Institut für Biologie

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Decoding the geography of natural TBEV microfoci in German: a geostatistical approach based on land-use patterns and climatological conditions
(2022) Borde, Johannes P.; Glaser, Rüdiger; Braun, Klaus; Riach, Nils; Hologa, Rafael; Kaier, Klaus; Chitimia-Dobler, Lidia; Dobler, Gerhard
Background: Tickborne-encephalitis (TBE) is a potentially life-threating neurological disease that is mainly transmitted by ticks. The goal of the present study is to analyze the potential uniform environmental patterns of the identified TBEV microfoci in Germany. The results are used to calculate probabilities for the present distribution of TBEV microfoci in Germany based on a geostatistical model. Methods: We aim to consider the specification of environmental characteristics of locations of TBEV microfoci detected in Germany using open access epidemiological, geographical and climatological data sources. We use a two-step geostatistical approach, where in a first step, the characteristics of a broad set of environmental variables between the 56 TBEV microfoci and a control or comparator set of 3575 sampling points covering Germany are compared using Fisher’s Exact Test. In the second step, we select the most important variables, which are then used in a MaxEnt distribution model to calculate a high resolution (400 × 400 m) probability map for the presence of TBEV covering the entire area of Germany. Results: The findings from the MaxEnt prediction model indicate that multi annual actual evapotranspiration (27.0%) and multi annual hot days (22.5%) have the highest contribution to our model. These two variables are followed by four additional variables with a lower, but still important, explanatory influence: Land cover classes (19.6%), multi annual minimum air temperature (14.9%), multi annual sunshine duration (9.0%), and distance to coniferous and mixed forest border (7.0%). Conclusions: Our findings are based on defined TBEV microfoci with known histories of infection and the repeated confirmation of the virus in the last years, resulting in an in-depth high-resolution model/map of TBEV microfoci in Germany. Multi annual actual evapotranspiration (27%) and multi annual hot days (22.5%) have the most explanatory power in our model. The results may be used to tailor specific regional preventive measures and investigations.
Echinococcus multilocularis and other taeniid metacestodes of muskrats in Luxembourg: prevalence, risk factors, parasite reproduction, and genetic diversity
(2022) Martini, Matilde; Dumendiak, Sonja; Gagliardo, Anna; Ragazzini, Francesco; La Rosa, Letizia; Giunchi, Dimitri; Thielen, Frank; Romig, Thomas; Massolo, Alessandro; Wassermann, Marion
Muskrats (Ondatra zibethicus) are competent intermediate hosts for Echinococcus multilocularis, are frequently infected with this zoonotic cestode, and have even been proposed as a target species to monitor endemicity levels of this parasite. However, their contribution to maintaining the parasitic lifecycle is still unclear. To obtain data on infection frequency and reproductive potential, 280 muskrats from the Grand Duchy of Luxembourg were examined for cestode larvae in the years 2013–2017. Based on morphological and molecular identification, Echinococcus multilocularis was found at a prevalence of 14.6%. Other metacestodes were Hydatigera kamiyai, with a prevalence of 45.7%, Taenia martis with 8.9%, Taenia polyacantha with 5.0%, and Versteria mustelae, which was found in 0.7% of all muskrats. More than 80% of E. multilocularis-infected muskrats contained fertile metacestodes with a mean number of >300,000 (and up to 1,609,816) protoscoleces, which is by far the highest reproductive potential known from any intermediate host species in Europe. Temporal analysis of E. multilocularis prevalence within the study period (and in comparison with earlier data) strongly indicates a robust increase in the studied area. Host age seemed to be an important risk factor for infection, as well as co-infections with Hydatigera kamiyai. A preference for the right medial lobe of the liver as the location of E. multilocularis metacestode was observed. Intraspecific genetic variation among 89 discrete E. multilocularis metacestodes was non-existent based on 300–1590 bp sections of cox1. This is a stark contrast to H. kamiyai, of which nine haplotypes were found on a short 318 bp section of cox1, resulting in genetic diversity in the small country of Luxembourg at a similar level than previously reported from large stretches of Europe and northern Asia.
