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Seasonal dynamics of biting midges (Diptera: Ceratopogonidae: Culicoides), the potential vectors of bluetongue virus, in Sweden
Veterinary Parasitology 184 (2012) 59–67
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Seasonal dynamics of biting midges (Diptera: Ceratopogonidae: Culicoides), the potential vectors of bluetongue virus, in Sweden M. Ander a , R. Meiswinkel b , J. Chirico a,∗ a b
Department of Virology, Immunology and Parasitology, National Veterinary Institute, 751 89 Uppsala, Sweden Loc Santa Maria del Monte, Via Pratarone 14, 00030 Rocca di Cave, Rome, Italy
a r t i c l e
i n f o
Article history: Received 11 October 2010 Received in revised form 28 July 2011 Accepted 8 August 2011 Keywords: Culicoides Seasonal abundance Bluetongue Onderstepoort Veterinary Institute trap
a b s t r a c t The outbreak of bluetongue (BT) in northern Europe 2006 initiated the monitoring of vectors, biting midges of the genus Culicoides in Sweden. In order to determine the diversity, distribution and seasonal dynamics of Culicoides, weekly collections were made during 2008 and during March–December 2009 using the Ondestepoort Veterinary Institute black light trap. Twenty sampling sites were selected in 12 provinces. In total of 30,704 Culicoides were collected in 2008 and 32,252 in 2009. The most abundant species were the potential vectors of BTV Culicoides obsoletus/C. scoticus that comprised of 77% of the total catches. Other biting midges collected were Culicoides impunctatus (9%), Culicoides grisescens (3%), Culicoides punctatus (2%), Culicoides chiopterus (2%) and Culicoides pulicaris (2%). Culicoides obsoletus/C. scoticus were most abundant during May–June and August–September. The majority of the species were active from March to November in 2008 and April to October in 2009. Species considered as potential vectors of bluetongue virus (BTV) occurred as far north as latitude 65◦ N (Kalix). © 2011 Elsevier B.V. All rights reserved.
1. Introduction Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are the insect vector of economically important veterinary diseases such as African horse sickness virus (AHSV), epizootic hemorrhagic disease virus (EHDV) and the most important in northern Europe bluetongue virus (BTV) (Mellor et al., 2000). Bluetongue is caused by a double stranded RNA virus belonging to the genus Orbivirus of the family Reoviridae and infects ruminants (Purse et al., 2008). Since the discovery of bluetongue in northern Europe in August 2006 the BTV serotype 8 (BTV-8) affected tens of thousands holdings and caused production losses throughout the region (Wilson et al., 2007; Carpenter et al., 2009). Consecutive outbreaks also included Denmark, United Kingdom, Switzerland and Czech Republic. During
∗ Corresponding author. Tel.: +46 18 67 41 59; fax: +46 18 67 44 67. E-mail address: [email protected] (J. Chirico). 0304-4017/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.vetpar.2011.08.009
2008 the virus was spread into new countries including Sweden, Hungary, Austria and Italy (Hofmann et al., 2008; Carpenter et al., 2009; Rasmussen et al., 2010; Sternberg Lewerin et al., 2010; Szmaragd et al., 2010). In September 2008, the first case of BTV in Sweden was detected in a dairy herd in the county of Halland. After the initial detection more animals were tested positive for BTV by serology or PCR. A total of 30 infected cattle herds and three sheep flocks were tested positive in six counties in southern Sweden during September 2008–February 2009 (Sternberg Lewerin et al., 2010). Atmospheric transport modelling showed that the potential source of infection could have been wind-borne from Culicoides from Denmark or Germany in August 2008 (Agren et al., 2010). Sweden initiated vaccination directly after the outbreak, which in combination with establishment of restriction zones preventing animal movements probably restricted further spread of the disease (Sternberg Lewerin et al., 2010). Consequently, Sweden was declared free from BTV in late 2010.
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M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Table 1 Locations of traps operated during 2008–2009 where site 1 is the most southern location and site 20 most northern one. No.
Site
Province
Livestock
Coordinates (lat, long)
No. of collection made 2008/2009
Total Culicoides collected 2008/2009
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
Bara1 Löberöd Svalov1 Svängsta1 Vittsjö Torsås Ljungbyholm2 Halmstad1 Falkenberg1 Hemse Romakloster Torestorp Axvalla Uppsalaa Uppsalab , 2 Orsa Oviken Lycksele1 Kalix Abisko
Skåne Skåne Skåne Blekinge Skåne Småland Småland Halland Halland Gotland Gotland Västergötland Västergötland Uppland Uppland Dalarna Jämtland Västerbotten Norrbotten Lappland
Horses Cattle Cattle Cattle Sheep Sheep Horses Cattle Cattle Sheep Sheep Cattle Sheep Sheep, horses Sheep, horses Cattle Horses Horses Horses No livestock
55◦ 35 22.41 N; 13◦ 12 41.51 E 55◦ 45 9.11 N; 13◦ 32 28.15 E 55◦ 54 28.84 N; 13◦ 5 25.34 E 56◦ 18 5.54 N; 14◦ 43 6.61 E 56◦ 19 56.49 N; 13◦ 42 2.10 E 56◦ 21 26.44 N; 15◦ 49 19.16 E 56◦ 31 46.11 N; 16◦ 12 49.70 E 56◦ 40 20.10 N; 13◦ 10 36.20 E 56◦ 54 56.42 N; 12◦ 32 3.74 E 57◦ 12 22.18 N; 18◦ 22 12.66 E 57◦ 23 55.36 N; 18◦ 30 56.70 E 57◦ 23 34.84 N; 12◦ 39 27.64 E 58◦ 27 10.37 N; 13◦ 43 45.90 E 59◦ 48 58.63 N; 17◦ 46 27.49 E 59◦ 47 56.33 N; 17◦ 45 22.40 E 61◦ 08 50.34 N; 14◦ 37 53.46 E 63◦ 4 44.40 N; 14◦ 28 14.15 E 64◦ 35 4335 ; 18◦ 40 10.84 E 65◦ 44 45.13 N; 23◦ 3 55.62 E 68◦ 20 56.40 N; 18◦ 44 56.07 E
22/0 25/13 8/0 12/0 12/9 24/22 0/3 4/0 6/1 35/18 24/21 29/26 27/19 4/7 0/14 6/6 4/17 5/0 4/12 9/14
632/0 1134/53 2338/0 1462/0 11,605/18,123 8197/4939 0/485 15/0 252/0 167/33 545/17 1046/652 314/105 443/208 0/502 1154/45 249/7077 5/0 1033/8 113/5
Trap sites, numbers 1–20 refers to trap locations in Fig. 2. a Uppsala, Linneus Hammarby. b Uppsala, Funbo. 1 Trap operated only during 2008. 2 Trap operated only during 2009.
The potential vectors of BTV in northern Europe belong to the Obsoletus and Pulicaris groups, namely Culicoides obsoletus, Culicoides scoticus, Culicoides dewulfi, Culicoides chiopterus and Culicoides pulicaris. Their vector potential were determined either by viral isolation or virus detection by reverse transcriptase quantitative
polymerase chain reaction (RT-qPCR) (Caracappa et al., 2003; De Liberato et al., 2005; Savini et al., 2005; Meiswinkel et al., 2007; Dijkstra et al., 2008). Moreover, experimental studies showed that C. obsoletus and C. scoticus may replicate both BTV-8 and BTV-9 (Carpenter et al., 2008a).
