Acta Tropica 141 (2014) 32–36
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Response of the sand fly Phlebotomus papatasi to visual, physical and chemical attraction features in the field Günter C. Müller a, *, Jerome A. Hogsette b , Daniel L. Kline b , John C. Beier c, Edita E. Revay d, Rui-De Xue e a Department of Microbiology and Molecular Genetics, IMRIC, Kuvin Center for the Study of Infectious and Tropical Diseases, Faculty of Medicine, Hebrew University, Jerusalem 91120 Israel b United States Department of Agriculture-Agricultural Research Service-Center for Medical, Agricultural, and Veterinary Entomology, Gainesville, FL 32608, USA c Department of Epidemiology and Public Health, University of Miami Miller School of Medicine, Miami, FL 33136, USA d Bruce Rappaport Faculty of Medicine, Technion, Haifa 31096, Israel e Anastasia Mosquito Control District, 500 Old Beach Road, St. Augustine, FL 32080, USA
A R T I C L E I N F O
A B S T R A C T
Article history: Received 22 March 2013 Received in revised form 10 April 2014 Accepted 18 June 2014 Available online 27 June 2014
In this study, 27 CDC traps were modified with various attractive features and compared with a CDC trap with no light source or baits to evaluate the effects on attraction to Phlebotomus papatasi (Scopoli) north of the Dead Sea near Jericho. Attractive features included CO2, lights, colored trap bodies, heat, moisture, chemical lures and different combinations of the same. Traps were placed 20 m apart and rotated from one trap location to the next after 24 h trapping periods. The most significant attractive feature was CO2, which attracted more sand flies than any other feature evaluated. Ultraviolet light was the next most attractive feature, followed by incandescent light. When evaluated alone, black or white trap bodies, heat and moisture, all influenced trap catch but effects were greater when these attractive features were used together. The results of this study suggest that traps with CO2 and UV light could be used in batteries as control interventions if suitable CO2 sources become available. ã 2014 Elsevier B.V. All rights reserved.
Keywords: Sand flies CDC traps UV light CO2 Visual preference
1. Introduction Phlebotomine sand flies are distributed mainly in the tropics and subtropics of the Old World (Adler and Theodor, 1957). They are the major nuisance and known vectors of leishmaniases, bartonellosis (Birtles, 2001) and numerous viruses including phleboviruses, flaviviruses, orbiviruses and vesiculoviruses (Ashford, 2000, 2001; Comer and Tesh, 1991). Leishmaniasis remains a severe global public health problem with an estimated 12 million patent cases and a yearly incidence of 1.5–2 million cases. Of these, most are of the cutaneous form (Desjeux, 2001). Today, leishmaniases undoubtedly have a wider geographical distribution than before and are now being reported in areas that were previously non-endemic (Ashford, 2000; Oumeish, 1999). Increasing risk factors related to natural and man-made environmental changes are making leishmaniasis a growing public health concern for many countries (Daulaire, 1999; WHO, 1990). A major risk factor is the worldwide phenomenon of urbanization
* Corresponding author. E-mail address: [email protected] (G.C. Müller). http://dx.doi.org/10.1016/j.actatropica.2014.06.007 0001-706X/ ã 2014 Elsevier B.V. All rights reserved.
(Desjeux, 2001). Leishmaniasis is a major military concern for troops in endemic areas (Burkett et al., 2007). Phlebotomus papatasi (Scopoli) is one of the most important vectors of cutaneous leishmaniasis, and the only vector of Leishmania major Yakimimoff and Schokhornin. P. papatasi is widespread in Israel and the Middle East, commonly biting humans indoors and outdoors from sunset to sunrise. It is an important vector of the parasite in the Jordan Valley and southern Israel where large sand fly populations are found in the burrows of the rodent reservoir, the fat sand rat (Psammomys obesus Cretzschmar) (Jainini et al., 1995; Schlein et al., 1982, 1984). P. papatasi is also common in Mediterranean habitats and a nuisance species in many of the settlements and larger cities. However, in the absence of sand rats,P. papatasi does not transmit Leishmania. The constant need for blood by female sand flies for egg development is the reason for frequent contacts between vectors and hosts (Killick-Kendrick, 1999). Sticky papers and CDC type traps are the standard sampling methods for sand flies (Alexander, 2000; Killick-Kendrick, 1987) but catches without additional attractants like CO2 are often small (Kline, 2006). There is a large body of literature discussing all kinds of attractive features for biting flies but comparatively, little in this
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aspect is known about sand flies (Allan et al., 1987; Gibson and Torr, 1999). The purpose of this study was to explore optical, physical and chemical features that might lead to more effective trapping methods for monitoring and possibly controlling sand flies in the future. 2. Materials and methods 2.1. Study site The study was conducted in mid-autumn near Jericho, about 10 km north of the Dead Sea at an altitude of about 300 m below sea level. This region is an extreme desert and belongs to the Saharo-Arabian phyto-geographical zone (Danin, 1988). The annual precipitation of 50–100 mm is restricted to short winter rains. The average temperature ranges from around 20 C at the end of September through early April to more than 30 C from May through August (Ashbel, 1951). Attraction features were evaluated in a neglected date plantation where a large population of P. papatasi was associated with colonies of fat sand rats and Wagner’s gerbils (Gerbillus dasyurus) (Wagner, 1842). Zoonotic cutaneous Leishmaniasis is endemic in this area (Schlein et al., 1982, 1986) and P. papatasi is the dominant sand fly species comprising almost 100% of the catch, while other species are rare or absent (Faiman et al., 2009; Müller and Schlein, 2004; Schlein et al., 2001). During autumn when the experiment was conducted, the annual winter and spring vegetation was already dry and some scattered shrubs and semi-shrubs dominated the little remaining natural vegetation found inside the plantation. About 20% of these were Suaeda asphaltica (Boiss.), Salvia fruticosa (Forsk.), Atriplex halimus (L), Aglaia leucoclada (Boiss.): Chenopodiaceae and Prosopis farcta (Macbride): Mimosaceae. On the periphery of the oasis, some groups of Tamarix nilotica (Ehrenb.): Tamaricaceae trees and shrubs, like Alhagi graecorum (Boiss.): Papilionaceae and Salsola tetranda (Forssk.): Chenopodiaceae, were restricted to small water catchments. Flowering plants, honeydew or honeydew producing insects of any kind were not found in the area at the time of the experiment. 2.2. Experimental design The study was conducted for 10 consecutive days from late September to mid-October 2007 in the date plantation. During this time, the weather conditions were stable with clear sky during daytime and few to no clouds at night without any precipitation. Night temperatures ranged from 25–27 C in the early evening to 20 –22 C in the early morning and RH from 64 to 75%, respectively. To evaluate attraction features, CDC traps (Model 512, John Hock, Gainesville FL, USA) were operated without a lid and with their original white net bags, but were modified by adding attraction features like lights, black or white colored cardboard, heat, moisture, several chemical lures and CO2 in different combinations described in detail below. The baited traps, placed 20 m apart, were operated simultaneously and continuously (from late afternoon to early morning), along an unpaved road crossing the plantation. Traps were suspended from bamboo tripods with the top of the trap body 50 cm above the ground. Traps were rotated clockwise daily at 17:00 through the trap sites (in intervals of three positions) to eliminate areal bias. Insects drawn to the traps were collected promptly at 07:00 h to prevent degradation; the catch was stored in a freezer and processed at a later point of time. The CDC traps were powered by 6 V motorcycle batteries, which were recharged daily.
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2.3. Evaluating attraction features Altogether, we evaluated 28 features or combinations of features for their attractiveness to P. papatasi (Table 1). For a non-attractive control, we used CDC traps with no light. To test the effect of light, the original CDC light trap configuration with incandescent light (bulb model: model CM-47) was maintained. To test the effect of UV light, the incandescent bulb was removed and a small portable money checker (Tragbarer Geldschein-Prüfer mit Leuchte, model 751778–62, Conrad Munich, Germany) equipped with a 4 W, 6 V UV tube (model: F4T5 BLB) was attached horizontally 3 cm above the cylindrical body of the trap (very much like the CDC Model 1212, John Hock, Gainesville FL, USA). The UV unit was connected to a separate 6 V motorcycle battery. For color modification, if needed, the original transparent cylinders of the CDC traps were wrapped either in white or non-glossy black cardboard. Heat was generated by heat film (MDS Heating Industries Ltd., Nazareth, Israel) beneath the metal jacket of a 4 mm iron sheet that fit tightly around the trap cylinders. The modified cylinders were then wrapped with cardboard (either white or black) as mentioned above. The temperature was adjusted to 41 C on the outside surface of the cylinders, which was measured with a laser-sited infrared thermometer (model OS546, Omega Engineering Inc., Canada). Though the surface temperature of potential hosts is usually lower mosquito traps are usually utilizing heaters ranging from 40 to 45 C because of a significantly higher attraction rate of biting flies.
