Browsing by Person "Mackenstedt, Ute"

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Blood parasites of vangas and other corvoidea on Madagascar
(2022) Magaña Vázquez, Regina; Woog, Friederike; Dinkel, Anke; Mackenstedt, Ute; Musa, Sandrine
Madagascar hosts a great diversity of bird species. This study focuses on the description of the diversity and prevalence of blood parasites (Haemosporida, trypanosomes and filarioid nematodes) in 131 blood samples of 14 species of Corvoidea, namely vangas (Vangidae), Coracina cinerea (Campephagidae), Dicrurus forficatus (Dicruridae) and Terpsiphone mutata (Monarchidae) found in primary rainforests on Madagascar. Blood parasites were detected using both molecular and microscopic methods. Multiplex PCR was used to detect mixed haemosporidian infections and nested PCR was used to describe a 479 bp fragment of the haemosporidian cytochrome b (cytb) gene. Furthermore, a 770 bp SSU rRNA fragment of trypanosomes, and, for microfilariae, a 690 bp fragment of 28S rRNA, as well as a 770 bp fragment of 28S rRNA, were amplified for identification using nested PCRs. Phylogenetic analyses were carried out for all sequences obtained from all blood parasite taxa. Over half of the samples (54.2%; n = 71) were infected with Haemosporida, whereas only 21.4% (n = 28) were infected with Trypanosoma and 5.3% (n = 7) contained filarioid nematode DNA. Fourteen of 56 blood smears contained some of the above-mentioned parasite taxa. The results corroborate the great diversity of blood parasites in the different bird species studied, especially in vangas. Vangas had the greatest diversity of parasites found, as well as the highest number of multiple infections, which may be due to their morphological diversity and resulting habitat use. Fifteen haemosporidian lineages, seven Trypanosoma and five filarioid nematode isolates were newly discovered in the avian species studied, particularly in the vangas. Members of the other Corvoidea families on Madagascar showed a lower susceptibility for avian haemosporidian parasites than vangas, which could be attributed to possible resistance against those parasites. The study confirmed the host specificity of some Haemosporida and microfilariae; however, it demonstrated that this was not the case for Trypanosoma.
The emergence and dynamics of tick-borne Encephalitis Virus in a new endemic region in Southern Germany
(2022) Lang, Daniel; Chitimia-Dobler, Lidia; Bestehorn-Willmann, Malena; Lindau, Alexander; Drehmann, Marco; Stroppel, Gabriele; Hengge, Helga; Mackenstedt, Ute; Kaier, Klaus; Dobler, Gerhard; Borde, Johannes
Tick-borne encephalitis (TBE) is the most important viral tick-borne infection in Europe and Asia. It is emerging in new areas. The mechanisms of emergence are fairly unknown or speculative. In the Ravensburg district in southern Germany, TBE emerged, mainly over the last five years. Here, we analyzed the underlying epidemiology in humans. The resulting identified natural foci of the causal TBE virus (TBEV) were genetically characterized. We sampled 13 potential infection sites at these foci and detected TBEV in ticks (Ixodes ricinus) at eight sites. Phylogenetic analysis spurred the introduction of at least four distinct TBEV lineages of the European subtype into the Ravensburg district over the last few years. In two instances, a continuous spread of these virus strains over up to 10 km was observed.