Fig. 1. Total numbers of Culicoides species collected at 20 sites during 2008–2009. Gray, no collection was made, white, 0 Culicoides; yellow, 1–10 Culicoides; orange, 10–100 Culicoides; brown, 100–1000 Culicoides and red, 1000–10,000 Culicoides. A point in box means collection made inside. Black horizontal bar between figures represent time of year during 2008 when the Swedish outbreak of BTV occurred.
197 (31) 250 (21) 164 (7) 43 (3) 77 (0.3) 454 (3) 17 (4) 0 (0) 13 (5) 21 (10) 376 (67) 32 (2) 105 (25) 134 (20) 57 (11) 4 (0) 6 (0) 1 (20) 617 (59) 4 (3) 0 (0) 1 (0) 0 (0) 3 (0) 82 (0.3) 112 (1) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 2 (0) 25 (6) 29 (5) 2 (0) 575 (48) 555 (8) 3 (60) 337 (33) 113 (96) 1 (0) 1 (0) 0 (3) 79 (5) 3934 (13) 1527 (12) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 129 (8) 33 (8) 0 (0) 0 (0) 105 (9) 2 (0) 0 (0) 3 (0) 1 (1)
Other species C. grisescens N (%) C. impunctatus N (%)
61
During the emergence of the epidemic of BTV in Northern Europe most EU countries initiated monitoring of biting midges (Meiswinkel et al., 2008; Sehnal et al., 2008; Casati et al., 2009; Horbrand and Geier, 2009; Kiel et al., 2009; Vorsprach et al., 2009). In July 2007, Sweden implemented a vector-monitoring program. Thirty new species of Culicoides was recorded in the Swedish fauna, where the potential vectors, C. obsoletus, C. scoticus, C. chiopterus and C. dewulfi in the Obsoletus group and C. pulicaris, Culicoides punctatus in the Pulicaris group, were included (Nielsen et al., 2010). Still, the knowledge and role of potential vectors for BTV in Sweden is very limited. The aim of the present study was to determine the diversity, distribution and seasonal dynamics of Culicoides species in different regions of Sweden during 2008–2009. 2. Materials and methods
56 (9) 60 (5) 25 (3) 62 (4) 127 (0.4) 135 (1) 31 (6) 4 (27) 26 (10) 19 (10) 79 (14) 516 (30) 6 (1.4) 29 (5) 166 (33) 7 (1) 43 (1) 0 (0) 33 (3) 0 (0) 27 (4) 76 (6) 43 (5) 116 (8) 317 (1) 133 (1) 0 (0) 4 (27) 25 (10) 26 (13) 14 (2.5) 143 (8) 10 (2.4) 2 (0) 9 (2) 4 (0) 258 (3) 0 (0) 3 (0) 0 (0) 1 (0) 454 (38) 194 (2) 44 (3) 278 (1) 29 (0) 68 (14) 0 (0) 4 (2) 2 (1) 25 (4.5) 0 (0) 1 (0.2) 1 (0) 4 (1) 1 (0) 178 (2) 0 (0) 9 (1) 0 (0) 22 38 8 12 21 46 3 4 7 53 45 55 46 11 14 12 21 5 16 23
632 1188 2338 1536 29,728 13,136 485 15 252 200 562 1698 419 651 508 1199 7326 5 1041 118
350 [95, 7, 29, 32, 187] (55) 345 [291, 7, 7, 11, 29] (29) 1, 912 [1644, 215, 16, 2611] (82) 1, 189 [537, 4, 10, 618, 18] (77) 24, 913 [15553, 41, 103, 9191, 25] (84) 10, 746 [8514, 98, 173, 1829, 132] (82) 369 [98, 205, 2, 62, 2] (76) 7 [4, 0, 0, 3, 0] (47) 184 [115, 2, 5, 51, 11] (73) 132 [83, 6, 23, 17, 3] (66) 68 [45, 0, 13, 6, 4] (12) 876 [531, 7, 87, 240, 11] (52) 239 [23, 8, 22, 160, 26] (57) 456 [253, 6, 45, 97, 55] (70) 270 [169, 1, 16, 80, 4] (53) 503 [393, 1, 13, 96, 0] (42) 6284 [5422, 11, 108, 737, 6] (86) 1 [0, 0, 0, 1, 0] (20) 39 [30, 0, 2, 7, 0] (4) 0 [0, 0, 0, 0, 0] (0) 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
C. punctatus N (%) C. chiopterus N (%)
C. pulicaris N (%)
2.1. Sampling sites
Site no.No. of Total C. obsoletus/scoticus N collections collections [Nu, B, G, P, M] (%)
Table 2 Percentage of the seven most prevalent Culicoides species captured in site 1–20 in Sweden 2008–2009. The proportion of species per trap site, physiological stages and sex determination are given for C. obsoletus/C. scoticus. Nu, nulliparous; B, freshly blood fed; G, gravid; P, parous; M, male.
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Nineteen of the twenty sampling sites were farms with a minimum herd size of ten animals of cattle, sheep or horses. Site 20, Abisko research station, did not harbour any livestock. The locations of sites are shown in Table 1. These were also studied by Nielsen et al. (2010). 2.2. Collection of midges Collection was performed according to a protocol adapted after Goffredo and Meiswinkel (2004) using the Onderstepoort Veterinary Institute black light suction trap described by Venter and Meiswinkel (1994). This device consists of a fluorescent black light and a down-drought suction fan powered with a 12 V DC car battery. On each sampling occasion, the minimum–maximum temperatures were recorded. Traps were operated weekly beginning in the late afternoon over night for a duration of 18 h on any given weekday when suitable conditions prevailed, i.e. no rain or wind speeds more than 5 m/s. Traps were placed 1.5–2 m above ground, adjacent to pastures, stables or animal sheds. Such collections were carried out during 2008 at sites 1–6, 8–14, 16–20 and during March–December in 2009 at sites 2, 5–7, 10–15, 17, 19–20, respectively. In order to record any indoor activity, collections were performed in animal stables out of reach from the animals, from January to the end of May in 2008 at sites 1–3, 5, 10–13 and were again initiated at sites 10–11 in November. To establish and compare first occurrence of Culicoides during the years, data-sets from Nielsen et al. (2010) were used, i.e. during January–May 2008 from sites 1–13 and from August–October from sites 16–20. In 2009, indoor collections were initiated at sites 10, 12–13 in March and ceased in May, at sites 10, 12 and in April at site 13. Corresponding catches were only performed at site 7 during October and at site 10 during December this year. In order to establish first and last occurrence of midges, data-sets were selected from sites where both the on-set of the activity was detected
62
Table 3 Presence of Culicoides species in 18 sites located throughout Sweden during 2008–2009. Species are sorted into subgenus starting with the most distributed species. Site no. Sample year
1b
2a/b/c
3b
4a/b
5a/b/c