Table 1 Mean numbers (SE) of Phlebotomus papatasi adults (both males and females) captured in CDC traps with 28 different combinations of attractant features (n = 10). NO.
Lighta
CO2 (ml/min)
Luresb
Otherc
Mean SE
28 26 23 25 27 24 22 21 20 3 19 16 18 14 17 10 2 15 7 11 8 12 5 6 9 1 13 4
UV UV UV Incand. – – Incand. – UV UV Incand. – – – – – Incand. – – – – – – – – – – –
750 250 250 250 750 250 250 250 250 250 250 250 – – – – – – – – – – – – – – – –
– – – – – – – – – – – Kaz Oct. – BG – – – – – – – Kaz BG – – – Oct.
B, H, M B, H, M – B, H, M B, H, M B, H, M – – B, H, M – B, H, M B, H, M B, H, M B, H, M B, H, M B, M – W, H, M M W, M B B, H – – W – W, H –
1554.79 0.10a 1300.22 0.09a 696.11 0.08b 570.61 0.13b 491.32 0.16b 466.63 0.14b 278.45 0.11c 194.93 0.15c 30.62 0.13d 21.96 0.12d 12.76 0.124e 9.05 0.19ef 8.67 0.13ef 8.14 0.18ef 6.67 0.20fg 6.30 0.10fg 5.99 0.09fg 5.69 0.18fg 4.96 0.10fgh 4.16 0.13gh 3.57 0.10ghi 3.01 0.19hij 2.84 0.16hijk 1.98 0.13ijk 1.58 0.13jk 1.41 0.13jk 1.36 0.18k 1.34 0.14k
Means followed by the same letter are not significantly different [P < 0 . 05; Ryan–Einot–Gabriel–Welsch Multiple Range Test (SAS Institute, 2003)]. a Incand.: incandescent bulb supplied with trap; UV: BLB-T5/4 W fluorescent tube (peak wave length = 365 nm), Conrad Electronic SE, Hirschau, Germany. b Kaz: 2-in-1 Power Bait (1-Octen-3-ol, 6.531% AI and Lactic acid, 5.331% AI), Kaz-Inc., Southborough, MA; Oct: Octenol lure (1-Octen-3-ol, 6.531% AI), AgriSense BCS Ltd., Pontypridd, UK; BG: BG-Lure (lactic acid, 13% AI, caproic acid, 1% AI, and ammonium bicarbonate, 4% AI), Biogents, Regensburg, Germany. c B: black cardboard over cylindrical trap body; W: white cardboard over cylindrical trap body, H: heat film over cylindrical trap body; M: moisture.
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Moisture was supplied from sheets of 80 80 cm filter papers folded fan-like with their tightly folded side inserted in beakers of water (Müller and Schlein, 2006). The three following commercially available lures in slow release cartridges were used to evaluate the effects of chemical attractants: the BG-Lure (Biogents, Regensburg, Germany) is a combination of lactic acid (13% AI), caproic acid (1% AI), ammonium bicarbonate (4% AI) and fatty acids packed in a mesh dispenser; the Octenol lure (AgriSense BCS Ltd., Pontypridd, UK) contains only 1-octen-3-ol (6.531% AI); the 2-in-1 Power Bait (Kaz-Inc., Southborough, MA) is a combination of lactic acid (5.331% AI) and 1-octen-3-ol (6.531% AI). Lure cartridges were suspended and CO2 was released 10 cm below the trap cylinders in the airflow. CO2 was released from compressed CO2 tanks at rates of 250 ml or 750 ml per min. 2.4. Statistical analysis Insect collection data were subjected to the general linear models procedure (GLM) (SAS, 2003) to determine the effects of treatment (attraction features), trap location and trap day. Means were separated with the Ryan–Einot–Gabriel–Welsh multiple range test (SAS, 2003) and unless otherwise stated P = 0.05. Insect collection data were transformed with log10 (n + 1) prior to analysis but back transformed numbers are shown in text and tables. 3. Results Over 10 trapping nights, 126,769 sand flies, 79,316 females and 47,453 males, were caught by 27 modified CDC 512 traps plus the suction-only control trap (Trap 1). The only sand fly species collected was P. papatasi.
CDC traps with CO2 captured significantly more sand flies than CDC traps without CO2 despite other modifications. This resulted in significant differences in sand fly captures among these groups of traps (Table 1). The two CDC traps with UV light, CO2 released at 750 and 250 ml/min, respectively (Traps 28 and 26), plus black cardboard over the trap body (B) with the heat film (H) and moisture (M) captured significantly more sand flies than did the other 26 traps (Table 1). The above combinations of modifications increased sand fly means between 2 and 1000 X over less attractive traps including the untreated control (Trap 1). Removal of the BHM modifications from trap 26 but maintaining the UV light and a CO2 flow rate of 250 ml/min created trap 23. This trap dropped into the next lowest significance grouping along with traps using incandescent or no light, either level of CO2 plus the BHM modifications (Traps 25, 27 and 24). Sand fly means in this group were 2 X higher than the trap captures of the next lower significance group (Table 1). Removing the BHM modifications from traps 25 and 24 above but maintaining the incandescent or no light modification and the CO2 flow rate of 250 ml/min created traps 22 and 21, respectively, but dropped them into the next lowest significance grouping. Sand fly means in this group were 6X higher than trap captures of the next lower significance groups (Table 1). Traps with UV or incandescent light plus CO2 captured significantly more sand flies than traps equipped with either light source but without CO2 despite other modifications. Mean numerical differences in these instances were 20 X (Table 1). With the exception of trap 19, mean numbers of sand flies captured by traps using incandescent or no light and no CO2 dropped into single digits. When the BHM modifications were evaluated with or without the Kaz (Trap 16), Oct (Trap 18) and BG (Trap 17) lures, the mean numbers of sand flies captured increased
Table 2 Mean numbers (SE) of Phlebotomuspapatasi adults captured by sex in CDC traps with 28 different combinations of attractant features (n = 10). Mean SE No.
Lighta
CO2 (ml/min)
Luresb
Otherc
Females
Males
28 26 27 23 25 24 22 21 20 3 19 16 14 18 17 15 10 2 12 7 5 11 8 6 1 13 9
UV UV – UV Incand. – Incand. – UV UV Incand. – – – – – – Incand. – – – – – – – – –
750 250 750 250 250 250 250 250 – – – – – – – – – – – – – – – – – – –
– – – – – – – –
B, H, M B, H, M B, H, M – B, H, M B, H, M – – B, H, M B, H, M B, H, M B, H, M B, H, M B, H, M B, H, M B,H, M B, H, M B, H, M B, H, M B, H, M – w, M B – – W, H W
1913.26 0.11aX 1503.18 0.14abX 852.10 0.12bcX 813.33 0.11bc 760.55 0.16cX 701.59 0.15cX 379.80 0.10dX 308.53 0.11dX 40.47 0.14eX 22.35 0.19efX 17.23 0.18fgX 16.49 0.14fgX 14.47 0.13fgh 13.02 0.16fgh 12.35 0.14fghiX 9.18 0.18ghijX 7.47 0.14ijkly 6.86 0.13hijkX 5.78 0.14hijkX 5.28 0.16jklX 4.95 0.14klx 4.55 0.17jklm 3.47 0.16klmnX 2.81 0.18lmnx 1.96 0.18mnX 1.69 0.30nX 1.43 0.20nX
1263.45 0.13aY 1124.64 0.11abY 283.12 0.14cdY 595.76 0.11bcY 428.04 0.17cdY 310.24 0.13cdY 204.07 0.13deY 123.02 0.17eY 23.11 0.18fY 21.57 0.18fX 9.39 0.13gY 4.78 0.23ghiY 4.40 0.21ghijY 5.67 0.13ghY 3.40 0.25hijklY 3.40 0.26hijklY 5.30 0.13ghX 5.22 0.14ghX 1.38 0.25klmY 4.65 0.13ghiX 1.48 0.19jklmY 3.79 0.22ghijkX 3.68 0.14ghijkX 1.32 0.14klmY 0.97 0.18mX 1.07 0.24lmX 1.73 0.17ijklmX
Kaz Oct. BG
Kaz
BG
Means by sex in columns (lowercase) and between sex in rows (uppercase) followed by the same letter are not signficantly different [P < 0.05; Ryan–Einot–Gabriel–Welsch Multiple Range Test (SAS Institute, 2003)]. a Incand.: incandescent bulb supplied with trap; UV: BLB-T5/4W fluorescent tube (peak wave length = 365 nm), Conrad Electronic SE, Hirschau, Germany. b Kaz: 2-in-1 Power Bait (1-Octen-3-ol, 6.531% and Lactic acid, 5.331% AI), Kaz, Inc., Southborough, MA; Oct: Octenol lure (1-Octen-3-ol, 6.531% AI), AgriSense BCS Ltd., Pontypridd, UK; BG: BG-Lure (lactic acid, caproic acid and ammonia AI), Biogents, Regensburg, Germany. c B: black cardboard over cylindrical trap body; W: white cardboard over cylindrical trap body; H: heat film over cylindrical trap body; M: moisture.
