The efficacy and pharmacokinetics of long-term low-level intraruminal administration of tricalbendazole in buffalo with induced fasciolosis

A study was conducted on the pharmacokinetics and therapeutic efficacy of triclabendazole at three low dose rates of 0.5, 1.0 and 1.5 mg/kg body weight in buffaloes experimentally infected with Fasciola gigantica. The pharmacokinetics were compared with the effects of a single intraruminal dose at 24.0 mg/kg body weight in uninfected buffaloes. At all three dose rates, an equilibrium between the absorption of triclabendazole and the disposition of its metabolites was observed by days 3 and 4 and remained almost unchanged thereafter. Continuous daily dosing at 1.5 mg/kg body weight proved to be efficacious against liver fluke infection in buffaloes.Abbreviations TCBZ triclabendazole - p.i. post-infection - HPLC high-performance liquid chromatography - TCBZ-SO triclabendazole sulphoxide - TCBZ-SO2 triclabendazole sulphone - C _max peak concentration in plasma - T _max time to reach C _max - AUC area under the concentration-time curve - t _1/2 elimination half-life - epg eggs per gram     

Assessment of the main metabolism pathways for the flukicidal compound triclabendazole in sheep

Triclabendazole (TCBZ) is an halogenated benzimidazole (BZD) compound worldwide used to control immature and adult stages of the liver fluke Fasciola hepatica. The purpose of this investigation was to characterize in vitro the patterns of hepatic and ruminal biotransformation of TCBZ and its metabolites in sheep. TCBZ parent drug was metabolized into its sulphoxide (TCBZSO), sulphone (TCBZSO2) and hydroxy derivatives by sheep liver microsomes. The same microsomal fraction was also able to oxidize TCBZSO into TCBZSO2 and hydroxy-TCBZSO (HO-TCBZSO). TCBZ sulphoxidation was significantly (P < 0.001) inhibited after inactivation of the flavin-monooxygenase (FMO) system (77% inhibition) as well as in the presence of the FMO substrate methimazole (MTZ) (71% inhibition). TCBZ sulphoxidative metabolism was also reduced (24% inhibition, P < 0.05) by the cytochrome P450 inhibitor piperonyl butoxide (PB). The rate of TCBZSO conversion into TCBZSO2 was also significantly inhibited by PB (55% inhibition), MTZ (52% inhibition) and also following FMO inactivation (58% inhibition). The data reported here indicate that the FMO is the main enzymatic pathway involved in TCBZ sulphoxidation (ratio FMO/P450 = 3.83 +/- 1.63), although both enzymatic systems participate in a similar proportion in the sulphonation of TCBZSO to form the sulphone metabolite (ratio FMO/P450 = 1.31 +/- 0.23). Additionally, ketoconazole (KTZ) did not affect TCBZ sulphoxidation but decreased (66% inhibition, P < 0.05) the formation of TCBZSO2. Similarly, inhibition of TCBZSO2 production was observed after incubation of TCBZSO in the presence of KTZ and erythromycin (ETM). Conversely, thiabendazole (TBZ) and fenbendazole (FBZ) did not affect the oxidative metabolism of both incubated substrates. The sheep ruminal microflora was able to reduce the sulphoxide (TCBZSO) into the parent thioether (TCBZ). The ruminal sulphoreduction of the HO-TCBZSO derivative into HO-TCBZ was also demonstrated. The rate of sulphoreduction of HO-TCBZSO was significantly (P < 0.05) higher than that observed for TCBZSO. The metabolic approach tested here contributes to the identification of the different pathways involved in drug biotransformation in ruminant species. These findings on the pattern of hepatic and ruminal biotransformation of TCBZ and its main metabolites are a further contribution to the understanding of the pharmacological properties of widely used anthelmintics in ruminants. Comprehension of TCBZ metabolism is critical to optimize its flukicidal activity.     

