Echinococcus multilocularis infection affects risk-taking behaviour in Microtus arvalis: adaptive manipulation?

Manipulation of host behaviour by parasites to enhance transmission to the next host is a fascinating phenomenon that has interested scientists since the 1970s. It has been proposed that infection with the cestode Echinococcus multilocularis produces an impairment of the antipredatory behaviour in the rodent intermediate host common vole, Microtus arvalis, which may facilitate transmission of the tapeworm to the canid final host. In this study, we observed the behaviour of infected common voles at 12 weeks post-infection, when protoscoleces production and maturation commonly occurs, in order to assess behavioural changes compared to uninfected controls, that might ease predation in the wild. Infected and uninfected voles were monitored for 24 h to observe their spontaneous activity. In addition, the next day, both infected and uninfected voles were subjected to 4 different behavioural tests: open field test, barrier test, platform test and air-puff test in a running wheel. No significant difference between uninfected and infected voles emerged during the behavioural tests. However, observation of spontaneous activity revealed that infected voles increased their feeding frequency and spent significantly more time above bedding even when not eating, compared to the uninfected controls. In the wild, these behavioural changes increase the animals exposure to predators, raising their chance of becoming prey. These findings are the first direct evidence consistent with behavioural manipulation by E. multilocularis on common voles.

Intestinal infection with Echinococcus multilocularis in a dog.

OBJECTIVE: To raise veterinary awareness of a newly recognized parasitic threat to canine and human health, highlight the increasing availability of molecular parasitological diagnostics and the need to implement best practices of cestocidal use in high-risk dogs. ANIMAL: A young Boxer dog with vomiting and bloody diarrhea, suspected diagnosis of inflammatory bowel disease. CLINICAL PRESENTATION, PROGRESSION, AND PROCEDURES: Bloodwork revealed inflammation, dehydration, and protein loss, addressed with supportive therapy. Fecal culture revealed only Escherichia coli. On centrifugal flotation, tapeworm eggs (which could be Taenia or Echinococcus spp) and, unusually, adult cestodes of Echinococcus were observed. The referring veterinarian was contacted to initiate immediate treatment with a cestocide due to zoonotic potential. Diagnosis was confirmed with a coproPCR which has higher sensitivity for Echinococcus spp than fecal flotation alone. DNA was identical to an introduced European strain of E multilocularis currently emerging in dogs, people, and wildlife. Since dogs can also self-infect and develop hepatic alveolar echinococcosis (severe and often fatal), this was ruled out using serology and abdominal ultrasound. TREATMENT AND OUTCOME: Following cestocidal treatment, fecal flotation and coproPCR were negative for eggs and DNA of E multilocularis; however, coccidia were detected and diarrhea resolved following treatment with sulfa-based antibiotics. CLINICAL RELEVANCE: This dog was serendipitously diagnosed with E multilocularis, acquired through ingestion of a rodent intermediate host likely infected from foxes and coyotes. Therefore, as a dog at high risk of reexposure from eating rodents, regular (ideally monthly) treatment with a labeled cestocide is indicated going forward.

Current and future distribution of a parasite with complex life cycle under global change scenarios: Echinococcus multilocularis in Europe.

Global change is expected to have complex effects on the distribution and transmission patterns of zoonotic parasites. Modelling habitat suitability for parasites with complex life cycles is essential to further our understanding of how disease systems respond to environmental changes, and to make spatial predictions of their future distributions. However, the limited availability of high quality occurrence data with high spatial resolution often constrains these investigations. Using 449 reliable occurrence records for Echinococcus multilocularis from across Europe published over the last 35 years, we modelled habitat suitability for this parasite, the aetiological agent of alveolar echinococcosis, in order to describe its environmental niche, predict its current and future distribution under three global change scenarios, and quantify the probability of occurrence for each European country. Using a machine learning approach, we developed large-scale (25 × 25 km) species distribution models based on seven sets of predictors, each set representing a distinct biological hypothesis supported by current knowledge of the autecology of the parasite. The best-supported hypothesis included climatic, orographic and land-use/land-cover variables such as the temperature of the coldest quarter, forest cover, urban cover and the precipitation seasonality. Future projections suggested the appearance of highly suitable areas for E. multilocularis towards northern latitudes and in the whole Alpine region under all scenarios, while decreases in habitat suitability were predicted for central Europe. Our spatially explicit predictions of habitat suitability shed light on the complex responses of parasites to ongoing global changes.