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significantly compared with the control trap (Trap 1). When the three lures (Traps 4–6) were used alone, differences between their mean captures and those of the control trap (Trap 1) were no longer significant. Changing the color of the cardboard over the trap body from black (Traps 14, 10, 8, and 12) to white (Traps 15, 11, 9, and 13) significantly decreased mean numbers of sand flies captured only when the cardboard was used alone (Traps 8 and 9) or in combination with heat (Traps 12 and 13). There were no significant differences between the mean numbers of males and females captured by 10 of the traps, including the untreated control (Trap 1). The highest mean number of both sexes was captured by the trap equipped only with UV light (Trap 3) (22 males and females) (Table 2). The remaining 18 traps captured significantly more females than males (Table 2). 4. Discussion The relative attraction of sand flies to CO2 and light is clearly demonstrated by our results and several relationships between UV light, incandescent light, or no light at all, used with or without CO2, can be seen in Table 1. Traps equipped with UV light plus CO2 released at 250 ml/min (Trap 23) captured significantly more sand flies than traps equipped only with CO2 released at 250 ml/min (Trap 21). These were followed at a lower significance level by traps equipped with UV light but no CO2 (Trap 3). The same order was observed (Traps 22, 21, and 2) when incandescent light was substituted for UV light. The addition of UV light to traps equipped only with CO2 (Traps 21 and 23) significantly increased the sand fly catch but the addition of incandescent light to traps equipped only with CO2 (Traps 21 and 22) did not (Table 1). It is obvious from the results that CO2 produced major increases in captures of P. papatasi (Table 1) and was thus, a significant trapping component. When CO2 was removed from the traps, mean catches were reduced from 3–4 digits to 1– 2 digits. The largest trap catches when CO2 was not used were produced with UV light with (Trap 20) and without (Trap 3) the BHM modifications. Without either CO2 or UV light, 11 traps captured significantly more P. papatasi than the un-baited control trap, although mean captures were 13 flies (Table 1). It is well known that the use of CO2 can be essential for the trapping of mosquitoes (Kline, 2006) and the same is true for P. papatasi. Carbon dioxide seems to be a universal attractant for sand flies so our results were not unexpected. Carbon dioxide is often used in surveillance projects when large numbers of individuals are required (Müller et al., 2011). In average humans are emitting while breathing 320 ml of CO2 per min (Fox, 2008). P. papatasi is very sensitive to CO2 and uses the minute amounts released by plants to guide them to sugar-rich foliage (Schlein and Jacobson, 2008). Numerous publications have suggested that CO2 is a long-range attractant for mosquitoes (Takken, 1991). In experiments with ramp traps, mosquitoes were attracted to hosts and CO2 that were at least 13.5 m away (Gilles and Wilkes, 1974) and Kline (2006) formed a mosquito barrier with CO2-baited traps spaced 16.5 m apart. It can, therefore, be concluded that CO2 is a long-range attractant for mosquitoes. Our CO2-baited trap means ranged from 195 to 1500 individuals which were 150–1000 times higher for female and male P. papatasi, respectively, than those captured in the un-baited control trap. This suggests that CO2 is also a long-range attractant for sand flies. It is noteworthy that a 3-fold release of CO2 (from 250 ml/min to 750 ml/min) did not result in a significant increase in the numbers of sand flies trapped. This indicates the existence of an optimum CO2 concentration threshold for sand flies, above which attraction does not increase. Many dipterans, including sand flies, mosquitoes and stable flies, are attracted to UV light because they have ocular UV receptors which are photo-sensitive in the 340–360 nm range
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(Agee and Patterson, 1983; Green and Cosens, 1983; Mellor et al., 1996; Stark and Tan, 1982). However, the performance of our traps with UV, incandescent or natural light was greatly increased by the concomitant use of CO2 (Table 1). Light, especially in the UV region, has been assumed to be a strong lure for many nocturnally active flying insects. Actually, the light merely eliminates orientation and the insects, thus, deprived of integer vision, are drawn in direction of the light source (Junnila et al., 2011; Nowinszky, 2004). But based on our data, the attraction of CO2 enhances any attractive effects of the evaluated light sources. This demonstrates the importance of CO2 when it is available, and the importance of UV light for maximizing trap performance when CO2 cannot be used. The same trend is observed when trap catches are separated by sex (Table 2). Our traps were placed at 20 m intervals so they would independently attract sand flies without competitive interference from each other. The fact that some traps had lights was a concern. However, the documented range of attraction to light by sand flies is from 2–6 m depending on the experimental design and the sand fly species (Campbell-Lendrum et al., 1999; Killick-Kendrick et al., 1985; Valenta et al., 1995). Mosquitoes are visually attracted to suction traps from a distance of 15 to 20 m (Bidlingmayer and Hem, 1980), but it is doubtful that the same is possible for sand flies. In our experiments, traps with a black body (Trap 8) attracted significantly more sand flies than traps with white (Trap 9) or clear bodies (Trap 1). The evaluations of traps with black, white, or clear cylindrical bodies demonstrate that sand flies have visual preferences. The eyes of tsetse, mosquito and sand flies show similarities in anatomical structure (Mellor et al., 1996; Muir et al., 1992), but surprisingly little is known about the visual responses of sand flies (Gibson and Torr, 1999) except for their attraction to artificial UV and incandescent light. The significant response of sand flies to colored LEDs (Hoel et al., 2007; Rajinder et al., 2009) and colored chemical light sticks (Burkett et al., 2007) might indicate the possible existence of color vision but otherwise little is known on the visual ecology of sand flies (Allan et al., 1987) The addition of moisture to traps with clear (Trap 7) or white (Trap 11) bodies significantly increased the catch (Traps 1 and 9, respectively), but trap catches were not significantly affected by the addition of moisture to traps with black bodies (Traps 8 and 10, respectively) (Table 1). In experiments in arid southern Israel numerous non-flowering plants and moisture proved to be significantly attractive to sand flies and it was speculated that moisture might guide sand flies in their search for plant meals, shelter, and breeding sites (Schlein and Yuval, 1987). The addition of heat to traps with black (Traps 8 and 