Untersuchung zur Culicidenfauna in Deutschland mit besonderer Berücksichtigung des Culex pipiens-Komplexes und der Vektorkompetenz für Nematoden und Protozoen

The present study was performed aiming to investigate a large number and a wide variety of mosquitoes from various regions of Germany for the presence of defined groups of parasites. Accordingly, during three successive years between 2010 and 2012, more than 140.000 mosquitoes, trapped at various sites in Germany, were characterized and analysed. The selected trapping sites, most of them located in southwest and northeast Germany, are known tor their abundant occurrence of mosquitoes, such as the foodplains of the river Isar. In addition, some of them represent important stepping stones for migrating birds, such as the Lake Chiemsee or the Lake Constance. The mosquitoes were characterized on the basis of morphological criteria. The results indicated, that the main autochthonous species generally present in Germany were trapped, in particular abundant species such as Aedes vexans and Culex pipiens s. l.. As morphological criteria do not allow the discrimination of members of the Culex pipiens complex or between females of Culex pipiens and Culex torrentium, a multiplex real-time PCR was developed for the rapid genetic differentiation of the various Culex species and biotypes. Using this method, hybrids of Cx. pipiens pipiens biotype pipiens and biotype molestus were detected in Germany for the first time. These hybrids are known as important bridge vectors for the transmission of West Nile virus from birds to humans. To fascilitate the detection of parasites in such a large number of mosquio samples within a reasoonable period of time, further multiplex real-time PCR assays were developed to allow analyses of respective microbial DNAs by high-throughput screening. The results indicated that a considerable number of German mosquitoes are carrying filariae. In this context, a new filarial species was detected, occuring primarily in southern Germany. The reservoir of this new filarial species is probably birds, as it was detected mainly in ornithophilic mosquitoes. Studies on Dirofilaria immitis, Dirofilaria repens as well as Onchocerca lupi, three zoonotic filarial species endemis in southern Europe and pathofenic for humans, revealed the detection of a focus of stable transmission of D. repens in the federal state of Brandenburg. This rilarial species was detected in several mosquito catches from 2011 and 2012, along the Oder Valley in the vincinity of the town of Eberswalde. As there were reports on cases of avian malaira in the zoo of Heidelberg in 2010 screening of mosquitoes was extended for the presence of Haemosporidia, such as Haemoproteus sp., Leukocytozoon sp. or Plasmodium sp.. A multiplex real-time PCR was developed able to detect nematodes as well as blood protozoans. Depending on the catchment area, more than 70 % of the mosquitoes were positive for at least one of the two groups of parasites

Molecular detection of Setaria tundra (Nematoda: Filarioidea) and an unidentified filarial species in mosquitoes in Germany

<p>Abstract</p> <p>Background</p> <p>Knowledge of the potential vector role of Culicidae mosquitoes in Germany is very scanty, and until recently it was generally assumed that they are not involved in the transmission of anthroponotic or zoonotic pathogens in this country. However, anticipated changes in the course of global warming and globalization may alter their status.</p> <p>Methods</p> <p>We conducted a molecular mass screening of mosquitoes for filarial parasites using mitochondrial 12S rRNA-based real-time PCR.</p> <p>Results</p> <p>No parasites causing disease in humans such as <it>Dirofilaria </it>spp. were detected in about 83,000 mosquitoes tested, which had been collected in 2009 and 2010 in 16 locations throughout Germany. However, minimum infection rates of up to 24 per 1000 mosquitoes were revealed, which could be attributed to mosquito infection with <it>Setaria tundra </it>and a yet unidentified second parasite. <it>Setaria tundra </it>was found to be widespread in southern Germany in various mosquito species, except <it>Culex </it>spp. In contrast, the unidentified filarial species was exclusively found in <it>Culex </it>spp. in northern Baden-Württemberg, and is likely to be a bird parasite.</p> <p>Conclusions</p> <p>Although dirofilariasis appears to be emerging and spreading in Europe, the absence of <it>Dirofilaria </it>spp. or other zoonotic filariae in our sample allows the conclusion that the risk of autochthonous infection in Germany is still very low. Potential vectors of <it>S. tundra </it>in Germany are <it>Ochlerotatus sticticus</it>, <it>Oc. cantans</it>, <it>Aedes vexans </it>and <it>Anopheles claviger</it>. Technically, the synergism between entomologists, virologists and parasitologists, combined with state-of-the-art methods allows a very efficient near-real-time monitoring of a wide spectrum of both human and veterinary pathogens, including new distribution records of parasite species and the incrimination of their potential vectors.</p

Aedes koreicus, a vector on the rise: Pan-European genetic patterns, mitochondrial and draft genome sequencing

