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Item Widespread occurrence of benzimidazole resistance single nucleotide polymorphisms in the canine hookworm, Ancylostoma caninum, in Australia(Elsevier Ltd on behalf of the Australian Society for Parasitology Incorporated, 2025-03) Abdullah S; Stocker T; Kang H; Scott I; Hayward D; Jaensch S; Ward MP; Jones MK; Kotze AC; Šlapeta JCanine hookworm (Ancylostoma caninum), a gastrointestinal nematode of domestic dogs, principally infects the small intestine of dogs and has the potential to cause zoonotic disease. In greyhounds and pet dogs in the USA, A. caninum has been shown to be resistant to multiple anthelmintics. We conducted a molecular survey of benzimidazole resistance in A. caninum from dogs at veterinary diagnostic centers in Australia and New Zealand. First, we implemented an internal transcribed spacer (ITS)-2 rDNA deep amplicon metabarcoding sequencing approach to ascertain the species of hookworms infecting dogs in the region. Then, we evaluated the frequency of the canonical F167Y and Q134H isotype-1 β-tubulin mutations, which confer benzimidazole resistance, using the same sequencing approach. The most detected hookworm species in diagnostic samples was A. caninum (90%; 83/92); the related Northern hookworm (Uncinaria stenocephala) was identified in 11% (10/92) of the diagnostic samples. There was a single sample with coinfection by A. caninum and U. stenocephala. Both isotype-1 β-tubulin mutations were present in A. caninum, 49% and 67% for Q134H and F167Y, respectively. Mutation F167Y in the isotype-1 β-tubulin mutation was recorded in U. stenocephala for the first known time. Canonical benzimidazole resistance codons 198 and 200 mutations were absent. Egg hatch assays performed on a subset of the A. caninum samples showed significant correlation between 50% inhibitory concentration (IC50) to thiabendazole and F167Y, with an increased IC50 for samples with > 75% F167Y mutation. We detected 14% of dogs with > 75% F167Y mutation in A. caninum. Given that these samples were collected from dogs across various regions of Australia, the present study suggests that benzimidazole resistance in A. caninum is widespread. Therefore, to mitigate the risk of resistance selection and further spread, adoption of a risk assessment-based approach to limit unnecessary anthelmintic use should be a key consideration for future parasite control.Item Veterinary anthelmintics : their efficacy and effects on abomasal physiology : a thesis presented in partial fulfilment of the requirements for the degree of Master of Veterinary Science in Veterinary Clinical Pharmacology at Massey University(Massey University, 1998) Whelan, Nicholas CharlesPART 1. A Review of the Veterinary Anthelmintic Literature A comprehensive review was undertaken of the pharmacology, efficacy, side effects and toxicity of veterinary anthelmintics used against nematode parasites. Anthelmintics reviewed for use in cattle, sheep, goats, horses, dogs and cats include copper, nicotine, arsenic, tetrachlorethylene, phenothiazine, diethylcarbamazine, piperazine, toluene, cyacethydrazide, bephenium, thenium, organophosphates, and methyridine. The review was limited to cattle for the benzimidazoles, pyrantel, morantel, tetramisole, levamisole, avermectin and milbemycins anthelmintics. Efficacy data is provided in a tabular format which classifies each anthelmintic according to method of administration and dose. PART 2 Efficacy of two formulations of moxidectin pour-on and the effects of treatment on serum pepsinogen and gastrin levels and tissue gastrin in cattle Three groups of eight yearling Friesian bulls were used to compare the efficacy of two 5 g/L pour-on formulations of moxidectin applied at 1 ml/10kg (500 mcg moxidectin per kg bodyweight) in removing naturally acquired gastrointestinal parasites. At slaughter, 14-16 days after treatment, the burdens of Ostertagia spp. and Trichostrongylus axei were significantly lower in both the treated groups versus the controls (P<0.01). Anthelmintic efficacies (based on geometric mean worm burdens of treated and control groups) were all greater than 99.8% for Ostertagia spp. and Trichostrongylus axei in both treated groups compared with the controls. A significantly lower burden of adult Cooperia spp. was recorded for Formulation 1 (P<0.05). The anthelmintic efficacy of the two treatments against Cooperia adults, early L4 and late L4 were 96.25% (P<0.05), 97.31% and 91.08% respectively in calves treated with Formulation 1, and 71.44%, 67.14% and 64.29% respectively for calves treated with Formulation 2. Low numbers of large intestinal worms, Trichuris ovis and Oesophagostomum spp. in the control cattle precluded any valid efficacy assessment of these species. Based on these results, Formulation 1 is distinguished from Formulation 