12) or white (Traps 9 and 13) bodies did not significantly increase the mean numbers of female sand flies captured. It was surprising that for males the combination of a black body and heat (Trap 12) did not significantly increase the catch compared to the control (Trap 1) but a black body (Trap 8) alone did. Heat was supplied by a heat film and a metal jacket covering the transparent trap body so an evaluation of this feature was only possible in conjunction with some kind of optical target. The same is true for the evaluation of moisture and heat (Table 1). Our data suggest that attraction significantly increased if heat and moisture were combined with black color. Males on the other hand did not respond to heat alone and an increased attraction to moisture was not further increased if heat was added. The cues used by biting flies to land near or on hosts are the least well understood (Gibson and Torr, 1999). When close to a host, mosquitoes use moistures and temperature gradients in association with convection currents to initiate landing (Eiras and Jepson, 1994; Takken, 1991). Little is known about this behavior in sand flies. In contrast to mosquitoes (Bernier et al., 2007; Eiras and Jepson, 1994; Knols, 1996) artificial chemical lures and natural human odor
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increased sand fly attraction little if at all (Andrade et al., 2008; Bernier et al., 2008; Bray and Hamilton, 2007; Cameron et al., 1991; Christensen and Herrer, 1980; Pinto et al., 2001; Hamilton and Ramsoondar, 1994; Rebollar-Tellez et al., 1999). Of the three lures evaluated, only the Kaz lure (Trap 5) attracted significantly more females than the control (Trap 1). Although the lures may have attracted fewer sand flies than expected, it is obvious that the lures and other attractive features worked much more effectively when used in combination with each other (Traps 16, 18, 14, and 17). In summary, CDC traps baited with CO2 and UV light attract tremendous amounts of sand flies per night. Batteries of traps with large catch bags or toxin-impregnated cloth targets baited with CO2 could be employed as barriers against P. papatasi and should over time greatly reduce the local adult populations. Without the use of CO2, this type of control trapping would not be a viable possibility. References Adler, S., Theodor, O., 1957. Transmission of disease agents by Phlebotomine sand flies. Annu. Rev. Entomol. 2, 203–226. Agee, H.R., Patterson, R.S., 1983. Spectral sensitivity of stable, face, and horn flies and behavioral responses of stable flies to visual traps (Diptera: Muscidae). Environ. Entomol. 12, 1823–1827. Alexander, B., 2000. Sampling methods for phlebotomine sand flies. Med. Vet. Entomol. 14, 109–122. Allan, S.A., Day, J.F., Edman, J.D., 1987. Visual ecology of biting flies. Ann. Rev. Entomol. 32, 297–316. Andrade, A.J., Andrade, M.R., Dias, E.S., Pinto, M.C., Eiras, A.E., 2008. Are light traps baited with kairomones effective in the capture of Lutzomyia longipalpis and Lutzomyia intermedia? An evaluation of synthetic human odor as an attractant for phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae). Mem. Inst. Oswaldo Cruz 103, 337–343. Ashbel, D., 1951. 151pp. Bio-Climatic Atlas of Israel. Meteorology Dept. of the Hebrew University, Jerusalem (Hebrew and English). Ashford, R.W., 2000. The leishmaniases as emerging and reemerging zoonoses. Int. J. Parasitol. 30, 1269–1281. Ashford, R.W., 2001. In: Service, M.W. (Ed.), Phlebotomus Fevers. The Encyclopedia of Arthropod-Transmitted Infections. CABI Publishing, Wallingford, U.K, pp. 397–401. Bernier, U.R., Kline, D.L., Allan, S.A., Barnard, D.R., 2007. Laboratory comparison of Aedes aegypti attraction to human odors and to synthetic human odor compounds and blends. J. Am. Mosq. Contr. Assoc. 23, 288–293. Bernier, U.R., Hoel, D.F., Hogsette, J., Hanafi, H.A., Kline, D., 2008. Effect of lures and trap placement on sand fly and mosquito traps. Proceedings of the Sixths International Conference on Urban Pests. OOK Press, Budapest, Hungary, pp. 171–175. Bidlingmayer, W.L., Hem, D.G., 1980. The range of visual attraction and the effect of competitive visual attractants upon mosquito (Diptera: Culicidae) flight. Bull. Ent. Res. 70, 321–342. Birtles, R.J., 2001. Carrion’s disease. In: Service, M.W. (Ed.), The Encyclopedia of Arthropod-Transmitted Infections. CABI Publishing, Wallingford, U.K, pp. 104–106. Burkett, D.A., Knight, R., Dennett, J.A., Sherwood, V., Rowton, E., Coleman, R.E., 2007. Impact of phlebotomine sand flies on U. S. military operations at Tallil Air base, Iraq: 3. Evaluation of surveillance devices for the collection of adult sand flies. J. Med. Entomol. 44, 381–384. Cameron, M.M., Amerisinghe, F.P., Lane, R.P., 1991. The field response of Sri Lanka sandflies and mosquitoes to synthetic cattle-derived attractants. Parassitologia 33, 119–126. Campbell-Lendrum, D.H., Pinto, M.C., Brandão-Filho, S.P., de Souza, A.A., Ready, P.D., Davies, C.R., 1999. Experimental comparison of anthropophily between geographically dispersed populations of Lutzomyia whitmani (Diptera Psychodidae). Med. Vet. Entomol. 13, 299–309. Christensen, H.A., Herrer, A., 1980. Panamanian Lutzomyia (Diptera: Psychodidae) host attraction profiles. J. Med. Entomol. 17, 522–528. Comer, J.A., Tesh, R.B., 1991. Phlebotomine sand flies as vectors of vesiculovirus: a review. Parasitologia 33, 143–150. Danin, A., 1988. Flora and vegetation of Israel and adjacent areas. In: Yom-Tov, Tchernov (Eds.), The Zoogeography of Israel. Junk Publishers, Dordrecht Netherlands, pp. 251–276. Daulaire, N., 1999. Globalization and health. Development 4, 22–24.
Desjeux, P., 2001. The increase in risk factors for leishmaniasis worldwide. Trans. Royal Soc. Trop. Med Hyg. 95, 239–243. Eiras, A.E., Jepson, P.C., 1994. Host location by Aedes aegypti (Diptera: Culicidae): a wind tunnel study of chemical cues. Bull. Ent. Res. 81, 151–160. Faiman, R., Cuño, R., Warburg, A., 2009. Comparative efficacy of three suction traps for collecting phlebotomine sand flies (Diptera: Psychodidae) in open habitats. J. Vector Ecol. 34, 114–118. Fox, S.I., 2008. Fundamentals of Human Physiology. McGraw-Hill Publishing Company, pp. 449. Gibson, G., Torr, S.J., 1999. Visual and olfactory responses of haematophagous Diptera to host stimuli. Med. Vet. Entomol. 13, 2–23. Gilles, M.T., Wilkes, T.J., 1974. The range of attraction of birds as baits for some West African mosquitoes (Diptera: Culicidae). Bull. Entomol. Res. 63, 573–581. Green, C.H., Cosens, D., 1983. Spectral responses of the tsetse fly, Glossina morsitans morsitans. J. Insect Physiol. 29, 795–800. Hamilton, J.G.C., Ramsoondar, T.M.C., 1994. Attraction of Lutzomyia longipalpis to human host odours. Med. Vet. Entomol. 