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Most trematodes and some cestodes have obligate life history features that include an asexual developmental stage that can produce genetically-identical individuals (clonemates) followed by an adult stage with sexual reproduction. These life history features can influence the evolutionary mechanism of inbreeding in parasites, especially among self-compatible hermaphroditic endoparasites whose mating opportunities are restricted to within hosts. As clonemate mating in hermaphroditic species produces a genetic inbreeding signature identical to that of self-mating, it is important to understand how clonemates are transmitted through their life stages. A handful of prior studies compared clone richness (number of clones) across life cycle stages to infer transmission processes and to characterize clone abundance distributions (CADs) among hosts. Here we illustrate the use of the proportion of clonemate dyads (PC) within hosts to describe the CADs. PC has several advantages as an ecological metric in that it is unbiased by sample size, takes into account relative parasite burdens, and has a direct transmission interpretation, i.e., the probability of cotransmitting clonemates. Moreover, PC is also an evolutionary metric as it can be used to estimate a potential clonemate mating rate. We demonstrate the use of PC in comparing CADs within and across 2 trematode developmental stages in the lancet fluke Dicrocoelium dendriticum. Also, we show how genetic estimates of apparent selfing (true selfing plus clonemate mating) at larval and juvenile stages can be compared to PC estimated at the adult stage to assess the contribution of clonemate mating to apparent selfing. The eco-evolutionary links presented are generalizable to assess sibling cotransmission as well. Thus, the framework presented herein will facilitate future field-based studies on the transmission and mating systems of parasitic flatworms.

The transmission of many parasites, particularly those with complex life cycles, can only occur when infective stages that are present in one host are consumed by another host. Parasites can enhance rates of transmission by altering the behaviour, coloration or conspicuousness of infected hosts that require ingestion by a predator in which parasite reproduction occurs (i.e. the definitive host). But such enhancement will impose costs if it also facilitates the transmission of infective stages into nonhosts that are incompatible with parasite development and reproduction. We assessed these potential costs in fathead minnows, Pimephales promelas, exposed to larvae of a brain-encysting trematode, Ornithodiplostomum ptychocheilus. We evaluated minnow antipredator escape responses immediately after they received visual stimuli from simulated host and nonhost predators. Our results show that mean response latencies of infected minnows were 12% greater than uninfected minnows when they were threatened with horizontally approaching stimuli (simulating a nonhost predator) regardless of infection level and regardless of parasite development stage. Response latencies of minnows exposed to vertically approaching stimuli (simulating a host predator) were lower than in controls, but the differences were not significant. In this case of parasite-induced alteration of host behaviour, the alteration enhances transmission into nonhosts. Costs associated with enhanced transmission into nonhosts likely constrains the development of specific parasite-induced alterations of host phenotypes in other host–parasite interactions that require predation of infected hosts.

Host manipulation by parasites is a fascinating evolutionary outcome, but adaptive scenarios that often accompany even iconic examples in this popular field of study are speculative. Kin selection has been invoked as a means of explaining the evolution of an altruistic-based, host-manipulating behavior caused by larvae of the lancet fluke Dicrocoelium dendriticum in ants. Specifically, cotransmission of larval clonemates from a snail first host to an ant second host is presumed to lead to a puppeteer parasite in the ant’s brain that has clonemates in the ant abdomen. Clonal relatedness between the actor (brain fluke) and recipients (abdomen flukes) enables kin selection of the parasite’s host-manipulating trait, which facilitates transmission of the recipients to the final host. However, the hypothesis that asexual reproduction in the snail leads to a high abundance of clonemates in the same ant is untested. Clonal relationships between the manipulator in the brain and the nonmanipulators in the abdomen are also untested. We provide empirical data on the lancet fluke’s clonal diversity within its ant host. In stark contrast to other trematodes, which do not exhibit the same host-manipulating behavioral trait, the lancet fluke has a high abundance of clonemates. Moreover, our data support existing theory that indicates that the altruistic behavior can evolve even in the presence of multiple clones within the same ant host. Importantly, our analyses conclusively show clonemate cotransmission into ants, and, as such, we find support for kin selection to drive the evolution and maintenance of this iconic host manipulation.