Pharmacokinetic disposition of triclabendazole in cattle and sheep; discrimination of the order and the rate of the absorption process of its active metabolite triclabendazole sulfoxide

A comparative pharmacokinetic study was conducted to determine the order and the rate of absorption of triclabendazole (TCBZ) in cattle and sheep. A commercial suspension of TCBZ (Biofasiolex, Biogénesis S.A., Argentina) was administered at a dose rate of 10 mg/kg by the oral route to six Holstein female calves and six Corriedale female sheep. The plasma concentration profiles of the metabolites triclabendazole sulfoxide (TCBZ-SO) and triclabendazole sulfone (TCBZ-SO₂) were analysed by means of the non-compartmental method. The order of the absorption process of the active metabolite, TCBZ-SO, was determined by construction of curves of cumulative absorbed fraction of the drug by means of the Wagner-Nelson method. The appearance of TCBZ-SO in plasma of cattle and sheep resembles the entry of a constant quantity of drug into the organism per unit time. This is explained by the reservoir effect of the rumen, which acts as a biological slow-release system for TCBZ-SO and its precursor TCBZ to the posterior digestive tract where they are absorbed. The plasma concentration profiles of TCBZ-SO in both species were well described by a one-compartment open model with zero-order process of absorption and first-order process of elimination. The values of AUC_0-∞ and C _max of TCBZ-SO did not differ between species, while other kinetic parameters except for λ _z had higher values in calves than in sheep. In the case of TCBZ-SO₂, t _max was the only parameter that did not differ between species, while other kinetic parameters except for λ _z had higher values in calves than in sheep.     

Combined use of ivermectin and triclabendazole in sheep: in vitro and in vivo characterisation of their pharmacological interaction

This study evaluated the pharmacokinetic properties of ivermectin (IVM) and triclabendazole (TCBZ) given either separately or co-administered to sheep. Corriedale sheep received IVM alone, TCBZ alone or a combination of IVM and TCBZ intravenously. Ivermectin elimination was delayed and its plasma availability was 3-fold higher when co-administered with TCBZ. Similarly, plasma concentrations of TCBZ and its metabolites were influenced by the co-administration of IVM. Higher peak plasma concentrations of TCBZ metabolites were detected after the co-administration of TCBZ and IVM compared to those obtained following TCBZ treatment in isolation. Complementary in vitro assays were carried out to assess the influence of TCBZ on the P-glycoprotein-mediated intestinal transport of IVM, using the everted gut sac technique. Enhanced accumulation of IVM in the intestinal wall occurred after co-incubation with TCBZ.     

Enhancement of triclabendazole action in vivo against a triclabendazole-resistant isolate of Fasciola hepatica by co-treatment with ketoconazole

An in vivo study in the laboratory rat model was carried out to monitor morphological changes in adult Fasciola hepatica over a 4-day period resulting from combination treatment of triclabendazole (TCBZ) and the metabolic inhibitor, ketoconazole (KTZ). Rats were infected with the TCBZ-resistant Oberon isolate of F. hepatica and divided into 3 groups at 12 weeks post-infection. The first group was dosed orally with TCBZ at a dosage of 10mg/kg and KTZ at a dosage of 10mg/kg. Flukes were recovered at 24, 48, 72 and 96 h post-treatment (p.t.). A second group of rats was treated with TCBZ alone (10mg/kg) and sacrificed at 96 h p.t. The third group acted as untreated controls. Surface changes were monitored by scanning electron microscopy (SEM). In flukes from the TCBZ+KTZ-treated group, the results showed a progressive and time-dependent increase in the level of disruption to the tegumental syncytium. Swelling, furrowing, blebbing and sloughing of the syncytium increased with time p.t. Another feature seen was a thick layer of tegumental shedding in some fluke samples at different times p.t. By comparison, flukes treated with TCBZ alone remained unaffected. The results demonstrated that the Oberon isolate is only sensitive to drug action in the presence of ketoconazole, indicating that combining triclabendazole with a metabolic inhibitor could be used to preserve the effectiveness of the drug against TCBZ-resistant populations of F. hepatica.     