Echinococcus multilocularis and other cestodes in red foxes (Vulpes vulpes) of northeast Italy, 2012-2018.

BACKGROUND: Echinococcus multilocularis is a small tapeworm affecting wild and domestic carnivores and voles in a typical prey-predator life cycle. In Italy, there has been a focus of E. multilocularis since 1997 in the northern Italian Alps, later confirmed in red foxes collected from 2001 to 2005. In this study, we report the results of seven years of monitoring on E. multilocularis and other cestodes in foxes and describe the changes that occurred over time and among areas (eco-regions) showing different environmental and ecological features on a large scale. METHODS: Eggs of cestodes were isolated from feces of 2872 foxes with a sedimentation/filtration technique. The cestode species was determined through multiplex PCR, targeting and sequencing ND1 and 12S genes. Analyses were aimed to highlight variations among different eco-regions and trends in prevalence across the study years. RESULTS: Out of 2872 foxes, 217 (7.55%) samples resulted positive for cestode eggs at coproscopy, with differences of prevalence according to year, sampling area and age class. Eight species of cestodes were identified, with Taenia crassiceps (2.65%), Taenia polyacantha (1.98%) and E. multilocularis (1.04%) as the most represented. The other species, Mesocestoides litteratus, Taenia krabbei, T. serialis, T. taeniaeformis and Dipylidium caninum, accounted for < 1% altogether. Echinococcus multilocularis was identified in foxes from two out of six eco-regions, in 30 fecal samples, accounting for 1.04% within the cestode positives at coproscopy. All E. multilocularis isolates came from Bolzano province. Prevalence of cestodes, both collectively and for each of the three most represented species (T. crassiceps, T. polyacantha and E. multilocularis), varied based on the sampling year, and for E. multilocularis an apparent increasing trend across the last few years was evidenced. CONCLUSIONS: Our study confirms the presence of a focus of E. multilocularis in red foxes of northeast Italy. Although this focus seems still spatially limited, given its persistence and apparent increasing prevalence through the years, we recommend research to be conducted in the future on the ecological factors that, on a smaller scale, allow this zoonotic species to persist. On the same scale, we recommend a health education campaign to inform on the measures to prevent this zoonosis, targeted at people living in the area, especially hunters, dog owners, forestry workers and other potentially exposed categories.

Characterization of microRNAs expressed in the cystic legion of the liver of Mus musculus perorally infected with Echinococcus multilocularis Nemuro strain.

Alveolar echinococcosis (AE) is a zoonosis caused by the metacestode of Echinococcus multilocularis. The published genome of E. multilocularis showed that approximately 86% of its genome is non-coding. Micro RNAs (miRNAs) are small non-coding regulatory RNAs, and recent studies on parasitic helminths expect miRNAs as a promising target for drug development and diagnostic markers. Prior to this study, only a few studies reported the E. multilocularis miRNA profiles in the intermediate host. The primary objective of this study was to characterize miRNA profiles via small RNA-seq in E. multilocularis Nemuro strain, a laboratory strain of Asian genotype, using mice perorally infected with the parasite eggs. The data were then compared with two previously published small RNA-seq data. We identified 44 mature miRNAs as E. multilocularis origin out of the 68 mature miRNA sequences registered in the miRNA database miRbase. The highest quantities of miRNAs detected were miR-10-5p, followed by bantam-3p, let-7-5p, miR-61-3p, and miR-71-5p. The top two most abundant miRNAs (miR-10-5p and bantam-3p) accounted for approximately 80.9% of the total parasite miRNAs. The highly expressed miRNA repertoire is mostly comparable to that obtained from the previous experiment using secondary echinococcosis created by an intraperitoneal administration of metacestodes. A detailed characterization and functional annotations of these shared miRNAs will lead to a better understanding of parasitic dynamics, which could provide a basis for the development of novel diagnostic and treatment methods for AE.

