Colonization of the Mediterranean basin by the vector biting midge species Culicoides imicola: an old story.

Understanding the demographic history and genetic make-up of colonizing species is critical for inferring population sources and colonization routes. This is of main interest for designing accurate control measures in areas newly colonized by vector species of economically important pathogens. The biting midge Culicoides imicola is a major vector of orbiviruses to livestock. Historically, the distribution of this species was limited to the Afrotropical region. Entomological surveys first revealed the presence of C. imicola in the south of the Mediterranean basin by the 1970s. Following recurrent reports of massive bluetongue outbreaks since the 1990s, the presence of the species was confirmed in northern areas. In this study, we addressed the chronology and processes of C. imicola colonization in the Mediterranean basin. We characterized the genetic structure of its populations across Mediterranean and African regions using both mitochondrial and nuclear markers, and combined phylogeographical analyses with population genetics and approximate Bayesian computation. We found a west/east genetic differentiation between populations, occurring both within Africa and within the Mediterranean basin. We demonstrated that three of these groups had experienced demographic expansions in the Pleistocene, probably because of climate changes during this period. Finally, we showed that C. imicola could have colonized the Mediterranean basin in the Late Pleistocene or Early Holocene through a single event of introduction; however, we cannot exclude the hypothesis involving two routes of colonization. Thus, the recent bluetongue outbreaks are not linked to C. imicola colonization event, but rather to biological changes in the vector or the virus.

Parapatric distribution and sexual competition between two tick species, Amblyomma variegatum and A. hebraeum (Acari, Ixodidae), in Mozambique.

BACKGROUND: Amblyomma variegatum and A. hebraeum are two ticks of veterinary and human health importance in south-east Africa. In Zimbabwe they occupy parapatric (marginally overlapping and juxtaposed) distributions. Understanding the mechanisms behind this parapatry is essential for predicting the spatio-temporal dynamics of Amblyomma spp. and the impacts of associated diseases. It has been hypothesized that exclusive competition between these species results from competition at the levels of male signal reception (attraction-aggregation-attachment pheromones) or sexual competition for mates. This hypothesis predicts that the parapatry described in Zimbabwe could also be present in other countries in the region. METHODS: To explore this competitive exclusion hypothesis we conducted field surveys at the two species' range limits in Mozambique to identify areas of sympatry (overlapping areas) and to study potential interactions (communicative and reproductive interference effects) in those areas. At sympatric sites, hetero-specific mating pairs were collected and inter-specific attractiveness/repellent effects acting at long and short distances were assessed by analyzing species co-occurrences on co-infested herds and co-infested hosts. RESULTS: Co-occurrences of both species at sampling sites were infrequent and localized in areas where both tick and host densities were low. At sympatric sites, high percentages of individuals of both species shared attachment sites on hosts and inter-specific mating rates were high. Although cross-mating rates were not significantly different for A. variegatum and A. hebraeum females, attraction towards hetero-specific males was greater for A. hebraeum females than for A. variegatum females and we observed small asymmetrical repellent effects between males at attachment sites. CONCLUSIONS: Our observations suggest near-symmetrical reproductive interference between A. variegatum and A. hebraeum, despite between-species differences in the strength of reproductive isolation barriers acting at the aggregation, fixation and partner contact levels. Theoretical models predict that sexual competition coupled with hybrid inviability, greatly reduces the probability of one species becoming established in an otherwise suitable location when the other species is already established. This mechanism can explain why the parapatric boundary in Mozambique has formed within an area of low tick densities and relatively infrequent host-mediated dispersal events.

Exploiting the heterogeneous drop-off rhythm of Amblyomma variegatum nymphs to reduce pasture infestation by adult ticks.

Tick distribution depends on the drop-off rhythm of the previous stage and on the suitability for tick survival of the environment where they are disseminated. Studies were implemented in Burkina Faso to assess detachment pattern of engorged Amblyomma variegatum nymphs. Experiments were carried out with naturally infested cattle kept in a paddock or monitored when grazing community pasture. In the pasture, 80% of the nymphs detached between 14.00 h and 17.00 h while less than 25% did so in the paddock. Further investigation was implemented to assess whether the density of adult ticks might be modified by herd management. During the early dry season, zebus grazed in 4 plots fenced in natural savannahs and fallows. Two of the plots were used in the morning and the two others in the afternoon. Six months later, zebus were put in these plots, in turn, on 9 occasions. The number of A. variegatum adults picked up by the cattle in each plot was highly variable: they captured more ticks in the plots installed on good lush savannah and 3-fold more ticks in those where the herd had grazed in the afternoon during the previous dry season. An integrated tick control strategy taking these results into account is proposed.