The results of phylogeographic and biogeographic studies of organisms residing in isolated habitats provide key insights into processes of population differentiation, speciation, and endemicity. Several of the approximately 70 species of land snail in the genus Oreohelix Pilsbry, 1904 occur only on isolated sky islands on the North American Great Plains. The restricted distributions of these snails have led to concerns regarding their conservation status, particularly in western Canada where their systematics and distributions are poorly known. Cooper’s Rocky Mountain snail (Oreohelix cooperi (Binney, 1858)) has been reported from several sky islands in the northern United States. We evaluated morphological characteristics and sequence data for samples of putative O. cooperi collected from a sky island in the Cypress Hills area of southeastern Alberta and southwestern Saskatchewan. COI sequences matched O. cooperi from snails collected on sky islands in South Dakota and Wyoming and morphological analyses of shell shape and male genitalia were consistent with published descriptions of this species. COI and ITS2 sequences and morphological characteristics of these snails did not match other Oreohelix spp. found in the Cypress Hills and in the adjacent Rocky Mountains. Our results extend the distribution of O. cooperi into southern Canada and confirm its endemicity within sky islands of western North America.

Background Some patients have features that indicate possible difficulty with direct laryngoscopy for tracheal intubation. Prediction of the likely outcome and selection of patients for an enhanced management algorithm would reduce the possible harm from failed intubation attempts. Methods Adult elective patients were assessed for seven features associated with difficult direct laryngoscopy, ranked in difficulty from 0 to 3. For a patient with at least one Class 3 feature, or two or more features of class 1 or higher, the enhanced management used a channelled videolaryngoscope Airtraq™ instead of a Macintosh laryngoscope. A long flexible angulated stylet and a flexible fibrescope would be used as the second and third steps. For patients with lesser difficulty scores, a Macintosh laryngoscope was used. Outcomes of enhanced management were analysed. Logistic regression and Random Forest algorithm, using the ranks of the predictive features, were used to predict difficulty during enhanced management. Results We prospectively studied 16 695 patients. We selected 1501 (9%) for enhanced management, and tracheal intubation was successful in all of them. Of these, 73% were intubated in less than 30 s, and only 4.5% required more than 4 min for intubation. Progression to the second and third steps of enhanced management was predicted by restriction of mouth opening and reduced cervical spine mobility. Conclusions An enhanced management algorithm allowed successful tracheal intubation of all patients with anticipated difficult laryngoscopy. The need to combine the use of a stylet and a fibrescope with the Airtraq™ could be predicted with a high degree of certainty.

Indigenous peoples living in Canada’s north have long-valued the livers of Burbot (Lota lota) as a traditional food source; however, there has been concern relating to liver quality and potential contaminants. In this study, livers of Burbot collected in lower Mackenzie River were ranked using a traditional appearance-based assessment. These rankings were compared to a variety of biological and contaminant metrics. Livers ranked “most palatable” had a significantly higher mass and lipid content and were from younger fish with greater hepatosomatic index and total mass and had lower parasite intensities. There were no differences in the concentrations of persistent organic pollutants or metals, except copper, which although still well below consumption guidelines, was significantly higher in fish with livers that appeared most palatable. The results of this study demonstrated that traditional methods effectively assessed the quality of livers by selecting for the most nutritious (high lipid levels) and safest (low parasite loading) food. This method could be incorporated into a community-based monitoring framework as a rough index of overall fish and ecosystem health; however, would not be effective in screening food for anthropogenic contaminants. This study highlights the importance and value of linking traditional knowledge into scientific studies.

