An improved model for the population dynamics of cattle gastrointestinal nematodes on pasture: parameterisation and field validation for Ostertagia ostertagi and Cooperia oncophora in northern temperate zones.

Gastrointestinal nematodes (GIN) are amongst the most important pathogens of grazing ruminants worldwide, resulting in negative impacts on cattle health and production. The dynamics of infection are driven in large part by the influence of climate and weather on free-living stages on pasture, and computer models have been developed to predict infective larval abundance and guide management strategies. Significant uncertainties around key model parameters limits effective application of these models to GIN in cattle, however, and these parameters are difficult to estimate in natural populations of mixed GIN species. In this paper, recent advances in molecular biology, specifically ITS-2 rDNA 'nemabiome' metabarcoding, are synthesised with a modern population dynamic model, GLOWORM-FL, to overcome this limitation. Experiments under controlled conditions were used to estimate rainfall constraints on migration of infective L3 larvae out of faeces, and their survival in faeces and soil across a temperature gradient, with nemabiome metabarcoding data permitting species-specific estimates for Ostertagia ostertagi and Cooperia oncophora in mixed natural populations. Results showed that L3 of both species survived well in faeces and soil between 0 and 30 °C, and required at least 5 mm of rainfall daily to migrate out of faeces, with the proportion migrating increasing with the amount of rainfall. These estimates were applied within the model using weather and grazing data and use to predict patterns of larval availability on pasture on three commercial beef farms in western Canada. The model performed well overall in predicting the observed seasonal patterns but some discrepancies were evident which should guide further iterative improvements in model development and field methods. The model was also applied to illustrate its use in exploring differences in predicted seasonal transmission patterns in different regions. Such predictive modelling can help inform evidence-based parasite control strategies which are increasingly needed due climate change and drug resistance. The work presented here also illustrates the added value of combining molecular biology and population dynamics to advance predictive understanding of parasite infections.

Seasonal epidemiology of gastrointestinal nematodes of cattle in the northern continental climate zone of western Canada as revealed by internal transcribed spacer-2 ribosomal DNA nemabiome barcoding.

BACKGROUND: Gastrointestinal nematode (GIN) epidemiology is changing in many regions of the world due to factors such as global warming and emerging anthelmintic resistance. However, the dynamics of these changes in northern continental climate zones are poorly understood due to a lack of empirical data. METHODS: We studied the accumulation on pasture of free-living infective third-stage larvae (L3) of different GIN species from fecal pats deposited by naturally infected grazing cattle. The field study was conducted on three organic farms in Alberta, western Canada. Grass samples adjacent to 24 fecal pats were collected from each of three different pastures on each farm. Internal transcribed spacer-2 nemabiome metabarcoding was used to determine the GIN species composition of the harvested larvae. The rotational grazing patterns of the cattle ensured that each pasture was contaminated only once by fecal pat deposition. This design allowed us to monitor the accumulation of L3 of specific GIN species on pastures under natural climatic conditions without the confounding effects of pasture recontamination or anthelmintic treatments. RESULTS: In seven out of the nine pastures, grass L3 counts peaked approximately 9 weeks after fecal deposition and then gradually declined. However, a relatively large number of L3 remained in the fecal pats at the end of the grazing season. Nemabiome metabarcoding revealed that Cooperia oncophora and Ostertagia ostertagi were the two most abundant species on all of the pastures and that the dynamics of larval accumulation on grass were similar for both species. Daily precipitation and temperature across the whole sampling period were similar for most of the pastures, and multiple linear regression showed that accumulated rainfall 1 week prior to sample collection had a significant impact on the pasture L3 population, but accumulated rainfall 3 weeks prior to sample collection did not. CONCLUSIONS: The results suggest that the pasture L3 population was altered by short-term microclimatic conditions conducive for horizontal migration onto grass. Overall, the results show the importance of the fecal pat as a refuge and reservoir for L3 of cattle GIN on western Canadian pastures, and provide an evidence base for the risk assessment of rotational grazing management in the region.

Oestrid myiasis at a cross-road.