Epidemiologie von Coxiella burnetii, Rickettsia spp., FSME- und Hantaviren in Süddeutschland unter Berücksichtigung klimatischer Veränderungen
(2011) Pluta, Silvia; Mackenstedt, Ute
Zoonoses, which are defined as human infections caused by animal pathogens, are of considerable medical importance. Therefore, it is important to evaluate the distribution of tick- and rodent borne pathogens and the risk of infections for humans. Hence, the epidemiology of specific human pathogens (Coxiella burnetii, Rickettsia spp., TBE-virus and Hantavirus) was studied. As climatic conditions are known to have a significant influence on ticks and rodents, the role of climate change in the epidemiology of the examined pathogens was considered. The role of natural foci in the epidemiology of Coxiella burnetii, the causative agent of Q fever, is widely unknown in Germany. Therefore, ticks of the genus Dermacentor and rodents from Q fever endemic areas in Southern Germany were examined for infections with C. burnetii. Altogether, 1120 ticks and 119 rodents were tested by PCR. However, no infections were detected. Rodent blood samples were additionally tested for antibodies against coxiellae, but likewise, all samples were found to be negative. Hence, no natural foci of Q fever were identified in the examined areas. A role of natural foci in the epidemiology of Q fever in Southern Germany is therefore unlikely. Both Dermacentor marginatus and D. reticulatus act as vectors for Rickettsia spp. To elucidate if human pathogenic rickettsiae are distributed in Southern Germany, Dermacentor ticks were investigated for infections by PCR. Moreover, rodents were screened by molecular and serological methods to identify potential reservoir hosts. The overall prevalence of Rickettsia spp. in 1120 ticks was 33.6%. By sequencing of the rOmpA-gene, R. raoultii was identified in 32.8% and R. slovaca in 0.8% of all examined ticks. Both species are pathogenic for humans; thus, there might be a considerable risk of infection. This was confirmed by the detection of an autochthonous case of R. slovaca-infection in Rhineland-Palatinate. Rickettsia spp. was not detected in any rodents by PCR. However, 48 animals (20.2%), muridae as well as arvicolidae, showed antibodies against rickettsiae. It was, therefore, shown that rodents can actually act as reservoir hosts for Rickettsia spp. To determine the current prevalence of TBE virus in ticks, 7842 Ixodes ricinus from Baden-Württemberg were examined by molecular methods. TBE virus was found in four out of 11 examined areas. Prevalences ranged from 0.13% to 0.66%. No infected ticks were found in other areas. The determined prevalences were lower compared to studies from the 1990ies in the same areas. However, reliable comparisons are difficult due to the low numbers of infected ticks. The viruses of ticks from Gengenbach, Hagnau am Bodensee and Hödinger Tobel were cultured and subsequently characterized by sequencing of the E and NS2a genes. Phylogenetic studies were conducted to identify the TBE virus strains present in Baden-Württemberg. The isolate from Hagnau am Bodensee was identified as strain Salem, whereas the isolates of the other two areas differed substantially from all strains found in the database. Therefore, no identification was possible for these isolates. In 2007, the number of notified hantavirus infections exceeded by far the average case numbers of the preceding years. To determine if the steep rise in human infections is associated with an unusual high virus prevalence in reservoir hosts, red bank voles from the endemic area of the Swabian Albs were examined for infections by molecular and serological methods. The overall prevalence of Puumala hantavirus was 57.9%. In a study in 2001 in the same area, the prevalence was only 10%. Therefore, there was a sixfold rise in virus prevalence which reflects the high number of human cases in the study year. Sequence comparisons showed a close relationship of the hantaviruses from the Swabian Alb to hantaviruses detected in other areas of Baden-Württemberg. The results presented in this study provide the basis for further continuous studies. These are necessary to estimate the actual influence of changes in climate conditions on the examined zoonoses and, thereby, the prospective infection risk for humans.
Genetische Diversität von Echinococcus granulosus sensu stricto
(2017) Ebi, Dennis; Mackenstedt, Ute
The dog tapeworm Echinococcus granulosus sensu stricto, distributed worldwide, is the most important agent of human Cystic Echinococcosis, a zoonotic disease which is considered as one of the priority ‘neglected zoonotic diseases’ by the World Health Organisation. The parasite is known from a large number of host species and is conventionally subdivided into several genotypes. The epidemiological consequences of the variation, however, are still unknown. In the last years several studies on the intraspecific variation of this parasite were published. Most of these studies were based on small numbers of isolates from geographically restricted regions, so that the relevance of that diversity concerning pathogenicity, host specificity, transmission cycles and biogeography is still little understood. To achieve a comprehensive estimate of its genetic diversity, 1085 isolates of E. granulosus s.s. from humans and animal hosts originating from widely different regions of the world were analyzed for the sequence polymorphism of a 1609 bp fragment of the mitochondrial cox1 gene. For analysis haplotype networks, diversity and neutrality indices of subpopulations of different geographic origin were compared. To improve the analytical efficiency of the sequence-data, a LabVIEW application was written. It could be shown that most diverse sub-population is found in the Middle Eastern region and that the genetic diversity is lower in the other analyzed regions, which confirms a previous tentative hypothesis on the origin of the domestic cycle of E. granulosus s.s. in that region. Records of identical haplotypes in different regions, inclusion of published data, calculation of parameters of population genetics and haplotype network analysis led to hypotheses explaining the distribution history of E. granulosus s.s.. This in line with fossil and molecular data on domestic sheep as the principal final host. A set of new and easily calculable genetic diversity indices were developed and their added value was explained. Additionally, a statistical resampling experiment was programmed which showed that at least 40 well distributed samples are necessary to provide a reliable estimate of the genetic diversity of the parasite in a region. Using an extension for that software, it was shown that the newly developed diversity indices do not strongly depend on the number of analyzed samples, and provide appropriate tools to assess the genetic diversity of parasite populations.