Echinococcus multilocularis and other zoonotic helminths in red foxes (Vulpes vulpes) from a southern German hotspot for human alveolar echinococcosis
(2023) Schneider, Carina; Kratzer, Wolfgang; Binzberger, Andreas; Schlingeloff, Patrycja; Baumann, Sven; Romig, Thomas; Schmidberger, Julian
Background: We describe the spatial distribution of Echinococcus multilocularis in its main definitive host, the red fox, and the distribution of human cases of alveolar echinococcosis (AE) within a highly endemic focus in southern Germany (13.7–19.9/100,000 in 1992–2018). Human cases were unequally distributed within the endemicity focus. The purpose of the study was to test whether this is reflected in the small-scale distribution of E. multilocularis in foxes. Methods: Three areas with contrasting numbers of human cases were selected within the counties of Ravensburg and Alb-Donau, Baden-Württemberg, Germany. From 2018 to 2020, a total of 240 fox carcasses were obtained from traditional hunters in these areas. Carcasses were necropsied and examined for the presence of intestinal helminths. The statistical analysis was performed with SAS version 9.4, and the geo-mapping with QGIS version 3.16.0 Hannover. Results: The prevalence of E. multilocularis in foxes was 44/106 (41.5%) in area I (commune Leutkirch and environs), 30/59 (50.8%) in area II (commune Isny and environs), and 31/75 (41.3%) in area III (commune Ehingen and environs). From 1992 to 2018, a total of nine human cases of alveolar echinococcosis were recorded in area I, five cases were recorded in study area III, and no cases were recorded in area II. No statistically significant differences between the areas were observed ( P > 0.05) for intestinal infections with E. multilocularis , and no apparent spatial correlation with the small-scale distribution of human cases was found. Concerning other zoonotic helminths, Toxocara spp. were equally common, with prevalence of 38.7%, 47.4% and 48.0%, respectively, while the frequency of Alaria alata varied among the study areas (0.0–9.4%), probably reflecting the specific habitat requirements for the establishment of its complex life cycle. Conclusions: Echinococcus multilocularis is highly prevalent in foxes in all the studied areas. The varying number of human AE cases within these areas should therefore be caused by factors other than the intensity of parasite transmission in foxes.
Exploring ND-011992, a quinazoline-type inhibitor targeting quinone reductases and quinol oxidases
(2023) Kägi, Jan; Sloan, Willough; Schimpf, Johannes; Nasiri, Hamid R.; Lashley, Dana; Friedrich, Thorsten
Bacterial energy metabolism has become a promising target for next-generation tuberculosis chemotherapy. One strategy to hamper ATP production is to inhibit the respiratory oxidases. The respiratory chain of Mycobacterium tuberculosis comprises a cytochrome bcc:aa3 and a cytochrome bd ubiquinol oxidase that require a combined approach to block their activity. A quinazoline-type compound called ND-011992 has previously been reported to ineffectively inhibit bd oxidases, but to act bactericidal in combination with inhibitors of cytochrome bcc:aa3 oxidase. Due to the structural similarity of ND-011992 to quinazoline-type inhibitors of respiratory complex I, we suspected that this compound is also capable of blocking other respiratory chain complexes. Here, we synthesized ND-011992 and a bromine derivative to study their effect on the respiratory chain complexes of Escherichia coli. And indeed, ND-011992 was found to inhibit respiratory complex I and bo3 oxidase in addition to bd-I and bd-II oxidases. The IC50 values are all in the low micromolar range, with inhibition of complex I providing the lowest value with an IC50 of 0.12 µM. Thus, ND-011992 acts on both, quinone reductases and quinol oxidases and could be very well suited to regulate the activity of the entire respiratory chain.