Assessment of the pharmacological interactions between the nematodicidal fenbendazole and the flukicidal triclabendazole: In vitro studies with bovine liver microsomes and slices

Parasitic diseases have a significant impact on livestock production. Nematodicidal drugs, such as fenbendazole (FBZ) or its oxidized metabolite oxfendazole (OFZ), can be used along with the trematodicidal triclabendazole (TCBZ), to broaden the spectrum of anthelmintic activity. However, co‐exposure to these compounds could lead to drug–drug (D‐D) interactions and eventually alter the clinical profile of each active principle. The aim of this study was to assess the presence of such interactions by means of two in vitro models, namely bovine liver microsomal fractions and bovine precision‐cut liver slices (PCLSs). To this end, an in vitro assessment involving incubation of FBZ and TCBZ or a combination of FBZ and TCBZ was carried out. Results with microsomal fractions showed a 78.4% reduction (p = .002) in the rate of OFZ production upon co‐incubation, whereas the sulfoxide metabolite of TCBZ (TCBZSO) exhibited a decreasing tendency. With PCLS, OFZ accumulation in the incubation medium increased 1.8‐fold upon co‐incubation, whereas TCBZSO accumulation decreased by 28%. The accumulation of FBZ and OFZ in the liver tissue increased upon 2‐hr co‐incubation, from 2.1 ± 1.5 to 18.2 ± 6.1 (p = .0009) and from 0.4 ± 0.1 to 1.3 ± 0.3 nmol (p = .0005), respectively. These results confirm the presence of D‐D interactions between FBZ and TCBZ. Further studies are needed to determine the extent of involvement of drug‐metabolizing enzymes and membrane transporters in interactions between compounds largely used in livestock production systems.     

Some Pharmacokinetic Parameters of Pefloxacin in Lactating Goats

The aim of this study was to elucidate some of the pharmacokinetic parameters of pefloxacin in lactating goats (n = 5) following intravenous (i.v.) or intramuscular (i.m.) injections of 10 mg/kg bw. Serially obtained serum, milk and urine samples were collected at precise time intervals, and the drug concentrations were assayed using a microbiological assay. A two-compartment open model best described the decrease of pefloxacin concentration in the serum after intravenous administration. The maximum serum concentration (C _p ⁰ ) was 8.4±0.48 g/ml; elimination half-life (t _1/2) was 1.6±0.3 h; total body clearance (Cl_tot) was 3.6±0.3 L/kg/h; steady-state volume of distribution (V _dss) was 5.14±0.21 L/kg; and the area under the curve (AUC) was 2.78±0.22 g.ml/h. Pefloxacin was absorbed rapidly after i.m. injection with an absorption half-life (t _1/2ab) of 0.32±0.02 h. The peak serum concentration (C _max) of 0.86±0.08 g/ml was attained at 0.75 h (T _max). The absolute bioavailability after i.m. administration was 70.63±1.13% and the serum protein-bound fraction ranged from 7.2% to 14.3%, with an average value of 9.8±1.6%. Penetration of pefloxacin from the blood into the milk was rapid and extensive, and the pefloxacin concentration in milk exceeded that in serum from 1 h after administration. The drug was detected in milk and urine for 10 and 72 h, respectively; no samples were taken after 72 h.     

