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<ul><li><p>Parasitology</p><p>AdditionalservicesforParasitology:</p><p>Emailalerts:ClickhereSubscriptions:ClickhereCommercialreprints:ClickhereTermsofuse:Clickhere</p><p>Developmentandapplicationofadelayedreleaseanthelminticintraruminalbolussystemforexperimentalmanipulationofnematodewormburdens</p><p>ANJAM.CARLSSON,KENNETHWILSONandR.JUSTINIRVINE</p><p>Parasitology/Volume139/Issue08/July2012,pp10861092DOI:10.1017/S0031182012000406,Publishedonline:15March2012</p><p>Linktothisarticle:</p><p>Howtocitethisarticle:ANJAM.CARLSSON,KENNETHWILSONandR.JUSTINIRVINE(2012).Developmentandapplicationofadelayedreleaseanthelminticintraruminalbolussystemforexperimentalmanipulationofnematodewormburdens.Parasitology,139,pp10861092doi:10.1017/S0031182012000406</p><p>RequestPermissions:Clickhere</p><p>Downloadedfrom,IPaddress:</p></li><li><p>Development and application of a delayed-releaseanthelmintic intra-ruminal bolus system for experimentalmanipulation of nematode worm burdens</p><p>ANJA M. CARLSSON1,2,3*, KENNETH WILSON1 and R. JUSTIN IRVINE2</p><p>1Lancaster Environment Centre, Lancaster University, Lancaster LA1 4YQ, UK2The James Hutton Institute, Craigiebuckler, Aberdeen AB15 8QH, UK3Department of Arctic Biology, University Centre in Svalbard, PO Box 156, NO-9171 Longyearbyen, Norway</p><p>(Received 7 December 2011; revised 9 February 2012; accepted 12 February 2012; first published online 15 March 2012)</p><p>SUMMARY</p><p>In order to quantify the impact of parasites on host population dynamics, experimental manipulations that perturb theparasite-host relationship are needed but, logistically, this is difficult for wild hosts. Here, we describe the use of a delayed-release anthelmintic delivery system that can be administered when the hosts can be captured and its activity delayed until amore appropriate period in the host-parasite cycle. Our model system is Svalbard reindeer infected with a nematodeparasite, Marshallagia marshalli, which appears to accumulate during the Arctic winter. To determine the extent to whichthis occurs and the effect on host fitness, reindeer need to be treated with anthelmintics in late autumn but they can only becaught and handled in April. To solve this problem, we devised an intra-ruminal capsule that releases the anthelmintic fromup to 6 months after being administered. The capsule was trialed in cannulated sheep and red deer to determine optimumcapsule orifice size and release rates. Capsules were estimated to release placebo for 100153 days followed by abamectin for2234 days. To test the efficacy of treatment in reindeer, capsules were administered in April and retrieved in October. Allcapsules had fully released the anthelmintic and treated reindeer had significantly lower worm burdens than controls. Thus,success of this system allows repeated treatment over several years to test the effect of winter parasitism on host fitness.</p><p>Key words: anthelmintic experiment, nematode, ruminant, parasite-host interaction.</p><p>INTRODUCTION</p><p>Theoretical studies have shown that parasites havethe potential to regulate host abundance througheffects on reproduction and survival of the host(Anderson and May, 1978; May and Anderson,1978). In spite of the increasing recognition of theimportance of parasites in population ecology andwildlife conservation, knowledge of host-parasiteinteractions in the wild remains limited (Smithet al. 2009). In domesticated livestock it is well rec-ognized that gastrointestinal nematodes cause sig-nificant disease through negative effects on growthand survival of the host, resulting in major economiclosses (Perry and Randolph, 1999). Consequently,the ecology of parasites in agricultural systems hasbeen studied in detail in order to understand howbestto control and limit disease (Alison, 2003; OConnoret al. 2006). Traditionally, the study of parasites inwild animals has relied on making assumptions basedon domesticated animals. However, the epidemiol-ogy of host-parasite dynamics in thewild is often verydifferent. At best, cross-sectional studies of carcassesand fecal egg counts can be carried out (Morgan et al.</p><p>2005; Newey et al. 2005; Irvine et al. 2006); however,the consequences of parasitism are difficult to deter-mine through these types of studies (Gulland, 1995;Stien et al. 2002; Wilson et al. 2002). To investigatethe effects of parasitism on the host, experiments thatperturb the host-parasite relationship are essential(Stien et al. 2002). One of the major constraintsin conducting such experiments is the logisticaldifficulties associated with accessing, treating andhandling wild animals.