Functionality of the Na+-translocating NADH:quinone oxidoreductase and quinol:fumarate reductase from Prevotella bryantii inferred from homology modeling
(2024) Hau, Jann-Louis; Schleicher, Lena; Herdan, Sebastian; Simon, Jörg; Seifert, Jana; Fritz, Günter; Steuber, Julia
Members of the family Prevotellaceae are Gram-negative, obligate anaerobic bacteria found in animal and human microbiota. In Prevotella bryantii , the Na + -translocating NADH:quinone oxidoreductase (NQR) and quinol:fumarate reductase (QFR) interact using menaquinone as electron carrier, catalyzing NADH:fumarate oxidoreduction. P. bryantii NQR establishes a sodium-motive force, whereas P. bryantii QFR does not contribute to membrane energization. To elucidate the possible mode of function, we present 3D structural models of NQR and QFR from P. bryantii to predict cofactor-binding sites, electron transfer routes and interaction with substrates. Molecular docking reveals the proposed mode of menaquinone binding to the quinone site of subunit NqrB of P. bryantii NQR. A comparison of the 3D model of P. bryantii QFR with experimentally determined structures suggests alternative pathways for transmembrane proton transport in this type of QFR . Our findings are relevant for NADH-dependent succinate formation in anaerobic bacteria which operate both NQR and QFR.
Hitze als Schlüssel zur Aufklärung der Lysogenie beim SPß-Phagen
(2025) Kohm, Katharina
Identification of novel genes including NAV2 associated with isolated tall stature
(2023) Weiss, Birgit; Ott, Tim; Vick, Philipp; Lui, Julian C.; Roeth, Ralph; Vogel, Sebastian; Waldmüller, Stephan; Hoffmann, Sandra; Baron, Jeffrey; Wit, Jan M.; Rappold, Gudrun A.
Very tall people attract much attention and represent a clinically and genetically heterogenous group of individuals. Identifying the genetic etiology can provide important insights into the molecular mechanisms regulating linear growth. We studied a three-generation pedigree with five isolated (non-syndromic) tall members and one individual with normal stature by whole exome sequencing; the tallest man had a height of 211 cm. Six heterozygous gene variants predicted as damaging were shared among the four genetically related tall individuals and not present in a family member with normal height. To gain insight into the putative role of these candidate genes in bone growth, we assessed the transcriptome of murine growth plate by microarray and RNA Seq. Two (Ift140, Nav2) of the six genes were well-expressed in the growth plate. Nav2 (p-value 1.91E-62) as well as Ift140 (p-value of 2.98E-06) showed significant downregulation of gene expression between the proliferative and hypertrophic zone, suggesting that these genes may be involved in the regulation of chondrocyte proliferation and/or hypertrophic differentiation. IFT140, NAV2 and SCAF11 have also significantly associated with height in GWAS studies. Pathway and network analysis indicated functional connections between IFT140, NAV2 and SCAF11 and previously associated (tall) stature genes. Knockout of the all-trans retinoic acid responsive gene, neuron navigator 2 NAV2, in Xenopus supports its functional role as a growth promotor. Collectively, our data expand the spectrum of genes with a putative role in tall stature phenotypes and, among other genes, highlight NAV2 as an interesting gene to this phenotype.
Morphological and molecular identification of sarcocystis arctica in captive cheetahs (Acinonyx jubatus) in China helps clarify phylogenetic relationships with Sarcocystis caninum and Sarcocystis felis
(2025) Liao, Zhe; Zhu, Niuping; Yang, Yurong; Deng, Shuangsheng; Jäkel, Thomas; Hu, Junjie; Salvarani, Felipe M.; Domingues, Sheyla Farhayldes Souza; Da Silveira, Júlia Angélica Gonçalves
To date, only one case is known where protozoan parasites of the genus Sarcocystis were found to infect cheetahs ( Acinonyx jubatus ); the cysts in the musculature were morphologically identified as S. felis . Here, we characterized sarcocysts by morphological and molecular methods that were observed in cheetahs who died in zoos in China. Only one type of sarcocyst was present in two of six cheetahs. By light microscopy, the sarcocyst wall was striated, 1.4–2.1 μm thick. Ultrastructurally, the wall had irregular-shaped, small villar protrusions, resembling wall type 9c, similar to those of S. arctica , S. caninum , and S. felis . The samples shared their highest molecular identity values with those of S. arctica and S. caninum : 99.9–100% and 99.8–100% (18S rRNA), 99.5% and 99.3–99.5% (28S rRNA), 95.9–97.5% and 96.3–97.3% (ITS-1), and 99.6% and 99.2–99.7% ( cox 1), respectively. Compared with ITS-1 of S. felis , identities ranged between 87.5% and 88.9%. Phylogenetic reconstruction revealed that the newly sequenced Sarcocystis clustered with S. arctica and S. caninum , whereas S. felis (ITS-1) and S. canis (ITS-1, 18S rRNA, cox 1) were sister species. Thus, we addressed the Sarcocystis species from the cheetahs as S. arctica , which is the first record of a Sarcocystis species believed to be specific for canids as intermediate hosts to infect a feline host.