2 by its significantly greater efficacy against adult Cooperia spp. Serum pepsinogen and gastrin levels were monitored in the three groups of calves after treatment. All groups showed a steady decline in levels of both pepsinogen and gastrin until termination on day 14. On only one occasion was there a significant difference in serum pepsinogen between treated and untreated calves. Following treatment there was a more rapid and significant decrease in gastrin levels in calves treated with Formulation 1 than Formulation 2. At no stage was there a significant difference between the controls and calves treated with Formulation 2. There was no correlation between pepsinogen and gastrin levels for any of the groups. Neither was there a correlation between numbers of Ostertagia spp. and serum pepsinogen or gastrin. It is suggested that the decreases seen in the control group were due to lack of larval challenge and normal loss of adult worms resulting insufficient numbers to sustain a hypergastrinaemia. Gastrin concentrations were also measured in tissue samples from the pyloric antrum or the proximal duodenum. There was no significant difference in antral and proximal duodenal gastrin concentrations between the treatment and the control groups. Antral levels were between 1148- 1323 pM/g which were 25-35 times those found in duodenal tissue (32.3-50.9 pM/g).Item Aspects of anthelmintic resistance in nematodes in sheep : this thesis is presented in partial fulfillment of the requirements of the degree of Master of Veterinary Science at Massey University(Massey University, 1989) Butler, Andrew RoyThe increasing prevalence of anthelmintic resistance in nematodes of sheep is causing concern among animal scientists and farmers. In Australia anthelmintic resistance has become widespread since the first case was reported in 1968, and in some districts up to 68% of farms are affected. Benzimidazole resistance is most common, but levamisole and morantel resistance also occurs, and some farms have nematodes resistant to both major anthelmintic groups. Strains of the following species have shown resistance: Haemonchus contortus. Ostertagia circumcincta. Trichostrongylus spp. and, least commonly, Nematodirus spp. In New Zealand, anthelmintic resistance was first reported in 1980, and subsequent surveys found that its prevalence was generally low except on farms with above average anthelmintic usage. The same anthelmintics and nematodes as in Australia were implicated, although resistant Nematodirus spp. were reported more frequently in New Zealand. The most common test used to identify anthelmintic resistance - the faecal egg count reduction (or depression) test - was used in the survey of 15 Manawatu sheep farms which is the subject of this report. On each farm the anthelmintics oxfendazole, ivermectin and levamisole were given by mouth to groups of 12 young sheep, at dose rates of 5.0, 0.2 and 8.0 mg/kg, respectively. Twelve additional sheep were designated as untreated controls. Faecal samples for egg counting were taken from all sheep on the day of treatment and 7 days later. Composite faecal samples from each farm were cultured for identification of larvae present before treatment and, when anthelmintic treatments were deemed unsatisfactory, post-treatment faeces were also cultured for larval identification. Using the simplest method of calculation of faecal egg count reduction, and a cut-off point of 90% or below, there were (respectively) 4,2 and 0 farms with oxfendazole, ivermectin and levamisole resistant strongylate nematodes (excluding Nematodirus spp.). However, supporting evidence for the existence of ivermectin resistant nematodes was weak, and no claim is made that true ivermectin resistance has been detected. On the 4 farms with oxfendazole resistant nematodes, resistant Trichostrongylus spp. were the most common (4 farms), followed by H. contortus (3 farms), 0. circumcincta (2 farms) and Oesophagostomum spp. (2 farms). Oxfendazole resistant Nematodirus spp. were very common, but it was not possible to reach any conclusion about the susceptibility of the Nematodirus spp. present on each farm because of the low numbers of sheep passing Nematodirus spp. eggs and the small numbers of those eggs. Resistant Nematodirus spp. have probably been overlooked in the past and methods to reduce the likelihood of this are suggested. The faecal egg count reduction test is a useful field screening test, but it is difficult to interpret when only a slight degree of anthelmintic resistance is encountered. This is partly due to uncertainty in where the cut-off point should be, and partly to variation in calculation methods. The acceptance of recently formulated standard test protocols for Australia and New Zealand should reduce this problem, but supplementary tests will still be needed in many