Helminths in invasive raccoons (Procyon lotor) from Southwest Germany
(2023) Reinhardt, Nico P.; Wassermann, Marion; Härle, Jessica; Romig, Thomas; Kurzrock, Lina; Arnold, Janosch; Großmann, Ernst; Mackenstedt, Ute; Straubinger, Reinhard K.
As hosts of numerous zoonotic pathogens, the role of raccoons needs to be considered in the One Health context. Raccoons progressively expand their range as invasive alien species in Europe. This study aimed to investigate the intestinal helminth fauna of raccoons in Baden-Wuerttemberg, Germany, as no such screening had ever been conducted there. In total, we obtained 102 animals from hunters in 2019 and 2020. Intestinal helminths were retrieved using the SSCT (segmented sedimentation and counting technique) and identified morphologically and by PCR-based Sanger sequencing. Fecal samples were assessed using the ELISA PetChekTM IP assay (IDEXX, Germany) and flotation technique. The artificial digestion method was employed for analyzing muscle tissue. We detected species of four nematode genera (Baylisascaris procyonis, Toxocara canis, Capillaria spp., and Trichuris spp.), three cestode genera (Atriotaenia cf. incisa/procyonis, Taenia martis, and Mesocestoides spp.), and three trematode genera (Isthmiophora hortensis/melis, Plagiorchis muris, and Brachylaima spp.). Echinococcus spp. and Trichinella spp. were not found. The invasive behavior and synanthropic habits of raccoons may increase the infection risk with these helminths in wildlife, domestic and zoo animals, and humans by serving as a connecting link. Therefore, it is crucial to initiate additional studies assessing these risks.
Increased vaccination diversity leads to higher and less-variable neutralization of TBE viruses of the European subtype
(2023) Bestehorn-Willmann, Malena; Girl, Philipp; Greiner, Franziska; Mackenstedt, Ute; Dobler, Gerhard; Lang, Daniel
Tick-borne encephalitis (TBE) is an infectious disease of the central nervous system. The causative agent is the tick-borne encephalitis virus (TBEV), which is most commonly transmitted by tick bites, but which may also be transmitted through the consumption of raw dairy products or, in rare instances, via infected transfusions, transplants, or the slaughter of infected animals. The only effective preventive option is active immunization. Currently, two vaccines are available in Europe—Encepur® and FSME-IMMUN®. In Central, Eastern, and Northern Europe, isolated TBEV genotypes belong mainly to the European subtype (TBEV-EU). In this study, we investigated the ability of these two vaccines to induce neutralizing antibodies against a panel of diverse natural TBEV-EU isolates from TBE-endemic areas in southern Germany and in regions of neighboring countries. Sera of 33 donors vaccinated with either FSME-IMMUN®, Encepur®, or a mixture of both were tested against 16 TBEV-EU strains. Phylogenetic analysis of the TBEV-EU genomes revealed substantial genetic diversity and ancestry of the identified 13 genotypic clades. Although all sera were able to neutralize the TBEV-EU strains, there were significant differences among the various vaccination groups. The neutralization assays revealed that the vaccination using the two different vaccine brands significantly increased neutralization titers, decreased intra-serum variance, and reduced the inter-virus variation.