Nucleo-cytoplasmic shuttling of murine RBPJ by Hairless protein matches that of Su(H) protein in the model system Drosophila melanogaster
(2021) Wolf, Dorina B.; Maier, Dieter; Nagel, Anja C.
CSL transcription factors are central to signal transduction in the highly conserved Notch signaling pathway. CSL acts as a molecular switch: depending on the cofactors recruited, CSL induces either activation or repression of Notch target genes. Unexpectedly, CSL depends on its cofactors for nuclear entry, despite its role as gene regulator. In Drosophila, the CSL homologue Suppressor of Hairless (Su(H)), recruits Hairless (H) for repressor complex assembly, and eventually for nuclear import. We recently found that Su(H) is subjected to a dynamic nucleo-cytoplasmic shuttling, thereby strictly following H subcellular distribution. Hence, regulation of nuclear availability of Su(H) by H may represent a new layer of control of Notch signaling activity. Here we extended this work on the murine CSL homologue RBPJ. Using a ‘murinized’ fly model bearing RBPJwt in place of Su(H) at the endogenous locus we demonstrate that RBPJ protein likewise follows H subcellular distribution. For example, overexpression of a H*NLS3 protein variant defective of nuclear import resulted in a cytosolic localization of RBPJ protein, whereas the overexpression of a H*NES protein variant defective in the nuclear export signal caused the accumulation of RBPJ protein in the nucleus. Evidently, RBPJ is exported from the nucleus as well. Overall these data demonstrate that in our fly model, RBPJ is subjected to H-mediated nucleo-cytoplasmic shuttling as is Su(H). These data raise the possibility that nuclear availability of mammalian CSL proteins is likewise restricted by cofactors, and may hence present a more general mode of regulating Notch signaling activity.
Red foxes harbor two genetically distinct, spatially separated Echinococcus multilocularis clusters in Brandenburg, Germany
(2021) Herzig, Mandy; Maksimov, Pavlo; Staubach, Christoph; Romig, Thomas; Knapp, Jenny; Gottstein, Bruno; Conraths, Franz J.
Background: Alveolar echinococcosis (AE) is a clinically serious zoonosis caused by the fox tapeworm Echinococcus multilocularis. We studied the diversity and the distribution of genotypes of E. multilocularis isolated from foxes in Brandenburg, Germany, and in comparison to a hunting ground in North Rhine-Westphalia. Methods: Echinococcus multilocularis specimens from 101 foxes, 91 derived from Brandenburg and 10 derived from North Rhine-Westphalia, were examined. To detect potential mixed infections with different genotypes of E. multilocularis, five worms per fox were analyzed. For genotyping, three mitochondrial markers, namely cytochrome c oxidase subunit 1 (Cox1), NADH dehydrogenase subunit 1 (Nad1), and ATP synthase subunit 6 (ATP6), and the nuclear microsatellite marker EmsB were used. To identify nucleotide polymorphisms, the mitochondrial markers were sequenced and the data were compared, including with published sequences from other regions. EmsB fragment length profiles were determined and confirmed by Kohonen network analysis and grouping of Sammon’s nonlinear mapping with k-means clustering. The spatial distribution of genotypes was analyzed by SaTScan for the EmsB profiles found in Brandenburg. Results: With both the mitochondrial makers and the EmsB microsatellite fragment length profile analyses, mixed infections with different E. multilocularis genotypes were detected in foxes from Brandenburg and North Rhine-Westphalia. Genotyping using the mitochondrial markers showed that the examined parasite specimens belong to the European haplotype of E. multilocularis, but a detailed spatial analysis was not possible due to the limited heterogeneity of these markers in the parasite population. Four (D, E, G, and H) out of the five EmsB profiles described in Europe so far were detected in the samples from Brandenburg and North Rhine-Westphalia. The EmsB profile G was the most common. A spatial cluster of the E. multilocularis genotype with the EmsB profile G was found in northeastern Brandenburg, and a cluster of profile D was found in southern parts of this state. Conclusions: Genotyping of E. multilocularis showed that individual foxes may harbor different genotypes of the parasite. EmsB profiles allowed the identification of spatial clusters, which may help in understanding the distribution and spread of the infection in wildlife, and in relatively small endemic areas.
