Lamb eimeriosis: Applied treatment protocols in dairy sheep production systems

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Veterinary Parasitology 196 (2013) 56– 63

Contents lists available at SciVerse ScienceDirect

Veterinary Parasitology

jou rn al h om epa ge: www.elsev ier .com/ locate /vetpar

amb eimeriosis: Applied treatment protocols in dairy sheeproduction systems

nastasios Saratsisa,b, Isidoros Karagiannisc, Christos Brozosc, Evagellosiossis c, Nikolaos Tzanidakisa,c, Anja Joachimb, Smaragda Sotirakia,∗

Veterinary Research Institute, Hellenic Agricultural Organisation-Demeter (former NAGREF), NAGREF Campus, PO Box 60272, 57001hermi, GreeceInstitute of Parasitology, Department of Pathobiology, University of Veterinary Medicine Vienna, Veterinaerplatz 1, 1210 Wien, AustriaClinic of Farm Animals, Veterinary School, Aristotle University, 54627 Thessaloniki, Greece

a r t i c l e i n f o

rticle history:eceived 7 August 2012eceived in revised form 19 January 2013ccepted 24 January 2013

eywords:imeriaamboccidiosisiclazuriloltrazurilreatment

a b s t r a c t

The effect of different treatment protocols using the triazinone compounds diclazuril andtoltrazuril on Eimeria oocyst excretion, diarrhoea and weight gain was evaluated in thepresent study. The protocols were designed in order to best fit to common managementpractices in dairy production systems as applied in Greece. During the first two trialscomparative preventive (11 days after birth) and therapeutic (18 days after birth) singletreatments using either drug were performed on an intensive farm. In Trial 3 the efficacyof a repeated diclazuril treatment after weaning (at the start and 4 weeks later) of the ani-mals in a semi-intensive farm was tested. The last trial was performed in order to assess theeffect of a single oral dose of toltrazuril after weaning of the animals on the same farm. Dur-ing an observation period of 6–7 weeks after treatment animals in all trials were clinicallyexamined for diarrhoea and faecal samples were regularly monitored for Eimeria oocystsat weekly intervals. Body weight was also determined at the start and end of each trial. A

single treatment with toltrazuril resulted in a significant reduction of oocyst excretion overthe study period compared to the control, with very high efficacy values during the first2–3 weeks after treatment, irrespective of the treatment protocol and the managementsystem applied. This in general could not be confirmed in the protocols using diclazuril,which showed a much lower and mostly variable efficacy pattern.

. Introduction

By invading and destroying the intestinal epithelialells coccidia of the genus Eimeria are a major contributoro enteric disease, especially in intensively reared lambs.f the 15 species described to infect sheep worldwide

Pellérdy, 1974), Eimeria crandallis and E. ovinoidallis arehe most pathogenic (Taylor et al., 2007).

∗ Corresponding author. Tel.: +30 2310365373; fax: +30 2310365371.E-mail addresses: smaro [email protected], [email protected]

S. Sotiraki).

304-4017/$ – see front matter © 2013 Elsevier B.V. All rights reserved.ttp://dx.doi.org/10.1016/j.vetpar.2013.01.020

© 2013 Elsevier B.V. All rights reserved.

Typically, lambs between 3 and 8 weeks of age are atparticular risk of getting infected (Taylor and Catchpole,1994). In heavily infected lambs the mucosa becomescompletely denuded, resulting in severe haemorrhage andimpaired water reabsorption, leading to diarrhoea, dehy-dration and occasionally death (Taylor et al., 2007).

Animals acquire immunity as a result of infection, whichprotects against clinical disease and which is maintainedby repeated exposure to oocysts. However, immunity doesnot lead to total prevention of parasite shedding (Catchpole

et al., 1993).

Resistance to coccidial infection can be reduced bystress factors such as weaning, high stocking densi-ties, multiple lambing with lower colostrum/milk intake,

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A. Saratsis et al. / Veterinary Parasitology 196 (2013) 56– 63 57

Table 1Study design.

