Georg Gerhard Duscher

Genetic diversity of the cestode Echinococcus multilocularis in red foxes at a continental scale in Europe

Background Alveolar echinococcosis (AE) is a severe helminth disease affecting humans, which is caused by the fox tapeworm Echinococcus multilocularis. AE represents a serious public health issue in larger regions of China, Siberia, and other regions in Asia. In Europe, a significant increase in prevalence since the 1990s is not only affecting the historically documented endemic area north of the Alps but more recently also neighbouring regions previously not known to be endemic. The genetic diversity of the parasite population and respective distribution in Europe have now been investigated in view of generating a fine-tuned map of parasite variants occurring in Europe. This approach may serve as a model to study the parasite at a worldwide level. Methodology/Principal Findings The genetic diversity of E. multilocularis was assessed based upon the tandemly repeated microsatellite marker EmsB in association with matching fox host geographical positions. Our study demonstrated a higher genetic diversity in the endemic areas north of the Alps when compared to other areas. Conclusions/Significance The study of the spatial distribution of E. multilocularis in Europe, based on 32 genetic clusters, suggests that Europe can be considered as a unique global focus of E. multilocularis, which can be schematically drawn as a central core located in Switzerland and Jura Swabe flanked by neighbouring regions where the parasite exhibits a lower genetic diversity. The transmission of the parasite into peripheral regions is governed by a “mainland–island” system. Moreover, the presence of similar genetic profiles in both zones indicated a founder event.

A molecular survey of vector-borne pathogens in red foxes (Vulpes vulpes) from Bosnia and Herzegovina.

BackgroundRed foxes (Vulpes vulpes) have recently been recognized as potential reservoirs of several vector-borne pathogens and a source of infection for domestic dogs and humans, mostly due to their close vicinity to urban areas and frequent exposure to different arthropod vectors. The aim of this study was to investigate the presence and distribution of Babesia spp., Hepatozoon canis, Anaplasma spp., Bartonella spp., ‘Candidatus Neoehrlichia mikurensis’, Ehrlichia canis, Rickettsia spp. and blood filaroid nematodes in free-ranging red foxes from Bosnia and Herzegovina.MethodsSpleen samples from a total of 119 red foxes, shot during the hunting season between October 2013 and April 2014 throughout Bosnia and Herzegovina, were examined for the presence of blood vector-borne pathogens by conventional PCRs and sequencing.ResultsIn the present study, three species of apicomplexan parasites were molecularly identified in 73 red foxes from the entire sample area, with an overall prevalence of 60.8%. The DNA of B. canis, B. cf. microti and H. canis was found in 1 (0.8%), 38 (31.9%) and 46 (38.6%) spleen samples, respectively. In 11 samples (9.2%) co-infections with B. cf. microti and H. canis were detected and one fox harboured all three parasites (0.8%). There were no statistically significant differences between geographical region, sex or age of the host in the infection prevalence of B. cf. microti, although females (52.9%; 18/34) were significantly more infected with H. canis than males (32.9%; 28/85). The presence of vector-borne bacteria and filaroid nematodes was not detected in our study.ConclusionThis is the first report of B. canis, B. cf. microti and H. canis parasites in foxes from Bosnia and Herzegovina and the data presented here provide a first insight into the distribution of these pathogens among the red fox population. Moreover, the relatively high prevalence of B. cf. microti and H. canis reinforces the assumption that this wild canid species might be a possible reservoir and source of infection for domestic dogs.

Ticks and haemoparasites of dogs from Praia, Cape Verde.

In February 2008 an epidemiological field study on arthropod-borne infections in dogs was carried out in Praia, the capital city of Cape Verde. For this purpose 130 dogs were included in the study. Of these, 94.6% were infested with ticks. Altogether, 1293 ticks of the genus Rhipicephalus (in all evaluated cases R. sanguineus) were collected. Examination for haemotropic parasites was performed via polymerase chain reaction (PCR). Lymph node fine-needle aspirates were screened by PCR for Leishmania infantum infections in 20 dogs with enlarged lymph nodes. Our investigation revealed two species of protozoa (Babesia canis vogeli and Hepatozoon canis) and two species of rickettsiae (Anaplasma platys and Ehrlichia canis). In 101 dogs (77.7%) DNA of one or more pathogens was detected. The PCR examination for H. canis was positive in 83 dogs (63.8%), for E. canis in 34 dogs (26.2%), for A. platys in 10 dogs (7.7%) and for B. canis in five dogs (3.8%), whereas neither B. gibsoni nor L. infantum DNA could be detected. Of the infected dogs, 71.3% had a monoinfection, 27.7% had infections with two pathogens and 1.0% with three pathogens. B. canis, H. canis, E. canis, A. platys and their vector tick R. sanguineus are endemic to Cape Verde and can be present in dogs in high prevalences. These results outline the risk of importing tropical canine diseases when Capeverdian stray dogs are taken to Europe.