The effect of fasting on the plasma disposition of triclabendazole following oral administration in goats

This study was designed to investigate the effect of feeding on the plasma disposition of triclabendazole (TCBZ) in goats following oral administration. A total of eight goats, aged 14–16 months and weighing 20–30 kg were used in this study. The animals were allocated into two groups (fasted and fed groups) of four animals each. The goats in fed group were fed ad libitum but the animals in fasted group were not fed 24 h before and 6 h after drug administration. Commercial oral drench formulation of TCBZ (Endex-K, 5%) was administered orally to animals in two groups at dose of 10 mg/kg bodyweight. Heparinized blood samples were collected between 1 and 192 h after treatment and the plasma samples were analysed by high performance liquid chromatography (HPLC) for TCBZ, TCBZ sulphoxide (TCBZ–SO), and TCBZ sulphone (TCBZ–SO₂). Relatively very low concentration of TCBZ parent drug was detected between 2 and 48 h, but TCBZ–SO and TCBZ–SO₂ metabolites were present between 2 and 192 h in the plasma samples of fed and fasted animals. Fasting significantly enhanced the plasma concentration of TCBZ and its metabolites. The availability of TCBZ, TCBZ–SO and TCBZ–SO₂ in the plasma samples of fasted goats were markedly greater compared to those of fed goats. It was concluded that fasting decreases the digesta flow rate and prolongs the retention of the drug into the gastrointestinal tract, resulting in enhanced quantitative gastrointestinal absorption or systemic availability of TCBZ and its metabolites in fasted goats.     

Disruption of egg formation by Fasciola hepatica following treatment in vivo with triclabendazole in the sheep host

Eight indoor-reared cross-bred sheep with no prior exposure to Fasciola hepatica were infected by oral gavage with 200 metacercarial cysts of the triclabendazole (TCBZ)-susceptible Cullompton isolate of F. hepatica. Twelve weeks after infection, sheep were treated with 10mg/kg triclabendazole. Two sheep were euthanised per time period; at 48 h, 72 h and 96 h post-treatment (pt). Two untreated control sheep were euthanised at 96 h pt. Flukes were recovered from the liver and, if present, from the gall bladder of the sheep. They were processed for whole mount analysis, histology and transmission electron microscopy of the female reproductive system; specifically, the uterus, vitelline follicles, Mehlis' gland and ovary. Over the 4-day post-treatment period, there was a progressive reduction in the number of oogonia and oocytes in the ovary and evidence of apoptosis. Vacuolation and a decrease in the number of Mehlis' gland cells were observed from 48 h pt onwards and disruption of the normal role of the gland in egg formation was evident. The vitelline follicles showed a gradual decrease in size and became vacuolated; the population structure in each follicle changed to be one consisting mainly of mature cells and the production of shell protein material declined. The follicle became disorganised as the cells broke down and released their contents into the lumen of the follicle. While the uterus appeared to contain eggs at 48 h pt in whole-mount specimens, no properly-formed eggs were observed in histological sections. By 96 h pt, the uterus was completely devoid of eggs. Overall, egg production was seen to be severely affected by TCBZ treatment and flukes were incapable of producing normal eggs within 2 days of treatment. The implications of this in terms of the epidemiology of the disease are discussed.     

Digestibility of vegetative and mature tall fescue greenchop with or without alfalfa greenchop substitution.

Nine ruminally and duodenally cannulated steers (average BW 355 kg) were used to evaluate effects of prebloom alfalfa greenchop substitution at 20% of DMI on utilization of late-May (high quality; HQ; Period 1) and mid-August (low quality; LQ; Period 2) tall fescue greenchop. Alfalfa inclusion did not influence (P greater than .10) diet ad libitum DMI during Period 1 but it decreased (P less than .10) DMI during Period 2. Ruminal and total tract DM, cell wall, and GE digestibility of HQ were unaffected (P greater than .10) by alfalfa inclusion; however, digestibility of these constituents in LQ was increased (P less than .03) by alfalfa substitution. Alfalfa substitution did not influence (P greater than .10) dietary cell wall monosaccharide disappearance. Ruminal CP digestibility was greater (P less than .10) when steers received alfalfa, but microbial efficiency (grams of bacterial N/kilogram of OM truly digested in the rumen) was not enhanced (P greater than .10) by alfalfa inclusion in either HQ or LQ diets. There was a trend (P = .15) for greater microbial efficiency with alfalfa substitution to LQ. Ruminal particulate passage rate did not differ (P greater than .10) between treatments for either stage of maturity. Fluid passage rate was faster (P less than .10) in steers that received only LQ (7.1%/h) than in those fed LQ substituted with 20% alfalfa (5.0 %/h). Our data suggest that alfalfa inclusion in a low-quality fescue diet enhanced cell wall and GE digestibility.     