Trial Farm Managementsystem

Age at the startof trial (days)

Treated with Age at treatment(days)

Treatment

11 Intensive

11Diclazuril

11 Before oocyst excretionToltrazuril

2 18Diclazuril

18 At the beginning of oocyst excretionToltrazuril

32 Semi-intensive

27 Diclazuril

nt)

30 and 58 At the beginning of oocyst excretion

ril

(doubletreatme

4 45 Toltrazu

adverse conditions such as dietary changes, prolongedtransport, rapid weather changes, or concurrent infections(Gregory, 1990; Catchpole et al., 1993; Taylor, 1995; Alzieuet al., 1999). When lambs are weaned during the first monthof their life and then reared indoors, as in Mediterraneandairy systems, coccidiosis is seen 2–3 weeks post-weaning(Yvoré and Esnault, 1987; Saratsis et al., 2011).

Although hygienic measures can in principle reduceinfection pressure, they are difficult to implement once sus-ceptible animals are housed together, especially indoorswhere oocysts rapidly accumulate. Therefore, control withanticoccidial drugs is frequently necessary to preventclinical outbreaks and production losses. Toltrazuril anddiclazuril have previously been applied and evaluated forthis purpose (Gjerde and Helle, 1986, 1991; Taylor andKenny, 1988; Le Sueur et al., 2009; Platzer et al., 2005;Mundt et al., 2009; Taylor et al., 2011). The majority of thestudies dealing with the efficacy of chemical treatmentshave so far mainly focused on lamb fattening systems,either pasture or indoor based (Gjerde and Helle, 1986,1991; Taylor and Kenny, 1988; Alzieu et al., 1999; Mundtet al., 2009; Taylor et al., 2011). The purpose of the presentstudy was to determine the effects of chemical treatmentagainst coccidiosis in lambs kept under dairy husbandrysystems applied in the Southern Mediterranean region. Tri-als included several possible application schemes under themost common practices and were performed in lambs nat-urally infected with Eimeria spp., comparing the effect oftreated to untreated control groups on clinical and para-sitological parameters.

2. Materials and methods

2.1. Study animals and management systems

Trials were performed on farms where the presenceof Eimeria was previously confirmed to ensure naturalinfection. Earlier studies in the area showed that animalsacquire infection within the first 3 weeks after birth witha prevalence of 61.7%. The overall cumulative prevalenceof E. ovinoidallis and E. crandallis from birth until wean-ing ranged between 20.7% and 47.1%, depending on thelambing period (Saratsis et al., 2011).

In dairy husbandry two different systems in terms of

lamb management exist; the intensive system, where thelambs are separated from their dams just after birth andreared on milk supplement, and the more common semi-intensive system where the lambs stay with their mothers

45 At the beginning of oocyst excretion

mostly indoors and are weaned at 4–8 weeks after birth(Zervas et al., 1999; Zygoyiannis et al., 1999; Stefanakiset al., 2007; Volanis et al., 2007). Due to the more indus-trial farming type, stocking densities are almost alwaysmuch higher in the intensively managed farms than in thesemi-intensive ones. The trials in the current study wereperformed on two commercial farms belonging to eitherone of the mentioned systems as follows:

Farm 1 applied an intensive management system withlambings all year round (every two months). Lambs wereseparated from their dams one day after birth and offereda commercially available milk replacer. The farm had intotal 400 ewes of the local Chios breed. The animals werekept on straw.Farm 2 applied a semi-intensive husbandry system, withlambs reared with the ewes for 4–6 weeks before beingweaned. Lambing took place twice a year, which is a com-mon practice for the majority of such farms. The farm hadin total 300 ewes of the local Sfakia breed and the animalswere also kept on straw.

No anticoccidials were used on these farms for at leastone year before our first visit.