6a/b/c
7a/c
9a/b
10a/b/c
11a/b/c
12a/b/c
13a/b/c
14a/b/c
15c
16b/c
17b/c
19b/c
Avaritia
C. obsoletus/C. scoticus C. dewulfi C. chiopterus C. pulicaris C. punctatis C. lupicaris C. grisescens C. newsteadi C. impunctatus C. subfasciipennis C. pallidicornis C. achrayi C. fascipennis C. kibunensis C. duddingstoni C. pictipennis C. festivipennis C. clastrieri C. truncurum C. alazanicus C. albicans C. circumscriptus C. salinarius C. sphagnumensis C. segnis C. reconditus C. riethi C. stigma C. nubeculosus 30
b
a, b, c a, c a, b, c a, b, c a, b, c a a, b
b b b b b
a, b a, b b a, b a, b
a, b, c a, b a, b, c a, b, c a, b, c b a, b, c
a b, c a, b, c a, b, c a, b, c a, b, c b a, b, c
a, c
a, b b a, b a, b a, b a, b
a, b, c b b a, b, c a, b, c
a, b, c a a, b, c a, b, c a, b, c
a, b, c a, b a, c a, b, c a, b, c c a, b, c a a, b, c a, b a, c a, b, c a, c
a, b, c a a, b a, b, c a, b, c a a, b, c
a, b, c a, b a, c a, b, c a, b, c
c c c c c c c
b*, c
b*, c
b*, c
c b* b*, c
b* b* b*
b*, c
c b*, c b*, c c b*, c
b*
c
Culicoides
Silvaticulicoides
Oecacta
Beltranmyia
Wirthomyia Monoculicoides No. species
b b b b
a, b
a, c a a, c
a, c b b b
a a, b a, c
b b b b
a, b a b a, b
a, b b b
b
b
b b
a
a, b, c a a, b, c b b
b
a, b a, b, c a a, b, c c b a, b, c
a
a
a, b a, b a, c
a, b a
a
a
a, b, c a, b
a
a, c
a, b, c
a, b, c
a, b, c
b b b b
17
a
a, c a a
19
a, b, c a a, c a, b a, c a, b, c a, b, c a, b, c
a
a, b, c
a, c b a, c a, c a, b, c c c a, b, c a, b, c
c c c c
20b/c
b*, c b*, c b
b*, c b* c
b*, c
b*
b*
c c
a
b
a a b
a, b
c c
a, b, c
a, b, c
a, b, c
a, b, c a
a, c a, b, c a, b, c
b*
c
c b*, c
b
11
12
15
17
11
9
10
14
19
19
b c 18
c c 19
b*
c
9
10
10
Total 17 12 17 17 17 9 12 5 8 5 10 11 8 10 10 5 11 2 3 1 2 8 5 1 9 4 2 4 2
5
Sample year: a, 2007 (Nielsen et al., 2010); b, 2008; c, 2009. A letter means presence of at least one Culicoides of that species during the year. Site 8 and site 18 are excluded due to low abundance of Culicoides N = 15, N = 5 respectively. b*, collections from 2008 at site 16–20 were previously presented by Nielsen et al. (2010).
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Subgenus
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Fig. 2. Location of trapping sites in Sweden during 2008 and 2009. The names of the locations are given in Table 1. Species diversity in eight geographic areas.
in spring and the absence of Culicoides was recorded in autumn. 2.3. Identification and age grading All Culicoides were identified to species and sexdetermined using the keys of Campbell and Pelham-Clinton (1960) and Delécolle (1985). Collections were initially
sorted to group level under a stereo light microscope (Wild M3, Switzerland) at low magnification. Parts of the collection were selected where head, lower part of abdomen and one wing was slide-mounted and determined to species. The rest of the body were then homogenized and determined to species according to a protocol modified after Nolan et al. (2007) (Ander et al., submitted). However, it is difficult to distinguish females of C. obsoletus from
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Fig. 3. Seasonal dynamics of C. obsoletus/C. scoticus, C. chiopterus, C. impunctatus, C. punctatus and C. pulicaris at sites 5, 6, 12 and 17.
C. scoticus. Thus, the vast majority of these two dominant species were therefore identified on morphological characters and recorded as C. obsoletus/C. scoticus. Based on abdominal pigmentation females were agegraded to nulliparous (Nu), parous (P) and recorded as gravid (G) or freshly blood-fed (B) (Dyce, 1969). We determine vector-free periods as stipulated by the European Commission (Anonymous, 2007), i.e. when less than five
parous females of potential vector species are collected in a trap during one night. 3. Results 3.1. Diversity A total of 261 light trap collections caught 30,704 Culicoides from 18 sites during 2008 and in 2009 a total of 202
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
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Fig. 4. Seasonal dynamics of C. grisescens and remaining Culicoides species collected at sites 5, 6, 12 and 17.
collections from 14 traps collected 32,252 Culicoides (Fig. 1). In total 30 Culicoides species were recorded, 28 in 2008 and 26 in 2009, respectively (Table 3). The dominating species were C. obsoletus/C. scoticus, a grand total of 48,852 (77%), they were most abundant from latitude 55◦ N to 63 ◦ N (sites 1–17). Other midges caught were mainly C. impunctatus 5815 (9%), C. grisescens 1839 (3%), C. punctatus 1424 (2%), C. chiopterus 1293 (2%) and C. pulicaris 1210 (2%) (Fig. 2). These seven species comprised 95% of the midges captured (Table 2), less than 5% belonged to the other 25 Culicoides species recorded in 2008–2009 (Table 3). The number of midges collected varied much between sites and time of year ranging from none or very few Culicoides to more than 11,000 specimens.
3.2. Distribution The most widespread species in the present study were C. obsoletus, C. scoticus, C. chiopterus, C. pulicaris and C. punctatus. They were recorded in all the sites investigated, except in the most northern one, site 20 (Abisko). Culicoides dewulfi was present in the southern regions, occurring between latitudes 55◦ N–59◦ N, sites 2–6 and 9–15. Culicoides grisescens was recorded from 12 of the sites and was the dominant species at site 20. Culicoides impunctatus was abundant but restricted to sites 4, 5, 12, 13, 16, 17, 19 and 20. The remaining species occurred at 11 sites (Table 3). Out of the 30 species recorded in the present study, 28 were also found by Nielsen et al. (2010). However, Culicoides alazanicus (site 1) and Culicoides nubeculosus (sites 14–15) were recorded for the first time in Sweden.