Analysis of Amblyomma surveillance data in the Caribbean: lessons for future control programmes.

Amblyomma variegatum, the Tropical Bont Tick (TBT), is the principal vector of heartwater and is associated with dermatophilosis, major causes of losses in animal production and mortality in Caribbean livestock. From 1995 to 2007, the Caribbean Amblyomma Programme (CAP) supported treatment and surveillance activities in 11 islands of the Eastern Caribbean with an initial objective of eradicating TBT. In addition to control activities, surveillance data were collected between 1997 and 2006 in a unique regional database. We report the analysis of the surveillance data from four islands (Nevis, St Kitts, St Lucia, and Barbados) where control and surveillance followed the initial protocol and where enough data were collected. We describe the evolution of TBT infestation levels and the efforts carried out throughout the surveillance period. Logistic regression identified factors associated with herds found infested with TBT. Overall, treatment programmes were associated with a decrease in proportion of TBT-infested farms. High surveillance efforts were carried out throughout the 1997-2007 period for all island of interest, but inadequate level of surveillance was observed in several quarters especially for St Kitts. Third quarter of the year, as indication of adult TBT seasonality on livestock, was significantly associated with the risk of detecting TBT in Nevis and St Kitts livestock farms. Also, presence of cattle in Nevis farms was shown associated with an increasing probability of farms being declared TBT-infested. Outcomes of these analyses provide basis for recommendations to improve future national and regional control and surveillance activities. This analysis demonstrates the usefulness of long term and adequate surveillance data for control programmes and identification of factors associated with risk of having infested herds.

Control of bovine trypanosomosis by restricted application of insecticides to cattle using footbaths.

African animal trypanosomoses are the main parasitological constraints to livestock production in many sub-Saharan African countries infested with tsetse flies. A prospective survey was implemented in Dafinso (Burkina Faso) to assess the effect of deltamethrin 0.005% (Vectocid(ND), CEVA Santé Animale) impregnation of cattle on trypanosomes transmission in cattle. Two herds were involved in the survey. They were watered at two different waterpoints located on the same river harboring a Guinean riparian forest infested with two different species of tsetse flies (Diptera: Glossinidae), Glossina palpalis gambiensis Vanderplank and G. tachinoides Westwood. Animals belonging to one of the herds were impregnated with deltamethrin applied with a footbath whereas the other herd was used as control. The overall incidence of cattle trypanosomoses was reduced (p=0.01) from 0.76 (control group) to 0.11 (footbath-treated group). A positive effect of the footbath treatment on packed-cell volume was observed (p<0.001). The conditions requested to use a footbath to prevent cattle trypanosomoses are discussed.

Poor inheritance of low attractiveness for Amblyomma variegatum in cattle.

Because of the high variability of cattle infestation, selective breeding has been proposed to control three-host ticks in Africa in the same way as it has been implemented to control the one-host tick Boophilus microplus in Australia. Interbreeding (divergent selection experiment) between Gudali zebu cattle was performed at the Wakwa Research Centre, Adamawa, Cameroon to assess the feasibility and impact of such a selective breeding programme. Fifty cows (chosen out of 129) and 2 bulls (chosen out of 10) were selected according to their greater or lesser attractiveness for the tick Amblyomma variegatum, assessed by their infestation degree (ID, ratio between individual animal infestation and mean herd infestation). Half of the animals displayed a high ID (H) and the remainder a low ID (L). The presence of a suckling calf had a significant effect on the infestation of its dam, the udder of lactating cows being significantly less infested (P<0.001) because of a tick removal effect by the calves. Observed infestation data were therefore corrected to allow comparison between lactating and non-lactating cows. The ID of the cows was assessed on five occasions from 1992 to 1994: the consistency of the infestation hierarchy was good overall (P<0.001) despite high variability seen in some animals which would complicate the selection of cattle with low infestation. Four breeding groups were set up (LxL, LxH, HxL, HxH) and 40 calves were born in 1993 and 1994. There was a correlation (P<0.02-0.001) between liveweight (or age) and ID when the younger calves of the groups were 6 months old, but the correlation disappeared by the time the calves were all older than 1 year. There was no difference between the average ID of the calves from the four breeding groups, and no correlation was observed between dam ID and calf ID. The average ID of the 10 calves born of the dams with the lowest ID was however significantly lower than that of the 10 calves born of the dams with the highest ID (0.82 versus 1.15, P=0.03), but the difference was much less than that observed between the dams (0.35 versus 2.30, P=7x10(-9)). A positive correlation between dam ID and calf ID was found when only these 20 cows were taken into consideration (r=0.57, P<0.01). These results indicate that selective breeding of Gudali zebu cattle to produce animals with low infestation by the tick A. variegatum would be uncertain but that a more drastic selection of the breeders than that implemented in the present study may perhaps have some positive impact.

