Novel approaches to the control of cyathostomins in equids

Cyathostomins, are clade 5 gastrointestinal (GI) nematodes infecting equids. They are associated with a range of pathologies, the most serious of which is larval cyathostominosis, a protein losing enteropathy with a 50 % mortality rate. Importantly, they are the most abundant GI nematode of equids in the developed world and, as they do not induce protective immunity, equids remain at risk of infection throughout their lives. The effective control of cyathostomins is currently threatened by the development of anthelmintic resistance (AR) to all three major classes of anthelmintic licenced for use in equids. Of primary concern is the emerging resistance to the macrocyclic lactones (MLs), which are the mainstay of cyathostomin control. It is therefore important that novel options for controlling cyathostomins are explored. In this study in vitro tests were used to explore a range of options for developing novel treatments against AR cyathostomins and improving the efficacy of the widely used ML, ivermectin (IVM). First the potential use of ethnoveterinary medicines for treatment of cyathostomins in equids in Ethiopia and the UK was explored (Chapters 3, 4 and 5). A participatory rural appraisal was performed in the Oromia region of Ethiopia to identify plants currently used as anthelmintics in equids and other livestock. A total of 37 species of plant were identified, data on dosing and side effects were also recorded. These data were triangulated with a literature review to identify five plants for in vitro screening for anthelmintic activity against cyathostomins. A literature review was used to identify plants for in vitro screening in the UK. A total of 138 publications, reporting the anthelmintic activity of plant extracts against nematodes, were found. The data collected was used to design a ranking system to provide a shortlist of five plant candidates. Shortlisted plants were collected, dried and chemically extracted to give crude extracts, which were screened against cyathostomins in the egg hatch test (EHT) and larval migration inhibition test (LMIT). A total of 7/9 plants screened in this way showed anthelmintic activity in either the EHT and/or the LMIT with median effective concentrations (EC-50s) in the range 0.18-8.90 mg/ml. For some of the most efficacious extracts there were reports of in vivo efficacy against GI nematodes of other veterinary parasitic nematodes, making these good candidates for future in vivo trials. One of the plants shortlisted for the UK, for which there was extensive evidence of anthelmintic activity in the literature, was the cysteine proteinase (CP) containing Carica papaya. Extract from this species, papaya latex supernatant (PLS), could not be processed and tested using the same protocols as the other plant species, as the active compound is an enzyme. The EHT and LMIT were optimised for use with PLS and its efficacy evaluated in chapter 5. It was found that PLS had a potent anthelmintic effect against cyathostomins in the EHT, which was attributable to the action of CPs, with EC-50 values in repeats ranging between 0.12-0.22 µM. PLS also showed efficacy in the LMIT, although this appeared due to another active compound. To the authors knowledge this is the first report of efficacy of PLS against the free living stages of any parasitic nematode, which may indicate that cyathostomins are particularly susceptible to CPs. In chapter 6 the role of P-glycoproteins (P-gps) in IVM resistance in cyathostomins was investigated, with the more specific aim of assessing whether P-gp inhibitors could potentially be used in combination with IVM to improve its efficacy against AR parasites. The transcription of pgp-9, which is putatively associated with IVM resistance in Teladorsagia circumcincta and Haemonchus contortus, was measured after exposure to IVM in in ML resistant (AR) versus anthelmintic naïve (AN) cyathostomin third stage larvae (L3). Pgp-9 expression was significantly increased after incubation with IVM in the AR, but not the AN, parasite population. Two in vitro tests, the LMIT and the larval development test (LDT), were used to assess the effect of a range of P-gp inhibitors (ketoconazole, pluronic 85 and ivermectin aglycone) on IVM efficacy in ML resistant (AR) versus anthelmintic naïve (AN) cyathostomins. In the LMIT ketoconazole and ivermectin aglycone conferred a significant increase in IVM efficacy in the AR parasites, but had less effect on the AN parasites. Pluronic 85 also had a profound effect on IVM efficacy which depended on IVM concentration, however there was no differential effect dependent on parasite population. In the LDT, ketoconazole and pluronic 85 both significantly increased IVM efficacy in both AR and AN parasites. These results suggest that there is a lifecycle stage-specific association of P-gps with AR cyathostomins, and that P-gp inhibitors could potentially be used to improve IVM efficacy in AR cyathostomin populations. In summary in this study has identified a range of potential novel treatments for cyathostomins. The data also suggest a role for P-gps in the emerging resistance to MLs, which could potentially be exploited to improve ML efficacy in AR cyathostomin populations. Importantly, this work will underpin the financial and ethical justification for future in vivo trials on the extracts/compounds tested here

This Gut Ain't Big Enough for Both of Us. Or Is It? Helminth-Microbiota Interactions in Veterinary Species

Gastrointestinal helminth parasites share their habitat with a myriad of other organisms, that is, the commensal microbiota. Increasing evidence, particularly in humans and rodent models of helminth infection, points towards a multitude of interactions occurring between parasites and the gut microbiota, with a profound impact on both host immunity and metabolic potential. Despite this information, the exploration of the effects that parasite infections exert on populations of commensal gut microbes of veterinary species is a field of research in its infancy. In this article, we summarise studies that have contributed to current knowledge of helminth-microbiota interactions in species of veterinary interest, and identify possible avenues for future research in this area, which could include the exploitation of such relationships to improve parasite control and delay or prevent the development of anthelmintic resistance

The first report of macrocyclic lactone resistant cyathostomins in the UK.