Echinococcus multilocularis in red foxes in North Belgium: Prevalence and trends in distribution.

A cross-sectional systematic sampling was carried out during three consecutive winters from 2012 to 2015, to update the knowledge on the fox tapeworm (Echinococcus multilocularis) distribution in the red fox (Vulpes vulpes) in Flanders. Earlier studies reported the low endemicity status of this tapeworm in the northern region of Belgium, in contrast to the south of the country and neighbouring countries. Using a modified Segmental Sedimentation and Counting Technique, followed by PCR-RFLP and sequencing, 923 foxes' intestines were examined for the presence of E. multilocularis. Based on microscopic examination, 38 out of 923 foxes were suspected to be infected with either E. multilocularis or Amoebotaenia spp., of which 19 were molecularly confirmed to be E. multilocularis, 18 were found positive for Amoebotaenia spp. and one was negative. The overall prevalence for E. multilocularis of 2.1% confirms the low endemicity of the fox tapeworm in Flanders. However, in one area in the most eastern part of Flanders (Voeren), neighbouring the Netherlands and Wallonia, a prevalence of 57% (12/21) was observed. Continuous monitoring of the fox tapeworm remains needed to assess spatio-temporal trends in distribution and to assess the risk of this zoonotic infection in Europe. The challenging differential diagnosis of E. multilocularis and Amoebotaenia spp. based on microscopic examination calls for attention.

Spatial distribution and incidence trend of human alveolar echinococcosis in southwest Germany: increased incidence and urbanization of the disease?

Ecology and epidemiology of Echinococcus multilocularis and human alveolar echinococcosis (AE) are changing in Central Europe. Our data from a regional referral center for AE in southwest Germany suggest rising regional incidence for AE (annual incidence per 100,000 population 2004-2011: 0.12; 2012-2019: 0.20) and emerging urban AE (of 7 cases of AE in Freiburg city dwellers none was diagnosed before 2012) calling for an intensification of E. multilocularis and AE surveillance and of AE prevention measures.

Regulation of hepatic microRNAs in response to early stage Echinococcus multilocularis egg infection in C57BL/6 mice.

We present a comprehensive analysis of the hepatic miRNA transcriptome at one month post-infection of experimental primary alveolar echinococcosis (AE), a parasitic infection caused upon ingestion of E. multilocularis eggs. Liver tissues were collected from infected and non-infected C57BL/6 mice, then small RNA libraries were prepared for next-generation sequencing (NGS). We conducted a Stem-loop RT-qPCR for validation of most dysregulated miRNAs. In infected mice, the expression levels of 28 miRNAs were significantly altered. Of these, 9 were up-regulated (fold change (FC) ≥ 1.5) and 19 were down-regulated (FC ≤ 0.66) as compared to the non-infected controls. In infected livers, mmu-miR-148a-3p and mmu-miR-101b-3p were 8- and 6-fold down-regulated, respectively, and the expression of mmu-miR-22-3p was reduced by 50%, compared to non-infected liver tissue. Conversely, significantly higher hepatic levels were noted for Mus musculus (mmu)-miR-21a-5p (FC = 2.3) and mmu-miR-122-5p (FC = 1.8). In addition, the relative mRNA expression levels of five genes (vegfa, mtor, hif1-α, fasn and acsl1) that were identified as targets of down-regulated miRNAs were significantly enhanced. All the five genes exhibited a higher expression level in livers of E. multilocularis infected mice compared to non-infected mice. Finally, we studied the issue related to functionally mature arm selection preference (5p and/or 3p) from the miRNA precursor and showed that 9 pre-miRNAs exhibited different arm selection preferences in normal versus infected liver tissues. In conclusion, this study provides first evidence that miRNAs are regulated early in primary murine AE. Our findings raise intriguing questions such as (i) how E. multilocularis affects hepatic miRNA expression;(ii) what are the alterations in miRNA expression patterns in more advanced AE-stages; and (iii) which hepatic cellular, metabolic and/or immunologic processes are modulated through altered miRNAs in AE. Thus, further research on the regulation of miRNAs during AE is needed, since miRNAs constitute an attractive potential option for development of novel therapeutic approaches against AE.

