International Journal for Parasitology: Drugs and Drug Resistance最新文献

Surveillance data for Ancylostoma spp. and the A. caninum benzimidazole treatment resistance associated F167Y polymorphism using molecular diagnostics was obtained in a large population of dogs from the United States and Canada. Real-time PCR (qPCR) for Ancylostoma spp. and allele-specific qPCR detecting a single nucleotide polymorphism (SNP) F167Y was used in 262,872 canine stool samples collected between March and December of 2022. Ancylostoma spp. was found at an overall prevalence of 2.5% (6538/262,872), with the highest prevalence in the Southern US, 4.4% (4490/103,095), and the lowest prevalence in Canada 0.6% (101/15,829). The A. caninum F167Y polymorphism was found with the highest prevalence (13.4%, n = 46/343) in the Western US and the lowest in Canada at 4.1% (4/97). The F167Y polymorphism was detected every month over the 10-month collection period. Seasonal distribution showed a peak in June for both Ancylostoma spp. (3.08%, 547/17,775) and A. caninum F167Y (12.25%, 67/547). However, the A. caninum F167Y polymorphism prevalence was highest in September (13.9%, 119/856). Age analysis indicates a higher prevalence of both hookworm infections and occurrence of resistant isolates in puppies. The breeds with the highest F167Y polymorphism prevalence in Ancylostoma spp. detected samples were poodles (28.9%), followed by Bernese Mountain dogs (25%), Cocker spaniels (23.1%), and greyhounds (22.4%). Our data set describes widespread geographic distribution of the A. caninum benzimidazole resistance associated F167Y polymorphism in the United States and Canada, with no clear seasonality compared to the Ancylostoma spp. prevalence patterns. The F167 polymorphism was present in all geographic areas with detected hookworms, including Canada. Our study highlights that the F167Y polymorphism is represented in many dog breeds, including greyhounds.

Rhipicephalus (Boophilus) microplus is one of the most successful ticks infesting cattle around the world. This highly-invasive species transmits cattle parasites that cause cattle fever leading to a high socio-economic burden. Tick eradication programs have often failed, due to the development of acaricide resistance. Here we characterize acaricide resistance in a large number of tick isolates from regions in South Africa (KwaZulu Natal, Mpumalanga, Western & Eastern Cape provinces) and two Brazilian regions.

By means of Larval Packet Tests (LPT's) acaricide resistance was evaluated against five commonly used acaricides (chlorfenvinphos, fipronil, deltamethrin, amitraz, and ivermectin). Furthermore, the coding region containing the knock down resistance (kdr) mutation, known to result in pyrethroid resistance, was sequenced.

Resistance to at least one acaricide class was reported in each of the five regions, and a high proportion of tick isolates exhibited multi-resistance to at least two acaricide classes (range: 22.2–80.0%). Furthermore, resistance ratios (RR) showed high spatial variation (intercontinental, as well as regional) but low regional spatial autocorrelation. Previous and current acaricide use correlated with current RR, and several combinations of acaricide RR were positively correlated. Moreover, fipronil resistance tended to be higher in farms with more intense acaricide use. The kdr-mutations provided the ticks a fitness advantage under the selection pressure of synthetic pyrethroids based on population (kdr-allele frequency) and individual level data (genotypes).

The data show the threat of acaricide (multi-)resistance is high in Brazil and South Africa, but acaricide specific levels need to be assessed locally. For this purpose, gathering complementary molecular information on mutations that underlie resistance can reduce costs and expedite necessary actions. In an era of human-caused habitat alterations, implementing molecular data-driven programs becomes essential in overcoming tick-induced socio-economic losses.