3.3. Seasonal dynamics Biting midges were recorded from March to November in 2008 and from April to October in 2009 (Fig. 1). Culicoides obsoletus/C. scoticus were the first midges recorded in spring and the last recorded in autumn, peaking in May–June and August–September. Culicoides chiopterus was recorded later in spring and in lower numbers with similar seasonal pattern as C. obsoletus/C. scoticus (Fig. 3 A–D). Culicoides impunctatus was abundant during June–July. Then it gradually declined and was only sporadically caught, while Culicoides pulicaris and C. punctatus were found in low numbers during the whole season without any significant change in abundance (Fig. 3 e–h). Culicoides grisescens appeared later in the season and was most prevalent in July–September (Fig. 4 a–d). Most species were collected during August, 26 species in 2008 and 23 species in 2009. In general, Culicoides were not collected in temperatures below 8 ◦ C. First nulliparous female were recorded at 55◦ N (site 1) in March, W13 in 2008 and in April, W16 in 2009 (Fig. 1). The majority of the first records of Culicoides in Southern Sweden (Sites 1–13) occurred both years in May (Fig. 1). Last nulliparous Culicoides was recorded at site 13, in November, W45 in 2008 and at site 15 in October, W41 in 2009 (Fig. 1). The vector free period ended in May, W19 in 2008 at sites 10–11 and W20 at sites 2, 12 and in June, W22 at sites 1, 13 and started in August, W32 at site 1 and in September, W35 at site 12, W36 at site 2, 13, W37 at site 11 and W38 at site 10 (Fig. 1). Then the vector free period ended again in 2009 in May, W20 at sites 2, 5, 11–12, June, W22 at site 17 and July, W27 at sites 13–14 and started in September, W39 at sites 13 and 15 and October, W40 at sites 6, 10 and
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17 (Fig. 1). Sites 19 and 20 had vector free period all season (Fig. 1). 4. Discussion The results from the present study show that the most abundant and widespread species of biting midges in Sweden are C. obsoletus/C. scoticus. This was valid for all sites except site 20. However, if this was due to the latitude or the absence of livestock is not known. The seasonal dynamics of C. obsoletus/C. scoticus had a characteristic pattern, appearing among the first Culicoides species in April, W13-16, then peaked in May–June, W19-27, and lower abundance was observed during July followed by higher abundances during August–September, W31-38 (Fig. 3). This seasonal pattern was also described from southern Europe by Savini et al. (2005) and Dahl (1997). The last C. obsoletus/C. scoticus were active in October–November, W42-45. The observed seasonality also corresponded well with the BTV outbreak in Sweden in 2008, where confirmed cases were detected in September–December at latitude 55–57◦ N. Moreover, the activity of potential vector species was high during initial introduction of BTV into Sweden in August–September, but then rapidly declined by the end of October. Other species that have been incriminated as vectors of BTV are C. chiopterus and C. dewulfi (Meiswinkel et al., 2007; Dijkstra et al., 2008). Culicoides chiopterus is commonly found in small numbers, however Carpenter et al. (2008b) showed that this species may be underestimated as it is less attracted to the UV-trap. The low abundance of C. dewulfi (0.3%) in the present study implies that it is probably less important as a vector of BTV in Sweden compared to the Netherlands (Meiswinkel et al., 2008). Culicoides pulicaris and C. punctatus have been found in low abundances in all sites, except site 20, during May–October. Thus, potential vector species of BTV occurred as far north as latitude 65◦ N, locally with high abundance. The species richness was comparable to other parts of northern Europe where extensive outbreaks of BTV-8 have occurred (Meiswinkel et al., 2008; Casati et al., 2009; Horbrand and Geier, 2009; Kiel et al., 2009; Vorsprach et al., 2009). However, a major concern for laboratories is the labour-intensive efforts to identify specimens collected. Moreover, to distinguish females of C. obsoletus and C. scoticus require entomological expertise as well as molecular techniques (Nielsen and Kristensen, 2011). Therefore, traditional identification of Culicoides species along with the development of molecular techniques could be useful for future BTV vector surveillance. Acknowledgements We would like to thank the farmers and veterinarians involved for their cooperation and help in the collections. Funding for this research was provided by the Swedish Board of Agriculture. References Agren, E.C., Burgin, L., Lewerin, S.S., Gloster, J., Elvander, M., 2010. Possible means of introduction of bluetongue virus serotype 8 (BTV-8) to
Sweden in August 2008: comparison of results from two models for atmospheric transport of the Culicoides vector. Vet. Rec. 167, 484–488. Anonymous, 2007. Comission Regulation (EC) No. 1266/2007, V. http://eur-lex.europa.eu/LexUriServ/site/en/oj/2007/ Annex l 283/l 28320071027en00370052.pdf. Campbell, J.A., Pelham-Clinton, E.C., 1960. A taxonomic review of the British species of Culicoides Latreille (Diptera, Ceratopogonidae) 67, 181–302. Caracappa, S., Torina, A., Guercio, A., Vitale, F., Calabro, A., Purpari, G., Ferrantelli, V., Vitale, M., Mellor, P.S., 2003. Identification of a novel bluetongue virus vector species of Culicoides in Sicily. Vet. Rec. 153, 71–74. Carpenter, S., McArthur, C., Selby, R., Ward, R., Nolan, D.V., Luntz, A.J., Dallas, J.F., Tripet, F., Mellor, P.S., 2008a. Experimental infection studies of UK Culicoides species midges with bluetongue virus serotypes 8 and 9. Vet. Rec. 163, 589–592. Carpenter, S., Szmaragd, C., Barber, J., Labuschagne, K., Gubbins, S., Mellor, P., 2008b. An assessment of Culicoides surveillance techniques in northern Europe: have we underestimated a potential bluetongue virus vector? J. Appl. Ecol. 45, 1237–1245. Carpenter, S., Wilson, A., Mellor, P.S., 2009. Culicoides and the emergence of bluetongue virus in northern Europe. Trends Microbiol. 17, 172–178. Casati, S., Racloz, V., Delecolle, J.C., Kuhn, M., Mathis, A., Griot, C., Stark, K.D., Vanzetti, T., 2009. An investigation on the Culicoides species composition at seven sites in southern Switzerland. Med. Vet. Entomol. 23, 93–98. Dahl, C., 1997. Diptera Ceratopogonidae, biting midges. In: Nilsson, A. (Ed.), The Aquatic Insects of North Europe 2. Apollo Books, Stenstrup, pp. 243–263. De Liberato, C., Scavia, G., Lorenzetti, R., Scaramozzino, P., Amaddeo, D., Cardeti, G., Scicluna, M., Ferrari, G., Autorino, G.L., 2005. Identification of Culicoides obsoletus (Diptera: Ceratopogonidae) as a vector of bluetongue virus in central Italy. Vet. Rec. 156, 301–304. Delécolle, J.C., 1985. Nouvelle contribution à l’étude sysématiqueet iconographique des espéces du ganre Culicoides (Diptera: Ceratopogonidae) du Nord-Est de la France. PhD theses. Université Louis Pateur du Strasbourg, Strasbourg. Dijkstra, E., van der Ven, I.J., Meiswinkel, R., Holzel, D.R., Van Rijn, P.A., 2008. Culicoides chiopterus as a potential vector of bluetongue virus in Europe. Vet. Rec. 162, 422. Dyce, A.L., 1969. Recognition of nulliparous and parous Culicoides (Diptera: Ceratopogonidae) without dissection. J. Aust. Ent. Soc. 8, 11–15. Goffredo, M., Meiswinkel, R., 2004. Entomological surveillance of bluetongue in Italy: methods of capture, catch analysis and identification of Culicoides biting midges. Vet. Ital. 40, 260–265. Hofmann, M., Griot, C., Chaignat, V., Perler, L., Thur, B., 2008. Bluetongue disease reaches Switzerland. Schweiz. Arch. Tierheilkd. 150, 49–56. Horbrand, T., Geier, M., 2009. Monitoring of Culicoides at nine locations in Southern Germany (2007–2008). Parasitol. Res. 105, 387–392. Kiel, E., Liebisch, G., Focke, R., Liebisch, A., Werner, D., 2009. Monitoring of Culicoides at 20 locations in northwest Germany. Parasitol. Res. 105, 351–357. Meiswinkel, R., Goffredo, M., Leijs, P., Conte, A., 2008. The Culicoides ‘snapshot’: a novel approach used to assess vector densities widely and rapidly during the 2006 outbreak of bluetongue (BT) in The Netherlands. Prev. Vet. Med. 87, 98–118. Meiswinkel, R., van Rijn, P., Leijs, P., Goffredo, M., 2007. Potential new Culicoides vector of bluetongue virus in northern Europe. Vet. Rec. 161, 564–565. Mellor, P.S., Boorman, J., Baylis, M., 2000. Culicoides biting midges: their role as arbovirus vectors. Annu. Rev. Entomol. 45, 307–340. Nielsen, S.A., Kristensen, M., 2011. Morphological and molecular identification of species of the Obsoletus group (Diptera: Ceratopogonidae) in Scandinavia. Parasitol. Res.. Nielsen, S.A., Nielsen, B.O., Chirico, J., 2010. Monitoring of biting midges (Diptera: Ceratopogonidae: Culicoides Latreille) on farms in Sweden during the emergence of the 2008 epidemic of bluetongue. Parasitol. Res. 106, 1197–1203. Nolan, D.V., Carpenter, S., Barber, J., Mellor, P.S., Dallas, J.F., Mordue Luntz, A.J., Piertney, S.B., 2007. Rapid diagnostic PCR assays for members of the Culicoides obsoletus and Culicoides pulicaris species complexes, implicated vectors of bluetongue virus in Europe. Vet. Microbiol. 124, 82–94. Purse, B.V., Brown, H.E., Harrup, L., Mertens, P.P., Rogers, D.J., 2008. Invasion of bluetongue and other orbivirus infections into Europe: the role of biological and climatic processes. Rev. Off. Int. Epizoot. 27, 427–442.