Long-term (35 years) cryopreservation of Echinococcus multilocularis metacestodes.

The metacestode of Echinococcus multilocularis is the etiological agent of alveolar echinococcosis. The metacestode stage used for research is maintained in rodents by serial passages. In order to determine whether cryopreservation of E. multilocularis metacestodes would be suitable for long-term maintenance and replace serial passages, isolates of different geographic origin were cryopreserved in 1984-1986. The aim of the current study was to test the viability of cryopreserved isolates following long-term cryopreservation (up to 35 years) and to determine the phylogenetic clades these isolates belonged to. Cryopreserved isolates were tested for viability in vitro and in vivo in gerbils. In vitro results of 5 isolates indicated protoscolex survival in 13 of 17 experiments (76%) and metacestode survival in 5 of 12 (42%) in vivo experiments. In vivo results showed 'abortive lesions' in 13 of the 36 animals, 15 were negative and 8 harboured proliferating metacestode tissue containing protoscoleces. Genetic analysis confirmed the isolates belonged to European, Asian and North-American clades. In conclusion, the results of the current study indicate that metacestodes of E. multilocularis are able to survive long-term cryopreservation. Therefore, cryopreservation is a suitable method for long-term storage of E. multilocularis metacestode isolates and reduces the number of experimental animals.

A mathematical modelling approach for treatment and control of Echinococcus multilocularis.

Alveolar echinococcosis (AE) is a zoonotic parasitic diseases caused by a cestode parasite known as Echinococcus multilocularis. The parasite has a wildlife cycle with definitive hosts (foxes) and small mammals as intermediate hosts (rodents) while humans are the accidental hosts. Parasite infection pressure relation to time of the year and age dependent infection pressure for parasite abundance also depend on the urbanization. The aim of current work is forecasting the thresholds via the computational analysis of the disease spread which is a useful approach since it can help to design the experimental settings with better planning and efficiency. Network analysis when interlinked with the computational techniques provides better insight into the spatial and temporal heterogeneities. In the present study, a mathematical framework that describes the transmission dynamics and control measures of E. multilocularis in foxes is documented. We used treatment of foxes with baits for the prevention of the E. multilocularis infection. A novel approach of networking, called Petri net (PN), based on density dependent differential equations, is utilized during this research. The accurate description of the transmission of the parasite and the effect of drug on it is provided to the readers in this article. The transitions, which are difficult to analyse theoretically, are presented with the aid of the discrete approach of networking. A discrete mathematical framework can prove to be an accurate and robust tool to analyse and control the parasite dynamics.

Species specificity preliminary evaluation of an IL-4-based test for the differential diagnosis of human echinococcosis.

The diagnosis of cystic echinococcosis (CE) is based on imaging, while serology is a complementary test of particular use when imaging is inconclusive. Serology has several limitations. Among them, false-positive results are often obtained in subjects with alveolar echinococcosis (AE), rendering difficult the differential diagnosis. We set up an immune assay based on IL-4-specific production after stimulating whole blood with an antigen B (AgB)-enriched fraction from E granulosus that associates with CE and CE cysts in active stage. We aimed to evaluate potential cross-reactivity of this test using samples from patients with AE. Twelve patients with AE were recruited; IL-4 levels ranged from 0 to 0.07 pg/mL. Based on the previously identified cut-off of 0.39 pg/mL using samples from patients with CE, none of samples from AE patients scored positive. In contrast, almost 80% of samples from AE patients scored positive in serology tests based on different E granulosus-derived antigenic preparations. Our preliminary data show that this experimental whole-blood assay has no cross-reactivity in our cohort of patients with AE, in turn indicating a high specificity of the assay for CE diagnosis. This result supports further work towards the development of improved diagnostic tests for CE.