Pharmacokinetic comparison of six anthelmintics in sheep,goats, and cattle

This study was initiated to determine whether a comparative pharmacokinetic (PK) approach could be used to expand the pool of approved anthelmintics for minor ruminant species. Accordingly, the PK profiles of six anthelmintics (levamisole, albendazole, fenbendazole, moxidectin, doramectin, and ivermectin) in sheep, goats, and cattle were determined. The PK values determined for each anthelmintic included T_max, T_last, C_max, AUC, AUC/dose, and C_max/dose. The results of this study demonstrate that a comparative PK approach does not show commonality in the way these six anthelmintics are individually processed by these three ruminants. While some drugs demonstrated identical PK profiles between sheep and goats, none of these drugs demonstrated PK profiles in sheep and goats comparable to the PK profiles found in cattle. The results from this study suggest drug approval across these three ruminants is not a viable concept. However, the resulting PK profiles for each combination of drug and ruminant species represents a new dataset that can be used to support the US FDA Center for Veterinary Medicine's Minor Use/Minor Species indexing process for drug approvals in minor species such as sheep and goats.     

Inhibition of cytochrome P450 activity enhances the systemic availability of triclabendazole metabolites in sheep

Understanding the disposition kinetics and the pattern of metabolism is critical to optimise the flukicidal activity of triclabendazole (TCBZ) in ruminants. TCBZ is metabolised by both flavin-monooxygenase (FMO) and cytochrome P450 (P450) in the liver. Interference with these metabolic pathways may be useful to increase the systemic availabilities of TCBZ metabolites, which may improve the efficacy against Fasciola hepatica . The plasma disposition of TCBZ metabolites was evaluated following TCBZ co-administration with FMO [methimazole (MTZ)] and P450 [piperonyl butoxyde (PB) and ketoconazole (KTZ)] inhibitors in sheep. Twenty (20) healthy Corriedale x Merino weaned female lambs were randomly allocated into four experimental groups. Animals of each group were treated as follow: Group A, TCBZ alone (5 mg/kg, IV route); Group B, TCBZ (5 mg/kg, IV) + MTZ (3 mg/kg, IV); Group C, TCBZ (5 mg/kg, IV) + PB (30 mg/kg, IV) and Group D, TCBZ (5 mg/kg, IV) + KTZ (10 mg/kg, orally). Blood samples were taken over 240 h post-treatment and analysed by HPLC. TCBZ sulphoxide and sulphone were the main metabolites recovered in plasma. MTZ did not affect TCBZ disposition kinetics. TCBZ sulphoxide Cmax values were significantly increased ( P  < 0.05) after the TCBZ + PB (62%) and TCBZ + KTZ (37%) treatments compared to those measured in the TCBZ alone treatment. TCBZ sulphoxide plasma AUCs were higher ( P  < 0.05) in the presence of both PB (99%) and KTZ (41%). Inhibition of TCBZ P450-mediated oxidation in the liver accounted for the increased systemic availability of its active metabolite TCBZ sulphoxide. This work contributes to the search of different strategies to improve the use of this flukicidal drug in ruminants.     