8, 375–380. Hoel, D.F., Butler, J.F., Fawaz, A.Y., Watany, N., El-Hossary, S.S., Villinski, J., 2007. Response of phlebotomine sand flies to light-emitting diode-modified light traps in southern Egypt. J. Vetor Ecol. 32, 302–308. Junnila, A., Kline, D.L., Müller, G.C., 2011. Comparative efficacy of small commercial traps for the capture of adult Phlebotomus papatasi. J. Vector Ecol. 36, 172–178. Killick-Kendrick, R., 1987. Methods for the study of phlebotomine sand flies. In: Peters, W., Killick-Kendrick, R. (Eds.), The Leishmaniases in Biology and Medicine, vol. 1. Academic Press, London, pp. 473–497. Killick-Kendrick, R., 1999. The biology and control of phlebotomine sand flies. Clin. Dermatol. 17, 279–289. Killick-Kendrick, R., Wilkes, T.J., Alexander, J., Bray, R.S., Rioux, J.A., Bailly, M., 1985. The distance of attraction of CDC light traps to phlebotomine sandflies. Ann. Parasitol. Hum. Comp. 60, 763–767. Kline, D.L., 2006. Traps and trapping techniques for adult mosquito control. J. Am. Mosq Control Assn. 22, 490–496. Knols, G.G.J., 1996. 214pp. Odour-Mediated Host-Seeking Behavior of the AfroTropical Malaria Vector Anopheles Gambiae Giles. PhD Thesis. DUOTONE, Wagingen, The Netherlands, pp. 214. Mellor, H.E., Hamilton, J.G.C., Anderson, M., 1996. Spectral sensitivity in the eye of male and female Lutzomyia longipalpis sandflies. Med. Vet. Entomol. 10, 372–374. Muir, L.E., Thorne, M.J., Kay, D.H., 1992. Aedes aegypti (Diptera: Culicidae) vision spectral sensitivity and other perceptual parameters of the female eye. J. Med. Entomol. 29, 278–281. Müller, G.C., Schlein, Y., 2004. Nectar and honeydew feeding of Phlebotomus papatasi in a focus of Leishmania major in Neot-Hakikar oasis. J. Vector Ecol. 29, 154–158. Müller, G.C., Revay, E.E., Beier, J.C., 2011. Simplified and improved monitoring traps for sampling sand flies. J. Vector Ecol. 36, 454–457. Nowinszky, L., 2004. Nocturnal illumination and night flying insects. Appl. Ecol. Environ. Res. 2, 17–52. Oumeish, Y.O., 1999. Cutaneous Leishmaniasis: a historical perspective. Clin. Dermatol. 17, 249–254. Pinto, M.C., Campbell-Lendrum, Lozovei, D.H., Teodoro, U., 2001. Phlebotomine sandfly responses to carbon dioxide and human odour in the field. Med. Vet. Entomol. 15, 132–139. Rajinder, S.M., Kaufmann, P.E., Butler, J.F., 2009. Lutzomyia spp. (Diptera: Psychodidae) response to olfactory attractant- and light emitting diodemodified Mosquito Magnet X (MM-X) traps. J. Med. Entomol. 46, 1052–1061. Rebollar-Tellez, E.A., Hamilton, J.G.C., Ward, D., 1999. Response of female Lutzomyia longipalpis to host odour kairomones from human skin. Physiol Entomol. 24, 220–226. Institute, S.A.S., 2003. SAS version 9 .1 for Windows. SAS Institute, Cary, NC, pp. 9. Schlein, Y., Jacobson, R.L., 2008. High nocturnal CO2 emanation guides the sand fly Phlebotomus papatasi to sugar-rich plants. Physiol. Entomol. 33, 353–359. Schlein, Y., Yuval, B., 1987. Leishmaniasis in the Jordan Valley IV. Attraction of Phlebotomus papatasi (Diptera: Psychodidae) to plants in the field. J. Med. Entomol. 24, 87–90. Schlein, Y., Jacobson, R.L., Müller, G.C., 2001. Sand fly feeding on noxious plants: a potential method for the control of leishmaniasis. Am. J. Trop. Med. Hyg. 65, 300–303. Stark, W.S., Tan, K.E.W.P., 1982. Ultraviolet light photosensitivity and other effects on the visual system. Photochem. Photobiol. 36, 371–380. Takken, W., 1991. The role of olfaction in host seeking of mosquitoes: a review. Int. J. Trop. Insect Sci. 12, 287–295. Valenta, Tang, J., Anez, N., 1995. A new method to determine the distance at which phlebotomine sand flies are attracted to light under field conditions. Merida VenezuelaProc. II Intern. Symp. Phlebot. Sandflies (ISOPS II), 35. , pp. 353–358 Boletin de la Direccion de Malariologia y Saneamiento Ambiental. WHO, 1990. Control of the leishmaniases. Report of a WHO Expert Committee. Tech. Rep. Ser (WHO, Geneva).
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Response of the sand fly Phlebotomus papatasi to visual, physical and chemical attraction features in the field Günter C. Müller a, *, Jerome A. Hogsette b , Daniel L. Kline b , John C. Beier c, Edita E. Revay d, Rui-De Xue e a Department of Microbiology and Molecular Genetics, IMRIC, Kuvin Center for the Study of Infectious and Tropical Diseases, Faculty of Medicine, Hebrew University, Jerusalem 91120 Israel b United States Department of Agriculture-Agricultural Research Service-Center for Medical, Agricultural, and Veterinary Entomology, Gainesville, FL 32608, USA c Department of Epidemiology and Public Health, University of Miami Miller School of Medicine, Miami, FL 33136, USA d Bruce Rappaport Faculty of Medicine, Technion, Haifa 31096, Israel e Anastasia Mosquito Control District, 500 Old Beach Road, St. Augustine, FL 32080, USA
A R T I C L E I N F O
A B S T R A C T
Article history: Received 22 March 2013 Received in revised form 10 April 2014 Accepted 18 June 2014 Available online 27 June 2014
In this study, 27 CDC traps were modified with various attractive features and compared with a CDC trap with no light source or baits to evaluate the effects on attraction to Phlebotomus papatasi (Scopoli) north of the Dead Sea near Jericho. Attractive features included CO2, lights, colored trap bodies, heat, moisture, chemical lures and different combinations of the same. Traps were placed 20 m apart and rotated from one trap location to the next after 24 h trapping periods. The most significant attractive feature was CO2, which attracted more sand flies than any other feature evaluated. Ultraviolet light was the next most attractive feature, followed by incandescent light. When evaluated alone, black or white trap bodies, heat and moisture, all influenced trap catch but effects were greater when these attractive features were used together. The results of this study suggest that traps with CO2 and UV light could be used in batteries as control interventions if suitable CO2 sources become available. ã 2014 Elsevier B.V. All rights reserved.
Keywords: Sand flies CDC traps UV light CO2 Visual preference
1. Introduction Phlebotomine sand flies are distributed mainly in the tropics and subtropics of the Old World (Adler and Theodor, 1957). They are the major nuisance and known vectors of leishmaniases, bartonellosis (Birtles, 2001) and numerous viruses including phleboviruses, flaviviruses, orbiviruses and vesiculoviruses (Ashford, 2000, 2001; Comer and Tesh, 1991). Leishmaniasis remains a severe global public health problem with an estimated 12 million patent cases and a yearly incidence of 1.5–2 million cases. Of these, most are of the cutaneous form (Desjeux, 2001). Today, leishmaniases undoubtedly have a wider geographical distribution than before and are now being reported in areas that were previously non-endemic (Ashford, 2000; Oumeish, 1999). Increasing risk factors related to natural and man-made environmental changes are making leishmaniasis a growing public health concern for many countries (Daulaire, 1999; WHO, 1990). A major risk factor is the worldwide phenomenon of urbanization
* Corresponding author. E-mail address: [email protected] (G.C. Müller). http://dx.doi.org/10.1016/j.actatropica.2014.06.007 0001-706X/ ã 2014 Elsevier B.V. All rights reserved.