Morphological and molecular studies on cerebral and non-cerebral coenurosis in sheep and goats
(2017) Christodoulopoulos, Georgios; Mackenstedt, Ute
In dieser Forschungsarbeit wurden Isolate von Taenia multiceps als Erreger der zerebralen und nicht-zerebralen Coenurose bei Schafen und Ziegen vergleichend untersucht. Erreger von nicht-zerebraler Coenurose aus einem breiten geographischen Bereich in Afrika und Westasien wurden verglichen mit Erregern zerebraler Coenuruszysten von gesammelten Drehkrankheitsfällen aus Griechenland, wo zerebrale Coenurose häufig vorkommt, wo aber nicht-zerebrale Coenurose noch nie beschrieben wurde. Diese Forschungsarbeit enthält eine feld- und eine laborexperimentelle Komponente und beschäftigt sich mit Aspekten wie: (i) das Vorhandensein oder Fehlen von nicht-zerebraler Coenurose bei Schafen; (ii) die Beschreibung von nicht-zerebraler Coenurose im Zwischenwirt (Schafe und Ziegen); und (iii) die phylogenetische Auflösung des T. multiceps-Clusters sowie eine mögliche Erklärung, warum nicht-zerebrale Coenurose in bestimmten geographischen Gebieten unbekannt ist. Im Rahmen der Felduntersuchung wurde eine Gesamtzahl von 90,415 geschlachtete Schafe und 2,284 geschlachtete Ziegen aus Schlachthöfen der Vereinigten Arabischen Emirate (UAE) und Ägypten untersucht, die aus verschiedenen tropischen und subtropischen Ländern wie Indien, Pakistan, Iran, Oman, Sudan, Somalia und Äthiopien stammten. Die Feldarbeit umfaßte außerdem die Sammlung von zerebralen Coenurosezysten von 20 Schafen und sechs Ziegen, die vom griechischen Festland und Umgebung stammten. Im Labor wurden vier Teilstudien durchgeführt: (1) eine morphologische Untersuchung der Merkmale von nicht-zerebralen Coenurosezysten, deren Cluster und der Protoskolizes, (2) ein morphologischer Vergleich der Rostellarhaken der zerebralen und nicht-zerebralen Coenurosezysten, (3) ein morphologischer Vergleich von erwachsenen Würmern aus Hunden, die experimentell mit Protoskolizes von zerebralen und nicht-zerebralen Zysten von Schafen und Ziegen infiziert worden waren, und (4) eine molekulare Analyse von Teilsequenzen dreier mitochondrialer Gene (nad1, cox1 und 12S rRNA) der oben genannten Isolate von zerebralen und nicht-zerebralen Zysten. Die Prävalenz von nicht-zerebraler Coenurose betrug bei Ziegen 1,75% und nur 0,008% bei Schafen. Die Zysten wurden in verschiedenen Muskeln sowie an den Nieren, am Mesenterium und am Herz gefunden. Sechsundsiebzig nicht-zerebrale Coenuruszysten von Ziegen wurden gesammelt und statistisch ausgewertet. Die Anzahl der Protoskolizes korrelierte signifikant positiv mit dem Volumen von Zysten und die Anzahl von Clustern korrelierte signifikant positiv mit der Anzahl der Protoskolizes, was auf positives allometrisches Wachstum hinweist. Die Anzahl von Clustern korrelierte mit der Anzahl von Zysten, was auf ein negatives allometrisches Wachstum hinweist. Die biologische Bedeutung dieser Allometrie ist nicht bekannt, aber der Parasit investiert offenbar seine Ressourcen mehr in das Wachstum von Protoskolizes und weniger in das Wachstum von Zysten und Clustern. Die morphologischen Untersuchungen der Rostellarhaken von Protoskolizes aus zerebralen und nicht-zerebralen Zysten und die in Hunden experimentell erzeugten adulten Würmer zeigten keine Unterschiede in Bezug auf die Wirtstierart (Schafe oder Ziegen). Im Gegensatz dazu wiesen adulte Würmer, die aus der Inokulation zerebraler und nicht-zerebraler Zysten hervorgingen, deutliche morphologische Unterschiede auf. Die meisten Meßwerte der Haken und Proglottiden unterschieden sich signifikant, aber die Form der kleinen Haken, die Verteilung der Hoden in den reifen Proglottiden und das Aussehen der Spiralen des Vas deferens waren die Merkmale mit