The kinetics of triclabendazole disposition in sheep

To investigate whether the disposition of triclabendazole (TCBZ) and its metabolites in blood or bile influenced its flukicidal potency, TCBZ was administered intraruminally at 10 mg kg-1 to sheep surgically fitted with a permanent re-entrant bile duct cannula. The profiles of TCBZ metabolites in peripheral plasma and bile were determined using high performance liquid chromatography. In plasma, only TCBZ sulphoxide (TCBZ-SO) and TCBZ sulphone were present and reached their maximum concentrations (greater than 13 micrograms ml-1) at 18 and 36 h, respectively, after administration. TCBZ metabolites were specifically bound to plasma albumin, which is believed to exert a major influence on the duration of plasma TCBZ metabolite concentrations and consequent exposure of liver fluke. In bile, the major TCBZ metabolites were hydroxylated in the 4' position and secreted predominantly as sulphate esters with lesser proportions as glucuronide conjugates. The major biliary metabolite was conjugated hydroxy TCBZ-SO which reached a maximum concentration in excess of 40 micrograms ml-1 and contributed almost half the total conjugated metabolites. The major free biliary metabolite was TCBZ-SO. Of the administered TCBZ dose, 9.7% was secreted as free metabolites in bile whereas 35.8% was secreted as conjugated metabolites. Approximately 6.5% of the dose was excreted in urine.     

Bioequivalence Study of Two Long-acting Oxytetracycline Formulations in Sheep

Two commercially available long-acting oxytetracycline hydrochloride formulations (Primamycin LA (Pfizer) and Terralent 20% LA (İ.E. Ulagay)) were administered by the intramuscular route to 20 clinically healthy sheep at a dose of 20 mg/kg. The study was performed in a two-period crossover design. Plasma samples were analysed by high-pressure liquid chromatography. The mean maximum concentrations (C _max) was 8.00 ± 2.05 μg/mland 8.61 ± 1.42 μg/ml, respectively. The mean area under the concentration time curve (AUC) values were 154.95 ± 50.37(μg h)/ml and 161.70 ± 47.02(μg h)/ml, respectively. The 90%confidence intervals for the ratio of C _max and AUC values for the test and reference product are with in the interval 70−143% for C _max and interval 80-−125% for AUC proposed by EMEA. It was concluded that Primamycin LA and Terralent 20% LA formulations are bioequivalent in their rate and extent of drug absorbtion. Ozdemir N. and Yıldırım, M., 2006. Bioequivalence study of two long-acting oxytetracycline formulations in sheep. Veterinary Research Communications, 30(8), 929–934     

Plasma bupivacaine concentrations following orbital injections in cats

Objective

To determine plasma bupivacaine concentrations after retrobulbar or peribulbar injection of bupivacaine in cats.

Pharmacokinetics and oral bioavailability of amoxicillin in chicken infected with caecal coccidiosis

Chicken infected with caecal coccidiosis (Eimeria tenella) was used to evaluate the effect of coccidiosis on the pharmacokinetic and bioavailability of amoxicillin. The level of amoxicillin was estimated by high‐performance chromatography (HPLC) to calculate the pharmacokinetic parameters and oral bioavailability. For i.v. injection of amoxicillin, Vd and CL were 0.29 and 0.27 (mg/kg)/(μg/mL)/h, respectively. Compared with healthy chicken, intravenous injection of amoxicillin in the infected chicken showed higher distribution and elimination constants, delayed clearance and statistically significant higher AUC and MRT. Oral administration in healthy chicken was accompanied by rapid absorption and high bioavailability with T_max, C_max and F about 1.03 h, 3.26 μg/mL and 40.2, respectively. Furthermore, oral administration in the infected chicken produced higher mean absorption time, delayed T_max, lower C_max, smaller AUC value and lower bioavailability (16.76). Based on these results, monitoring and adjustment of amoxicillin dosing could be practiced during the presence of coccidiosis. The measured C_max values suggest the administration of 1.3‐folds of the normal dose to maintain the normal maximal serum concentrations of amoxicillin in chicken infected with caecal coccidiosis.     