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67 Rasmussen, L.D., Rasmussen, T.B., Belsham, G.J., Strandbygaard, B., Botner, A., 2010. Bluetongue in Denmark during 2008. Vet. Rec. 166, 714–718. Savini, G., Goffredo, M., Monaco, F., Di Gennaro, A., Cafiero, M.A., Baldi, L., de Santis, P., Meiswinkel, R., Caporale, V., 2005. Bluetongue virus isolations from midges belonging to the Obsoletus complex (Culicoides, Diptera: Ceratopogonidae) in Italy. Vet. Rec. 157, 133–139. Sehnal, P., Schweiger, S., Schindler, M., Anderle, F., Schneemann, Y., 2008. Bluetongue: Vector surveillance in Austria in 2007. Wien. Klin. Wochenschr. 120, 34–39. Sternberg Lewerin, S., Hallgren, G., Mieziewska, K., Berndtsson, L.T., Chirico, J., Elvander, M., 2010. Infection with bluetongue virus serotype 8 in Sweden in 2008. Vet. Rec. 167, 165–170.
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Szmaragd, C., Wilson, A.J., Carpenter, S., Wood, J.L., Mellor, P.S., Gubbins, S., 2010. The spread of bluetongue virus serotype 8 in Great Britain and its control by vaccination. PLoS One 5, e9353. Venter, G.J., Meiswinkel, R., 1994. The virtual absence of Culicoides imicola (Diptera: Ceratopogonidae) in a light-trap survey of the colder, high-lying area of the eastern Orange Free State, South Africa, and implications for the transmission of arboviruses. Onderstepoort J. Vet. Res. 61, 327–340. Wilson, A., Carpenter, S., Gloster, J., Mellor, P., 2007. Re-emergence of bluetongue in northern Europe in 2007. Vet. Rec. 161, 487–489. Vorsprach, B., Meiser, C.K., Werner, D., Balczun, C., Schaub, G.A., 2009. Monitoring of ceratopogonidae in southwest Germany. Parasitol. Res. 105, 337–344.
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Veterinary Parasitology journal homepage: www.elsevier.com/locate/vetpar
Seasonal dynamics of biting midges (Diptera: Ceratopogonidae: Culicoides), the potential vectors of bluetongue virus, in Sweden M. Ander a , R. Meiswinkel b , J. Chirico a,∗ a b
Department of Virology, Immunology and Parasitology, National Veterinary Institute, 751 89 Uppsala, Sweden Loc Santa Maria del Monte, Via Pratarone 14, 00030 Rocca di Cave, Rome, Italy
a r t i c l e
i n f o
Article history: Received 11 October 2010 Received in revised form 28 July 2011 Accepted 8 August 2011 Keywords: Culicoides Seasonal abundance Bluetongue Onderstepoort Veterinary Institute trap
a b s t r a c t The outbreak of bluetongue (BT) in northern Europe 2006 initiated the monitoring of vectors, biting midges of the genus Culicoides in Sweden. In order to determine the diversity, distribution and seasonal dynamics of Culicoides, weekly collections were made during 2008 and during March–December 2009 using the Ondestepoort Veterinary Institute black light trap. Twenty sampling sites were selected in 12 provinces. In total of 30,704 Culicoides were collected in 2008 and 32,252 in 2009. The most abundant species were the potential vectors of BTV Culicoides obsoletus/C. scoticus that comprised of 77% of the total catches. Other biting midges collected were Culicoides impunctatus (9%), Culicoides grisescens (3%), Culicoides punctatus (2%), Culicoides chiopterus (2%) and Culicoides pulicaris (2%). Culicoides obsoletus/C. scoticus were most abundant during May–June and August–September. The majority of the species were active from March to November in 2008 and April to October in 2009. Species considered as potential vectors of bluetongue virus (BTV) occurred as far north as latitude 65◦ N (Kalix). © 2011 Elsevier B.V. All rights reserved.
1. Introduction Biting midges of the genus Culicoides (Diptera: Ceratopogonidae) are the insect vector of economically important veterinary diseases such as African horse sickness virus (AHSV), epizootic hemorrhagic disease virus (EHDV) and the most important in northern Europe bluetongue virus (BTV) (Mellor et al., 2000). Bluetongue is caused by a double stranded RNA virus belonging to the genus Orbivirus of the family Reoviridae and infects ruminants (Purse et al., 2008). Since the discovery of bluetongue in northern Europe in August 2006 the BTV serotype 8 (BTV-8) affected tens of thousands holdings and caused production losses throughout the region (Wilson et al., 2007; Carpenter et al., 2009). Consecutive outbreaks also included Denmark, United Kingdom, Switzerland and Czech Republic. During
∗ Corresponding author. Tel.: +46 18 67 41 59; fax: +46 18 67 44 67. E-mail address: [email protected] (J. Chirico). 0304-4017/$ – see front matter © 2011 Elsevier B.V. All rights reserved. doi:10.1016/j.vetpar.2011.08.009
2008 the virus was spread into new countries including Sweden, Hungary, Austria and Italy (Hofmann et al., 2008; Carpenter et al., 2009; Rasmussen et al., 2010; Sternberg Lewerin et al., 2010; Szmaragd et al., 2010). In September 2008, the first case of BTV in Sweden was detected in a dairy herd in the county of Halland. After the initial detection more animals were tested positive for BTV by serology or PCR. A total of 30 infected cattle herds and three sheep flocks were tested positive in six counties in southern Sweden during September 2008–February 2009 (Sternberg Lewerin et al., 2010). Atmospheric transport modelling showed that the potential source of infection could have been wind-borne from Culicoides from Denmark or Germany in August 2008 (Agren et al., 2010). Sweden initiated vaccination directly after the outbreak, which in combination with establishment of restriction zones preventing animal movements probably restricted further spread of the disease (Sternberg Lewerin et al., 2010). Consequently, Sweden was declared free from BTV in late 2010.