Tsetse control in cattle from pyrethroid footbaths.

In Burkina Faso, we assessed the efficacy of treating cattle with a footbath containing aqueous formulations of pyrethroids to control two tsetse-fly species, Glossina tachinoides Westwood, 1850 (Diptera, Glossinidae) and Glossina palpalis gambiensis Vanderplank 1949. Legs were the most targeted parts of the body for tsetse-fly blood meals: 81% (95% CI: 73, 89) for G. tachinoides and 88% (81, 95) for G. palpalis. The in-stable efficacy of footbath treatments was compared with manual full spraying with a 0.005% alphacypermethrin (Dominex, FMC, Philadelphia, USA) formulation (250mL versus 2L). The proportions of knocked-down flies were the same with footbath and full spray but the latter was more protective against fly bites. In field use, the efficacy of both methods should be similar given the recommended treatment frequency: 3 days for footbath versus 7 days for full spray. Among 96 cattle drinking at the same water point in Dafinso (Burkina Faso), 68 (71%) were treated with a footbath containing a 0.005% deltamethrin formulation (Vectocid, CEVA SA, Libourne, France). We observed the effect of this live-bait technique on the one hand on released cohorts of reared, irradiated flies, and on the other hand on wild tsetse flies. In both cases, the footbath treatment was associated with a reduction of the apparent fly density probably related to an increased mortality.

Attachment kinetics of the adult tick Amblyomma variegatum to cattle.

At the beginning of the 1999 rainy season, three traditional cattle herds were monitored for 48 days while grazing in the bushy savannah of southwestern Burkina Faso. Cattle in each herd were caught on several occasions each day and the attached ticks were counted. This confirmed that Amblyomma variegatum Fabricius (Acari: Ixodidae) adults picked up in the pastures mainly attach to the interdigital areas (87% of the 791 ticks captured), and reach the predilection sites later (chest and udder/inguinal area) when the animals lie down. As many females as males attached to the hosts, but the seasonal distribution was very heterogeneous, with only a few females attaching as long as the humidity rate remained low. It is suggested that this prevents eggs from being laid when conditions are not optimal for their survival and that of the larvae. Ticks attached all day but the number picked up hourly and daily varied greatly according to their density on the pasture. As a general trend, confirmed by another study carried out in 2005, the number of ticks picked up daily increased from less than one tick/animal/day, before the onset of the rainy season, to 6.5 (+/- 1.5) ticks/animal/day on average during the infestation peak, which lasted 6-8 weeks, until early or mid-July. The number then decreased to less than one tick/animal/day from the end of July onwards. The infestation on the predilection sites followed the same trend, with a daily tick burden increase of three to five A. variegatum adults, depending on herd and year, during the infestation peak.

Footbath acaricide treatment to control cattle infestation by the tick Amblyomma variegatum.