Echinococcus multilocularis inoculation induces NK cell functional decrease through high expression of NKG2A in C57BL/6 mice.

BACKGROUND: Alveolar echinococcosis (AE) is caused by the larval stage of Echinococcus multilocularis (E. multilocularis), and considered as public health issue. Parasite-host immune interaction is pivotal during infection. As a subset of innate lymphoid cells, NK cells are known to play an important role during virus, bacteria, intra/extracellular parasitic infections and tumor progression. However, the possible role of NK cells in E. multilocularis infection in both human and murine is little known. Herein, the functional alteration of hepatic NK cells and their related molecules in E. multilocularis infected mice were studied. METHODS: 2000 protoscoleces (PSCs) were injected to C57BL/6 mice via the portal vein to establish secondary E. multilocularis infection. NK cells population and their related molecules (CD69, Ly49D, Ly49G2, Ly49H, Ly49I, NKG2A, NKG2D, granzyme B, IFN-γ, TNF-α) were assessed by using fluorescence-activated cell sorter (FACS) techniques and qRT-PCR. NK cell depletion was performed for further understanding the possible function of NK cells during infection. RESULTS: The total frequencies of NK cells and NK-derived IFN-γ production were significantly reduced at designated time points (2, 4, 12 weeks). The liver resident (CD49a+DX5-) NK cells are decreased at 4 weeks after inoculation and which is significantly lower than in control mice. Moreover, in vivo antibody-mediated NK cell depletion increased parasitic load and decreased peri-parasitic fibrosis. Expression of the inhibitory receptor NKG2A was negatively related to NK- derived IFN-γ secretion. CONCLUSIONS: Our study showed down regulates of NK cells and upper regulates of NKG2A expression on NK cells during E. multilocularis infection. Reduction of NK cell frequencies and increased NKG2A might result in low cytotoxic activity through decreased IFN-γ secretion in E. multilocularis infection. This result might be helpful to restore NK cell related immunity against E. multilocularis infection to treat alveolar echinococcosis.

Surgical approaches for definitive treatment of hepatic alveolar echinococcosis: results of a survey in 178 patients.

Hepatic alveolar echinococcosis (HAE) is a potentially fatal disease caused by the larval growth of Echinococcus multilocularis. We analysed the clinical data of 178 consecutive HAE patients treated with definitive radical surgery at our institution. According to the surgical approach: group A patients underwent direct radical hepatic resection; group B patients first underwent percutaneous puncture external drainage, followed by radical hepatic resection 2 months later; group C patients underwent a two-step hepatic resection; and group D patients underwent liver transplantation. The baseline characteristics, mortality, postoperative complications and recurrence rates were evaluated. Symptoms were present in 79.8% (142/178) patients. Bi-lobar lesion was found in 34 (19.1%, 34/178) patients, 47.2% (84/178) of whom had ⩾2 lesions each. There were no intraoperative deaths. The postoperative mortality was 2.29% in group A, 8.62% in group D and 0% in groups B and C. The main cause of death was a serious postoperative complication (Clavien-Dindo grades III-V). Patients were followed-up systematically for a median of 35.8 months (8-72) without recurrence. Active HAE should be treated by radical liver resection, and the complicated alveolar echinococcosis of the liver has been managed whenever possible using principles of radical liver resection by experienced hepatic surgeons.

Echinococcosis transmission on the Tibetan Plateau.

Since the mid-1990s detailed studies and field investigations on the Tibetan Plateau have revealed human echinococcosis to be an under-reported major public health problem, particularly in the dominant pastoral communities in the eastern and central regions. Human prevalence surveys showed that cystic echinococcosis (CE, caused by Echinococcus granulosus) and alveolar echinococcosis (AE, caused by Echinococcus multilocularis) are co-endemic with higher burdens of each disease than other endemic world regions. Epidemiological investigations identified some major risk factors for human CE and AE including dog ownership, husbandry practices and landscape features. Dogs appear to be the major zoonotic reservoir for both E. granulosus and E. multilocularis, but the latter is also transmitted in complex wildlife cycles. Small mammal assemblages especially of vole and pika species thrive on the Plateau and contribute to patterns of E. multilocularis transmission which are influenced by landscape characteristics and anthropogenic factors. Tibetan foxes are a principal definitive host for both E. multilocularis and E. shiquicus. In 2006 a national echinococcosis control programme was initiated in Tibetan communities in northwest Sichuan Province and rolled out to all of western China by 2010, and included improved surveillance (and treatment access) of human disease and regular deworming of dogs with annual copro-testing. Control of echinococcosis in Tibetan pastoral communities poses a difficult challenge for delivery and sustainability.