den auffälligsten Unterschieden. Diese morphologischen Unterschiede fallen allerdings in den Bereich der Variationen von T. multiceps. Die phylogenetische Analyse der mitochondrialen Haplotypen ergab drei distinkte Cluster: eines, das sowohl zerebrale Isolate aus Griechenland als auch nicht-zerebrale Isolate aus tropischen und subtropischen Ländern umfaßte, und zwei Cluster, die ausschließlich aus zerebralen Isolaten aus Griechenland bestanden. Die meisten der nicht-zerebralen Proben gruppierten zusammen, bildeten aber keine monophyletische Gruppe. Dasselbe gilt für geographische Aspekte, obwohl Proben aus derselben Region zu Clustern tendierten. Die Daten zeigten eine hohe intraspezifische Diversität. Die Ergebnisse dieser Studie unterstützen den Zusammenhang zwischen genetischer Identität der T. multiceps-Isolate und der geographischen Herkunft, und führten zum Vorschlag einer neuen Hypothese zum Vorkommen von zerebralen und nicht-zerebralen Formen. Unsere phylogenetische Analyse legt nahe, daß die Entwicklung der zerebralen Coenuri bei Schafen eine ursprüngliche Eigenschaft von T. multiceps sein könnte. Alle Varianten wären damit in der Lage, zerebrale Coenurose bei Schafen zu verursachen, wogegen nur einige Varianten, vor allem aus einem genetischen Cluster, sekundär die Fähigkeit erworben haben, das Gehirn von anderen Arten (Ziegen und Rindern) zu befallen, sowie nicht-zerebrale Formen (vor allem in Ziegen) hervorzubringen. Unsere phylogenetische Analyse zeigt somit eindeutig eine molekulare Basis für nicht-zerebrale Pathogenität innerhalb der Art T. multiceps. Spezifische Biosicherheitsmaßnahmen sollten daher erwogen werden, um die Einführung von T. multiceps Varianten mit einer solchen Pathogenität zu verhindern.
Raising the bar: genus-specific nested PCR improves detection and lineage identification of avian haemosporidian parasites
(2024) Musa, Sandrine; Hemberle, Theo; Bensch, Staffan; Palinauskas, Vaidas; Baltrūnaitė, Laima; Woog, Friederike; Mackenstedt, Ute
Avian haemosporidian parasites are useful model organisms to study the ecology and evolution of parasite-host interactions due to their global distribution and extensive biodiversity. Detection of these parasites has evolved from microscopic examination to PCR-based methods, with the mitochondrial cytochrome b gene serving as barcoding region. However, standard PCR protocols used for screening and identification purposes have limitations in detecting mixed infections and generating phylogenetically informative data due to short amplicon lengths. To address these issues, we developed a novel genus-specific nested PCR protocol targeting avian haemosporidian parasites. The protocol underwent rigorous testing utilizing a large dataset comprising blood samples from Malagasy birds of three distinct Passeriformes families. Furthermore, validation was done by examining smaller datasets in two other laboratories employing divergent master mixes and different bird species. Comparative analyses were conducted between the outcomes of the novel PCR protocol and those obtained through the widely used standard nested PCR method. The novel protocol enables specific identification of Plasmodium, Haemoproteus (Parahaemoproteus), and Leucocytozoon parasites. The analyses demonstrated comparable sensitivity to the standard nested PCR with notable improvements in detecting mixed infections. In addition, phylogenetic resolution is improved by amplification of longer fragments, leading to a better understanding of the haemosporidian biodiversity and evolution. Overall, the novel protocol represents a valuable addition to avian haemosporidian detection methodologies, facilitating comprehensive studies on parasite ecology, epidemiology, and evolution.