Portable user-friendly diagnostic tests can benefit detection and surveillance of wildlife diseases. Here, the performance of a compact POCKIT™ Nucleic Acid Analyzer for detection of Ambystoma tigrinum virus (ATV), an emerging Iridovirus that is associated with high host mortality in the western tiger salamander (Ambystoma mavortium) in North America was assessed. Tissue samples from 188 larval tiger salamanders collected from sites in Alberta, Canada were tested by both iiPCR and by conventional PCR. Results of the two assays showed 96.3% agreement. All 176 samples that tested positive by conventional PCR were also positive by iiPCR, while 12 of the samples that were negative by conventional PCR were positive by iiPCR. Comparison of the limits of detection of the two assays shows that the iiPCR assay was more sensitive than conventional PCR and had a LOD95 of 20 copies per reaction. The instrument automatically analyzes and displays results within 40 min following nucleic acid extraction. The novel technology could enhance detection of, and response to, wildlife pathogens, particularly those that occur sporadically, cause rapid outbreaks, and/or occur within isolated geographical regions.

Complete genome sequences of six Ambystoma tigrinum viruses (ATV) were determined directly from tail clips of western tiger salamanders (Ambystoma mavortium) from 2013 (high-mortality year) and 2014 (low-mortality year) in Alberta, Canada. The genome lengths ranged from 106,258 to 106,915 bp and contained 108 open reading frames encoding predicted proteins larger than 50 amino acids.

The control of emerging parasites requires a fundamental knowledge of where and when rates of transmission are high. Data on spatiotemporal patterns of infection are challenging to obtain, particularly for complex life cycle parasites that involve transmission into multiple obligate hosts. The lancet liver fluke, Dicrocoelium dendriticum, has a long history of colonization outside its native host and geographical range in continental Europe. Infection patterns involving adult and metacercarial stages have been characterized for this trematode in a region of emergence in western Canada within co-grazing herbivores and ants, but infection patterns in snail intermediate hosts in this region are unknown. We combined spatiotemporal prevalence surveys with sequence analyses of the cytochrome c oxidase subunit 1 ( COI) barcoding gene from samples of sporocyst tissue in infected snails to confirm that D. dendriticum utilizes 3 sympatric species of Oreohelid land snail ( Oreohelix subrudis, Oreohelix sp., and Oreohelix cooperi) as first intermediate host. Mean prevalence within a total sample of 900 adult snails collected over 1 field season from 6 sites was 9.9 ± 2.4%. For each species of snail, prevalence ranged between 5-30% within monthly samples, with peaks in mid-summer followed by declines in fall. Between-site variation in prevalence was low and non-significant, implying that rates of transmission of D. dendriticum miracidia from domestic stock and wildlife into snails are similar within localized sites, despite high variation in local habitat characteristics and in the structure of the definitive host community.

Introduction Postoperative ileus(POI) and anastomotic leakage(AL) are highly associated but the direction of this link is unknown. Indeed, it has traditionally been stated that AL leads to POI(secondary POI). The objective was to statistically determine the best model of link between POI and AL by comparing different Structural Equation Models(SEM). Methods This is a retrospective analysis of a prospective multicentric database from the Francophone Group for Enhanced Recovery after surgery(GRACE) including patients undergoing colorectal resection or Hartmann reversal between 2014 and 2017. The main outcome measure was the occurrence of postoperative ileus defined by the absence of Gastrointestinal (GI)-3 recovery in or after postoperative day 4 and the occurrence of AL. Results Among the 2227 patients included, 223 patients experienced POI (10 %) and 72 patients experienced anastomotic leakage (3.2 %). In all the models, POI was significantly increased in case of male gender, older age, opioids consumption, right colectomy, chronic obstructive bronchopneumopathy disease and performance of a stoma, and reduced by Enhanced Recovery Programs (ERP) compliance ≥ 70 % and intake of Non-Steroidal Anti-Inflammatory Drugs. Open surgery was not significantly associated with POI in all the models. In contrast, the male gender was the only factor increasing AL in the model drawn with ERP risk factors for AL. In all the models, POI was highly associated with AL and statistical analyses advocate for POI leading to AL. Conclusion The findings of the present large study suggest that the more statistically probable model is the one in which POI come first before AL.