The oestrids are a genuinely unique group of Diptera that represent many species whose larvae parasitize mammals, feeding on live tissues of a wide range of hosts, from mice to elephants, causing obligatory myiases. Those have had greatest impact on the activities of humans and their use of animals (e.g., as food, recreation, in the conduct of wars). To a large extent their impact has been controlled through the use of therapeutic application of synthetic chemical treatments. While in some limited cases eradication has been achieved, it is unlikely to become the norm as these parasites seem well adapted to survive at low population densities and small smoldering foci are likely to present a challenge in the future. Should organic production eschew the use of parasiticides, other than for rescue of seriously ill animals, the foci of infection by these parasites will remain. In this article, we explore current scientific information on this group of parasites that may have become an anachronism in modern livestock parasitology, identify gaps in our knowledge as well as the suggestions for the management of these parasites in the era of genomics and proteomics.

High levels of third-stage larvae (L3) overwinter survival for multiple cattle gastrointestinal nematode species on western Canadian pastures as revealed by ITS2 rDNA metabarcoding.

BACKGROUND: The ability of infective larvae of cattle gastrointestinal nematode (GIN) species to overwinter on pastures in northerly climatic zones with very cold dry winters is poorly understood. This is an important knowledge gap with critical implications for parasite risk assessment and control. METHODS: Infective third-stage larvae (L3) were quantified in samples of fecal pats, together with adjacent grass and soil, before and after winter on three farms in southern, central and northern Alberta. Nemabiome ITS2 metabarcoding was then performed on the harvested L3 populations to determine the species composition. Finally, parasite-free tracer calves were used to investigate if the L3 surviving the winter could infect calves and develop to adult worms in spring. RESULTS: Farm level monitoring, using solar powered weather stations, revealed that ground temperatures were consistently higher, and less variable, than the air temperatures; minimum winter air and ground temperatures were - 32.5 °C and - 24.7 °C respectively. In spite of the extremely low minimum temperatures reached, L3 were recovered from fecal pats and grass before and after winter with only a 38% and 61% overall reduction over the winter, respectively. Nemabiome ITS2 metabarcoding assay revealed that the proportion of L3 surviving the winter was high for both Cooperia oncophora and Ostertagia ostertagi although survival of the former species was statistically significantly higher than the latter. Nematodirus helvetinaus and Trichostrongylus axei could be detected after winter whereas Haemonchus placei L3 could not overwinter at all. Adult C. oncophora, O. ostertagi and N. helvetianus could be recovered from tracer calves grazing after the winter. CONCLUSIONS: The largest proportion of L3 were recovered from fecal pats suggesting this is important refuge for L3 survival. Results also show that L3 of several GIN parasite species can survive relatively efficiently on pastures even in the extreme winter conditions in western Canada. Tracer calf experiments confirmed that overwintered L3 of both C. oncophora and O. ostertagi were capable of establishing a patent infection in the following spring. These results have important implications for the epidemiology, risk of production impact and the design of effective control strategies. The work also illustrates the value of applying ITS2 nemabiome metabarcoding to environmental samples.

Conspecific infection threat rapidly biases the social responses of female mice: Involvement of oxytocin.

Pathogen threat affects social preferences and responses across species. Here we examined the effects of social context and the infection status of conspecific females and males on the social and mate responses of female mice. The responses of female mice to males were rapidly affected by the presence of infected female conspecifics and infected males. In mice odor cues drive appetitive and aversive social and mate responses. Brief (1 min) exposure to the fresh urinary odors of females infected with the murine nematode parasite, Heligmosomoides polygyrus, attenuated the responses of other uninfected females to the odors of naturally preferred unfamiliar males and enhanced their preferences for familiar males. Likewise exposure to the odors of a male either infected with H. polygyrus or treated with the bacterial endotoxin, lipopolysaccharide, reduced the responses of females to the odors of unfamiliar males. In addition, females displayed an avoidance of, and discrimination against, male mice whose odors had been associated with that of an infected female ("guilt by association") and a preference for the odors associated with an uninfected female ("mate copying"). These shifts in preferences for female associated male odors were attenuated in a dose-related manner by pre-treatment with the oxytocin receptor antagonist, L-368,899. These findings show that social information associated with the infection status of conspecifics can rapidly bias the mate preferences of female mice in an oxytocin receptor dependent manner.

Mitochondrial Genome Fragmentation Unites the Parasitic Lice of Eutherian Mammals.