2.2. Study design

Efficacy of treatment with toltrazuril (BAYCOX® 5% sus-pension, Bayer Animal Health) and diclazuril (VECOXAN®

2.5% suspension, Elanco) was assessed on both farms, asdescribed below. Both products were orally administeredaccording to the manufacturer’s recommendations (doserate of 20 mg/kg for BAYCOX® and 1 mg/kg for VECOXAN®).Animals were randomly assigned to the respective exper-imental group before the start of each trial based on theirsex, weight and presence of twins. An additional group ofanimals receiving no treatment served as a negative con-trol. In each trial animals were kept per group in a differentpen. Animals were weighed at the start and the end of eachtrial (Table 1).

The study design included separate trials per farmfor which different protocols were strategically plannedaccording to the management system.

2.2.1. Farm 1: intensive managementUnder such a management system, where the expected

infection pressure was high, two strategic treatmentswere designed; one to treat the lambs during prepatency

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8 A. Saratsis et al. / Veterina

prevention) and the other to treat lambs just after thenset of oocyst excretion (therapy) in the flock. Each ofhose treatment protocols was conducted in a separate trialt two different lambing periods.

.2.1.1. Trial 1: single preventive treatment. In total, 65ambs were enrolled in this trial, allocated to 3 exper-mental groups; 23 to the negative control, 20 to theoltrazuril-treated and 22 to the diclazuril-treated group.he average age of the lambs on treatment was 11 ± 2ays and the treatment (study day 1, SD1) was performedfter confirming that no animal was excreting Eimeriaocysts with two consecutive faecal examinations at days

and 9 after birth. From SD1 onwards faecal samples wereollected for 7 times at weekly intervals (SD1–SD7) and dif-erentiation of sporulated oocysts took place on SD2, SD4nd SD7.

.2.1.2. Trial 2: single treatment at the start of excretion.n all, 36 lambs were enrolled in this trial, allocated to 3xperimental groups, 12 to the negative control, 13 to theoltrazuril-treated and 11 to the diclazuril-treated group.he average age of the lambs on treatment was 18 ± 3 daysnd treatment (SD1) was performed after confirming thenset of Eimeria oocyst excretion in the flock (41.3% of thenimals were excreting oocysts on treatment day). FromD1 onwards faecal samples were collected for 8 times ateekly intervals (SD1–SD8) and differentiation of sporu-

ated oocysts took place on SD2, SD4 and SD7.

.2.2. Farm 2: semi intensive managementUnder such a management system the lambs are com-

ng in close contact with each other only after weaning,hen they are separated from their mothers and moved tootentially infected pens. The treatment design under suchonditions was to treat animals just after weaning. The twoifferent anticoccidials were evaluated in separated trialss follows.

.2.2.1. Trial 3: double therapeutic treatment with diclazurilfter weaning. Twenty-four lambs just after weaning werenrolled in this trial and allocated to two experimentalroups, 12 to the negative control, and 12 to the diclazuril-reated group. The age of the lambs at the beginning ofhe trial was 30 ± 2 days and treatment was applied at theeginning (SD1) and 4 weeks later (SD5). On SD1 95.8% ofhe animals were excreting Eimeria oocysts. Faecal exam-nations were performed on a weekly basis starting fromD1 for 7 weeks (SD8) and differentiation of sporulatedocysts took place on SD3, SD5 and SD8.

.2.2.2. Trial 4: single therapeutic treatment with toltrazurilfter weaning. Thirty-two lambs just after weaning werenrolled and allocated to two experimental groups, 16 tohe negative control and 16 to the toltrazuril-treated group.he average age of the lambs at treatment (SD1) was 45 ± 3ays. On SD1 100% of the animals were excreting Eimeria

ocysts. Faecal examinations were performed on a weeklyasis starting from SD1 for 6 weeks (SD7) and differen-iation of sporulated oocysts took place on SD3, SD5 andD7.