TRPL-Recycling in Photorezeptoren von Drosophila : Identifikation beteiligter Gene mittels tsCRISPR Mutagenesescreen
(2025) Zeger, Matthias; Huber, Armin
Die präzise Regulation des intrazellulären Transports von Membranproteinen ist essenziell für die Funktion neuronaler Zellen. Störungen in diesem Prozess, insbesondere im Proteinrecycling über den Retromerkomplex, stehen im Zusammenhang mit neurodegenerativen Erkrankungen wie Alzheimer oder Parkinson. In dieser Arbeit wurde die lichtabhängige Translokation des TRPL (Transient Receptor Potential Like)-Ionenkanals im Drosophila-Komplexauge als Modell genutzt, um molekulare Komponenten des Membranproteintransports zu identifizieren. Bei Dunkelheit ist TRPL in den Rhabdomeren lokalisiert und transloziert nach Belichtung, aus dem Rhabdomer ins Zellinnere. Bei anhaltender Belichtung wandert der Großteil des TRPLs innerhalb von 12 Stunden zu einem Speicherkompartiment, das bislang als das Endoplasmatische Retikulum identifiziert wurde. Bei ausbleibendem Lichtreiz bzw. bei einer erneuten Dunkeladaptation der Photorezeptorzellen von Drosophila wandert TRPL innerhalb von 90–120 min wieder zurück in die rhabdomerische Membran. Ein augenspezifisches CRISPR-Knockout-System für Drosophila wurde etabliert, um gezielt Rab-GTPasen – zentrale Regulatoren des intrazellulären Vesikeltransports – im Zusammenhang mit dem TRPL-Recycling zu untersuchen. Durch die Analyse der TRPL-Lokalisation unter verschiedenen Lichtbedingungen konnten mehrere Rab- Proteine mit potenzieller Funktion im Recyclingprozess identifiziert und immunhistochemisch charakterisiert werden. Besonders starke Effekte auf das TRPLRecycling zeigten die Rab3-, Rab4-, Rab7-, Rab32-, Rab40- und RabX2-Knock-out Mutanten. Rab3, das unter anderem an späten Endosomen lokalisiert ist, wurde erstmals in vivo als Regulator für das Proteinrecycling identifiziert. RabX2 hingegen ist am trans-Golgi lokalisiert; sein Knock-out führte zu einer Akkumulation von TRPL im trans-Golgi während des retrograden Transports zurück in das Rhabdomer. Die Ergebnisse legen nahe, dass TRPL auch über einen zweiten ER-unabhängigen Weg zurück in das Rhabdomer recycelt wird, da ein Knock-Out von Rab3, Rab7, Rab32 oder Rab40 die ER/TRPL-Kolokalisation stark verringert aber der Rücktransport weniger stark beeinflusst wird. Außerdem wurde eine weitere TRPL-Recyclingroute über den trans-Golgi identifiziert welche abhängig von RabX2 ist. VII Im zweiten Teil der Arbeit wurde mittels TurboID-basiertem Biotin-Proximity Labeling nach TRPL-assoziierten Interaktionspartnern gesucht. Eine Auswahl der dabei identifizierten Kandidaten wurde funktionell analysiert, um ihren Einfluss auf die TRPLTranslokation zu bewerten. In dem Screen nach TRPL Interaktionspartnern wurden Signaltransduktionsproteine wie INAD, TRP und NorpA durch TRPL::TbID identifiziert. Darüber hinaus wurden Proteine wie Rab32 und Vps35 detektiert, die mit dem TRPLRecycling in Verbindung stehen, sowie weitere Proteine wie Sec61 und HIP-R, die potenziell mit ER-Stress assoziiert sind. Zu den weiteren identifizierten TRPLInteraktionspartnern gehörten Vap33, PIP82, Orp8, Sec63 und Sec22, die detailliert immunhistochemisch untersucht wurden. Dabei zeigte der Knock-out von Sec22 und Sec63 eine reduzierte Kolokalisation von TRPL mit Calnexin (Cnx), jedoch blieb der Rücktransport weitgehend wildtypisch – ein Phänotyp, der dem von