Organelle genome fragmentation has been found in a wide range of eukaryotic lineages; however, its use in phylogenetic reconstruction has not been demonstrated. We explored the use of mitochondrial (mt) genome fragmentation in resolving the controversial suborder-level phylogeny of parasitic lice (order Phthiraptera). There are approximately 5000 species of parasitic lice in four suborders (Amblycera, Ischnocera, Rhynchophthirina, and Anoplura), which infest mammals and birds. The phylogenetic relationships among these suborders are unresolved despite decades of studies. We sequenced the mt genomes of eight species of parasitic lice and compared them with 17 other species of parasitic lice sequenced previously. We found that the typical single-chromosome mt genome is retained in the lice of birds but fragmented into many minichromosomes in the lice of eutherian mammals. The shared derived feature of mt genome fragmentation unites the eutherian mammal lice of Ischnocera (family Trichodectidae) with Anoplura and Rhynchophthirina to the exclusion of the bird lice of Ischnocera (family Philopteridae). The novel clade, namely Mitodivisia, is also supported by phylogenetic analysis of mt genome and cox1 gene sequences. Our results demonstrate, for the first time, that organelle genome fragmentation is informative for resolving controversial high-level phylogenies.

3D virtual histology at the host/parasite interface: visualisation of the master manipulator, Dicrocoelium dendriticum, in the brain of its ant host.

Some parasites are able to manipulate the behaviour of their hosts to their own advantage. One of the most well-established textbook examples of host manipulation is that of the trematode Dicrocoelium dendriticum on ants, its second intermediate host. Infected ants harbour encysted metacercariae in the gaster and a non-encysted metacercaria in the suboesophageal ganglion (SOG); however, the mechanisms that D. dendriticum uses to manipulate the ant behaviour remain unknown, partly because of a lack of a proper and direct visualisation of the physical interface between the parasite and the ant brain tissue. Here we provide new insights into the potential mechanisms that this iconic manipulator uses to alter its host's behaviour by characterising the interface between D. dendriticum and the ant tissues with the use of non-invasive micro-CT scanning. For the first time, we show that there is a physical contact between the parasite and the ant brain tissue at the anteriormost part of the SOG, including in a case of multiple brain infection where only the parasite lodged in the most anterior part of the SOG was in contact with the ant brain tissue. We demonstrate the potential of micro-CT to further understand other parasite/host systems in parasitological research.

Population genetic analysis informs the invasion history of the emerging trematode Dicrocoelium dendriticum into Canada.

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.

Life Cycle, Host Utilization, and Ecological Fitting for Invasive Lancet Liver Fluke, Dicrocoelium dendriticum, Emerging in Southern Alberta, Canada.

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.

Where's the risk? Landscape epidemiology of gastrointestinal parasitism in Alberta beef cattle.

BACKGROUND: Gastrointenstinal nematodes (GIN) present a serious challenge to the health and productivity of grazing stock around the globe. However, the epidemiology of GIN transmission remains poorly understood in northern climates. Combining use of serological diagnostics, GIS mapping technology, and geospatial statistics, we evaluated ecological covariates of spatial and temporal variability in GIN transmission among bovine calves pastured in Alberta, Canada. METHODS: Sera were collected from 1000 beef calves across Alberta, Canada over three consecutive years (2008-2010) and analyzed for presence of anti-GIN antibodies using the SVANOVIR Ostertagia osteragi-Ab ELISA kit. Using a GIS and Bayesian multivariate spatial statistics, we evaluated the degree to which variation in specific environmental covariates (e.g. moisture, humidity, temperature) was associated with variation in spatial and temporal heterogeneity in exposure to GIN (Nematodirus and other trichostrongyles, primarily Ostertagia and Cooperia). RESULTS: Variation in growing degree days above a base temperature of 5 °C, humidity, air temperature, and accumulated precipitation were found to be significant predictors of broad-scale spatial and temporal variation in serum antibody concentrations. Risk model projections identified that while transmission in cattle from southeastern and northwestern Alberta was relatively low in all years, rate of GIN transmission was generally higher in the central region of Alberta. CONCLUSIONS: The spatial variability in risk is attributed to higher average humidity, precipitation and moderate temperatures in the central region of Alberta in comparison with the hot, dry southeastern corner of the province and the cool, dry northwestern corner. Although more targeted sampling is needed to improve model accuracy, our projections represent an important step towards tying treatment recommendations to actual risk of infection.