The physiological significance of metabotropic acetylcholine receptors in parasitic nematodes remains largely unexplored. Here, three different Trichinella spiralis G protein-coupled acetylcholine receptors (TsGAR-1, -2, and -3) were identified in the genome of T. spiralis. The phylogenetic analyses showed that TsGAR-1 and -2 receptors belong to a distinct clade specific to invertebrates, while TsGAR-3 is closest to the cluster of mammalian-type muscarinic acetylcholine receptors (mAChR). The mRNA of TsGAR-1, -2, and -3 was detected in muscle larvae, newborn larvae, and adults. The functional aequorin-based assay in Chinese hamster ovary cells revealed that all three types of T. spiralis GARs trigger the G_q/11 pathway upon activation of the receptor with the acetylcholine ligand. TsGAR-1 and TsGAR-2 showed atypical affinity with classical muscarinic agonists, while TsGAR-3 was sensitive to all muscarinic agonists tested. High concentrations of propiverine antagonist blocked the activities of all three TsGARs, while atropine and scopolamine antagonists effectively inhibited only TsGAR-3. Our data indicate that the distinct pharmacological profile of TsGAR-1 and -2 receptors, as well as the phylogenetic distance between them and their mammalian orthologs, place them as attractive targets for the development of selective anthelmintic drugs interfering with nematodes’ cholinergic system.

Plasmodium falciparum is the deadliest causative agent of human malaria. This parasite has historically developed resistance to most drugs, including the current frontline treatments, so new therapeutic targets are needed. Our previous work on guanine quadruplexes (G4s) in the parasite's DNA and RNA has highlighted their influence on parasite biology, and revealed G4 stabilising compounds as promising candidates for repositioning. In particular, quarfloxin, a former anticancer agent, kills blood-stage parasites at all developmental stages, with fast rates of kill and nanomolar potency. Here we explored the molecular mechanism of quarfloxin and its related derivative CX-5461. In vitro, both compounds bound to P. falciparum-encoded G4 sequences. In cellulo, quarfloxin was more potent than CX-5461, and could prevent establishment of blood-stage malaria in vivo in a murine model. CX-5461 showed clear DNA damaging activity, as reported in human cells, while quarfloxin caused weaker signatures of DNA damage. Both compounds caused transcriptional dysregulation in the parasite, but the affected genes were largely different, again suggesting different modes of action. Therefore, CX-5461 may act primarily as a DNA damaging agent in both Plasmodium parasites and mammalian cells, whereas the complete antimalarial mode of action of quarfloxin may be parasite-specific and remains somewhat elusive.

Toxoplasma gondii is an obligate intracellular parasite in the phylum Apicomplexa that causes toxoplasmosis in humans and animals worldwide. Despite its prevalence, there is currently no effective vaccine or treatment for chronic infection. Although there are therapies against the acute stage, prolonged use is toxic and poorly tolerated. This study aims to explore the potential of repurposing topotecan and 10-hydroxycamptothecin (HCPT) as drugs producing double strand breaks (DSBs) in T. gondii. DSBs are mainly repaired by Homologous Recombination Repair (HRR) and Non-Homologous End Joining (NHEJ). Two T. gondii strains, RHΔHXGPRT and RHΔKU80, were used to compare the drug's effects on parasites. RHΔHXGPRT parasites may use both HRR and NHEJ pathways but RHΔKU80 lacks the KU80 protein needed for NHEJ, leaving only the HRR pathway. Here we demonstrate that topotecan and HCPT, both topoisomerase I venoms, affected parasite replication in a concentration-dependent manner. Moreover, variations in fluorescence intensity measurements for the H2A.X phosphorylation mark (γH2A.X), an indicator of DNA damage, were observed in intracellular parasites under drug treatment conditions. Interestingly, intracellular replicative parasites without drug treatment show a strong positive staining for γH2A.X, suggesting inherent DNA damage. Extracellular (non-replicating) parasites did not exhibit γH2A.X staining, indicating that the basal level of DNA damage is likely to be associated with replicative stress. A high rate of DNA replication stress possibly prompted the evolution of an efficient repair machinery in the parasite, making it an attractive target. Our findings show that topoisomerase 1 venoms are effective antiparasitics blocking T. gondii replication.