SUMMARYEpidemiological rate parameters of host generalist parasites are difficult to estimate, especially in cases where variation in parasite performance can be attributed to host species. Such cases are likely common for generalist parasites of sympatric grazing mammals. In this study, we combined data from experimental exposures in cattle and sheep and natural infections in elk to compare the recruitment, morphology and reproduction of adultDicrocoelium dendriticum, a generalist trematode that has emerged in sympatric grazing hosts in Cypress Hills Provincial Park, Alberta. Overall, there were no significant differences in the recruitment of metacercariae and in the pre-patency period of adults in experimentally exposed cattle and sheep. All flukes reached reproductive maturity and the degree of reproductive inequality between individual flukes within each infrapopulation was moderate and approximately equal among the three host species. Neither fluke size norper capitafecundity was constrained by density dependence. Thus, fitness parameters associated with growth and reproduction were approximately equivalent among at least three species of definitive host, two of which are sympatric on pastures in this Park. The generalist life-history strategy of this trematode, which is known to extend to other stages of its life cycle, has likely contributed to its invasion history outside its native range in Europe.

Hippoboscid flies (Diptera: Hippoboscidae) include species that are ectoparasites of birds in the Northern Hemisphere, but little is known regarding their taxonomy, parasites, avian host associations, or geographical distribution in North America. In late August of 2013 and 2014, we collected hippoboscid flies from live birds trapped in mist nets as part of a banding study in Cypress Hills Interprovincial Park in southeastern Alberta, Canada. A total of 113 birds comprising 9 species was examined in 2013. Of these, 18 individuals were infested with 1-3 Ornithomya anchineuria Speiser (n = 22 flies; prevalence = 15.9%). Eight of these flies carried 1-8 adult female epidermoptid mites anchored to their ventral, posterior abdomens. Each female was associated with clusters of up to 30 stalked eggs. The first pair of tarsi on adult female mites was highly modified as anchors, indicating permanent attachment through the host cuticle. Morphological traits identified these mites as Myialges cf. borealis Mironov, Skirnisson, Thorarinsdottier and Nielsen. Cytochrome c oxidase subunit 1 ( COX1) gene sequences obtained for 2 mites were distinct from those previously reported for species of Myialges, being most similar to Myialges trinotoni Cooreman. The paucity of available gene sequences for Myialges and related genera of epidermoptid mites prevents any further conclusions regarding taxonomy. These findings extend previous reports of O. anchineuria from Pacific and Atlantic coasts of Canada inland to the central migratory flyway of the Northern Great Plains and expand the limited information available for Myialges spp.

The expansion of parasite distributions outside of their native host and geographical ranges has occurred repeatedly over evolutionary time. Contemporary examples include emerging infectious diseases (EIDs), many of which pose threats to human, domestic animal, and wildlife populations. Theory predicts that parasites with complex life cycles will be rare as EIDs due to constraints imposed by host specialization at each life-cycle stage. In contrast to predictions of this theory, we report 2 new intermediate hosts in the 3-host life cycle of the liver fluke Dicrocoelium dendriticum in Cypress Hills Provincial Park, Alberta, Canada. Results of sequence analysis of the cytochrome oxidase 1 (cox1) mitochondrial gene identified the terrestrial snail Oreohelix subrudis and the ant Formica aserva as first and second intermediate hosts, respectively, in the region. Neither of these intermediate hosts, nor their suite of domestic and wild mammalian grazers used in the life cycle, occurs within the native range of D. dendriticum in Europe. Our results from host surveys show that the prevalence of D. dendriticum in samples of O. subrudis varied between 4% and 10%, whereas mean metacercariae intensity in F. aserva varied between 33 and 41 (n = 163, mean ± SD = 38 ± 35). These results are the first to describe the complete life cycle of emerging lancet fluke in western North America. The process of multi-level ecological fitting, in which the lancet fluke possesses pre-existing traits to utilize host resources, rather than host species, at each life-cycle stage provides a mechanism for the establishment of this complex life cycle in a novel habitat and in novel hosts.