Taeniidae in Namibian wildlife with emphasis on lion, cheetah, and African wild dog
(2024) Aschenborn, Ortwin; Mackenstedt, Ute
An opportunic survey for Echinococcus spp. in wild mammals was conducted in seven distinct stuy areas throughout Namibia, representing all major ecosystems, between 2012 and 2021. In total, 184 individually attributable faeces and 40 intestines were collected from eight species of carnivores, and 300 carcasses or organs of thirteen species of ungulates were examined for Echinococcus cysts. Nested PCR and sequencing of the mitochondrial nad1 gene led to the identification of five species of the Echinococcus granulosus sensu lato complex. Echinococcus canadensis G6/7 was found throughout Namibia at low frequency in lions, cheetahs, African wild dogs, black-backed jackals and oryx antelopes. Echinococcus equinus was present only in northern Namibia, locally at high frequency in lions, black-backed jackals and plains zebras. Echinococcus felidis was found only in one small area in the north-east of Namibia, but with high frequency in lions and warthogs. Echinococcus granulosus sensu stricto was identified only in two African wild dogs in the north-east of Namibia, and Echinococcus ortleppi occurred in central and southern Namibia in black-backed jackals and oryx antelopes. The development of fertile cysts indicated active intermediate host roles of oryx antelopes for E. canadensis and E. ortleppi, of warthogs for E. felidis, and of plains zebras for E. equinus. Our data support earlier hypotheses of exclusive or predominant wildlife life-cycles for E. felidis involving lions and warthogs, and – in Namibia – for E. equinus involving lions and/or black-backed jackals and plains zebras. Our data further support an interlink of wild and domestic transmission for E. ortleppi. A possible involvement of livestock and domestic dogs in transmission of E. canadensis G6/7 and E. granulosus s.s., the two parasite species with highest zoonotic potential, is uncertain for Namibia and needs further investigation. The present study was conducted in the isolated desert town of Oranjemund in the far south of Namibia. It is an extremely arid region where no livestock husbandry is practiced and only animals adapted to the desert can be found. However, in and around the city, artifi cial irrigation maintains lush green patches of grass that attract wild animals, in particular oryx antelopes (Oryx gazella). In 2015 four oryx antelopes were euthanised due to poor conditions and a post-mortem examination was conducted. Two were found positive for cystic echinococcosis and 16 cysts were collected for molecular analyses. In addition, faecal samples from black-backed jackals (n=5) and domestic dogs (n=9), which were regularly observed to feed on oryx carcasses, were collected and taeniid eggs isolated. Parasite species identifi cation of the cysts and eggs was done by amplifying and se- quencing the mitochondrial nad1 gene. Both oryx antelopes were found infected with E. ortleppi and one co-infected with E. canadensis G6/7. Both Echinococcus species were able to develop fertile cysts in oryx, making oryx antelopes competent hosts for these parasites. Therefore, the analysis of faecal samples was of high interest and although the numbers were quite small, taeniid eggs were found in three out of fi ve faecal samples of jackals and in all nine dog samples. However, species determination was only successful with two jackal and one dog sample. All three were positive for E. canadensis G6/7. The absence of E. ortleppi may be due to the low number of faecal samples examined. In our small study, we discovered a rather unique lifecycle of Echinococcus spp. between jackals and domestic dogs as defi nitive hosts and oryx antelopes as intermediate hosts. Here, the presence of E. canadensis G6/7 is of particular concern, as it is the second most important causative agent of CE in humans.
Untersuchung viraler, zoonotischer Erreger in Baden-Württemberg
(2019) Lindau, Alexander; Mackenstedt, Ute
Tick-borne encephalitis (TBE) is the most common tick-borne viral infection in Germany. Tick-borne encephalitis virus (TBEV) belongs to the family Flaviviridae and is transmitted either directly by tick bites or by the alimentary route, which means by contaminated food. The main reservoir hosts of TBEV are the yellow-necked mouse and the bank vole, which is also the main reservoir host for the rodent-borne Hanta viruses. Infections with Hanta virus are mainly acquired by inhaling of aerosols of urine and faeces of infected individuals. The most common type of Hanta viruses in Southern Germany is the Puumala type. Hanta-virus infections as well as TBE infections are notifiable diseases in Germany. For both diseases the amount of registered cases per year vary between years in a contrary pattern. To be able to evaluate possible risks of TBEV for humans, it is very important to know detailed information about the epidemiology, spreading of the viruses, as well as their hosts and the viruses themselves. Therefore, the presence of TBEV-foci, as well as the breakdown into micro- and macrofoci was addressed. Additionally, the prevalence of Hantavirus in rodents was studied.