This study quantified the extent of anthelmintic resistance (AR) in ascarid and strongylid nematodes against commonly used anthelmintics in Australian Thoroughbred horses. Faecal egg count reduction tests (FECRTs, n = 86) and egg reappearance period (ERP) tests were conducted on 22 farms across Australia. Faecal egg counts (FECs) were determined using the modified McMaster technique, and percent faecal egg count reduction (%FECR) was calculated using the Bayesian hierarchical model and hybrid Frequentist/Bayesian analysis method. The results were interpreted using old (published in 1992) and new (2023) research guidelines of the World Association for the Advancement of Veterinary Parasitology (WAAVP). The species composition of strongylid nematodes was detected utilising a DNA-metabarcoding method using pre- and post-treatment samples. Resistance was observed in strongylid nematodes to commonly used single-active and combination anthelmintics, including ivermectin (IVM %FECR range: 82%–92%; 95% lower credible interval (LCI) range: 80%–90%), abamectin (ABM: 73%–92%; 65%–88%), moxidectin (MOX: 89%–91%; 84%–89%), oxfendazole (OFZ: 0%–56%; 0%–31%) and its combination with pyrantel (OFZ + PYR: 0%–82%; 0%–78%). Resistance in Parascaris spp. was observed to IVM (10%–43%; 0%–36%), ABM (0%; 0%) and MOX (0%; 0%). When the new thresholds recommended by the WAAVP were used, AR was detected in six additional FECRTs for strongylids and three more tests for Parascaris spp., introducing resistance to OFZ and OFZ + PYR in the latter. Shortened ERPs (4–6 weeks) of strongylids were observed in 31 FECRTs in which AR was not detected at 2 weeks post-treatment for all the anthelmintics tested. Among cyathostomins, Cylicocyclus nassatus, Cylicostephanus longibursatus and Coronocyclus coronatus were the most prevalent species at 2 weeks post-treatment, whereas the main species appearing at five weeks following treatments with macrocyclic lactones were Cylicocyclus nassatus, Cylicostephanus longibursatus and Cylicocyclus ashworthi. After treatment with OFZ + PYR, the latter three, plus Coronocyclus coronatus and Cyathostomum catinatum, were detected at 5 weeks post-treatment. Overall, the study highlights the prevalence of AR in both ascarids and strongylid nematodes against commonly used anthelmintic products to control worms in Australian horses. The results indicate that ML combination products provided acceptable efficacy at 2 weeks. However, ERP calculations suggest that products work less effectively than previously measured. It is suggested to regularly monitor the efficacy of the anthelmintics and consider changing the worm control practices to better manage worms and AR in Australian horses.

Malaria, an infectious disease with a tremendous impact on human health is caused by Plasmodium parasites, and transmitted by Anopheles mosquitoes. New approaches to control the disease involve transmission blocking strategies aiming to target the parasite in the mosquito. Here, we investigated the putative inhibitory activity of essential oils and their components on the early mosquito stages of the parasite. We employed an in vitro assay of gametocyte-to-ookinete development of the rodent model parasite Plasmodium berghei combined with high content screening. 60 essential oils with known composition were tested. The results revealed that fifteen EOs had inhibitory activity. Furthermore, a machine learning approach was used to identify the putative inhibitory components. Five of the most important chemical components indicated by the machine learning-based models were actually confirmed by the experimental approach. This combined approach was used for the first time to identify the potential transmission blocking activity of essential oils and single components at the zygote and ookinete stages.