cases of anthelmintic resistance testing.Item Epidemiological studies of parasitism in sheep and reproduction in horses : a thesis presented in partial fulfilment of the requirements for the degree of Master of Veterinary Studies (Epidemiology) at Massey University(Massey University, 1997) Macchi, ChiaraAnthelmintic resistance is a well-recognised problem for control of nemotodes in sheep in most countries of the world. The climatic conditions in New Zealand are particularly favourable to the survival and development of gastrointestinal nemo=atodes. As a consequence, gastrointestinal parasitism is a major impediment to profitable sheep raising in New Zealand. A random postal survey of 300 sheep farmers in the southern North Island region of Manawatu was conducted with the purpose of examining current farming and drenching practices and investigating possible risk factors in the development of anthelmintic resistance. The results of this study, reported in Chapter 2, revealed a high degree of awareness and concern about the problem of resistance, but also a lack of understanding on how grazing management strategies should be combined in order to achieve integrated control over gastrointestinal nematodes while minimising the use of anthelmintic drugs. Only 31% of respondents had performed at least one drench test on their property. Amond testing farms, prevalence of resistance approached 70% and involved benzimidazole products in all but one case. Subsequently, a trial was undertaken to investigate the economic consequences of anthelmintic resistance in growing lambs on commercial farms (Chapters 3 and 4). Five farms with a history of resistance to benzimidazole drenches were selected. The effects of three treatment strategis on nematode egg counts, bodyweight gains and susceptibility to diarrhoea were compared between groups of ewe lambs. Suppressively treated lambs performed significantly better than effectively treated lambs, which in turn performed better than ineffectively treated lambs. However, a partial budgeting analysis carried out by means of a stochastic simulation model (Chapter 4) indicated that effective treatment yielded the highest net returns. The model also showed that the range of possible outcomes oscillated substantially around the mean, reflecting the degree of uncertainty about the outcome on any single farm due to variation between farms. Chapter 5 describes a study which was carried out with the objective of evaluating two management strategies for breeding mares after foaling. Mares were examined on day 7-9 postpartum by palpation and ultrasound. Pregnancy rates, pregnancy loss rates and time from foaling to conception in two groups were compared. The statistical power of the study as determined by power analysis, was insufficient for the observed differences to reach statistical significance.Item Development of a novel equine combination anthelmintic and a study of anthelmintic susceptibility using a larval development assay : a thesis presented in partial fulfilment of the requirements for the degree of Master of Philosophy in Veterinary Science at Massey University(Massey University, 2000) Islam, ZahidulThe occurrence of anthelmintic resistance to benzimidazoles and avermectin in cyathostome nematodes in New Zealand horses was investigated using a larval development assay (LDA) to conduct a small survey. For the benzimidazoles the analogue, thiabendazole was used and for avermectin the analogues, ivermectin and ivermectin aglycone were used in the LDA. The normal range of LD50 values was estimated by assaying eggs from Kaimanawa feral horses (n = 22) for each analogue used. From these the mean LD50 values + 2xstandard deviation was taken as the upper limit of normal. The survey involved domestic horses (n = 47) from several locations around New Zealand. For ivermectin and ivermectin aglycone 12% had LD50 values higher than normal with resistance factors up to 5.3 and 6.8 respectively. This represents horses from three separate farms. For the benzimidazoles 43% of domestic horses had higher than normal LD50 values with resistance factors up to 4.8. These results suggest that some nematodes in these domestic horses had an increased tolerance to avermectins and as well as to benzimidazoles. Two studies were conducted to assess the efficacy of a combination of abamectin (0.2mg/kg), oxibendazole (10mg/kg) and bithionol (5mg/kg or 7mg.kg). In the first study bithionol was included at 5mg/kg and in the second study it was included at 7mg/kg. These studies showed the efficacy of the combination was >98% against adult luminal stages of S. vulgaris, S. edentatus, cyathostomes, migratory stages of S. edentatus and third instar stages of Gasterophilus intestinalis. Efficacy against mucosal stages of cyathostomes (about 64%) and arterial stages of S. vulgaris (71%) was poor. Inclusion of bithionol at 7mg/kg achieved an efficacy of 100% against A. perfoliata whereas at 5mg/kg the efficacy was only 84.6%. In these studies the following species were identified: two species of Strongylus (S. vulgaris and S. edentatus); three species of Triodontophorus (T. serratus, T. minor and T. tenuicollis); and twelve species of cyathostomes (Cyathostomum coronatum, Cyathostomum labiatum, Cyathostomum catinatum, Cylicocyclus nassatus, Cylicocyclus leptostomus, Cylicocyclus radiatus, Cylicocyclus insigne, Cylicostephanus poculatus, Cylicostephanus minutus, Cylicostephanus calicatus, Cylicostephanus longibursatus and Cylicostephanus goldi).Item A survey of anthelmintic resistance and parasite management practices on sheep farms in New Zealand : a dissertation presented in partial fulfilment of the requirements for the degree of Master of Veterinary Studies (Veterinary Parasitology) at Massey University(Massey University, 2004) Sharma, Shyam SundarThe occurrence of anthelmintic resistance in sheep nematodes in New Zealand was conducted by using DrenchRite and Inhouse larval development assays (LDA) on 25 randomly selected farms. Samples from 6 farms were put onto both DrenchRite and Inhouse LDA plates and were available for comparison. Both showed a similar LC50 and LC50 well for benzimidazoles but not for levamisole. Including results from both assay systems it is concluded that anthelmintic resistance in Trichostrongylus (either suspected or high level) was demonstrated in 60% of the farms (9/15) to benzimidazole (BZ), resistance to levamisole (LEV) in 66% of farms (10/15), combination drench (BZ+LEV) on 43% of farms (3/7) and avermectin on 1 of 8 farms. For those farms where Trichostrongylus was the predominant genus there was resistance to at least one anthelmintic on all tested farms (n=12). A survey of parasite control procedures over 2002/2003 was conducted on 38 farms. The principle findings were: about 58% farmers (n=37) performed quarantine drenching of brought-in sheep for which a majority of farmers (52%) used macrocyclic lactones alone or in combination with other anthelmintics; about 78% (n=37) of farmers followed a 5-6 drench programme to lambs/hoggets; the annual frequency of anthelmintic treatments (n=38) were 6.1 drenches to lambs/hoggets; 1.4 to two-tooths and 1.8 to mature ewes; about 71% farmers used macrocyclic lactones for lambs/hoggets on at least one occasion either alone or in combination with other anthelmintics; a majority (61%) of farmers followed a planned annual drench family rotation; about 76.5% (29/38) farmers had never carried out a test for drench resistance and about 57%(4/7) of those farmers who had tested did identify resistance on their farms to either the benzimidazole or combination (benzimidazole + levamisole) drench. Worms were considered "very important" by 55% farmers (n=38) for the cause of economic losses in their farm, whereas 42% farmer considered it "important". Drench resistance was considered as a "serious problem" today for the industry by 47% farmers (n=38), whereas, 34% farmers believe it as a problem but "not serious one today" for the industry.Item Some investigations into the larval development assay and trichostrongylid nematodes of sheep : a thesis presented in partial fulfillment of the requirements for the degree of Master of Veterinary Science at Massey University(Massey University, 1998) Hoza, Shaaban BakariTwo experiments were conducted to investigate the change over time in LD50 values in an in vitro larval development assay. In each experiment, six field-reared Romney lambs were effectively treated with an anthelmintic and housed. In experiment I, six three month old lambs were given a single infection of 35000 infective larvae of T. colubriformis. From ten days post infection (DPI) three lambs (Group 1.1) were treated twice weekly with 0.5mg/kg of dexamethasone trimethylacetate whilst the other three (Group 1.2) served as controls and remained untreated. In experiment II, three lambs (Group 2.2), six month old were infected with a single dose of 22000 infective larvae of T. colubriformis whilst the other three (Group 2.1) of the same age were trickle-infected with 2000 infective larvae once weekly for 14 weeks. Larval development assays were conducted weekly for 14 weeks with ivermectin in Experiment I and ivermectin, avermectinB2 and levamisole in Experiment II. In Experiment I and II for ivermectin, the LD50 values rose to a 4x increase between 50-70 DPIand fell again. The general pattern seen following a single infection with all anthelmintics was for the LD50 values to be relatively constant from 21-35 DPI, then rose 2.5-7x increase to peak 49-56 DPI and declined at the same rate again to original starting values by 84 DPI where they remained until the end of the experiment. In experiments