(Desjeux, 2001). Leishmaniasis is a major military concern for troops in endemic areas (Burkett et al., 2007). Phlebotomus papatasi (Scopoli) is one of the most important vectors of cutaneous leishmaniasis, and the only vector of Leishmania major Yakimimoff and Schokhornin. P. papatasi is widespread in Israel and the Middle East, commonly biting humans indoors and outdoors from sunset to sunrise. It is an important vector of the parasite in the Jordan Valley and southern Israel where large sand fly populations are found in the burrows of the rodent reservoir, the fat sand rat (Psammomys obesus Cretzschmar) (Jainini et al., 1995; Schlein et al., 1982, 1984). P. papatasi is also common in Mediterranean habitats and a nuisance species in many of the settlements and larger cities. However, in the absence of sand rats,P. papatasi does not transmit Leishmania. The constant need for blood by female sand flies for egg development is the reason for frequent contacts between vectors and hosts (Killick-Kendrick, 1999). Sticky papers and CDC type traps are the standard sampling methods for sand flies (Alexander, 2000; Killick-Kendrick, 1987) but catches without additional attractants like CO2 are often small (Kline, 2006). There is a large body of literature discussing all kinds of attractive features for biting flies but comparatively, little in this
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aspect is known about sand flies (Allan et al., 1987; Gibson and Torr, 1999). The purpose of this study was to explore optical, physical and chemical features that might lead to more effective trapping methods for monitoring and possibly controlling sand flies in the future. 2. Materials and methods 2.1. Study site The study was conducted in mid-autumn near Jericho, about 10 km north of the Dead Sea at an altitude of about 300 m below sea level. This region is an extreme desert and belongs to the Saharo-Arabian phyto-geographical zone (Danin, 1988). The annual precipitation of 50–100 mm is restricted to short winter rains. The average temperature ranges from around 20 C at the end of September through early April to more than 30 C from May through August (Ashbel, 1951). Attraction features were evaluated in a neglected date plantation where a large population of P. papatasi was associated with colonies of fat sand rats and Wagner’s gerbils (Gerbillus dasyurus) (Wagner, 1842). Zoonotic cutaneous Leishmaniasis is endemic in this area (Schlein et al., 1982, 1986) and P. papatasi is the dominant sand fly species comprising almost 100% of the catch, while other species are rare or absent (Faiman et al., 2009; Müller and Schlein, 2004; Schlein et al., 2001). During autumn when the experiment was conducted, the annual winter and spring vegetation was already dry and some scattered shrubs and semi-shrubs dominated the little remaining natural vegetation found inside the plantation. About 20% of these were Suaeda asphaltica (Boiss.), Salvia fruticosa (Forsk.), Atriplex halimus (L), Aglaia leucoclada (Boiss.): Chenopodiaceae and Prosopis farcta (Macbride): Mimosaceae. On the periphery of the oasis, some groups of Tamarix nilotica (Ehrenb.): Tamaricaceae trees and shrubs, like Alhagi graecorum (Boiss.): Papilionaceae and Salsola tetranda (Forssk.): Chenopodiaceae, were restricted to small water catchments. Flowering plants, honeydew or honeydew producing insects of any kind were not found in the area at the time of the experiment. 2.2. Experimental design The study was conducted for 10 consecutive days from late September to mid-October 2007 in the date plantation. During this time, the weather conditions were stable with clear sky during daytime and few to no clouds at night without any precipitation. Night temperatures ranged from 25–27 C in the early evening to 20 –22 C in the early morning and RH from 64 to 75%, respectively. To evaluate attraction features, CDC traps (Model 512, John Hock, Gainesville FL, USA) were operated without a lid and with their original white net bags, but were modified by adding attraction features like lights, black or white colored cardboard, heat, moisture, several chemical lures and CO2 in different combinations described in detail below. The baited traps, placed 20 m apart, were operated simultaneously and continuously (from late afternoon to early morning), along an unpaved road crossing the plantation. Traps were suspended from bamboo tripods with the top of the trap body 50 cm above the ground. Traps were rotated clockwise daily at 17:00 through the trap sites (in intervals of three positions) to eliminate areal bias. Insects drawn to the traps were collected promptly at 07:00 h to prevent degradation; the catch was stored in a freezer and processed at a later point of time. The CDC traps were powered by 6 V motorcycle batteries, which were recharged daily.
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2.3. Evaluating attraction features Altogether, we evaluated 28 features or combinations of features for their attractiveness to P. papatasi (Table 1). For a non-attractive control, we used CDC traps with no light. To test the effect of light, the original CDC light trap configuration with incandescent light (bulb model: model CM-47) was maintained. To test the effect of UV light, the incandescent bulb was removed and a small portable money checker (Tragbarer Geldschein-Prüfer mit Leuchte, model 751778–62, Conrad Munich, Germany) equipped with a 4 W, 6 V UV tube (model: F4T5 BLB) was attached horizontally 3 cm above the cylindrical body of the trap (very much like the CDC Model 1212, John Hock, Gainesville FL, USA). The UV unit was connected to a separate 6 V motorcycle battery. For color modification, if needed, the original transparent cylinders of the CDC traps were wrapped either in white or non-glossy black cardboard. Heat was generated by heat film (MDS Heating Industries Ltd., Nazareth, Israel) beneath the metal jacket of a 4 mm iron sheet that fit tightly around the trap cylinders. The modified cylinders were then wrapped with cardboard (either white or black) as mentioned above. The temperature was adjusted to 41 C on the outside surface of the cylinders, which was measured with a laser-sited infrared thermometer (model OS546, Omega Engineering Inc., Canada). Though the surface temperature of potential hosts is usually lower mosquito traps are usually utilizing heaters ranging from 40 to 45 C because of a significantly higher attraction rate of biting flies.
Table 1 Mean numbers (SE) of Phlebotomus papatasi adults (both males and females) captured in CDC traps with 28 different combinations of attractant features (n = 10). NO.
Lighta
CO2 (ml/min)
Luresb
Otherc
Mean SE
28 26 23 25 27 24 22 21 20 3 19 16 18 14 17 10 2 15 7 11 8 12 5 6 9 1 13 4
UV UV UV Incand. – – Incand. – UV UV Incand. – – – – – Incand. – – – – – – – – – – –
750 250 250 250 750 250 250 250 250 250 250 250 – – – – – – – – – – – – – – – –
– – – – – – – – – – – Kaz Oct. – BG – – – – – – – Kaz BG – – – Oct.
B, H, M B, H, M – B, H, M B, H, M B, H, M – – B, H, M – B, H, M B, H, M B, H, M B, H, M B, H, M B, M – W, H, M M W, M B B, H – – W – W, H –
1554.79 0.10a 1300.22 0.09a 696.11 0.08b 570.61 0.13b 491.32 0.16b 466.63 0.14b 278.45 0.11c 194.93 0.15c 30.62 0.13d 21.96 0.12d 12.76 0.124e 9.05 0.19ef 8.67 0.13ef 8.14 0.18ef 6.67 0.20fg 6.30 0.10fg 5.99 0.09fg 5.69 0.18fg 4.96 0.10fgh 4.16 0.13gh 3.57 0.10ghi 3.01 0.19hij 2.84 0.16hijk 1.98 0.13ijk 1.58 0.13jk 1.41 0.13jk 1.36 0.18k 1.34 0.14k
Means followed by the same letter are not significantly different [P < 0 . 05; Ryan–Einot–Gabriel–Welsch Multiple Range Test (SAS Institute, 2003)]. a Incand.: incandescent bulb supplied with trap; UV: BLB-T5/4 W fluorescent tube (peak wave length = 365 nm), Conrad Electronic SE, Hirschau, Germany. b Kaz: 2-in-1 Power Bait (1-Octen-3-ol, 6.531% AI and Lactic acid, 5.331% AI), Kaz-Inc., Southborough, MA; Oct: Octenol lure (1-Octen-3-ol, 6.531% AI), AgriSense BCS Ltd., Pontypridd, UK; BG: BG-Lure (lactic acid, 13% AI, caproic acid, 1% AI, and ammonium bicarbonate, 4% AI), Biogents, Regensburg, Germany. c B: black cardboard over cylindrical trap body; W: white cardboard over cylindrical trap body, H: heat film over cylindrical trap body; M: moisture.
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Moisture was supplied from sheets of 80 80 cm filter papers folded fan-like with their tightly folded side inserted in beakers of water (Müller and Schlein, 2006). The three following commercially available lures in slow release cartridges were used to evaluate the effects of chemical attractants: the BG-Lure (Biogents, Regensburg, Germany) is a combination of lactic acid (13% AI), caproic acid (1% AI), ammonium bicarbonate (4% AI) and fatty acids packed in a mesh dispenser; the Octenol lure (AgriSense BCS Ltd., Pontypridd, UK) contains only 1-octen-3-ol (6.531% AI); the 2-in-1 Power Bait (Kaz-Inc., Southborough, MA) is a combination of lactic acid (5.331% AI) and 1-octen-3-ol (6.531% AI). Lure cartridges were suspended and CO2 was released 10 cm below the trap cylinders in the airflow. CO2 was released from compressed CO2 tanks at rates of 250 ml or 750 ml per min. 2.4. Statistical analysis Insect collection data were subjected to the general linear models procedure (GLM) (SAS, 2003) to determine the effects of treatment (attraction features), trap location and trap day. Means were separated with the Ryan–Einot–Gabriel–Welsh multiple range test (SAS, 2003) and unless otherwise stated P = 0.05. Insect collection data were transformed with log10 (n + 1) prior to analysis but back transformed numbers are shown in text and tables. 3. Results Over 10 trapping nights, 126,769 sand flies, 79,316 females and 47,453 males, were caught by 27 modified CDC 512 traps plus the suction-only control trap (Trap 1). The only sand fly species collected was P. papatasi.