itology 196 (2013) 56– 63

2.3. Parasitological methods

On each sampling date and for each sample faecal con-sistency score and oocyst excretion (expressed as oocystsper gram of faeces, OPG) were recorded. Faecal sampleswere scored based on a scale of 1–4 as 1, solid; 2, pasty;3, liquid diarrhoea and 4, bloody diarrhoea. The oocystquantification was performed by using a modified McMas-ter method with saturated sodium chloride solution plus500 g glucose per litre as flotation solution (Henriksen andChristensen, 1992; Meyer et al., 1999). In addition, Eimeriaoocysts were concentrated from individual faecal samples,on the above-mentioned selected sampling days, and iden-tified to species level based on their morphology. In detail,oocysts were concentrated from samples having OPG val-ues greater than 1000 and were sporulated in 2% potassiumdichromate solution. Differentiation was performed for100 sporulated oocysts per sample (Eckert et al., 1995;Saratsis et al., 2011).

2.4. Statistical analysis

Data were analysed by using the SPSS statistics soft-ware (version 19.0). The effect of treatment on OPG valueswas analysed by using the linear mixed-effects model(dependent variable: OPG; categorical predictor variable:treatment group) (Laird and Ware, 1982). The covariance‘structure’ for Trials 1, 2, 3 and 4 was first order autoregres-sive, correlation compound symmetry, ante-dependencefirst order and first order autoregressive, respectively. Theadjustment of p values between multiple comparisons wasdone using Sidak correction. Area under the curve (AUC)values were calculated by using the Graphpad Prism© soft-ware (version 5.01). AUC values were compared by eitherusing one-way analysis of variance (ANOVA) or t-tests forindependent samples. The same tests were also appliedfor the comparison of weight gain between the respectivegroups. The relative oocyst excretion reduction was calcu-lated based on the formula C − T/C × 100 (C = mean controlOPG values, T = mean treatment OPG values). Comparisonsbetween the mean oocyst reduction curves distributions ofboth drugs during Trials 1 and 2 were performed by usingthe non-parametric Mann–Whitney test.

3. Results

3.1. Efficacy

The mean oocyst reduction for each treatment duringthe course of each trial is shown in Fig. 1.

During Trial 1 the mean oocyst reduction aftertoltrazuril treatment was 100% and 99% for 1 (SD2) and 2weeks (SD3) after treatment, respectively, and then rangedfrom 86.5% to 71.9% until the end of the trial. The meanoocyst reduction after diclazuril treatment for weeks 4(SD5), 5 (SD6) and 6 (SD7) after treatment was 37.8%, 80%and 71.5%, respectively. For the period up to week 4 (SD5)

there was no reduction in oocyst output after diclazuriltreatment. The efficacy values between the comparativetreatment groups were statistically significantly different(p = 0.017).

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2 3 4 5 6 7

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020304050609092949698

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a. Trial 1 b. Trial 2

2 3 4 5 6 7 8

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atment

c. Trial 3

Fig. 1. % Eimeria oocyst reduction per tre

During Trial 2 the mean oocyst reduction aftertoltrazuril treatment was 99.8% and 99.7% for 1 (SD2) and 2weeks (SD3) after treatment respectively, 96.2% and 92.7%for weeks 3 (SD4) and 4 (SD5) after treatment droppinglater to 59.3%. The effect of the diclazuril treatment onoocyst excretion reduction was variable showing a signifi-cant oocyst reduction of 96.9% only during the second weekafter treatment (SD3). The overall comparison of the curvesdid not show any significant differences.

During Trial 3 the mean oocyst reduction after diclazuriltreatment was 92.6% 1 week (SD2) after the first treat-ment decreasing to 65% a week later and 95% 1 week (SD6)after the second treatment decreasing to 22.9% a week later(SD7). There was no effect on oocyst excretion during theother sampling dates.

During Trial 4 mean oocyst reduction after toltrazuriltreatment was 96.2% and 95.6% for 1 (SD2) and 2 weeksafter treatment (SD3), respectively, and 78% and 45.2% forweeks 3 (SD4) and 4 (SD5) after treatment.

