A. Königová

Benzimidazole resistance in equine cyathostomes in Slovakia.

The present study included 19 stud farms, including 243 horses, that were investigated for the occurrence of anthelmintic resistant cyathostomes. The number of horses on the farms varied from nine to more than 100, and horses of all ages were included. A minimum of seven horses were used for faecal egg count reduction (FECR) tests. The anthelmintics included were: fenbendazole (paste formulation), ivermectin (paste formulation) and pyrantel (powder). Resistance to benzimidazoles was detected on 14 farms, with FECR values ranging from 65.1 to 86.3%. Larval cultures after fenbendazole treatment revealed exclusively cyathostome larvae. Ivermectin was tested on eight farms and proved to be effective on all. Pyrantel was tested on two farms and FECR test indicated high efficacy (92-97%). Egg hatch assay (EHA) results showed that mean concentrations of thiabendazole that inhibited hatching in 50% of the eggs (ED(50)) in resistant populations were over 0.1 microg ml(-1). The results of our study suggest widespread resistance to fenbendazole in equine cyathostomes in Slovakia, and possible strategies to delay anthelmintic resistance are discussed briefly.

Anthelmintic resistance in parasites of small ruminants: sheep versus goats

SummaryAnthelmintic resistance among parasites of sheep and goats has been known to occur for at least four decades. Both species of host have similar genera of nematodes, but the nematodes in goat herds usually develop anthelmintic resistance more rapidly. In vitro tests show higher ED50 values in goats than in flocks of sheep. Sheep and goats differ in many ways; for example, goats have a higher metabolic rate and require higher dose rates for drugs. The immune system of goats is also different. Additionally, these animals are reared under different management systems, i.e. sheep graze pastures and goats browse bushes, and lambing/kidding periods are different. Most anthelmintics used in goats have not been licensed for this animal species, and correct dose rates have rarely been experimentally determined. Possible explanations for such differences are discussed.

Comparison of in vitro methods and faecal egg count reduction test for the detection of benzimidazole resistance in small strongyles of horses

The objective of the study was to compare the in vitro egg hatch test (EHT), larval development test (LDT) and in vivo faecal egg count reduction test (FECR test) for the detection of benzimidazole resistance in equine strongyles. The presence of resistant or susceptible strongyle populations was determined in 25 stud farms using the in vivo FECR test and in vitro EHT. On the basis of the FECR values, resistance to fenbendazole was detected on 15 of the 25 farms (60%). The ED50 value (anthelmintic concentration producing 50% inhibition of hatching) for suspected resistant populations varied from 0.110 to 0.222 μg/ml thiabendazole (TBZ). Final LD50 values (anthelmintic concentration inhibiting development of 50% of eggs into L3 infective larvae) above 0.029 μg/ml TBZ in the in vitro larval development test on samples from 11 stud farms revealed the presence of populations of small strongyles suspected of being benzimidazole-resistant.

Phenotypic and genotypic characterisation of benzimidazole susceptible and resistant isolates of Haemonchus contortus

The study was designed to compare the in vitro egg hatch test (EHT) and molecular allele-specific polymerase chain reaction PCR (AS-PCR) methods as tools for detection of benzimidazole resistance in Haemonchus contortus, a nematode parasite of small ruminants. Comparisons were made during the course of an experimental infection and changes in EHT and AS-PCR were monitored to measure the correlation between in vitro and molecular tests. Both methods were carried out according to World Association for the Advancement of Veterinary Parasitology (WAAVP) recommendations. The molecular test was used to discriminate TAC/TTC polymorphism in the beta-tubulin 200 codon of eight (four resistant and four susceptible) isolates of H. contortus. Using DNA from 100 third-stage larvae, the polymerase chain reaction (PCR)-based analysis revealed a decrease of the homozygous TTC/TTC genotype and an increase in heterozygous TTC/TAC and homozygous TAC/TAC individuals in all resistant isolates. Both methods showed comparable and reliable results with regard to detection of benzimidazole resistance. The molecular test has an advantage over the EHT because of its higher sensitivity. On the other hand, EHT is less time-consuming, allows reliable detection of <10% resistance allele frequency and is fairly reliable for the detection of benzimidazole resistance under field conditions.

Detection of ivermectin resistance by a larval development test--back to the past or a step forward?

The aim of this study was to evaluate the potential of the larval development test for the detection of ivermectin (IVM) resistance in Haemonchus contortus of sheep. Single infections with 5000 third-stage larvae of five resistant and two susceptible isolates of H. contortus were given to sheep. Fecal samples were collected four times during patency, and the micro-agar version of the larval development test (MALDT) was performed. Three macrocyclic lactone drugs (IVM, eprinomectin and IVM aglycone) were tested. The results of the tests are presented as LC50 and LC99 values. The MALDT was well able to distinguish between susceptible and resistant isolates. Resistance factors (RF) for the LC99 values were generally higher than those obtained by comparing LC50 values. The highly resistant isolates were readily distinguishable from the susceptible isolates, particularly when using IVM aglycone and eprinomectin, with RFs above 20.