I, the steroid treated group started with similar values but had a 5x fall by 42 DPI which was not seen in the single infection group. The trickle infection group in Experiment II generally resulted in a small increase of 1-1.5x from 42-77 DPI and then declined again to starting values until the remainder of the experiment. The study demonstrates that there is a similar change in the LD50 values with time for T. colubriformis with all three anthelmintics tested and that the change was prevented in steroid treated animals and was less apparent in trickle-infected animals.Item Studies of the pharmacodynamics and modes of action of anthelmintic drugs : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy at Massey University(Massey University, 1977) Min Soe, U.The aim of this work is to extend existing knowledge both with respect to the mode of action of anthelmintics and the biochemical and physiological mechanisms which may be disrupted by drug action. The helminth species examined include nematodes, Ascaris suum, Ascaridia galli and Trichuris ovis and cestodes, Moniezia, T. hydatigena, T. taeniæformis and Echinoccocus granulosus; the anthelmintics studied were methyridine, diethylcarbamazine, pyrantel, morantel, tetramisole, levamisole, dichlorvos, vincofos, cambendazole and mebendazole. The helminth characteristics selected for most intensive study are (a) the occurrence and properties of helminth cholinesterase and (b) the uptake of glucose. The breadth of the study was limited by the availability of fresh material and not all combinations of helminth and drug were investigated. The histochemical localisation of cholinesterase activity in whole mounts and sections of tapeworms using thiocholine esters revealed a complex network of tegumental receptors feeding a nervous system with efferents to suckers, rostellum and hook muscles. It is suggested that tapeworms have reflex arcsinvolving these structures allowing them to maintain their position in the host intestine in spite of peristaltic action. These arcs are susceptible to anticholinesterase anthelmintics. Other cholinesterase activity is associated with the scolex, cirrus, genital pore and sometimes the tegument. High cholinesterase specific activities against acetylthiocholine were measured in Echinococcus scoleces and tapeworms, but lower levels in nematodes. Differential centrifugation of homogenates was used to study their occurrence in the tissue and facilitate further characterisation. However, the enzyme was widely distributed in these species although somewhat higher in the particulate fractions. Activity was increased little, if any, by attempts to solubilise it with the detergent, Triton X-100. Cholinesterase in some fractions particularly from T. ovis, had a high temperature optimum around 60C, but never showed the phenomenon of autoinhibition by substrate at concentrations up to 10-2M. Cholinesterase in species of worm with high levels of enzyme was more sensitive to eserine inhibition than those with lower levels. In studies of glucose uptake from the medium by Ascaris and two tapeworms, it was confirmed that transport into Ascaris was strongly inhibited by certain benzimidazole anthelmintics. Transport into Ascaris, but not the cestodes, was also discovered to be sensitive to local anaesthetics such as procaine or lignocaine. Uptake into tapeworms was inhibited by the absence of sodium ions, phlorizin, iodoacetate and dinitrophenol. It was less inhibited by benzimidazoles and not at all by organophosphate anthelmintics, but was sensitive to phenolic drugs such as hexachlorophene and nitroxynil. In the dog and sheep, a number of anthelmintic drugs administered intravenously showed predominantly nicotinic effects on blood pressure and respiration supporting the cholinergic action of these drugs. Although sheep red-cell cholinesterase is more sensitive to inhibition than that of all helminths tested, the oral route of administration of anthelmintics remains safe for the host and effective against intestinal parasitic worms.Item The effect of host immunity on the development and survival of the free-living stages of common trichostrongylid parasites of sheep : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Veterinary Science at Massey University, Palmerston North, New Zealand(Massey University, 2000) Jørgensen, Lise Tønner; Jørgensen, Lise Tønner; Jorgensen, Lise TonnerThe effect of host immunity on the free-living stages of common trichostrongylid parasites was studied in a series of experiments, involving both artificially infected housed animals and naturally infected animals in the field. In Perendale ewes, bred for either enhanced or lowered resistance to nematodes, reduced developmental success of eggs to infective larvae was found in the resistant animals at some times of the year (p<0.01). This