3.2. Effects on oocyst excretion

During Trial 1 Eimeria oocysts were first detected in thefaeces of the control group on SD2 (lamb age: 18 days ± 2).The excretion rate on SD2 was 91.3% peaking to 100%on SD4. Animals were also infected in the two treatmentgroups reaching an excretion rate of 100% on SD6 and SD7for the diclazuril- and toltrazuril-treated groups, respec-tively. The onset of oocyst excretion was delayed for 1 weekcompared to the control in the toltrazuril-treated group,remaining low on SD3 and SD4 (mean OPG values of ≤600).In the diclazuril-treated group the onset of oocyst excretionwas the same as in the control group. The difference in OPGsbetween the three different treatment groups is shown in

Fig. 2a.

Comparison of logarithmic AUC (lnAUC) values revealedthat toltrazuril treatment resulted in a significant reduc-tion (p < 0.01; Fig. 2) of the total number of excreted

d. Trial 4

over time (study days – SD) in each trial.

oocysts compared to both untreated and diclazuril-treatedlambs.

The linear mixed effects model showed the effect oftreatment on OPG values. The estimates for the modelrevealed that the effect of the toltrazuril treatment onoocyst excretion (estimate: −44,629.6) was significant(p = 0.002), whereas the effect of diclazuril treatment (esti-mate: −23,537.5) was not.

During Trial 2 Eimeria oocysts were detected in the fae-ces of 10 out of 12 lambs in the control group (83.3%) onSD2 (average age: 25 days) and in 100% on SD4. A 100%cumulative incidence of oocyst excretion was reached onSD4 and SD5 in the diclazuril and toltrazuril treated groups,respectively. The level of oocyst excretion in the toltrazuril-treated group was close to zero for weeks 1 (SD2) and 2(SD3) after treatment, remaining also low for weeks 3 (SD4)and 4 (SD5) after treatment (mean OPG values of ≤9800). Asfar as the diclazuril-treated group was concerned, 81.8% ofthe animals excreted oocysts 1 week after treatment (SD2),which resulted in higher mean OPG values compared to thecontrol. The differences in mean OPG between the threedifferent treatment groups are shown in Fig. 2b.

The lowest mean AUC value (SD1–SD7) was observedafter toltrazuril treatment, which significantly differedcompared to both the control (p = 0.004) and the diclazuril-treated group (p = 0.014; Fig. 2).

The estimates for the linear mixed model revealed thatthe effect of toltrazuril treatment on the overall excretionwas significant (p = 0.014), whereas the effect of diclazuriltreatment was not. Toltrazuril treatment compared to thecontrol had a mean difference in OPG value of −44,326.0which was statistically significant (p = 0.04).

During Trial 3 all animals except one excreted oocystson SD1. The differences in mean OPG between the threedifferent treatment groups are shown in Fig. 2c.

The mean AUC values between the two groups dif-fered reaching borderline significance (p = 0.065) and werereduced from 205,488 (control) to 137,419 (diclazuril-treated group).

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0 1 2 3 4 5 6 70

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100,000

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diclazuril

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PG

0 1 2 3 4 5 6 7 80

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a. Trial 1 b. Trial 2

Trial 2 SD1 SD2 SD3 SD4 SD5 SD6 SD7 SD8 Mean AUC

Control 24.4(79.9) 76.3(112.2) 27.1(41.5) 99.7(103.5) 140.4( 214.8) 51.5(49.8) 161.1( 245.8) 64.0 (7 4.8) 600.7a

Toltra zuril 16.0(29.8) 0.1( 0.1) 0.0 7(0.1) 3.7( 8.3) 9.8( 12.5) 20.9(25.4) 126.6 (1 66.9) 126.5 (1 46.3) 228.3b

Diclazuril 1. 0(23.3) 180.2(303.5) 0.8(2.5) 102.3(245.5) 55.5(99.1) 150.4(170.8) 105.7 (90.8) 37.7 (25.2) 614.5a