CDC traps with CO2 captured significantly more sand flies than CDC traps without CO2 despite other modifications. This resulted in significant differences in sand fly captures among these groups of traps (Table 1). The two CDC traps with UV light, CO2 released at 750 and 250 ml/min, respectively (Traps 28 and 26), plus black cardboard over the trap body (B) with the heat film (H) and moisture (M) captured significantly more sand flies than did the other 26 traps (Table 1). The above combinations of modifications increased sand fly means between 2 and 1000 X over less attractive traps including the untreated control (Trap 1). Removal of the BHM modifications from trap 26 but maintaining the UV light and a CO2 flow rate of 250 ml/min created trap 23. This trap dropped into the next lowest significance grouping along with traps using incandescent or no light, either level of CO2 plus the BHM modifications (Traps 25, 27 and 24). Sand fly means in this group were 2 X higher than the trap captures of the next lower significance group (Table 1). Removing the BHM modifications from traps 25 and 24 above but maintaining the incandescent or no light modification and the CO2 flow rate of 250 ml/min created traps 22 and 21, respectively, but dropped them into the next lowest significance grouping. Sand fly means in this group were 6X higher than trap captures of the next lower significance groups (Table 1). Traps with UV or incandescent light plus CO2 captured significantly more sand flies than traps equipped with either light source but without CO2 despite other modifications. Mean numerical differences in these instances were 20 X (Table 1). With the exception of trap 19, mean numbers of sand flies captured by traps using incandescent or no light and no CO2 dropped into single digits. When the BHM modifications were evaluated with or without the Kaz (Trap 16), Oct (Trap 18) and BG (Trap 17) lures, the mean numbers of sand flies captured increased
Table 2 Mean numbers (SE) of Phlebotomuspapatasi adults captured by sex in CDC traps with 28 different combinations of attractant features (n = 10). Mean SE No.
Lighta
CO2 (ml/min)
Luresb
Otherc
Females
Males
28 26 27 23 25 24 22 21 20 3 19 16 14 18 17 15 10 2 12 7 5 11 8 6 1 13 9
UV UV – UV Incand. – Incand. – UV UV Incand. – – – – – – Incand. – – – – – – – – –
750 250 750 250 250 250 250 250 – – – – – – – – – – – – – – – – – – –
– – – – – – – –
B, H, M B, H, M B, H, M – B, H, M B, H, M – – B, H, M B, H, M B, H, M B, H, M B, H, M B, H, M B, H, M B,H, M B, H, M B, H, M B, H, M B, H, M – w, M B – – W, H W
1913.26 0.11aX 1503.18 0.14abX 852.10 0.12bcX 813.33 0.11bc 760.55 0.16cX 701.59 0.15cX 379.80 0.10dX 308.53 0.11dX 40.47 0.14eX 22.35 0.19efX 17.23 0.18fgX 16.49 0.14fgX 14.47 0.13fgh 13.02 0.16fgh 12.35 0.14fghiX 9.18 0.18ghijX 7.47 0.14ijkly 6.86 0.13hijkX 5.78 0.14hijkX 5.28 0.16jklX 4.95 0.14klx 4.55 0.17jklm 3.47 0.16klmnX 2.81 0.18lmnx 1.96 0.18mnX 1.69 0.30nX 1.43 0.20nX
1263.45 0.13aY 1124.64 0.11abY 283.12 0.14cdY 595.76 0.11bcY 428.04 0.17cdY 310.24 0.13cdY 204.07 0.13deY 123.02 0.17eY 23.11 0.18fY 21.57 0.18fX 9.39 0.13gY 4.78 0.23ghiY 4.40 0.21ghijY 5.67 0.13ghY 3.40 0.25hijklY 3.40 0.26hijklY 5.30 0.13ghX 5.22 0.14ghX 1.38 0.25klmY 4.65 0.13ghiX 1.48 0.19jklmY 3.79 0.22ghijkX 3.68 0.14ghijkX 1.32 0.14klmY 0.97 0.18mX 1.07 0.24lmX 1.73 0.17ijklmX
Kaz Oct. BG
Kaz
BG
Means by sex in columns (lowercase) and between sex in rows (uppercase) followed by the same letter are not signficantly different [P < 0.05; Ryan–Einot–Gabriel–Welsch Multiple Range Test (SAS Institute, 2003)]. a Incand.: incandescent bulb supplied with trap; UV: BLB-T5/4W fluorescent tube (peak wave length = 365 nm), Conrad Electronic SE, Hirschau, Germany. b Kaz: 2-in-1 Power Bait (1-Octen-3-ol, 6.531% and Lactic acid, 5.331% AI), Kaz, Inc., Southborough, MA; Oct: Octenol lure (1-Octen-3-ol, 6.531% AI), AgriSense BCS Ltd., Pontypridd, UK; BG: BG-Lure (lactic acid, caproic acid and ammonia AI), Biogents, Regensburg, Germany. c B: black cardboard over cylindrical trap body; W: white cardboard over cylindrical trap body; H: heat film over cylindrical trap body; M: moisture.
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significantly compared with the control trap (Trap 1). When the three lures (Traps 4–6) were used alone, differences between their mean captures and those of the control trap (Trap 1) were no longer significant. Changing the color of the cardboard over the trap body from black (Traps 14, 10, 8, and 12) to white (Traps 15, 11, 9, and 13) significantly decreased mean numbers of sand flies captured only when the cardboard was used alone (Traps 8 and 9) or in combination with heat (Traps 12 and 13). There were no significant differences between the mean numbers of males and females captured by 10 of the traps, including the untreated control (Trap 1). The highest mean number of both sexes was captured by the trap equipped only with UV light (Trap 3) (22 males and females) (Table 2). The remaining 18 traps captured significantly more females than males (Table 2). 4. Discussion The relative attraction of sand flies to CO2 and light is clearly demonstrated by our results and several relationships between UV light, incandescent light, or no light at all, used with or without CO2, can be seen in Table 1. Traps equipped with UV light plus CO2 released at 250 ml/min (Trap 23) captured significantly more sand flies than traps equipped only with CO2 released at 250 ml/min (Trap 21). These were followed at a lower significance level by traps equipped with UV light but no CO2 (Trap 3). The same order was observed (Traps 22, 21, and 2) when incandescent light was substituted for UV light. The addition of UV light to traps equipped only with CO2 (Traps 21 and 23) significantly increased the sand fly catch but the addition of incandescent light to traps equipped only with CO2 (Traps 21 and 22) did not (Table 1). It is obvious from the results that CO2 produced major increases in captures of P. papatasi (Table 1) and was thus, a significant trapping component. When CO2 was removed from the traps, mean catches were reduced from 3–4 digits to 1– 2 digits. The largest trap catches when CO2 was not used were produced with UV light with (Trap 20) and without (Trap 3) the BHM modifications. Without either CO2 or UV light, 11 traps captured significantly more P. papatasi than the un-baited control trap, although mean captures were 13 flies (Table 1). It is well known that the use of CO2 can be essential for the trapping of mosquitoes (Kline, 2006) and the same is true for P. papatasi. Carbon dioxide seems to be a universal attractant for sand flies so our results were not unexpected. Carbon dioxide is often used in surveillance projects when large numbers of individuals are required (Müller et al., 2011). In average humans are emitting while breathing 320 ml of CO2 per min (Fox, 2008). P. papatasi is very sensitive to CO2 and uses the minute amounts released by plants to guide them to sugar-rich foliage (Schlein and Jacobson, 2008). Numerous publications have suggested that CO2 is a long-range attractant for mosquitoes (Takken, 1991). In experiments with ramp traps, mosquitoes were attracted to hosts and CO2 that were at least 13.5 m away (Gilles and Wilkes, 1974) and Kline (2006) formed a mosquito barrier with CO2-baited traps spaced 16.5 m apart. It can, therefore, be concluded that CO2 is a long-range attractant for mosquitoes. Our CO2-baited trap means ranged from 195 to 1500 individuals which were 150–1000 times higher for female and male P. papatasi, respectively, than those captured in the un-baited control trap. This suggests that CO2 is also a long-range attractant for sand flies. It is noteworthy that a 3-fold release of CO2 (from 250 ml/min to 750 ml/min) did not result in a significant increase in the numbers of sand flies trapped. This indicates the existence of an optimum CO2 concentration threshold for sand flies, above which attraction does not increase. Many dipterans, including sand flies, mosquitoes and stable flies, are attracted to UV light because they have ocular UV receptors which are photo-sensitive in the 340–360 nm range