Fasciola hepatica – Monitoring the milky way? The use of tank milk for liver fluke monitoring in dairy herds as base for treatment strategies

In this study 595 lactating cows originating from 31 carinthian farms were investigated in accordance of liver fluke infection using individual and tank milk as well as individual blood and faecal samples. Two commercial ELISAs were used to test the milk and blood serum, and the results were compared with coproscopy and a commercial copro-antigen ELISA. Receiver operating characteristics (ROC) and two-graph operating characteristics (TG ROC) of tank milk results were conducted based on the individual milk to determine the minimum reliable in-herd antibody prevalence for the predominant condition in the investigation area. In 17.8% of the examined individuals located in 64.5% of the farms eggs were detected by coproscopy. The copro-antigen ELISA delivered 13.4% positive individuals from 54.8% of the farms. The milk ELISAs showed 42.7% (Euroclone) and 44.2% (Pourquier) positive cows on 90.3% of the farms. The blood samples were positive in 43% (Euroclone) and 45.2% (Pourquier) of the individuals from 90.3% to 96.8% of the herds, respectively. Based on the milk and the blood an average in-herd prevalence of 30-45% can be assumed. The serum and milk samples delivered correlating results with kappa values between 0.94 and 0.97, whereas the coproscopy and copro-antigen ELISA did not correlate well with the ELISA results. The two different ELISA tests highly correlated on individual and on herd level. Both showed a reliable minimum in-herd prevalence of ∼20%, meaning that one fifth of the individuals in a herd have to be positive to obtain a positive bulk tank milk result. In the investigated area a higher in-herd prevalence is expected, therefore the tank milk is useful as a monitoring tool and can be used as a basis for intervention strategies.

Detection of Dirofilaria in Austrian dogs

A recent increase in the occurrence of Dirofilaria repens and Dirofilaria immitis has been reported from the eastern neighbouring countries of Austria (Slovakia, Hungary). In order to obtain data from Austria--especially from the areas bordering these countries--blood samples of dogs from Gänserndorf (northeastern Austria, bordering Slovakia) and Neusiedl (eastern Austria, bordering Hungary) were examined for the presence of Dirofilaria spp.-DNA by quantitative real-time PCR (qPCR). For D. repens one (out of eight) and six (out of ninety) positive dogs were found, respectively. D. immitis-DNA was not detected. Most of the positive dogs had previously been abroad in known Dirofilaria endemic areas. Two dogs, however, had never been abroad and presumably became infected in Austria. To investigate an autochthonous focus, 1366 mosquitoes from the eastern part of Austria were also examined by qPCR. Although positive mosquitoes could not be found, an emerging endemic occurrence of D. repens in Austria due to positive dogs seems possible.

Fox on the run--molecular surveillance of fox blood and tissue for the occurrence of tick-borne pathogens in Austria.

BackgroundThe red fox (Vulpes vulpes) is a widespread species, harbouring many pathogens relevant for humans and pets. Indeed, Anaplasma spp., Ehrlichia canis and Rickettsia spp. among the bacteria and Hepatozoon canis as well as Babesia sp. among the parasites have been the focus of several studies.FindingsIn a cohort of 36 foxes shot on one day in the north-eastern part of Austria, Babesia microti-like pathogens were found in 50%, while H. canis was detected in 58.3% of the samples. The spleen was more useful for detection of H. canis, whereas B. microti-like parasites were more frequently found in the blood. Bacteria could not be confirmed in any of the cases to demonstrate the occurrence of such tick-borne pathogens using PCR and sequencing on blood and spleen samples.ConclusionsThe occurrence of B. microti-like and H. canis parasites raised many questions, because these infections have never been found autochthonously in dogs. Furthermore in the case of H. canis the main vector tick, Rhipicephalus sanguineus, is absent in the sampling area, leaving space for further hypotheses for transmission such as vertical transmission, transmission via ingestion of prey animals or other vector ticks. Further studies are needed to evaluate the risks for pets in this area. PCRs delivered differing results with the different tissues, suggesting the use of both spleen and blood to obtain an integral result.