0 1 2 3 4 5 6 7 80

20,000

40,000

60,000control

diclazuril

SD

mean

OP

G

0 1 2 3 4 5 6 70

20,000

40,000

60,000control

tolt raz uril

SD

mean

OP

G

c. Trial 3 d. Tr ial 4

Trial 3 SD1 SD2 SD3 SD4 SD5 SD6 SD7 SD8 Mean AUC

Control 12.1(20.3) 41.8(33.3) 55.7(82.1) 25.0(21.9) 21.9( 14.5) 35.5( 43.1) 11.3( 11.2) 16.0 (1 0.8) 205.4a

Diclazuril 6.3(11.8) 3.0( 6.4) 19.4(27.8) 43.9(77.9) 24.3( 28.2) 1.7( 3.9) 8.7( 5.7) 65.7(78.8) 137.4a

Trial 4 SD1 SD2 SD3 SD4 SD5 SD6 SD7 Mean AUC

Control 8.7(10.9) 12.2(10.1) 57.2(58.8) 60.6(68.5) 40.3(48.1) 6.3(10.1) 5.7(5.2) 185.0a

Toltra zuril 9.6 (7.8) 0.4(1.0) 2.5(3.5) 13.3(16.4) 22.1(22.6) 18.3(16.2) 6.5(4.9) 64.8b

Trial 1 SD1 SD2 SD3 SD4 SD5 SD6 SD7 Mean AUC

Control 0 (0) 62.6 (129.2) 1.4 (4.8) 2.4 (5.1) 42.8 (9 2.8) 108.3 (2 61.2) 150.2 (3 27.3) 292.9a

Toltrazuril 0 (0) 0 (0) 0.01(0.03) 0.6 (2.6) 5.7 (11.4) 18.0 (24.6) 31.9 (36.3) 38.2b

Diclazuril 0 (0) 103.1 (2 18.2) 8.3 (2 4.2) 2.3 (8.2) 26.6 (8 0.3) 21.7 (3 5.8) 42.8 (5 2.0) 183.6a

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ig. 2. Oocyst excretion patterns (mean OPG values) per trial and treatmean AUC values for the whole experimental period are also given in the

ifferences (p < 0.05).

The estimate for the linear mixed model revealed thathe effect of diclazuril treatment on OPG was significantestimate: −6875.7; p = 0.019). Compared to the controlhe diclazuril treatment group had a mean difference inPG value of −6875.7 which is statistically significant

p = 0.019).During Trial 4 the lambs in the control group were also

ighly infected showing maximum oocyst excretion lev-ls on SD3 and SD4. The level of oocyst excretion for theoltrazuril treatment group was constantly lower than thatf the control group up to SD6. The difference in mean OPGetween the two different treatment groups is shown in

ig. 2d.

A significant reduction (−64.9%; p = 0.003) could also behown for the mean AUC values, which were reduced from85,058 (control) to 64,892 (toltrazuril-treated).

p. Exact OPG values (×10 ), standard deviations (×10 ; in brackets) andtive table. Different superscripts in the same column indicate significant

The estimates for the linear mixed model revealedthat the effect of toltrazuril treatment was significant(p = 0.011). Toltrazuril treatment has a mean difference inOPG value of −14,861.4 compared to the control which isstatistically significant (p = 0.011).

3.3. Eimeria species

The species of Eimeria/trial/treatment identified areshown in details in Table 2.

Throughout Trial 1, E. ovinoidalis and E. crandallis werethe most common species identified in control samples

(67.2%). The estimated prevalences for the diclazuril andtoltrazuril treated groups were 55% and 20.2%, respectively.One week after treatment (SD2) in the diclazuril-treatedgroup, 56.8% and 28% of the differentiated oocysts belonged

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Table 2Percentage (pooled data considering SDs at which speciation was per-formed) of Eimeria species per trial and group.

Trial Group E. ovinoidalis E. crandallis Others

1Control 32.9 34.3 32.8Toltrazuril 9.2 11.0 79.8Diclazuril 35.1 19.9 45.0

2Control 32.6 13.7 53.7Toltrazuril 13.7 2.1 84.2Diclazuril 29.6 16.6 53.8

3Control 5.1 1.7 93.2Diclazuril 2.0 1.6 96.4

4Control 6.7 9.0 84.3Toltrazuril 7.4 9.4 83.2

to E. ovinoidalis and E. crandallis, whereas in the controlgroup the respective values where 46.4% and 45.1%. Noanimals excreted oocysts in the toltrazuril-treated groupat that time.