Vakzinierungsstrategien gegen eine Echinococcus multilocularis-Infektion zur Charakterisierung protektiver Immunantworten
(2010) Wassermann, Torsten; Mackenstedt, Ute
The larval stages of Echinococcus multilocularis are the causative agents of the human alveolar echinococcosis (AE), which is according to the WHO the most important parasite-induced zoonosis in middle Europe. Additional knowledge about the infection process and possible protective mechanisms against an infection with this cestode would be of great value. Almost all previous studies about Echinococcus multilocularis infections in the intermediate host were carried out by using the secondary echinococcosis as route of infection. This route of infection doesn?t correspond to the natural route, the oral uptake of Echinococcus eggs resulting in a primary alveolar echinococcosis. Thereby the resulting immune responses of the secondary AE cannot be converted without reservations to the primary AE, especially in the early stages of the infection. On that account the primary AE was carried out in this study. On the one hand a conservative vaccination with purified antigen EM95 and EMGAPDH in combination with an adjuvant was subcutaneous administered and on the other hand a system was developed in which the antigen EM95 were expressed by salmonellae and exported via the hemolysinA-transport system. A significant reduction of the formation of cysts could be achieved in two separate immunisation trials with EMGAPDH combined with the adjuvant Saponin which was administered subcutaneously. The manifestation of cysts was reduced by 76.4 % in the first and 86.1 % in the second trial compared to the infected control group. The subcutaneous immunisation with the antigen EM95 reduced the manifestation of cysts even by 96.9 % and 98.4 % respectively. Sixty percentages of the animals showed a complete protection against an infection with E. multilocularis in both trials. Based of the experiments which were carried out during this study, the influence of the secretory unit or the transmembrane domain of EM95 can be excluded. The subcutaneous immunisation with the recombinant antigens EM95 and EMGAPDH induced a high antibody titre against those proteins, which based predominantly on the IgG-subclasses IgG1 and IgG2a. The detection of specific antibodies against E. multilocularis crude antigen after duration of infection of 4 weeks was neither in protected animals nor in the infected control group possible. The vaccination trials with the recombinant antigens EM95 and EMGAPDH lead us to the conclusion that the murine immune system was stimulated to detect suitable target epitopes of the oncosphere surface and to destroy these oncospheres by classical complement activation. It seems very probable that cytotoxic T-cells and natural killer cells are involved in the protective immune mechanisms against an E. multilocularis infection owing to the finding of an increased release of IFNγ as a result of an immunisation with the recombinant antigens EM95 and EMGAPDH. Summarizing the investigated immunological parameters, the protective mechanisms are based on a Th1 associated immune response. Nevertheless, an involvement of Th2 components is conceivable due to the fact that at least in parts of the study the cytokine IL-10 and a distinct IgG1 antibody response could be detected past the immunisation. By the use of the plasmid pVDL9.3 we succeeded for the first time to manipulate Salmonella typhimurium to export the metazoan protein EM95 via the hemolysinA-system. The immunisation with ZpVDL9.3EM95 resulted in a reduction of manifested cysts by 78 % compared to the infected control group. Mice immunized with ZpVDL9.3EMGAPDH, where an export was not accomplished, showed a decrease in cysts manifestation by 73 %. The amount of cysts could be further reduced up to 87 % by the combined immunisation with the Salmonella vector ZpVDL9.3EMGAPDH and a subcutaneous application of EMGAPDH. A significant reduction of cysts (68 %) could be also observed by the control immunisation with plain S. typhimurium. Antibodies against the E. multilocularis antigens EM95 and EMGAPDH could not be detected after the immunisation with the Salmonella vectors. However, the antibody formation against S. typhimurium was strongly developed. A subcutaneous post infection immunisation trial with EM95 in combination with the adjuvant Saponin at the time of 4dpi, 7dpi, 24dpi and 60dpi led neither to a reduction of the amount of cysts nor to a change in size of the developed cysts. The gained insights of this study into the protective potential of EM95 and EMGAPDH on the basis of a natural infection rout expand the knowledge and understanding about the protective immune mechanisms against an Echinococcus multilocularis infection. The development of an immunisation system in form of a Salmonella typhimurium live vaccine, with the ability of exporting metazoan antigens, opens up new possibilities of immunisation strategies against diverse parasites in future.