Pharmacokinetics of amoxicillin trihydrate in cultured olive flounder (Paralichthys olivaceus)

The study was aimed at investigating the pharmacokinetics of amoxicillin trihydrate (AMOX) in olive flounder (Paralichthys olivaceus) following oral, intramuscular, and intravenous administration, using high‐performance liquid chromatography following. The maximum plasma concentration (C_max), following oral administration of 40 and 80 mg/kg body weight (b.w.), AMOX was 1.14 (T_max, 1.7 h) and 0.76 μg/mL (T_max, 1.6 h), respectively. Intramuscular administration of 30 and 60 mg/kg of AMOX resulted in C_max values of 4 and 4.3 μg/mL, respectively, with the corresponding T_max values of 29 and 38 h. Intravenous administration of 6 mg/kg AMOX resulted in a C_max of 9 μg/mL 2 h after administration. Following oral administration of 40 and 80 mg/kg AMOX, area under the curve (AUC) values were 52.257 and 41.219 μg/mL·h, respectively. Intramuscular 30 and 60 mg/kg doses resulted in AUC values of 370.274 and 453.655 μg/mL·h, respectively, while the AUC following intravenous administration was 86.274 μg/mL·h. AMOX bioavailability was calculated to be 9% and 3.6% following oral administration of 40 and 80 mg/kg, respectively, and the corresponding values following intramuscular administration were 86% and 53%. In conclusion, this study demonstrated high bioavailability of AMOX following oral administration in olive flounder.     

Tegumental surface changes in juvenile Fasciola hepatica in response to treatment in vivo with triclabendazole

Eight indoor-reared crossbred sheep with no pre-exposure to Fasciola hepatica were infected, by oral gavage, with 200 metacercarial cysts of the triclabendazole (TCBZ)-susceptible Cullompton isolate of F. hepatica. Anthelmintic dosing occurred at 4 weeks post-infection with 10 mg/kg triclabendazole. Two treated sheep were euthanized at 48 h, 72 h and 96 h post-treatment with triclabendazole. Two control sheep were euthanized alongside the 48 h triclabendazole-treated sheep. Juvenile flukes were recovered from each of the sheeps' liver and processed for scanning electron microscopy (SEM). Flukes were still active 48 h post-treatment and displayed limited morphological disruption. There was some blebbing and sloughing of the tegument around the oral sucker. In several of the specimens, an extra layer had been deposited on the fluke surface, giving it a flattened appearance. At 72 h post-treatment, only one fluke remained alive and the disruption varied in degree. In the majority of flukes, there was severe swelling of the tegument, accompanied by isolated areas of flattening along the lateral margins of the flukes and in the tail region. Limited areas of sloughing occurred in the tail region. In more seriously affected specimens, the syncytium had been stripped away to reveal the basal lamina and some deeper lesions were also observed. By 96 h post-treatment, all the flukes were dead and were grossly disrupted. They were totally devoid of tegument and deep lesions exposed the internal tissues of the fluke.     

Composite faecal egg count reduction test to detect resistance to triclabendazole in Fasciola hepatica

A faecal egg count reduction test, using composite samples, was developed in order to assess the efficacy of the flukicide, triclabendazole (TCBZ) on commercial sheep farms in England and Wales. First, a comparison between individual counts and composite counts was conducted using sheep on two farms with different levels of infection. Faecal samples were collected from 50 sheep on each farm at the time of TCBZ treatment and 21 days later. The results showed that a composite fluke egg count (CFEC) was as sensitive as using individual samples, and the test was subsequently validated on an additional 18 sheep farms. The pre- and post-treatment CFECs using five composite samples were subjected to bootstrap analysis. The variance in fluke egg counts of composite samples was high, but analyses indicated that 20 individual samples analysed as two composites was as sensitive as using five composites. The two-composite assay was evaluated on another five farms. On seven out of 25 farms sampled, egg counts either did not decrease significantly or increased following treatment, suggesting TCBZ resistance on these farms. This assay represents a practical field test that can be used in the first instance to evaluate the efficacy of TCBZ on sheep farms where resistance is suspected.