During the whole duration of Trial 2 E. ovinoidalisand E. crandallis were also found in high proportions ofspeciated oocysts in the control (46.3%), and the diclazuril-treated group (46.2%) and in a lower proportion in thetoltrazuril-treated animals (15.8%). On SD2 E. ovinoidalisand E. crandallis accounted for 97.5% of the speciatedoocysts in the diclazuril-treated group.

In Trials 3 and 4, E. ovinoidalis and E. crandallis werealso present but in much lower proportions. Other non-pathogenic species such as E. parva, E. bakuensis, E. ahsataand E. weybridgensis were dominating.

3.4. Weight gain

The total weight gain for the control, diclazuril-treatedand toltrazuril-treated groups respectively was 7.4 kg (SD2.0), 7.0 kg (SD 1.8) and 7.8 kg (SD 2.0) for Trial 1 and 10.5 kg(SD 2.2) 11.0 kg (SD 2.7) 11.2 kg (SD 2.1) for Trial 2, showingno significant difference between groups.

In Trial 3 the total weight gain was 4.3 kg (SD 1.9) for thecontrol and 5.6 kg (SD 1.0) for the diclazuril-treated group.In this trial there was a significant difference (p = 0.05)between groups. In Trial 4, the weight gain was 6.7 kg (SD1.7) for the control and 6.5 kg (SD 2.6) for the toltrazuril-treated group.

3.5. Diarrhoea

During Trial 1, 45% of the lambs for the diclazuril-treatedgroup and 45.5% of the lambs for the toltrazuril-treatedgroup did not show signs of diarrhoea (faecal scores 3 or4). In the control group the percentage of animals withoutdiarrhoea was 21.7%. The comparison between treatmentgroups and the control for the presence of diarrhoea duringthis trial revealed no significant differences (�2; p = 0.172).

Diarrhoea was more evident during Trial 2 where only9.1% of the lambs in the diclazuril-treated group and 15.4%

of the lambs in the toltrazuril-treated group did not showsigns of diarrhoea until the end of the trial. In the controlgroup only 8.3% of the animals did not show any sign ofdiarrhoea. The comparison between treatment groups and

itology 196 (2013) 56– 63 61

the control for at least 1 record of diarrhoea during this trialrevealed no significant differences (�2; p = 0.827).

No statistical analysis was performed for Trials 3 and 4,due to the low incidence of diarrhoea, which did not exceed2% for any of the experimental groups.

4. Discussion

During all trials animals were naturally infected withEimeria, more heavily on the intensively managed farm(Trials 1 and 2), confirming the high abundance of thoseparasites in lambs as shown previously, especially onindoor reared lambs (Le Sueur et al., 2009; Mundt et al.,2009; Dittmar et al., 2010; Saratsis et al., 2011).

Several studies have previously demonstrated the pos-itive impact of toltrazuril and diclazuril treatments onEimeria infections in lambs under various field conditions(Platzer et al., 2005; Le Sueur et al., 2009; Mundt et al.,2009; Chartier and Paraud, 2012). In the present paper,the focus was to evaluate the application protocols of suchtreatments in practice under common dairy husbandry sys-tems.

The application of the above drugs when employedin intensive dairy systems (Trials 1 and 2) showed thattoltrazuril treatment was more efficient demonstratinga more immediate effect after application, and a moreconcise efficacy pattern compared to diclazuril treatmentwhich had a delayed effect showing efficacy above 95% onlyonce 2 weeks after treatment in Trial 2.