</p><p>Experimental perturbations of the parasite com-munities in wild animals generally rely on the use ofanthelmintics to reduce parasite burdens (Murrayet al. 1996; Irvine, 2000; Craig et al. 2009). However,the ability to reduce worm burdens experimentally islimited to the time when anthelmintics can beadministered and the duration of the effective treat-ment period. Usually, a single oral dosing or injectionis used to deliver the anthelmintic in thewild (Murrayet al. 1997; Hudson et al. 1998; Irvine, 2000; Neweyet al. 2005; Pedersen and Greives, 2008) and theperiod of efficacy is generally a maximum of 35weeks. However, slow-release intra-ruminal deviceshave been developed for livestock which deliver atherapeutic dose over a period of up to 100 days(Cardinal, 1997;VandammeandEllis, 2004) reducingboth labour and material costs in the control ofnematode parasites. These devices have advantages</p><p>* Corresponding author: Lancaster Environment Centre,Lancaster University, Lancaster LA1 4YQ, UK. Tel:01524 93406. E-mail:</p><p>1086</p><p>Parasitology (2012), 139, 10861092. Cambridge University Press 2012doi:10.1017/S0031182012000406</p></li><li><p>for wildlife research, allowing a single treatment toprovide a lengthy parasite-free period in animals thatare difficult to catch or can be caught only at specifictimes of the year.The application of slow-release capsules has been</p><p>successfully utilized on Soay sheep (Gulland et al.1993) and Svalbard reindeer (Rangifer tarandusplatyrhynchus) (Stien et al. 2002). In the latter case,these capsules were administered to reindeer in April,which reduced worm burdens for approximately3 months and confirmed that Ostertagia gruehneritransmission occurs in spring (Albon et al. 2002).Furthermore, treatment also demonstrated that therewas a parasite-mediated reduction in host fecunditythat was sufficient to regulate the populationdynamics of Svalbard reindeer (Albon et al. 2002).However, cross-sectional studies have also indicatedthat the other major nematode species of Svalbardreindeer (Marshallagia marshalli) is almost absentduring the summer but increases during winter,suggesting over-winter transmission, but the conse-quence of this infection on reindeer performancehas not yet been quantified (Irvine et al. 2000). Todetermine the transmission dynamics ofM.marshalli,and the impact of the parasite on the fecundity andsurvival of the host, an experiment is needed in whichreindeer are treated at the start of thewinter.However,logistical and environmental constraints make itimpossible to catch and treat animals at this time ofyear.In this study, we describe the adaptation and</p><p>application of a delayed-release anthelminitic deliv-ery system. Domestic sheep and farmed red deer(Cervus elaphus) were used as model systems, and theefficacy was tested against the abomasal nematodes ofSvalbard reindeer. The aim was to develop a devicethat can be administered in April when Svalbardreindeer can be most easily caught, but to delaydelivery of the anthelmintic until autumn severalmonths later. This would then allow quantification ofthe effect of winter parasitism on host fitnessparameters in a later study. We predicted thatSvalbard reindeer treated with the delayed-releasebolus would have significantly lower worm burdensin October compared to controls.</p><p>MATERIALS AND METHODS</p><p>Capsule design</p><p>Capsules used in this study were small slow-releaseintra-ruminal capsules (Argenta ManufacturingLimited, NZ) that comprise a plastic hollow cylindercapped at both ends containing up to 11 tablets(identical in size and mode of action to the capsulesused by Stien et al. (2002)). One end is closed andincludes a spring that pushes the tablets towards theorifice in the opposite end. The orifice allows rumenfluid to enter the capsule and sequentially dissolves</p><p>the tablets within the cylinder; the resulting gel ispushed out of the capsule at a rate determined by thesize of the orifice. Retention of the capsule in therumen is achieved by polymeric wings attached tothe cylinder. During administration, the wings areconstrained to the cylinder with water-soluble tape,subsequently the tape dissolves and the wings open,preventing regurgitation of the device. Each capsulewas fitted with a unique identifying number to ensuretraceability and linking the capsule to batch number,formulation code, orifice size, spring strength,plunger type, capsule type and date of manufacture.All capsules were administered orally using a capsuleapplicator gun.</p><p>Rumen-cannulated sheep</p><p>The trial was carried out using 3 rumen-cannulatedsheep held at the Macaulay Land Use ResearchInstitutes Glensaugh research farm, Aberdeenshire,Scotland (56.8N, 2.6E), in July 2005. The durationof release from slow-release capsules fitted withorifices of 40, 37 and 35 mmdiameter and thicknessof 2 mm was determined. Each capsule contained9 tablets of placebo (35% hydrated aluminiumsilicate) followed by 2 tablets containing 32%abamectin. Each sheep was fitted with 2 capsules ofeach size (total of 6 capsules per sheep). The capsuleswere attached to silicon tubing using cable ties andtethered to the cannula bung. The capsules wereretrieved from the sheep rumens on days 0, 24, 50, 78and 106 and the core length of each capsule wasmeasured using vernier calipers from the lower edgeof the plunger (coloured red to aid visibility) to theinner edge of the orifice plate. Not all capsules