Scraping or shaking—a comparison of methods for the quantitative determination of Echinococcus multilocularis in fox intestines

A modified sedimentation technique (the “shaking in a vessel” technique; SVT) to examine intestines for smaller helminths such as Echinococcus multilocularis is described and compared with the intestinal scraping technique (IST). Out of 356 foxes, 26 were E. multilocularis positive, 19 using IST (sensitivity 73.1%) and 25 with SVT (sensitivity 96.2%) after IST. Especially low infection intensities went undetected with IST. The result is comparable to the established sedimentation and counting technique (SCT). SVT is recommended for the routine examination of fox intestines as an alternative to SCT.

A golden jackal (Canis aureus) from Austria bearing Hepatozoon canis--import due to immigration into a non-endemic area?

The protozoan Hepatozoon canis, which is transmitted via ingestion of infected ticks by canine hosts, is not endemic to mid-latitude regions in Europe. Its distribution is supposed to be linked to the occurrence of its primary tick vector Rhipicephalus sanguineus. A young male golden jackal (Canis aureus) found as road kill close to Vienna, Austria, was infected by this pathogen. Cloning and sequencing of the PCR product revealed 6 different haplotypes of H. canis. Based on the sequences, no clear relationship to the origin of infection could be traced. This is the first report of H. canis for Austria, and wild canines such as the currently found jackal may provide a source of natural spread of this parasite into non-endemic areas. This natural immigration of wild animals represents a way of pathogen introduction, which has to be considered in disease prevention in addition to human-made introduction due to animal import and export.

Seasonal and spatial distribution of ixodid tick species feeding on naturally infested dogs from Eastern Austria and the influence of acaricides/repellents on these parameters

BackgroundEffective control of tick infestation and pathogen transmission requires profound knowledge of tick biology in view of their vector function. The particular time of the year when the different tick species start to quest and the favoured sites on the canine host are of major interest. The efficacy of acaricides/repellents to control ticks in the field requires observation.MethodsTo address these issues, 90 dogs, grouped in “untreated”, “acaricide/repellent” (permethrin) and “acaricide only” (fipronil) animals and subjected to tick infestation under natural conditions in Burgenland (Eastern Austria), were examined. The number and species of ticks occurring during and outside the protection time was evaluated during a period of 11 months and the biting location on the dogs’ skin was recorded.ResultsOf the 700 ticks collected, the most common species in that particular walking area was Ixodes ricinus, followed by Dermacentor reticulatus and Haemaphysalis concinna. Regarding the on-host activity, D. reticulatus displayed more infestations in early spring and late autumn, whereas I. ricinus occurred almost one month later in spring and one month earlier in autumn. H. concinna followed a monophasic pattern of activity with a peak in summer. The preferred feeding sites of the ticks on the dogs were on the head, neck, shoulder and chest. This distribution over the dog’s body was not influenced by the use of the drugs, although on the whole fewer ticks (22.5% of all ticks) were found during the protection time. Interestingly, differences occurred with the use of drugs compared to non-protected dogs with regard to the infestation over the year. Acaricide-treated dogs displayed a higher prevalence in April, May and September, whereas dogs of the acaricide/repellent group showed a higher infestation in March, July, October and November.ConclusionThe different tick species display different on-dog activity peaks over the year, during which particular canine diseases can be expected and predicted, considering the specific incubation times for each pathogen.The tick species occurring in this study do not seem to choose particular sites on the dogs. Their arrival place seems to represent the attachment and consequently the feeding sites. The use of acaricides leads to a significantly (p<0.01) lower number of infesting ticks but no change of the distribution pattern on the dogs was observed.

Wildlife reservoirs for vector-borne canine, feline and zoonotic infections in Austria

Graphical Abstract