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M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Table 1 Locations of traps operated during 2008–2009 where site 1 is the most southern location and site 20 most northern one. No.
Site
Province
Livestock
Coordinates (lat, long)
No. of collection made 2008/2009
Total Culicoides collected 2008/2009
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
Bara1 Löberöd Svalov1 Svängsta1 Vittsjö Torsås Ljungbyholm2 Halmstad1 Falkenberg1 Hemse Romakloster Torestorp Axvalla Uppsalaa Uppsalab , 2 Orsa Oviken Lycksele1 Kalix Abisko
Skåne Skåne Skåne Blekinge Skåne Småland Småland Halland Halland Gotland Gotland Västergötland Västergötland Uppland Uppland Dalarna Jämtland Västerbotten Norrbotten Lappland
Horses Cattle Cattle Cattle Sheep Sheep Horses Cattle Cattle Sheep Sheep Cattle Sheep Sheep, horses Sheep, horses Cattle Horses Horses Horses No livestock
55◦ 35 22.41 N; 13◦ 12 41.51 E 55◦ 45 9.11 N; 13◦ 32 28.15 E 55◦ 54 28.84 N; 13◦ 5 25.34 E 56◦ 18 5.54 N; 14◦ 43 6.61 E 56◦ 19 56.49 N; 13◦ 42 2.10 E 56◦ 21 26.44 N; 15◦ 49 19.16 E 56◦ 31 46.11 N; 16◦ 12 49.70 E 56◦ 40 20.10 N; 13◦ 10 36.20 E 56◦ 54 56.42 N; 12◦ 32 3.74 E 57◦ 12 22.18 N; 18◦ 22 12.66 E 57◦ 23 55.36 N; 18◦ 30 56.70 E 57◦ 23 34.84 N; 12◦ 39 27.64 E 58◦ 27 10.37 N; 13◦ 43 45.90 E 59◦ 48 58.63 N; 17◦ 46 27.49 E 59◦ 47 56.33 N; 17◦ 45 22.40 E 61◦ 08 50.34 N; 14◦ 37 53.46 E 63◦ 4 44.40 N; 14◦ 28 14.15 E 64◦ 35 4335 ; 18◦ 40 10.84 E 65◦ 44 45.13 N; 23◦ 3 55.62 E 68◦ 20 56.40 N; 18◦ 44 56.07 E
22/0 25/13 8/0 12/0 12/9 24/22 0/3 4/0 6/1 35/18 24/21 29/26 27/19 4/7 0/14 6/6 4/17 5/0 4/12 9/14
632/0 1134/53 2338/0 1462/0 11,605/18,123 8197/4939 0/485 15/0 252/0 167/33 545/17 1046/652 314/105 443/208 0/502 1154/45 249/7077 5/0 1033/8 113/5
Trap sites, numbers 1–20 refers to trap locations in Fig. 2. a Uppsala, Linneus Hammarby. b Uppsala, Funbo. 1 Trap operated only during 2008. 2 Trap operated only during 2009.
The potential vectors of BTV in northern Europe belong to the Obsoletus and Pulicaris groups, namely Culicoides obsoletus, Culicoides scoticus, Culicoides dewulfi, Culicoides chiopterus and Culicoides pulicaris. Their vector potential were determined either by viral isolation or virus detection by reverse transcriptase quantitative
polymerase chain reaction (RT-qPCR) (Caracappa et al., 2003; De Liberato et al., 2005; Savini et al., 2005; Meiswinkel et al., 2007; Dijkstra et al., 2008). Moreover, experimental studies showed that C. obsoletus and C. scoticus may replicate both BTV-8 and BTV-9 (Carpenter et al., 2008a).
Fig. 1. Total numbers of Culicoides species collected at 20 sites during 2008–2009. Gray, no collection was made, white, 0 Culicoides; yellow, 1–10 Culicoides; orange, 10–100 Culicoides; brown, 100–1000 Culicoides and red, 1000–10,000 Culicoides. A point in box means collection made inside. Black horizontal bar between figures represent time of year during 2008 when the Swedish outbreak of BTV occurred.
197 (31) 250 (21) 164 (7) 43 (3) 77 (0.3) 454 (3) 17 (4) 0 (0) 13 (5) 21 (10) 376 (67) 32 (2) 105 (25) 134 (20) 57 (11) 4 (0) 6 (0) 1 (20) 617 (59) 4 (3) 0 (0) 1 (0) 0 (0) 3 (0) 82 (0.3) 112 (1) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 2 (0) 25 (6) 29 (5) 2 (0) 575 (48) 555 (8) 3 (60) 337 (33) 113 (96) 1 (0) 1 (0) 0 (3) 79 (5) 3934 (13) 1527 (12) 0 (0) 0 (0) 0 (0) 0 (0) 0 (0) 129 (8) 33 (8) 0 (0) 0 (0) 105 (9) 2 (0) 0 (0) 3 (0) 1 (1)
Other species C. grisescens N (%) C. impunctatus N (%)
61
During the emergence of the epidemic of BTV in Northern Europe most EU countries initiated monitoring of biting midges (Meiswinkel et al., 2008; Sehnal et al., 2008; Casati et al., 2009; Horbrand and Geier, 2009; Kiel et al., 2009; Vorsprach et al., 2009). In July 2007, Sweden implemented a vector-monitoring program. Thirty new species of Culicoides was recorded in the Swedish fauna, where the potential vectors, C. obsoletus, C. scoticus, C. chiopterus and C. dewulfi in the Obsoletus group and C. pulicaris, Culicoides punctatus in the Pulicaris group, were included (Nielsen et al., 2010). Still, the knowledge and role of potential vectors for BTV in Sweden is very limited. The aim of the present study was to determine the diversity, distribution and seasonal dynamics of Culicoides species in different regions of Sweden during 2008–2009. 2. Materials and methods
56 (9) 60 (5) 25 (3) 62 (4) 127 (0.4) 135 (1) 31 (6) 4 (27) 26 (10) 19 (10) 79 (14) 516 (30) 6 (1.4) 29 (5) 166 (33) 7 (1) 43 (1) 0 (0) 33 (3) 0 (0) 27 (4) 76 (6) 43 (5) 116 (8) 317 (1) 133 (1) 0 (0) 4 (27) 25 (10) 26 (13) 14 (2.5) 143 (8) 10 (2.4) 2 (0) 9 (2) 4 (0) 258 (3) 0 (0) 3 (0) 0 (0) 1 (0) 454 (38) 194 (2) 44 (3) 278 (1) 29 (0) 68 (14) 0 (0) 4 (2) 2 (1) 25 (4.5) 0 (0) 1 (0.2) 1 (0) 4 (1) 1 (0) 178 (2) 0 (0) 9 (1) 0 (0) 22 38 8 12 21 46 3 4 7 53 45 55 46 11 14 12 21 5 16 23
632 1188 2338 1536 29,728 13,136 485 15 252 200 562 1698 419 651 508 1199 7326 5 1041 118
350 [95, 7, 29, 32, 187] (55) 345 [291, 7, 7, 11, 29] (29) 1, 912 [1644, 215, 16, 2611] (82) 1, 189 [537, 4, 10, 618, 18] (77) 24, 913 [15553, 41, 103, 9191, 25] (84) 10, 746 [8514, 98, 173, 1829, 132] (82) 369 [98, 205, 2, 62, 2] (76) 7 [4, 0, 0, 3, 0] (47) 184 [115, 2, 5, 51, 11] (73) 132 [83, 6, 23, 17, 3] (66) 68 [45, 0, 13, 6, 4] (12) 876 [531, 7, 87, 240, 11] (52) 239 [23, 8, 22, 160, 26] (57) 456 [253, 6, 45, 97, 55] (70) 270 [169, 1, 16, 80, 4] (53) 503 [393, 1, 13, 96, 0] (42) 6284 [5422, 11, 108, 737, 6] (86) 1 [0, 0, 0, 1, 0] (20) 39 [30, 0, 2, 7, 0] (4) 0 [0, 0, 0, 0, 0] (0) 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20
C. punctatus N (%) C. chiopterus N (%)
C. pulicaris N (%)
2.1. Sampling sites
Site no.No. of Total C. obsoletus/scoticus N collections collections [Nu, B, G, P, M] (%)
Table 2 Percentage of the seven most prevalent Culicoides species captured in site 1–20 in Sweden 2008–2009. The proportion of species per trap site, physiological stages and sex determination are given for C. obsoletus/C. scoticus. Nu, nulliparous; B, freshly blood fed; G, gravid; P, parous; M, male.