Fasciola hepatica infections lead to severe health problems and production losses in sheep farming, if not treated effectively. Triclabendazole has been used extensively over decades due to its unique efficacy range against all definitive hostfluke stages but published data about the susceptibility of F. hepatica to anthelmintics in Germany are lacking. This study aimed to identify current F. hepatica infections in German sheep flocks by coproscopic examinations and to evaluate the efficacy of anthelmintics with a focus on triclabendazole in a field study conducted from 2020 to 2022. Initial screening included 71 sheep farms, many of them with known history of fasciolosis. In this highly biased sample set, the frequency of F. hepatica infection at individual sheep and farm level were 12.8% and 35.2%, respectively. Additionally, eggs of Paramphistominae were found at frequencies of 4.8% and 15.5% at individual sheep and farm level, respectively. Due to low egg shedding intensity, faecal egg count reduction (FECR) tests could only be conducted on a few farms. The efficacy of triclabendazole was tested on 11 farms and albendazole on one farm, including 3–53 sheep/farm. Individual faecal samples were collected before and two weeks after treatment to evaluate the FECR using the sedimentation or FLUKEFINDER® or a modified FLUKEFINDER® method. On all farms a coproantigen reduction test was conducted in parallel. Lacking efficacy of triclabendazole even at double dosage was shown on one farm associated with a high number of animal losses due to acute fasciolosis. On this farm, the Fasciola miracidium development test was additionally performed, revealing a high in vitro ovicidal activity of albendazole while closantel was effective in vivo. On all other farms, sufficient efficacy of triclabendazole was observed. In conclusion, triclabendazole resistance appears not to be widespread on German sheep farms but, when present, can have serious effects on animal health.

Consisting of approximately 50 different species, the cyathostomin parasites are ubiquitous in grazing horses. Co-infection with several species is common, and large burdens can cause the fatal disease of larval cyathostominosis. Due to intense anthelmintic drug use, cyathostomin resistance has developed to all available anthelmintic drug groups. Resistance to the anthelmintic drug pyrantel (PYR) has been documented in over 90% of studies published over the past two decades. In Sweden, a study performed in the early 2000s only confirmed resistance in 4.5% of farms. Further, prescription-only administration of equine anthelmintic drugs was enforced in Sweden in 2007. However, it is unknown if this conservative drug use has maintained PYR efficacy in cyathostomins. The aim of the present study was to investigate the effect of PYR on cyathostomin infection in Sweden using fecal egg count reduction tests (FECRTs). Further, the effect of PYR treatment on cyathostomin species composition was studied using metabarcoding.

Sixteen farms with at least six horses excreting a minimum of 100 eggs per gram feces were included. Using the current World Association for the Advancement of Veterinary Parasitology (WAAVP) guidelines, PYR resistance was demonstrated in nine of farms, with seven farms showing full susceptibility. Farms with low biosecurity measures had significantly lower efficacy of PYR treatment. The most common cyathostomin species were Cylicocyclus nassatus, Cyathostomum catinatum, Cylicostephanus longibursatus, Cys. calicatus, Cys. goldi, Cys. minutus, Coronocyclus coronatus and Cya. pateratum, accounting for 97% of all sequence reads prior to treatment. Of these, Cyc. nassatus and Cya. catinatum had the highest occurrence, accounting for 68% of all sequence reads prior to PYR treatment. Treatment did not significantly affect the species composition. The results highlight the importance of drug efficacy testing when using PYR to treat cyathostomin infection, even when selective anthelmintic treatment and thus low treatment intensity, is used on the farm.

East Coast Fever (ECF) is a disease affecting cattle in sub-Saharan Africa, caused by the tick-borne Apicomplexan pathogen Theileria parva. The disease is a major problem for cattle farmers in affected regions and there are few methods of control, including a complex infection and treatment vaccine, expensive chemotherapy, and the more widespread tick control through acaricides. New intervention strategies are, therefore, sorely needed. Benzoxaboroles are a versatile class of boron-heterocyclic compounds with demonstrable pharmacological activity against a diverse group of pathogens, including those related to T. parva. In this study, the in vitro efficacy of three benzoxaboroles against the intracellular schizont stage of T. parva was investigated using a flow cytometry approach. Of the benzoxaboroles tested, only one showed any potency, albeit only at high concentrations, even though there is high protein sequence similarity in the CPSF3 protein target compared to other protozoan pathogen species. This finding suggests that benzoxaboroles currently of interest for the treatment of African animal trypanosomiasis, toxoplasmosis, cryptosporidiosis and malaria may not be suitable for the treatment of ECF. We conclude that testing of further benzoxaborole compounds is needed to fully determine whether any lead compounds can be identified to target T. parva.