Previous studies have shown that about 90% of adult Amblyomma variegatum Fabricius (Acari: Ixodidae) picked up daily by grazing cattle are still attached to the interdigital areas in the evening, when the animals return from pasture. It was therefore postulated that a targeted treatment, designed to kill the ticks attached to the feet, would limit infestation of the predilection sites. Footbaths filled with various pyrethroid formulations were used over 3 years, at the beginning of the rainy season (from mid-May to the end of July), to assess the efficacy of such a control method. It proved efficient in preventing the ticks from attaching to the predilection sites. Although five to 12 A. variegatum adults attached to each treated animal daily, and although the tick burden of the predilection sites of control cattle increased each day by four to 10 ticks, the average infestation of the predilection sites of treated cattle that were initially highly infested (over 100 ticks/animal) continuously decreased to reach a level of about 10-30 ticks/animal after 6-8 weeks of treatment. In herds with a lower initial tick burden (40-70 ticks/animal) this level was obtained within 2-3 weeks and the mean infestation subsequently remained consistently low. Footbath treatment carried out every other day during the adult peak infestation period should therefore greatly limit losses due to ticks. This method was appreciated by traditional livestock farmers, essentially because it is not time-consuming and because it requires only c. 200 mL aqueous formulation per animal at each passage. The cost of the acaricide needed to treat one animal during the peak infestation period was assessed at c. euro 0.20. This control method might also have an impact on some species of tsetse flies and mosquitoes, thereby contributing to trypanosomiasis and malaria control.

Invasion of West African cattle by the tick Amblyomma variegatum.

Studies in Cameroon and Burkina Faso examined the invasion process of cattle by adult Amblyomma variegatum Fabricius (Acari: Ixodidae) ticks. Nearly all the ticks picked up in the pasture during grazing were found on the limb ends, near the hooves, where they temporarily attached. Then when the cattle lay down, the ticks moved from the feet towards the predilection sites, where they attached definitively. Many ticks disappeared during this displacement. All the female ticks and approximately 70% of the males were thus unable to attach to the predilection sites as long as the pioneer males had not attached and started to produce attraction-aggregation-attachment pheromones. Nevertheless, A. variegatum females and males attached to the feet in similar numbers during grazing, whether the cattle were already infested or not, indicating that stimuli originating from the hosts are powerful enough to attract both sexes. After attachment of the pioneer males, the number of ticks that successfully reached the predilection sites increased. However, even on infested animals, 40-50% of A. variegatum ticks found near the hooves after the grazing periods disappeared during the night following their capture. When moving from the temporary attachment sites towards the final ones, one-third of the ticks changed the individual host. Considering this two-stage infestation process, it is suggested that a targeted tick control, using a foot-bath, might greatly reduce cattle infestation. In particular, it could be effective in traditional herds, with animals grazing permanently during the day, lying down only once back in the night pen.

Impact of natural infestation of Amblyomma variegatum on the liveweight gain of male Gudali cattle in Adamawa (Cameroon).

The effect of natural tick infestation on the liveweight gain (LWG) of male Gudali zebu cattle was studied throughout a year by comparing the performances of two herds, one of which was submitted to weekly acaricidal treatment and the other was left untreated against ticks. Six species of ticks were identified on the untreated animals: Amblyomma variegatum, Boophilus decoloratus, Rhipicephalus lunulatus, Rhipicephalus turanicus, Hyalomma nitidum and Hyalomma marginatum rufipes. Most of the losses observed in the untreated herd during the rainy season were due to A. variegatum, and the loss in LWG was estimated to be 55-76 g per engorged female A. variegatum. The infestation also leads to wounds and to lesions of dermatophilosis. There was an interval between the peak infestation by A. variegatum and the appearance of weight loss owing to them. The control of ticks on the Gudali zebu in Adamawa, during the months of high infestation by A. variegatum adults, is economically profitable. On the other hand, the performances of the two herds during the dry season were similar, showing that infestation by larvae and nymphs of A. variegatum has no impact on the zebu LWG, and that tick control during that period is not profitable.

Variability of cattle infestation by Amblyomma variegatum and its possible utilisation for tick control.

A great variability of the individual infestation by Amblyomma variegatum adults was observed on naturally infested Gudali zebus. Some of the animals (called "attractive for A. variegatum") had a tick burden 10 to 16 times higher than that of the least parasitized cattle of the herd (called "non-attractive"). Ranking of the animals based on A. variegatum infestation was correlated for successive tick counts. Experiments were designed to determine if the "non-attractive" cattle remained lightly infested when the "attractive" ones are removed from the herd. When these two types of cattle grazed separately, it was observed that the "non-attractive" animals had a lower tick burden and that their infestation occurred more slowly than that of the "attractive" ones. The difference between the two groups was nevertheless smaller than that existing when the animals were in the same herd. The selection of the "non-attractive" cattle, on which a tick control programme should not exclusively be based, could however be used as a component of a tick control strategy, if this characteristic is hereditary. An experiment in progress will study the question.