wererecovered at all sampling dates, as some were lostwhen they became detached from the silicon tubingthat tethered them to the cannula bung.Release rates, defined as the reduction in tablet size</p><p>(mm per day since capsule administration), werecalculated from the core-length measurements usinglinear mixed effect models from the lme package inR version 2.13.0 (R Development Core Team, 2011),with sheep identity and capsule as random effects,and days since initiation of experiment and orificesize as fixed effects. Model fit was assessed usingdiagnostic plots of fitted values and residuals. Log-transforming the response variable improved modelfit and linearized the data. However, estimates fromthis model were unrealistic, predicting periods ofrelease on a log-scale (&gt;460 days) well in excess ofthose observed (e.g. we know that capsules fromreindeer culled 6 months after treatments were fullydischarged).</p><p>Red deer</p><p>From trials in sheep it was determined that thesmallest orifice size was the most appropriate design</p><p>1087Development of a delayed-release anthelmintic delivery system</p></li><li><p>and the release rate of capsules was tested in red deer.The trial was carried out using red deer at theGlensaugh research farm (see above). Six yearling reddeer were each given 1 capsule with 11 placebo (35%hydrated aluminum silicate) tablets and orificediameter of 35 mm and thickness of 2 mm on days0, 36, and 63 of the experiment. The animals werekept under normal management until they wereslaughtered on day 100 of the experiment and allcapsules recovered from the rumen. At the end of theexperiment, the core-length of each capsule wasmeasured as described above. All capsules weresuccessfully retrieved and identified by the end ofthe trial. Release rate, as defined above, was predictedusing linear mixed effect models using the lmepackage in R version 2.13.0 (R Development CoreTeam, 2011) with deer identity as a random effect anddays since capsule administration until cull as a fixedeffect.</p><p>Svalbard reindeer</p><p>Svalbard reindeer are a wild population living on thehigh Arctic archipelago of Svalbard (Halvorsen andBye, 1999). Female Svalbard reindeer have beenmarked and caught annually in the Colesdalen-Reindalen valley system, Nordenskildland,Spitsbergen (77507820N, 15001730E) since1994, allowing treatment and recapture of knownindividuals (Milner et al. 2003). Between 1126 April2006 and 2007, 62 adult female reindeer (36 monthsor older) were randomly allocated for treatment andfitted with an intra-ruminal controlled-release cap-sule with an orifice diameter of 35 mm and thicknessof 2 mm. Each capsule contained 9 tablets of placeboand 2 tablets abamectin, in order to delay treatmentuntil autumn. In 2006 the concentration of abamectinin tablets was 32%. In their original design, theestimated dose rate with abamectin of this concen-tration was 23 g/kg/day; however, these figures werebased on sheep weighing 80 kg, using capsules withorifice diameter of 45 mm. The orifice diameter ofthe reindeer capsules was smaller, and it is thereforelikely that the abamectin dose rate was lower. Thefollowing year the abamectin dose was doubled (to6%) to compensate for the smaller orifice size in thedelayed release capsules, with estimated delivery rateof 45 g/kg/day. Administration of capsules wasperformed as outlined above. To assess the efficacyof the treatment, 16 control and 4 treated animalswere culled in late October 2006 and 2007 and theabomasums recovered 186194 days after adminis-tration. Stien et al. (2002) have previously reportedon an experiment where animals were treated withimmediate-acting anthelmintic drugs in April, usingivermectin capsules (Ivomec Maximizer, MerialAnimal Health Ltd, Harlow Essex, UK) and/orinjectable moxidectin, to reduce worm burdens inSvalbard reindeer during the summer.Todemonstrate</p><p>the difference in efficacy between the immediate, anddelayed anthelmintic drug treatment in removingnematode burdens in October, a subset of these datawas analysed. Abundance data of M. marshalli andO. gruheneri adult worms from 7 Svalbard reindeertreated with immediate-acting anthelmintics and10 non-treated Svalbard reindeer culled October1999 were used. For more details of these exper-iments see the report by Stein et al. (2002).Abomasums were removed from culled reindeerand adult nematodes were extracted and counted asdescribed by Halvorsen et al. (1999) and Irvine et al.(2000). A subset of adult male nematodes wasidentified to species using morphology (Drozdz,1965), with minor and major morphs groupedaccording to recent molecular findings (Dallas et al.2000, 2001). Adult nematode abundance for eachspecies was estimated using the proportion ofidentified worms and scaling up to the populationlevel based on the worm counts. The effect of the 3different treatments (control, immediate-acting an-thelmintic and delayed-release anthelmintic capsule)on adult nematode worm burden was analysed usinggenera...</p></li></ul>


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