was consistent with the hypothesis of an adverse effect of increased host immunity on the development of the free-living stages of gastrointestinal nematodes. In lambs, this effect had been demonstrated previously and again in 1998, whereas results from 1997 were inconsistent. In fleece-weight selected and control lines of Romney lambs, exposed to the same level of pasture larval challenge, developmental success decreased with time (p<0.001), although the two lines did not differ. This was consistent with an increasing level of host immunity in both lines and provided strong support for the hypothesis of host immunity having an adverse effect on larval development. Nematode eggs from lambs in the field treated orally with either ivermectin or albendazole, did not differ in developmental success, providing no evidence that host immunity was influenced by the type of anthelmintic used. A lower developmental success of O. circumcincta in an LDA (p<0.001) was found in animals relatively immune to this parasite compared to control animals. In faecal cultures a significant difference was not demonstrated, but group sizes were very small. An effect of host immunity on the developmental success and infectivity of larvae of T. colubriformis, could not be demonstrated in trickle-infected groups of lambs that differed in their immunity to this parasite, one group being immunosuppressed with cortico-steroids. An adverse effect of small intestinal mucus and contents on larval development was demonstrated. This was more potent in intestinal contents than mucus. Although source animals differed greatly in their immunity to T. colubriformis, differences between immune and immunosuppressed animals in the magnitude of the effect of intestinal mucus and contents on larval development were not found. The results suggested that the effect of intestinal mucus and contents was not immunological but rather caused by some physical and non-specific properties. Overall, the results reported in this thesis further support the hypothesis of host immunity having an adverse effect on the development of the free-living stages of gastrointestinal nematodes of sheep. This was most evident in animals with naturally acquired infections and in housed animals infected with O. circumcincta. It is suggested that the failure to demonstrate this in experimental infections with T. colubriformis may have been due to the use of cortico-steroids to suppress immune responses.Item Larval paralysis factor of sheep : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Animal Science at Massey University, Palmerston North, New Zealand(Massey University, 2008) Soe, Aye KyawtThis study aimed to identify the origin and molecular nature of larval paralysis factor (LPF), an uncharacterised natural anthelmintic agent(s) known to be secreted by cells in the small intestine of nematode-parasite-immune sheep. The study first confirmed previous findings that gut mucus and small intestinal mucosal cell culture supernatants (CCSs) from parasite-immune sheep contain LPF. An in vitro larval migration inhibition (LMI) assay showed that LPF is active against numerous nematode parasite larvae of ruminants and adults of Trichostrongylus colubriformis. Lamina propria cells (LPCs) were a richer source of LPF than epithelial cells, and release of the factor was specifically triggered by parasite larval antigens. A series of trials was performed to optimise LPF production in vitro. LPF production was highest when LPCs were selectively extracted from the first three metres of small intestine of resistant-line or hyperimmune parasitized sheep three days after oral challenge with T. colubriformis larvae. LPF release in vitro appeared to be related to the percentage of eosinophils present in cell cultures, but not with mucosal mast cells or globule leucocytes. Subsequent in vitro experiments showed that mucus glycoproteins released from goblet cells enhanced and sustained the activity of LPF, which may point to a previously unrecognised effector link between goblet cell hyperplasia and mucosal immune responses to gut nematode parasites. Sequential purification and molecular analysis of LPF showed that the active agent has a molecular mass less than 1kDa, it is polar and heat and enzyme resistant. Further purification using solid phase extraction and HPLC established that LPF did not bind either C18 or ion exchangers, but LPF activity in the flow through from these sorbents was retained on aminopropyl HPLC columns and eluted as a single active peak after flushing with acetonitrile/water mixture (70:30). This peak, analysed by LC-MS, comprised three main compounds of interest: a UV absorbing compound at 254nm with no MS data, and two non UV absorbing compounds at 254nm: one was a component of m/z 104 (ESI+ve) and the other was detected at m/z 113 (ESI-ve).