Parasitic infections and pregnancy complications

SummaryInfections of various aetiology during pregnancy present a serious risk factor that can lead to abortion, premature birth, stillbirth, birth of newborn with genetic developmental defects, or seemingly healthy infant that may develop symptoms later in life. A total of 530 sera of patients from gynaecology departments were examined for antibodies against Toxoplasma gondii, Toxocara spp. and Listeria monocytogenes using ELISA (IgG, IgM). In women with habitual abortions we determined up to 42.1 % seroprevalence of IgG antibodies against T. gondii. The prevalence of antibodies was statistically significant (p < 0.0004) when compared to control group (25.1 %). In the patients diagnosed with sterility the prevalence of antibodies (26.3 %) was similar to that of control group. The prevalence of antibodies against Toxocara spp. (5.5 %) and Listeria monocytogenes (2.3 %) in patients with habitual abortions were similar to those of women in the general population.

Reduction of oxidative stress and liver injury following silymarin and praziquantel treatment in mice with Mesocestoides vogae (Cestoda) infection.

Oxidative stress is a common mechanism contributing to hepatic damage and fibrogenesis in a variety of liver disorders. The liver is the target organ for many parasitic infections, hence there is a great demand for the development of novel treatment strategies. In the present study conducted on mice infected with larval stage of Mesocestoides vogae, we investigated effects of therapy with praziquantel (PZQ) alone and in combination with silymarin on liver GSH content, lipid peroxidation and larval reduction. Proliferation of liver cells by means of BrdU incorporation into DNA and production of superoxide anions by peritoneal adherent cells was measured to assess the antioxidant activity of silymarin. Drug administration was carried on from day 15 post infection (p.i.) for ten consecutive days and examination was performed during 20 days of follow-up the therapy. Larval M. vogae infection caused liver damage and triggered extensive oxidative stress, resulting in the abolishment of GSH redox balance and ROS-induced lipid peroxidation. PZQ administration caused short-term decline of GSH levels in healthy mice. Low GSH levels in infected mice were elevated gradually in response to the drug, but respiratory burst in cells was not reduced. Silymarin in combination with PZQ showed strong direct antioxidant capacity and stimulated the larvicidal effect of praziquantel. Treatment with PZQ and silymarin downregulated the generation of superoxide anions, prevented lipid peroxidation, stimulated GSH synthesis and proliferation of hepatocytes in infected livers. These findings demonstrated that silymarin can markedly decrease the liver injury and its co-administration with PZQ potentiate effect of therapy, probably due to the down-regulation of fibrogenesis.

Nematode infections in Slovak children hospitalised during 2008-2009.

SummaryA study involved 1800 hospitalised children (age: 9 months to 16 years) examined by ovoscopic analyses and confirmed the occurrence of Ascaris lumbricoides and Trichuris trichiura in 46 patients (2.55 %). Of these, 30 patients had Ascaris infection, 13 were positive for both nematodes and 3 patients had Trichuris infection. The mean count of A. lumbricoides eggs in positive cases was 1050 eggs per gramme (EPG) in range 150–4450 EPG. The mean count of T. trichiura eggs was 150 EPG (50–250 EPG). The highest intensities of A. lumbricoides and T. trichiura infections occurred in children 3–5 years of age living in poor hygienic conditions. Most common clinical conditions in all the patients included anaemia combined with complicated bronchopneumonia, colitis and gastritis. The strongest correlation between the parasite burden and selected laboratory test data (eosinophil count, haemoglobin, total serum iron) was found in children of 2 years of age (P < 0.05) and decreased with age.

The first report of serratospiculiasis in Great Tit (Parus major) in Slovakia

SummarySerratospiculiasis is a parasitic disease caused by filariid nematodes of the genus Serratospiculum, the subfamily Dicheilonematinae. Serratospiculum spp. parasitizes the air sacs and members of this genus have been found in various species of Falcons, Bald Eagles and Cooper’s Hawk from all around the world. In the present study, infection with Serratospiculum was confirmed for the first time in the Great Tit in Slovakia. Nematode infestation was identified post mortem. Lesions in the respiratory system (airsacculitis, pneumonia) were associated with the presence of this nematode. Smears of the necrotic lesions in the crop and oesophagus contained the large numbers of embryonated eggs. Faecal samples were examined by flotation method and Serratospiculum eggs were found. Adult parasites were cleared in lactophenol solution and morphological analysis of male reproductive organs (shape of male nematode spicules) indicated the species Serratospiculum amaculata. Scattered inflammatory cells were seen in the mucosal and submucosal layers of infected oesophagus and inflammatory foci were found mainly in the stroma of the air sacs.The presence of the filariid nematode in the nonspecific species Great Tit (Parus major) common in Slovakia indicates the importance of monitoring of serratospiculiasis in the avian hosts. The parasite can cause serious health problems, even sudden death of their hosts, therefore suitable effective measures for their elimination should be implemented.

Comparison of two in vitro methods for the detection of ivermectin resistance in Haemonchus contortus in sheep

Summary Gastrointestinal parasitic nematodes in sheep cause severe economic losses. Anthelmintics are the most commonly used drugs for prophylaxis and therapy against parasitic helminths. The problem of drug resistance has developed for all commercially available anthelmintics in several genera and classes of helminths. In vitro and in vivo tests are used to detect anthelmintic resistance. Two in vitro methods (larval migration inhibition test and micromotility test) for the detection of ivermectin (IVM) resistance were compared using IVM-resistant and IVM-susceptible isolates of Haemonchus contortus. The degree of resistance for each test was expressed as a resistance factor (RF). The micromotility test was more sensitive for quantitatively measuring the degree of resistance between susceptible and resistant isolates. The RFs for this test for IVM and eprinomectin ranged from 1.00 to 108.05 and from 3.87 to 32.32, respectively.