Rab3-, Rab7-, Rab32- und Rab40-Knock-outs ähnelt. Die Orp8 Knock-out-Mutante zeigte nach erneuter Dunkeladaptation weiterhin eine TRPL/Cnx-Kolokalisation, was auf einen Defekt beim ER-Exit hinweist. Die PIP82-Nullmutante verursachte einen Internalisierungsdefekt sowie eine deutliche Abnahme des TRPL-Signals nach Lichtadaptation. Der Vap33 Knock-out führte bereits im Dunkelzustand zu anterograden Transportdefekt, da TRPL teilweise im Zytosol und ER lokalisiert war. Insgesamt konnte in der vorliegenden Arbeit gezeigt werden, dass neben dem ERabhängigen Recyclingweg von TRPL auch ein ER-unabhängiger Transportmechanismus existiert. Beide Routen sind auf die Funktion verschiedener Rab-GTPasen angewiesen. Besonders hervorzuheben ist der Nachweis einer Beteiligung von Rab3 am Proteinrecycling in Drosophila-Photorezeptorzellen – ein bisher nicht beschriebenes Phänomen, das über die bekannte Rolle von Rab3 in der synaptischen Transmission hinausgeht. Darüber hinaus wurde erstmals ein TRPLRecyclingweg über den trans-Golgi-Apparat identifiziert, der auf die Aktivität von RabX2 angewiesen ist. Ergänzend dazu konnten mehrere potenzielle Interaktionspartner von TRPL durch Biotin-Proximity Labelling identifiziert werden, darunter Vap33, PIP82, Orp8, Sec63 und Sec22. Funktionelle Analysen deuten darauf hin, dass diese Proteine einen Einfluss auf die zelluläre Lokalisation und Translokation von TRPL haben und somit eine Rolle in seiner dynamischen Regulation spielen könnten.
Unwanted souvenirs - import routes and pathogen detection of the non-endemic tick Rhipicephalus sanguineus s.l. in Germany
(2025) Fachet-Lehmann, Katrin; Lindau, Alexander; Mackenstedt, Ute
Ticks of the Genus Rhipicephalus occur worldwide. Especially members of Rh. sanguineus s.l. are primarily associated with dogs. As unwanted souvenirs, they are introduced by dogs into non-endemic areas such as Germany, where they can establish and reproduce indoors . A citizen-science study was conducted between 2019 and 2024, asking interested citizens to report tick infestations and send in travel related tick findings which were associated with dogs and were focused on Rhipicephalus species. Tick species were identified using the 16S rRNA gene and tested for pathogens associated with the genus Rhipicephalus . In addition, each tick introduction was considered as a case and categorized and analyzed individually. During the study period, 44 cases were reported. In 17 cases tick specimens were identified as Rh. sanguineus s.s., Rh. rutilus , Rh. linnaei, Rh. turanicus , and Rh. haemaphysaloides which were imported from other European countries and Sri Lanka. Neither Hepatozoon canis , Ehrlichia canis nor Babesia vogeli were detected in any of the 780 received specimens. In contrast, Rickettsia spp. was detected in 50 specimens from six independent cases, with Rickettsia massiliae being identified in 44 specimens. The import of dogs from abroad as well as travel with dogs lacking appropriate tick prophylaxis are responsible for more than 80% of cases. This Project highlights the risk of introductions of non-endemic tick species. Also, professionals such as veterinarians, animal welfare organizations and pest controllers need to be made aware of the possible introduction of Rhipicephalus spp. to ensure early recognition and rapid elimination of the ticks.

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