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Nineteen of the twenty sampling sites were farms with a minimum herd size of ten animals of cattle, sheep or horses. Site 20, Abisko research station, did not harbour any livestock. The locations of sites are shown in Table 1. These were also studied by Nielsen et al. (2010). 2.2. Collection of midges Collection was performed according to a protocol adapted after Goffredo and Meiswinkel (2004) using the Onderstepoort Veterinary Institute black light suction trap described by Venter and Meiswinkel (1994). This device consists of a fluorescent black light and a down-drought suction fan powered with a 12 V DC car battery. On each sampling occasion, the minimum–maximum temperatures were recorded. Traps were operated weekly beginning in the late afternoon over night for a duration of 18 h on any given weekday when suitable conditions prevailed, i.e. no rain or wind speeds more than 5 m/s. Traps were placed 1.5–2 m above ground, adjacent to pastures, stables or animal sheds. Such collections were carried out during 2008 at sites 1–6, 8–14, 16–20 and during March–December in 2009 at sites 2, 5–7, 10–15, 17, 19–20, respectively. In order to record any indoor activity, collections were performed in animal stables out of reach from the animals, from January to the end of May in 2008 at sites 1–3, 5, 10–13 and were again initiated at sites 10–11 in November. To establish and compare first occurrence of Culicoides during the years, data-sets from Nielsen et al. (2010) were used, i.e. during January–May 2008 from sites 1–13 and from August–October from sites 16–20. In 2009, indoor collections were initiated at sites 10, 12–13 in March and ceased in May, at sites 10, 12 and in April at site 13. Corresponding catches were only performed at site 7 during October and at site 10 during December this year. In order to establish first and last occurrence of midges, data-sets were selected from sites where both the on-set of the activity was detected
62
Table 3 Presence of Culicoides species in 18 sites located throughout Sweden during 2008–2009. Species are sorted into subgenus starting with the most distributed species. Site no. Sample year
1b
2a/b/c
3b
4a/b
5a/b/c
6a/b/c
7a/c
9a/b
10a/b/c
11a/b/c
12a/b/c
13a/b/c
14a/b/c
15c
16b/c
17b/c
19b/c
Avaritia
C. obsoletus/C. scoticus C. dewulfi C. chiopterus C. pulicaris C. punctatis C. lupicaris C. grisescens C. newsteadi C. impunctatus C. subfasciipennis C. pallidicornis C. achrayi C. fascipennis C. kibunensis C. duddingstoni C. pictipennis C. festivipennis C. clastrieri C. truncurum C. alazanicus C. albicans C. circumscriptus C. salinarius C. sphagnumensis C. segnis C. reconditus C. riethi C. stigma C. nubeculosus 30
b
a, b, c a, c a, b, c a, b, c a, b, c a a, b
b b b b b
a, b a, b b a, b a, b
a, b, c a, b a, b, c a, b, c a, b, c b a, b, c
a b, c a, b, c a, b, c a, b, c a, b, c b a, b, c
a, c
a, b b a, b a, b a, b a, b
a, b, c b b a, b, c a, b, c
a, b, c a a, b, c a, b, c a, b, c
a, b, c a, b a, c a, b, c a, b, c c a, b, c a a, b, c a, b a, c a, b, c a, c
a, b, c a a, b a, b, c a, b, c a a, b, c
a, b, c a, b a, c a, b, c a, b, c
c c c c c c c
b*, c
b*, c
b*, c
c b* b*, c
b* b* b*
b*, c
c b*, c b*, c c b*, c
b*
c
Culicoides
Silvaticulicoides
Oecacta
Beltranmyia
Wirthomyia Monoculicoides No. species
b b b b
a, b
a, c a a, c
a, c b b b
a a, b a, c
b b b b
a, b a b a, b
a, b b b
b
b
b b
a
a, b, c a a, b, c b b
b
a, b a, b, c a a, b, c c b a, b, c
a
a
a, b a, b a, c
a, b a
a
a
a, b, c a, b
a
a, c
a, b, c
a, b, c
a, b, c
b b b b
17
a
a, c a a
19
a, b, c a a, c a, b a, c a, b, c a, b, c a, b, c
a
a, b, c
a, c b a, c a, c a, b, c c c a, b, c a, b, c
c c c c
20b/c
b*, c b*, c b
b*, c b* c
b*, c
b*
b*
c c
a
b
a a b
a, b
c c
a, b, c
a, b, c
a, b, c
a, b, c a
a, c a, b, c a, b, c
b*
c
c b*, c
b
11
12
15
17
11
9
10
14
19
19
b c 18
c c 19
b*
c
9
10
10
Total 17 12 17 17 17 9 12 5 8 5 10 11 8 10 10 5 11 2 3 1 2 8 5 1 9 4 2 4 2
5
Sample year: a, 2007 (Nielsen et al., 2010); b, 2008; c, 2009. A letter means presence of at least one Culicoides of that species during the year. Site 8 and site 18 are excluded due to low abundance of Culicoides N = 15, N = 5 respectively. b*, collections from 2008 at site 16–20 were previously presented by Nielsen et al. (2010).
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
Subgenus
M. Ander et al. / Veterinary Parasitology 184 (2012) 59–67
63
Fig. 2. Location of trapping sites in Sweden during 2008 and 2009. The names of the locations are given in Table 1. Species diversity in eight geographic areas.
in spring and the absence of Culicoides was recorded in autumn. 2.3. Identification and age grading All Culicoides were identified to species and sexdetermined using the keys of Campbell and Pelham-Clinton (1960) and Delécolle (1985). Collections were initially
sorted to group level under a stereo light microscope (Wild M3, Switzerland) at low magnification. Parts of the collection were selected where head, lower part of abdomen and one wing was slide-mounted and determined to species. The rest of the body were then homogenized and determined to species according to a protocol modified after Nolan et al. (2007) (Ander et al., submitted). However, it is difficult to distinguish females of C. obsoletus from
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Fig. 3. Seasonal dynamics of C. obsoletus/C. scoticus, C. chiopterus, C. impunctatus, C. punctatus and C. pulicaris at sites 5, 6, 12 and 17.