Sera from calves originating on a 17,000 ha ranch in southeastern Alberta, were collected over three consecutive years (2008–2010) at weaning and analysed for the presence of anti-Ostertagia antibodies using the SVANOVIR® Ostertagia ostertagi-Ab ELISA kit. Faecal samples taken at the same time were analysed for presence of nematode eggs using the modified Wisconsin faecal flotation technique. Mean optical density ratio (ODR) values for all calves were similar in 2008 and 2010 while the 2009 values were significantly higher. Similar cumulative frequency distribution profiles were found for 2008 and 2010 while the 2009 profile had an increased number of values in the higher bin categories. This pattern indicates higher nematode transmission on pasture in 2009. There was no significant difference in ODR values between egg positive and egg-negative calves and no significant correlation was detected between individual ODR values and faecal egg output. Significantly higher ODR values in 2009 appear contradictory to general assumptions about the ecological requirements of larval development, as 2009 was a relatively dry year. However, examination of weather data identified 5–6 rainfall events that may have resulted in episodic enhanced survival of nematode larvae.

Parasite distributions are constantly changing due to climate change, local and global movement of animals and humans, as well as land use and habitat change. The trematode Dicrocoelium dendriticum is a relatively recent invader of Canada, being first reported in eastern Canada in the 1930s and western Canada in the 1970s. However, historical records are scarce and its emergence is poorly understood. The establishment of this parasite in Canada provides an interesting opportunity to explore the use of population genetic approaches to help elucidate the invasion history of a relatively recently established helminth parasite. In this study, we compare the genetic diversity and population structure of a number of D. dendriticum populations from western and eastern Canada, and compare these with much longer established European populations. Two independent genetic marker systems were used; a microsatellite marker panel and a cytochrome c oxidase 1 (cox1) mitochondrial (mt)DNA sequence marker. We found distinct differences in both genetic diversity and population structure of the different Canadian populations that provide insights into their invasion histories compared with the European populations. Two populations from British Columbia, Canada - Salt Spring and Vancouver Islands - are of low diversity, show evidence of a population bottleneck and are closely related to each other, suggesting a shared recent history of establishment. These west coast populations are otherwise most closely related to those from eastern Canada and western Europe, and in contrast are genetically divergent from those in Cypress Hills, Alberta, Canada. Although the Alberta parasite population is the most recently reported in Canada, being first identified there in the early 1990s, it was the most genetically diverse of those examined and showed a strong pattern of admixture of genotypes present in western and eastern Europe. Overall, our results are consistent with a model in which western Europe is likely the source of flukes on the east coast of Canada, which were then subsequently translocated to the west coast of Canada. The most recently reported D. dendriticum population in Canada appears to have a different history and likely has multiple origins.

This study characterizes polymorphic microsatellite loci from adults of the liver fluke Dicrocoelium dendriticum sampled from a population of sympatric beef cattle and wapiti in a region of emergence in southern Alberta, Canada. We also scrutinized the markers to validate their use in studying the population genetics of this complex life cycle parasite. Among the nine loci described, four deviated significantly from Hardy Weinberg Equilibrium (HWE) due to technical artefacts. The remaining five loci were in HWE. These five provided sufficient resolution to identify clonemates produced from the obligate asexual reproduction phase of the life cycle in snails and to assess the impact of non-random transmission of clonemates on measures of FIS, FST and genotypic disequilibrium. Excluding clonemates, we show that the sub-population of worms was in HWE, that average FIS within hosts was 0.003 (p=0.4922) and that there was no population genetic structure among hosts FST=0.001 (p=0.3243). These markers will be useful for studies of Dicrocoelium dendriticum ecology, transmission, and evolution.

Although results from field surveys have linked parasites to oxidative stress in their fish hosts, direct evidence involving experimentally infected hosts is lacking. We evaluated the effects of experimental infections with larval trematodes on induction of oxidative stress in fathead minnows, Pimephales promelas. Juvenile fish were exposed in the laboratory to the larvae (cercariae) of 2 species of trematode: Ornithodiplostomum sp. that develops in the liver, and O. ptychocheilus that develops in the brain. For Ornithodiplostomum sp., lipid peroxidation concentration in liver tissue increased 5 days after exposure and remained higher than controls until the end of the experiment at 28 days. For O. ptychocheilus, liver lipid peroxidation concentration was higher than controls at 5 days, but not thereafter. Sustained elevation in lipid peroxidation concentration for the liver trematode may be explained by direct tissue damage caused by developing larvae in the liver, or by an immune response. These experimental results support those from field studies, indicating that the lipid peroxidation assay may be an effective biomonitor for parasite-induced oxidative stress in fish, and that the nature of the oxidative stress response is species and/or tissue specific.