Subcutaneous Inoculation of Echinococcus multilocularis Induces Delayed Regeneration after Partial Hepatectomy.

Alveolar echinococcosis (AE) is caused by the larval stage of echinococcus multilocularis (E. multilocularis), and hepatectomy is the main modality in hepatic AE patients. Liver regeneration after partial hepatectomy (PHx) in such patients is challenging, and further investigation is needed. Thus far, knowledge regarding the possible impact of E. multilocularis on liver regeneration after PHx is limited. Herein, a subcutaneous infection model of E. multilocularis was developed in C57 BL/6 mice, and after 3 months, PHx was performed. Plasma and liver samples were harvested under inhalational isofluorane (2%) anaesthesia at designated post-PHx time points (0, 24, 48, 96 and 168 h). The parameters included the future remnant liver/body weight ratio (FLR/BW), liver function tests (AST and ALT) and related cytokines (TNF-α, IL-6, Factor V, HMGB1, TGF-ß, TSP-1, and TLR4) and proteins (MyD88 and STAT3). To assess the proliferation intensity of hepatocytes, BrdU, Ki67 and PAS staining were carried out in regenerated liver tissue. The FLR/BW in the infected group from 48 h after surgery was lower than that in the control group. The BrdU positive hepatocyte proportions reached their peak at 48 h in the control group and 96 h in the infected group and then gradually decreased. During the first 48 h after surgery, both the AST and ALT levels in the infected group were lower; however, these levels were altered from 96 h after surgery. In the infected group, the concentrations and mRNA expression levels of the pre-inflammatory cytokines TNF-α and IL-6 demonstrated a delayed peak. Moreover, post-operatively, the TGF-ß and TSP-1 levels showed high levels in the infected group at each different time-point compared to those in the control group; however, high levels of TGF-ß were observed at 96 h in the control group. The MyD88 and STAT3 protein expression levels in the infected group were markedly higher than those in the control group 96 h after surgery. Delayed liver regeneration after PHx was observed in the C57 BL/6 mice with the subcutaneous infection of E. multilocularis in the current study. This phenomenon could be partially explained by the alteration in the pro-inflammatory cytokines in the immunotolerant milieu induced by chronic E. multilocularis infection.

Effect of anacardic acid against echinococcosis through inhibition of VEGF-induced angiogenesis.

Echinococcosis is a zoonotic infection caused by cestode species of the genus Echinococcus, with limited treatment options. It is urgent to develop new anti-hydatid agent. In this paper, we reported anacardic acid (AA), a natural product isolated from the Brazilian cashew-nut shell liquid, which presented a high activity against metacestodes of Echinococcus multilocularis (E. multilocularis) and Echinococcus granulosus sensu stricto (E. granulosus s.s.) in vitro and in vivo. AA exerted a better efficacy on E. granulosus s.s. protoscoleces and E. multilocularis metacestodes than that of albendazole (ABZ) and dihydroartemisinin (DHA) in vitro, and an inhibition on the growth of Echinococcus metacestode as effective as ABZ in vivo. Moreover, we also found that one of the mechanisms of AA against Echinococcus could be the suppression of angiogenesis on/in the metacestode mass through inhibiting vascular endothelial growth factor (VEGF)-induced signalling pathways. This work finds that AA is a new promising potential candidate drug for echinococcosis treatment.

Health-related quality of life in patients with alveolar echinococcosis: a cross-sectional study.