C. scoticus. Thus, the vast majority of these two dominant species were therefore identified on morphological characters and recorded as C. obsoletus/C. scoticus. Based on abdominal pigmentation females were agegraded to nulliparous (Nu), parous (P) and recorded as gravid (G) or freshly blood-fed (B) (Dyce, 1969). We determine vector-free periods as stipulated by the European Commission (Anonymous, 2007), i.e. when less than five
parous females of potential vector species are collected in a trap during one night. 3. Results 3.1. Diversity A total of 261 light trap collections caught 30,704 Culicoides from 18 sites during 2008 and in 2009 a total of 202
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Fig. 4. Seasonal dynamics of C. grisescens and remaining Culicoides species collected at sites 5, 6, 12 and 17.
collections from 14 traps collected 32,252 Culicoides (Fig. 1). In total 30 Culicoides species were recorded, 28 in 2008 and 26 in 2009, respectively (Table 3). The dominating species were C. obsoletus/C. scoticus, a grand total of 48,852 (77%), they were most abundant from latitude 55◦ N to 63 ◦ N (sites 1–17). Other midges caught were mainly C. impunctatus 5815 (9%), C. grisescens 1839 (3%), C. punctatus 1424 (2%), C. chiopterus 1293 (2%) and C. pulicaris 1210 (2%) (Fig. 2). These seven species comprised 95% of the midges captured (Table 2), less than 5% belonged to the other 25 Culicoides species recorded in 2008–2009 (Table 3). The number of midges collected varied much between sites and time of year ranging from none or very few Culicoides to more than 11,000 specimens.
3.2. Distribution The most widespread species in the present study were C. obsoletus, C. scoticus, C. chiopterus, C. pulicaris and C. punctatus. They were recorded in all the sites investigated, except in the most northern one, site 20 (Abisko). Culicoides dewulfi was present in the southern regions, occurring between latitudes 55◦ N–59◦ N, sites 2–6 and 9–15. Culicoides grisescens was recorded from 12 of the sites and was the dominant species at site 20. Culicoides impunctatus was abundant but restricted to sites 4, 5, 12, 13, 16, 17, 19 and 20. The remaining species occurred at 11 sites (Table 3). Out of the 30 species recorded in the present study, 28 were also found by Nielsen et al. (2010). However, Culicoides alazanicus (site 1) and Culicoides nubeculosus (sites 14–15) were recorded for the first time in Sweden.
3.3. Seasonal dynamics Biting midges were recorded from March to November in 2008 and from April to October in 2009 (Fig. 1). Culicoides obsoletus/C. scoticus were the first midges recorded in spring and the last recorded in autumn, peaking in May–June and August–September. Culicoides chiopterus was recorded later in spring and in lower numbers with similar seasonal pattern as C. obsoletus/C. scoticus (Fig. 3 A–D). Culicoides impunctatus was abundant during June–July. Then it gradually declined and was only sporadically caught, while Culicoides pulicaris and C. punctatus were found in low numbers during the whole season without any significant change in abundance (Fig. 3 e–h). Culicoides grisescens appeared later in the season and was most prevalent in July–September (Fig. 4 a–d). Most species were collected during August, 26 species in 2008 and 23 species in 2009. In general, Culicoides were not collected in temperatures below 8 ◦ C. First nulliparous female were recorded at 55◦ N (site 1) in March, W13 in 2008 and in April, W16 in 2009 (Fig. 1). The majority of the first records of Culicoides in Southern Sweden (Sites 1–13) occurred both years in May (Fig. 1). Last nulliparous Culicoides was recorded at site 13, in November, W45 in 2008 and at site 15 in October, W41 in 2009 (Fig. 1). The vector free period ended in May, W19 in 2008 at sites 10–11 and W20 at sites 2, 12 and in June, W22 at sites 1, 13 and started in August, W32 at site 1 and in September, W35 at site 12, W36 at site 2, 13, W37 at site 11 and W38 at site 10 (Fig. 1). Then the vector free period ended again in 2009 in May, W20 at sites 2, 5, 11–12, June, W22 at site 17 and July, W27 at sites 13–14 and started in September, W39 at sites 13 and 15 and October, W40 at sites 6, 10 and
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17 (Fig. 1). Sites 19 and 20 had vector free period all season (Fig. 1). 4. Discussion The results from the present study show that the most abundant and widespread species of biting midges in Sweden are C. obsoletus/C. scoticus. This was valid for all sites except site 20. However, if this was due to the latitude or the absence of livestock is not known. The seasonal dynamics of C. obsoletus/C. scoticus had a characteristic pattern, appearing among the first Culicoides species in April, W13-16, then peaked in May–June, W19-27, and lower abundance was observed during July followed by higher abundances during August–September, W31-38 (Fig. 3). This seasonal pattern was also described from southern Europe by Savini et al. (2005) and Dahl (1997). The last C. obsoletus/C. scoticus were active in October–November, W42-45. The observed seasonality also corresponded well with the BTV outbreak in Sweden in 2008, where confirmed cases were detected in September–December at latitude 55–57◦ N. Moreover, the activity of potential vector species was high during initial introduction of BTV into Sweden in August–September, but then rapidly declined by the end of October. Other species that have been incriminated as vectors of BTV are C. chiopterus and C. dewulfi (Meiswinkel et al., 2007; Dijkstra et al., 2008). Culicoides chiopterus is commonly found in small numbers, however Carpenter et al. (2008b) showed that this species may be underestimated as it is less attracted to the UV-trap. The low abundance of C. dewulfi (0.3%) in the present study implies that it is probably less important as a vector of BTV in Sweden compared to the Netherlands (Meiswinkel et al., 2008). Culicoides pulicaris and C. punctatus have been found in low abundances in all sites, except site 20, during May–October. Thus, potential vector species of BTV occurred as far north as latitude 65◦ N, locally with high abundance. The species richness was comparable to other parts of northern Europe where extensive outbreaks of BTV-8 have occurred (Meiswinkel et al., 2008; Casati et al., 2009; Horbrand and Geier, 2009; Kiel et al., 2009; Vorsprach et al., 2009). However, a major concern for laboratories is the labour-intensive efforts to identify specimens collected. Moreover, to distinguish females of C. obsoletus and C. scoticus require entomological expertise as well as molecular techniques (Nielsen and Kristensen, 2011). Therefore, traditional identification of Culicoides species along with the development of molecular techniques could be useful for future BTV vector surveillance. Acknowledgements We would like to thank the farmers and veterinarians involved for their cooperation and help in the collections. Funding for this research was provided by the Swedish Board of Agriculture. References Agren, E.C., Burgin, L., Lewerin, S.S., Gloster, J., Elvander, M., 2010. Possible means of introduction of bluetongue virus serotype 8 (BTV-8) to
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