Comparative recruitment, morphology and reproduction of a generalist trematode, Dicrocoelium dendriticum, in three species of host.

Epidemiological 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 adult Dicrocoelium 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 nor per capita fecundity 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.

Fluke abundance versus host age for an invasive trematode (Dicrocoelium dendriticum) of sympatric elk and beef cattle in southeastern Alberta, Canada.

Epidemiological parameters such as transmission rate, rate of parasite-induced host mortality, and rate of development of host defenses can be assessed indirectly by characterizing the manner in which parasite burdens change with host age. For parasites that are host generalists, estimates of these important parameters may be host-species dependent. In a cross-sectional study, we determined age-abundance profiles of infection in samples of sympatric free-ranging elk and domestic cattle infected with the lancet liver fluke, Dicrocoelium dendriticum. This parasite was introduced into Cypress Hills Provincial Park in southeastern Alberta, Canada in the mid 1990s, and now occurs in 60-90% of co-grazing elk and beef cattle examined at necropsy. The livers of 173 elk were made available by hunters during the 1997-2011 hunting seasons and livers from 35 cattle were purchased from ranchers. In elk, median worm abundance peaked in 6-24 month-olds (median = 72, range = 0-1006) then significantly declined to <10 worms/host in 10-16 year olds. The decline in fluke burden with age is not consistent with an age-related decline in exposure to metacercariae in intermediate hosts and high rates of fluke-induced host mortality are unlikely. Rather, the pattern of peak fluke burdens in elk calves and juveniles, followed by a decline in older animals is consistent with the development of a protective immune response in older hosts. There was no pattern of worm accumulation or decline in sympatric cattle, although statistical power to detect a significant effect was low. These results highlight the complexity and context-dependent nature of epidemiological processes in multi-host systems.

Annual variation in serum antibody concentrations against gastrointestinal nematodes in beef calves from semi-arid rangelands of western Canada.

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.

Acaricide resistance in cattle ticks and approaches to its management: the state of play.

Cattle ticks are an important constraint on livestock production, particularly in tropical and subtropical areas. Use of synthetic acaricides is the primary method of tick control; therefore, it would be imperative to develop strategies to preserve the efficacy of existing acaricides. This paper summarizes the status of acaricide resistance in cattle ticks from different parts of the world and reviews modes of action of currently used acaricides, mechanism of resistance development, contributory factors for the development and spread of resistance, management of resistant strains and strategies to prolong the effect of the available acaricides. Use of vaccines, synthetic and botanical acaricides and educating farmers about recommended tick control practices are discussed, along with the integration of currently available options for the management of drug resistance and, ultimately, the control of cattle ticks.

Out of sight but not gone: Sero-surveillance for cattle grubs, Hypoderma spp., in western Canada between 2008 and 2010.

Despite the lack of evidence of clinically infected cattle in western Canada since the late 1990s there is persistent evidence from serological sampling that warble fly infections remain endemic. As part of a general survey of cattle parasites conducted on calves entering a feedlot in southern Alberta sera were tested for the presence of anti-Hypoderma antibodies using an established ELISA. Calves for the study were drawn from auction markets in Manitoba, Saskatchewan, Alberta and British Columbia and serum samples were collected, according to a randomization scheme, as the calves entered a single feedlot in October through December (Survey) of three consecutive years (2008-2010). In addition, a sub-sample of calves originating on a ranch in southeastern Alberta (Ranch 1) that had been the subject of previous surveillance was tested. Sero-prevalence of anti-Hypoderma antibodies of the Survey calves ranged from 0.0% to 100.0% depending on origin and overall varied among the three years with the sero-prevalence being 45.6%, 26.9%, 28.5% in 2008, 2009 and 2010 respectively. Sero-prevalence in Ranch 1 calves varied among years, with one year being much higher and was different from overall Survey sero-prevalence in two of three years (31.7%, 64.1%, 38.5% in 2008, 2009, 2010 respectively). In the Survey population there was a trend to lower sero-prevalence over the three years while on Ranch 1 there was no clear trend. In the samples with ELISA adjusted absorbance values above the positive cut-off an examination of the cumulative relative frequency showed that in all years approximately 90% of the samples were in the lower 3 of 14 bins in both populations and in two of three years in Ranch 1 population. The clustering of the positive ELISA data at the lower end of the range is suggestive of low level grub infestations. The increased frequency of high positive ELISA results from Ranch 1 in 2009 potentially indicate an 'outbreak year' in which pupal and adult fly survival was higher than for other years of this study resulting in more infested animals with higher burdens. Examination of precipitation and temperature profiles from the subject ranch support this suggestion.