The better and longer lasting efficacy of toltrazuril overdiclazuril in sheep coccidiosis is in agreement with previ-ous surveys (Mundt et al., 2009; Le Sueur et al., 2009) and ispossibly due to differences in the pharmacokinetic proper-ties (e.g. totlrazuril has a much longer elimination half-lifein plasma) and the spectrum of activity against the vari-ous parasitic stages of Eimeria (Haberkorn and Stoltefuss,1987; Maes et al., 1989; Taylor et al., 2003). However, therecorded delayed response of diclazuril treatment was notexpected since, according to the literature, high efficacywas commonly evident during the first week after treat-ment (Alzieu et al., 1999; Taylor et al., 2011). This could bepossibly attributed to the high infection pressure recordedin these trials or any other concurrent infection that mighthave affected its desorption in the gut which, in any case,is already low in lambs (Scholtysik and Steuber, 2002;Steuber and Kroker, 2003).

Comparing the post-treatment effect we observed thatin Trial 1 toltrazuril delayed the onset of excretion foralmost 2 weeks (efficacy rate 100% and 98.98% for 1 and2 weeks after treatment, respectively). When toltrazurilwas administered just after oocyst excretion onset theeffect was even more evident, from almost no excretionfor 2 weeks after treatment (efficacy 99.85% and 99.73%)to very low excretion rates for the following 2 weeks (effi-cacy 96.24% and 92.96%). Similar conclusions can be drawnwhen comparing the effect of the two drugs on oocystexcretion.

As regard the different administration time points oftoltrazuril it seems that both had a similar effect onoocyst excretion reduction rates, which makes both treat-ment protocols equally effective. This is consistent with

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revious findings (Mundt et al., 2009; Le Sueur et al., 2009).owever, when drug admission is made after the onsetf oocyst excretion, environmental contamination and theubsequent risk of re-infection is still high and this is whatauses oocyst shedding at a later stage when the treatmentfficacy declines (Gjerde and Helle, 1986; Le Sueur et al.,009).

It should be noted that in all groups and both trials theroportion of pathogenic species declined at the end of therial (SD7–SD8), possibly due to the developing immunity.his was already shown to also occur after administrationf either drug (Greif, 2000; Mundt et al., 2009; Taylor et al.,011).

The precise proportion of diarrhoea attributable to coc-idia was not determined, since this could only have beenone if specific diagnosis of all other gut pathogens hadeen carried out for all faecal samples. In any case from ourbservations, it seemed that diarrhoea scores for Trials 1nd 2 improved with treatment, regardless of the admin-stered medications; although this speculation remainso be proven since the presence of diarrhoea could beue to various other reasons than coccidiosis. In casef more severe diarrhoea, such as in Trial 2, toltrazurilhowed a better effect against the appearance of diarrhoeahan diclazuril, even though, again, not statistically signif-cant.

When treatment was applied in older animals underemi-intensive management practices (Trials 3 and 4),gain the overall anticoccidial effect of toltrazuril was moreoticeable than that of diclazuril even when the latter wasdministered twice. The effect of diclazuril treatment wasostly limited to the first week after treatment. When

he oocyst reduction was taken into account it seemedhat both treatments significantly reduced the total oocystsxcreted, thus reducing the environmental contamination.he infection severity was much lower on Farm 2 ando clinical diarrhoea was present. The group of animalsreated with diclazuril in Trial 3 showed a significantlyigher mean weight gain compared to the control as haseen shown previously (Alzieu et al., 1999; Platzer et al.,005). However, this was the only time for all treat-ents applied where a significant difference in weight

ain was recorded, the reasons for which we are unableo explain.

To conclude, the results of the current study show thatn all protocols used, under different management systems,he effect of toltrazuril against Eimeria spp. in lambs wasuperior to that of diclazuril. Moreover, in cases of highnfection pressure, if the treatment is applied before thenset of excretion and hygiene is appropriate, the efficacyf preventive treatment can be optimised.

cknowledgements

We acknowledge the excellent assistance of Dr Alexan-

ros Stefanakis, Mr Nikolaos Voutzourakis and Mr Nikolaoshristodoulakis who significantly contributed during sam-le collection. Moreover we especially thank the farmersor participating in our study.

itology 196 (2013) 56– 63

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