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(Agee and Patterson, 1983; Green and Cosens, 1983; Mellor et al., 1996; Stark and Tan, 1982). However, the performance of our traps with UV, incandescent or natural light was greatly increased by the concomitant use of CO2 (Table 1). Light, especially in the UV region, has been assumed to be a strong lure for many nocturnally active flying insects. Actually, the light merely eliminates orientation and the insects, thus, deprived of integer vision, are drawn in direction of the light source (Junnila et al., 2011; Nowinszky, 2004). But based on our data, the attraction of CO2 enhances any attractive effects of the evaluated light sources. This demonstrates the importance of CO2 when it is available, and the importance of UV light for maximizing trap performance when CO2 cannot be used. The same trend is observed when trap catches are separated by sex (Table 2). Our traps were placed at 20 m intervals so they would independently attract sand flies without competitive interference from each other. The fact that some traps had lights was a concern. However, the documented range of attraction to light by sand flies is from 2–6 m depending on the experimental design and the sand fly species (Campbell-Lendrum et al., 1999; Killick-Kendrick et al., 1985; Valenta et al., 1995). Mosquitoes are visually attracted to suction traps from a distance of 15 to 20 m (Bidlingmayer and Hem, 1980), but it is doubtful that the same is possible for sand flies. In our experiments, traps with a black body (Trap 8) attracted significantly more sand flies than traps with white (Trap 9) or clear bodies (Trap 1). The evaluations of traps with black, white, or clear cylindrical bodies demonstrate that sand flies have visual preferences. The eyes of tsetse, mosquito and sand flies show similarities in anatomical structure (Mellor et al., 1996; Muir et al., 1992), but surprisingly little is known about the visual responses of sand flies (Gibson and Torr, 1999) except for their attraction to artificial UV and incandescent light. The significant response of sand flies to colored LEDs (Hoel et al., 2007; Rajinder et al., 2009) and colored chemical light sticks (Burkett et al., 2007) might indicate the possible existence of color vision but otherwise little is known on the visual ecology of sand flies (Allan et al., 1987) The addition of moisture to traps with clear (Trap 7) or white (Trap 11) bodies significantly increased the catch (Traps 1 and 9, respectively), but trap catches were not significantly affected by the addition of moisture to traps with black bodies (Traps 8 and 10, respectively) (Table 1). In experiments in arid southern Israel numerous non-flowering plants and moisture proved to be significantly attractive to sand flies and it was speculated that moisture might guide sand flies in their search for plant meals, shelter, and breeding sites (Schlein and Yuval, 1987). The addition of heat to traps with black (Traps 8 and 12) or white (Traps 9 and 13) bodies did not significantly increase the mean numbers of female sand flies captured. It was surprising that for males the combination of a black body and heat (Trap 12) did not significantly increase the catch compared to the control (Trap 1) but a black body (Trap 8) alone did. Heat was supplied by a heat film and a metal jacket covering the transparent trap body so an evaluation of this feature was only possible in conjunction with some kind of optical target. The same is true for the evaluation of moisture and heat (Table 1). Our data suggest that attraction significantly increased if heat and moisture were combined with black color. Males on the other hand did not respond to heat alone and an increased attraction to moisture was not further increased if heat was added. The cues used by biting flies to land near or on hosts are the least well understood (Gibson and Torr, 1999). When close to a host, mosquitoes use moistures and temperature gradients in association with convection currents to initiate landing (Eiras and Jepson, 1994; Takken, 1991). Little is known about this behavior in sand flies. In contrast to mosquitoes (Bernier et al., 2007; Eiras and Jepson, 1994; Knols, 1996) artificial chemical lures and natural human odor
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increased sand fly attraction little if at all (Andrade et al., 2008; Bernier et al., 2008; Bray and Hamilton, 2007; Cameron et al., 1991; Christensen and Herrer, 1980; Pinto et al., 2001; Hamilton and Ramsoondar, 1994; Rebollar-Tellez et al., 1999). Of the three lures evaluated, only the Kaz lure (Trap 5) attracted significantly more females than the control (Trap 1). Although the lures may have attracted fewer sand flies than expected, it is obvious that the lures and other attractive features worked much more effectively when used in combination with each other (Traps 16, 18, 14, and 17). In summary, CDC traps baited with CO2 and UV light attract tremendous amounts of sand flies per night. Batteries of traps with large catch bags or toxin-impregnated cloth targets baited with CO2 could be employed as barriers against P. papatasi and should over time greatly reduce the local adult populations. Without the use of CO2, this type of control trapping would not be a viable possibility. References Adler, S., Theodor, O., 1957. Transmission of disease agents by Phlebotomine sand flies. Annu. Rev. Entomol. 2, 203–226. Agee, H.R., Patterson, R.S., 1983. Spectral sensitivity of stable, face, and horn flies and behavioral responses of stable flies to visual traps (Diptera: Muscidae). Environ. Entomol. 12, 1823–1827. Alexander, B., 2000. Sampling methods for phlebotomine sand flies. Med. Vet. Entomol. 14, 109–122. Allan, S.A., Day, J.F., Edman, J.D., 1987. Visual ecology of biting flies. Ann. Rev. Entomol. 32, 297–316. Andrade, A.J., Andrade, M.R., Dias, E.S., Pinto, M.C., Eiras, A.E., 2008. Are light traps baited with kairomones effective in the capture of Lutzomyia longipalpis and Lutzomyia intermedia? An evaluation of synthetic human odor as an attractant for phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae). Mem. Inst. Oswaldo Cruz 103, 337–343. Ashbel, D., 1951. 151pp. Bio-Climatic Atlas of Israel. Meteorology Dept. of the Hebrew University, Jerusalem (Hebrew and English). Ashford, R.W., 2000. The leishmaniases as emerging and reemerging zoonoses. Int. J. Parasitol. 30, 1269–1281. Ashford, R.W., 2001. In: Service, M.W. (Ed.), Phlebotomus Fevers. The Encyclopedia of Arthropod-Transmitted Infections. CABI Publishing, Wallingford, U.K, pp. 397–401. Bernier, U.R., Kline, D.L., Allan, S.A., Barnard, D.R., 2007. Laboratory comparison of Aedes aegypti attraction to human odors and to synthetic human odor compounds and blends. J. Am. Mosq. Contr. Assoc. 23, 288–293. Bernier, U.R., Hoel, D.F., Hogsette, J., Hanafi, H.A., Kline, D., 2008. Effect of lures and trap placement on sand fly and mosquito traps. Proceedings of the Sixths International Conference on Urban Pests. OOK Press, Budapest, Hungary, pp. 171–175. Bidlingmayer, W.L., Hem, D.G., 1980. The range of visual attraction and the effect of competitive visual attractants upon mosquito (Diptera: Culicidae) flight. Bull. Ent. Res. 70, 321–342. Birtles, R.J., 2001. Carrion’s disease. In: Service, M.W. (Ed.), The Encyclopedia of Arthropod-Transmitted Infections. CABI Publishing, Wallingford, U.K, pp. 104–106. Burkett, D.A., Knight, R., Dennett, J.A., Sherwood, V., Rowton, E., Coleman, R.E., 2007. Impact of phlebotomine sand flies on U. S. military operations at Tallil Air base, Iraq: 3. Evaluation of surveillance devices for the collection of adult sand flies. J. Med. Entomol. 44, 381–384. Cameron, M.M., Amerisinghe, F.P., Lane, R.P., 1991. The field response of Sri Lanka sandflies and mosquitoes to synthetic cattle-derived attractants. Parassitologia 33, 119–126. Campbell-Lendrum, D.H., Pinto, M.C., Brandão-Filho, S.P., de Souza, A.A., Ready, P.D., Davies, C.R., 1999. Experimental comparison of anthropophily between geographically dispersed populations of Lutzomyia whitmani (Diptera Psychodidae). Med. Vet. Entomol. 13, 299–309. Christensen, H.A., Herrer, A., 1980. Panamanian Lutzomyia (Diptera: Psychodidae) host attraction profiles. J. Med. Entomol. 17, 522–528. Comer, J.A., Tesh, R.B., 1991. Phlebotomine sand flies as vectors of vesiculovirus: a review. Parasitologia 33, 143–150. Danin, A., 1988. Flora and vegetation of Israel and adjacent areas. In: Yom-Tov, Tchernov (Eds.), The Zoogeography of Israel. Junk Publishers, Dordrecht Netherlands, pp. 251–276. Daulaire, N., 1999. Globalization and health. Development 4, 22–24.
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