Summary 1. Individuals that live in groups benefit from increased foraging success and decreased predation. Protection from some types of parasites may provide an additional benefit of groupliving. For fish, the extent to which shoaling can reduce an individual’s risk of exposure to the infective stages of parasites is unknown. 2. We tested for antiparasite benefits of shoaling in fathead minnows exposed to larvae (cercariae) of two of their most common species of trematode, Ornithodiplostomum ptychocheilus and Posthodiplostomum minimum. As developing stages (metacercariae) of these trematodes cause reductions in minnow activity, growth and survival, natural selection should favour the evolution of cercariae-avoidance behaviours. 3. We evaluated shoal dimensions in groups of minnows exposed to O. ptychocheilus and to other chemical/physical stimuli within aquaria. To compare risk of exposure in shoaling vs. non-shoaling fish, we confined groups of minnows into mesh cages in outdoor mesocosms, exposed them to cercariae, then compared mean worm numbers in grouped vs. solitary fish. Lastly, we tested whether fish located within the centre of an artificial shoal reduced their risk of cercariae exposure compared with those along peripheral edges. 4. Minnows distinguished infective cercariae from other potential aquatic threats and responded with activity that reduced the 2-dimensional area of their shoals 15-fold compared with water-only controls. Fish confined within artificial shoals had 3-fold fewer worms than single fish and minnows located within the centre of artificial shoals had significantly fewer worms than those without peripheral minnows. 5. These results show that shoaling reduces a minnows’ risk of exposure to cercariae, either directly via detection of cercariae in the water column followed by behavioural avoidance or indirectly via behaviour-mediated differences in exposure between shoaling vs. non-shoaling fish.

Apparent changes in the distribution of Dicrocoelium dendriticum in western Canada and the absence of other liver flukes in the area prompted an examination of the humoral responses in naturally infected cattle sharing pasture with heavily infected wild cervids. Western blots using sera from the infected cattle indicated that the cattle had antibodies to various fluke antigens, however the pattern of response varied among isotypes. Isotype analysis indicated that Dicrocoelium-infected cattle produced IgG1, and IgM antibodies, but not IgG2 antibodies. Subsequent comparison of the efficiency of coprological and serological approaches for diagnosis showed that while 43.5% of a separate group of 26 yearlings had eggs in their faeces, the immunoblots indicated that all had detectable antibodies. A group of cattle necropsied for confirmation of fluke status was used to evaluate humoral immune responses and to provide baseline information in the preliminary development of an indirect ELISA for detection of anti-fluke antibodies.

Epidermal club cells of fishes in the superorder ostariophysi have puzzled evolutionary biologists because they were historically linked to chemical alarm signalling and relied on group selectionist explanations. Alternative hypoth- eses to explain the existence of these cells include the possibility of an anti-pathogenic or anti-parasitic function. If this is so, individual fish should invest in increased numbers of club cells after exposure to parasites, and club cells should contain components that reduce the infectivity of skin-penetrating larvae. Infectivity of cercariae of the trematode Ornithodiplostomum sp. was significantly reduced when exposed to the skin extract of fathead minnows (an ostariophysan), but also to skin extract of mollies (a non-ostariophysan that lacks club cells), respectively, compared to controls. Moreover, club cell density was not affected by exposure to cercariae. Taken together, these results are inconsistent with an anti-parasite function for these cells and instead suggest a generic role in response to injury. © 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 98, 884-890. ADDITIONAL KEYWORDS: alarm substance - anti-predation - Ornithodiplostomum - Ostariophysi - infectivity - Pimephales.