. 10:349-360. NORPaS, K.R. 1966. Notes on the ecology of the bush fly, Musca vetustissima Walker (Diptera: Muscidae), in the Canberra district. Aut. J. Zool. 14:1139-1156. PdOSDILL-SMITH,T.J. 1981. Some effects of three species of dung beetles (Coleoptera: Scarabaeidae) in southwestern Australia on the survival of the bush fly Musca vetustissima Walker (Diptera: Muscidae), in dung pads. Bull. Entomol. Res. 71:425-433. RIDSDILL-SMITH,T.J., and MATTIESSEN,J.N. 1984. Field assessments of the impact of night-flying dung beetles (Coleoptera: Scarabaeidae) on the bush fly, Musea vetustissima Walker (Diptera: Muscidae) in south-western Australia. Bull. Ent. Res. 74:191-195. VOt3T, W.G., RUNKOE, S. and STAPaCK, N.T. 1985. A wind-oriented fly trap with a bait that maintains a constant level of attractiveness for quantitative sampling of adult bush fly, Musca vetustissima Walker. J. Aust. Ent. Soc. : In press. VOGT, W.G., WOODBURN,T.L., and COMMON, G.W. 1981. Estimating absolute densities of the bush fly, Musca vetustissima Walker (Diptera: Muscidae) using West Australian blowfly traps. Bull. Entomol. Res. 71:329-337.