In recent years, resistance to the benzimidazole (BZ) and tetrahydropyrimidine (PYR) anthelmintics in global cyathostomin populations, has led to reliance on the macrocyclic lactone drugs (ML-of which ivermectin and moxidectin are licensed in horses) to control these parasites. Recently, the first confirmed case of resistance to both ivermectin (IVM) and moxidectin (MOX) was reported in the USA in yearlings imported from Ireland. This suggests that ML resistance in cyathostomins has emerged, and raises the possibility that regular movement of horses may result in rapid spread of ML resistant cyathostomins. Resistance may go undetected due to a lack of surveillance for ML efficacy. Here, we report anthelmintic efficacies in cyathostomins infecting UK Thoroughbreds on four studs. Faecal egg count reduction tests (FECRT) were performed to define resistance (resistance = FECR <95% lower credible interval (LCI) < 90%). Stud A yearlings had FECRs of 36.4-78.6% (CI:15.7-86.3) after three IVM treatments, 72.6% (CI: 50.8-85.2) after MOX, and 80.8% (CI: 61.9-90.0) after PYR. Mares on stud A had a FECR of 97.8% (CI: 93.3-99.9) and 98% (95.1-99.4) after IVM and MOX treatment, respectively. Resistance to MLs was not found in yearlings or mares on studs B, C or D with FECR after MOX OR IVM treatment ranging from 99.8 to 99.9% (95.4-100); although yearlings on studs B, C and D all had an egg reappearance period (ERP) of six weeks for MOX and stud C had a four-week ERP for IVM. This study describes the first confirmed case of resistance to both licensed ML drugs on a UK Thoroughbred stud and highlights the urgent need for a) increased awareness of the threat of ML resistant parasites infecting horses, and b) extensive surveillance of ML efficacy against cyathostomin populations in the UK, to gauge the extent of the problem

Infections by human gastrointestinal helminths are associated with changes in faecal microbiota diversity and composition

Investigations of the impact that patent infections by soil-transmitted gastrointestinal nematode parasites exert on the composition of the host gut commensal flora are attracting growing interest by the scientific community. However, information collected to date varies across experiments, and further studies are needed to identify consistent relationships between parasites and commensal microbial species. Here, we explore the qualitative and quantitative differences between the microbial community profiles of cohorts of human volunteers from Sri Lanka with patent infection by one or more parasitic nematode species (H+), as well as that of uninfected subjects (H-) and of volunteers who had been subjected to regular prophylactic anthelmintic treatment (Ht). High-throughput sequencing of the bacterial 16S rRNA gene, followed by bioinformatics and biostatistical analyses of sequence data revealed no significant differences in alpha diversity (Shannon) and richness between groups (P = 0.65, P = 0.13 respectively); however, beta diversity was significantly increased in H+ and Ht when individually compared to H-volunteers (P = 0.04). Among others, bacteria of the families Verrucomicrobiaceae and Enterobacteriaceae showed a trend towards increased abundance in H+, whereas the Leuconostocaceae and Bacteroidaceae showed a relative increase in H- and Ht respectively. Our findings add valuable knowledge to the vast, and yet little explored, research field of parasite - microbiota interactions and will provide a basis for the elucidation of the role such interactions play in pathogenic and immune-modulatory properties of parasitic nematodes in both human and animal hosts.This work was supported by grants by University of Peradeniya, grant no. URG/2016/88/S to RSR and PKP; the Wellcome Trust/ Isaac Newton Trust/ University of Cambridge to CC; a Postgraduate Award by the Biotechnology and Biological Sciences Research Council to TPJ

The dynamic geophysical environment of (101955) Bennu based on OSIRIS-REx measurements

The top-shaped morphology characteristic of asteroid (101955) Bennu, often found among fast-spinning asteroids and binary asteroid primaries, may have contributed substantially to binary asteroid formation. Yet a detailed geophysical analysis of this morphology for a fast-spinning asteroid has not been possible prior to the Origins, Spectral Interpretation, Resource Identification, and Security-Regolith Explorer (OSIRIS-REx) mission. Combining the measured Bennu mass and shape obtained during the Preliminary Survey phase of the OSIRIS-REx mission, we find a notable transition in Bennu’s surface slopes within its rotational Roche lobe, defined as the region where material is energetically trapped to the surface. As the intersection of the rotational Roche lobe with Bennu’s surface has been most recently migrating towards its equator (given Bennu’s increasing spin rate), we infer that Bennu’s surface slopes have been changing across its surface within the last million years. We also find evidence for substantial density heterogeneity within this body, suggesting that its interior is a mixture of voids and boulders. The presence of such heterogeneity and Bennu’s top shape are consistent with spin-induced failure at some point in its past, although the manner of its failure cannot yet be determined. Future measurements by the OSIRIS-REx spacecraft will provide insight into and may resolve questions regarding the formation and evolution of Bennu’s top-shape morphology and its link to the formation of binary asteroids

Evidence for widespread hydrated minerals on asteroid (101955) Bennu

Early spectral data from the Origins, Spectral Interpretation, Resource Identification, and Security-Regolith Explorer (OSIRIS-REx) mission reveal evidence for abundant hydrated minerals on the surface of near-Earth asteroid (101955) Bennu in the form of a near-infrared absorption near 2.7 µm and thermal infrared spectral features that are most similar to those of aqueously altered CM-type carbonaceous chondrites. We observe these spectral features across the surface of Bennu, and there is no evidence of substantial rotational variability at the spatial scales of tens to hundreds of metres observed to date. In the visible and near-infrared (0.4 to 2.4 µm) Bennu’s spectrum appears featureless and with a blue (negative) slope, confirming previous ground-based observations. Bennu may represent a class of objects that could have brought volatiles and organic chemistry to Earth