PURPOSE: The Alveolar echinococcosis (AE) is a rare zoonosis caused by the parasite Echinococcus multilocularis. To date, nothing is known about the health-related quality of life (HRQoL) in patients with AE. The aim of the study was to evaluate the HRQoL in patients with AE in comparison of the healthy population. METHODS: We used the 36-Item Short Form Health Survey (SF-36) questionnaire to evaluate the HRQoL. The SF-36 scales have a range of 0-100 (0 represents the worst and 100 the most favourable state of health). SAS Version 9.2 was used for the statistical analysis of AE-cases (n = 30) and the healthy control group (n = 35). RESULTS: The analysis showed that the HRQoL in people with AE is reduced in comparison with the control population. The study group consisted of 15 (50.0%) men and 15 (50.0%) women; the control group of 16 (45.7%) men and 19 (54.3%) women. The mean age was 55.73 ± 16.65 years, while that of the control group was 54.57 ± 15.34 years. The physical quality of life in patients with AE (45.21 ± 11.42) was not significantly less than that of the control group (50.54 ± 10.52); p = 0.0568. Nevertheless, AE-patients show lower SF-36 scores for the physical quality of life. For the mental quality of life, patients with AE had a significantly lower score (45.46 ± 10.57) than the control group (51.57 ± 9.04); p = 0.0154. CONCLUSIONS: The HRQoL in people with AE is reduced in comparison with a control population. Assessment of the physical and mental quality of life in patients with AE may help to evaluate the patient outcome.

Helminth parasites and zoonotic risk associated with urban coyotes (Canis latrans) in Alberta, Canada.

Coyotes (Canis latrans) are resilient, adaptable, cosmopolitan omnivores that are increasingly prevalent in urban environments, where they interact with both humans and domestic dogs. Coyotes potentially transmit zoonotic parasites, including the tapeworm Echinococcus multilocularis, which appears to be increasing in prevalence in western North America. In this study, we analysed the carcasses of 23 urban coyotes in Edmonton, Alberta, Canada. Focusing primarily on the helminth community, we recovered three tapeworm species (E. multilocularis, Taenia pisiformis, T. serialis), four nematodes (Toxascaris leonina, Uncinaria stenocephala, Capillaria sp., Physaloptera sp.), and two trematodes (Alaria arisaemoides and A. americana). Compared to previous studies of urban coyotes conducted in North America, we report one of the highest levels of E. multilocularis infection in North America: 65.2% infection prevalence. These results amplify concerns expressed by others about the increasing prevalence of this zoonotic parasite and the role coyotes may play in parasite transmission. More research is needed to better understand how various ecological factors, urbanization and wildlife management practices influence the transmission of potentially zoonotic parasites such as E. multilocularis.

Immunotherapy of alveolar echinococcosis via PD-1/PD-L1 immune checkpoint blockade in mice.

The growth potential of the tumour-like Echinococcus multilocularis metacestode (causing alveolar echinococcosis, AE) is directly dependent upon the nature/function of the periparasitic adaptive host immune-mediated processes. PD-1/PD-L1 pathway (programmed cell death 1), which inhibits lymphocytic proliferation in tumour development, is over-expressed at the chronic stage of AE. We tested the impact of a PD-1/PD-L1 pathway blockade on the outcome of both chronic AE (intraperitoneal metacestode inoculation, secondary AE and SAE) and acute AE (peroral egg infection, primary AE and PAE). To assess the parasite proliferation potential, we measured parasite mass weight for SAE and liver lesion number for PAE. In both models, the parasite load was significantly decreased in response to anti-PD-L1 antibody treatment. In SAE, anti-PDL1 administration was associated with increased Th1 response parameters and decreased Treg responses, while in PAE anti-PDL1 administration was associated with fewer lesions in the liver and decreased Treg/Th2 responses. Our findings highly suggested that a PD-1/PD-L1 pathway blockade triggered the host immune responses in favour of an immune-mediated control of E. multilocularis proliferation. Based on this, future studies that combine PD-1/PD-L1 blockade with a parasitostatic albendazole medication may yield in a putatively curative therapeutic approach to control alveolar echinococcosis.