Vector-borne parasitic zoonoses: emerging scenarios and new perspectives.

Changing climate is not the only driver for alterations in the dynamic interaction between arthropod vectors of zoonotic parasites and their hosts, including humans. A suite of other factors ranging from urbanization and deforestation to changing demographics in both developing and developed countries, the impact of the recent economic crisis, increased global movement of people and animals and follow-on effects of major catastrophes. This article reviews the most important vector-borne parasites of zoonotic concern that are changing/expanding their distribution patterns in both endemic and/or previously non-endemic areas. We include the discussion of the changing aspects of malaria, leishmaniasis, babesiosis, Chagas disease as well as of some spirurid and filarioid nematodes.

Hidden antigens from third instar Hypoderma lineatum: impact of immunization on larval survival in artificial infestations.

Soluble fractions of Hypoderma lineatum third instar fat body, haemocytes and haemolymph were formulated with Quil A and used to immunize four groups of calves while a fifth group remained untreated. Calves received two subcutaneous injections of the soluble fractions, or adjuvant only delivered two weeks apart. Two weeks after the last injection the calves were exposed to 50 newly hatched larvae of H. lineatum which were placed on the skin and allowed to penetrate. Survival of larval stages was monitored by weekly palpation and collection of emergent third instars. Antibody responses to the immunogens were evaluated by immunoblots and following infestation antibody responses to first instar antigens were evaluated by an ELISA. Non-immunized calves and calves injected with adjuvant were all palpation positive for cattle grubs. In groups immunized with fat body, haemocyte and haemolymph components 100%, 33% and 33% were palpation positive for grubs respectively. First instar mortality, as reflected in palpable grubs, was high in the groups receiving injections with tissue components (99.3%, 95.1%, 95.8%, 83.9 and 80.4% mortality for those groups receiving fat body, haemocyte, haemolymph, adjuvant or control respectively). Second and third instar mortality was also higher in the immunized groups (100.0%, 91.7%, 91.7% for fat body, haemocyte, and haemolymph respectively) in comparison to the adjuvant only (14.0%) and unvaccinated (33.3%) groups. No viable flies emerged from pupae originating from larvae emergent from any of the immunized groups. Calves receiving the tissue extracts developed antibodies to several protein components following the second immunization which were still present 13 weeks post-infestation. Several proteins appeared to be common among the three tissue extracts and were recognized by antibodies from the immunized calves. All groups of calves became positive for antibodies to first instar antigens, although in some immunized calves the antibodies were transient.

Dicrocoelium dendriticum in cattle from Cypress Hills, Canada: humoral response and preliminary evaluation of an ELISA.

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.

Analysis of a mitochondrial noncoding region for the identification of the most diffused Hypoderma species (Diptera, Oestridae).

Hypoderma spp. larvae cause internal myiasis in domestic and wild animals characterized by subcutaneous warbles. Their differentiation is usually performed at species level based on the morphology of third stage larvae. The recent release of the whole mtDNA of Hypoderma lineatum and the retrieval of a 102 bp noncoding region occurring between tRNA(Ser) and NADH dehydrogenase subunit 1 (nd1) genes represented the foundation for this study. The noncoding region and the two flanking mitochondrial genes (i.e., tRNA(Ser) and nd1) of the most diffused Hypoderma spp. (i.e., Hypoderma actaeon, Hypoderma bovis, Hypoderma diana, H. lineatum, Hypodermasinense and Hypoderma tarandi) were analysed. Interspecific variations in amplicon size (from 20 to 102 bp in H. tarandi and H. lineatum, respectively) and nucleotide sequences allowed a genetic discrimination of the examined species providing information instrumental to a rapid molecular identification of Hypoderma spp.