25openYesBackground The mosquito Aedes koreicus (Edwards, 1917) is a recent invader on the European continent that was introduced to several new places since its first detection in 2008. Compared to other exotic Aedes mosquitoes with public health significance that invaded Europe during the last decades, this species’ biology, behavior, and dispersal patterns were poorly investigated to date. Methodology/Principal findings To understand the species’ population relationships and dispersal patterns within Europe, a fragment of the cytochrome oxidase I (COI or COX1) gene was sequenced from 130 mosquitoes, collected from five countries where the species has been introduced and/or established. Oxford Nanopore and Illumina sequencing techniques were combined to generate the first complete nuclear and mitochondrial genomic sequences of Ae. koreicus from the European region. The complete genome of Ae. koreicus is 879 Mb. COI haplotype analyses identified five major groups (altogether 31 different haplotypes) and revealed a large-scale dispersal pattern between European Ae. koreicus populations. Continuous admixture of populations from Belgium, Italy, and Hungary was highlighted, additionally, haplotype diversity and clustering indicate a separation of German sequences from other populations, pointing to an independent introduction of Ae. koreicus to Europe. Finally, a genetic expansion signal was identified, suggesting the species might be present in more locations than currently detected. Conclusions/Significance Our results highlight the importance of genetic research of invasive mosquitoes to understand general dispersal patterns, reveal main dispersal routes and form the baseline of future mitigation actions. The first complete genomic sequence also provides a significant leap in the general understanding of this species, opening the possibility for future genome-related studies, such as the detection of ‘Single Nucleotide Polymorphism’ markers. Considering its public health importance, it is crucial to further investigate the species’ population genetic dynamic, including a larger sampling and additional genomic markers.Kurucz, Kornélia; Zeghbib, Safia; Arnoldi, Daniele; Marini, Giovanni; Manica, Mattia; Michelutti, Alice; Montarsi, Fabrizio; Deblauwe, Isra; Van Bortel, Wim; Smitz, Nathalie; Pfitzner, Wolf Peter; Czajka, Christina; Jöst, Artur; Kalan, Katja; Šušnjar, Jana; Ivović, Vladimir; Kuczmog, Anett; Lanszki, Zsófia; Tóth, Gábor Endre; Somogyi, Balázs A; Herczeg, Róbert; Urbán, Péter; Bueno-Marí, Rubén; Soltész, Zoltán; Kemenesi, GáborKurucz, K.; Zeghbib, S.; Arnoldi, D.; Marini, G.; Manica, M.; Michelutti, A.; Montarsi, F.; Deblauwe, I.; Van Bortel, W.; Smitz, N.; Pfitzner, W.P.; Czajka, C.; Jöst, A.; Kalan, K.; Šušnjar, J.; Ivović, V.; Kuczmog, A.; Lanszki, Z.; Tóth, G.E.; Somogyi, B.A.; Herczeg, R.; Urbán, P.; Bueno-Marí, R.; Soltész, Z.; Kemenesi, G

Epizootic Emergence of Usutu Virus in Wild and Captive Birds in Germany

This study aimed to identify the causative agent of mass mortality in wild and captive birds in southwest Germany and to gather insights into the phylogenetic relationship and spatial distribution of the pathogen. Since June 2011, 223 dead birds were collected and tested for the presence of viral pathogens. Usutu virus (USUV) RNA was detected by real-time RT-PCR in 86 birds representing 6 species. The virus was isolated in cell culture from the heart of 18 Blackbirds (Turdus merula). USUV-specific antigen was demonstrated by immunohistochemistry in brain, heart, liver, and lung of infected Blackbirds. The complete polyprotein coding sequence was obtained by deep sequencing of liver and spleen samples of a dead Blackbird from Mannheim (BH65/11-02-03). Phylogenetic analysis of the German USUV strain BH65/11-02-03 revealed a close relationship with strain Vienna that caused mass mortality among birds in Austria in 2001. Wild birds from lowland river valleys in southwest Germany were mainly affected by USUV, but also birds kept in aviaries. Our data suggest that after the initial detection of USUV in German mosquitoes in 2010, the virus spread in 2011 and caused epizootics among wild and captive birds in southwest Germany. The data also indicate an increased risk of USUV infections in humans in Germany

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts

First record and morphological characterization of an established population of Aedes (Hulecoeteomyia) koreicus (Diptera: Culicidae) in Germany

Abstract Background The East Asian mosquito species Aedes koreicus was recorded out of its native range for the first time in Belgium in 2008. Since then, several other European populations or single individuals have been observed throughout Europe with reports from Italy, Switzerland, European Russia, Slovenia, Germany and Hungary. The Italian population seems to be the only one that is expanding rapidly, so the Swiss population very likely derives from it. Results In a surveillance program for invasive mosquito species, a single larva of Ae. koreicus was found in a cemetery vase in 2016 in the city of Wiesbaden, Germany. In the following year the finding was confirmed and an established population could be proven over an area of about 50 km2. The morphological identification of the first larva was confirmed by sequencing of a region within the nad4 sequence. A study of adult females showed that the morphological characteristics of this population are not identical to the populations from Belgium and Italy. The eggs and larvae were found together with Aedes j. japonicus in the same breeding sites and ovitraps, as well as with other indigenous mosquito species such as Culex pipiens/Culex torrentium, Aedes geniculatus and Anopheles plumbeus. Conclusions Since the newly discovered population in Germany shows different morphological characteristics to the populations in Belgium and Italy, it seems to originate from an independent introduction. It remains unknown how the introduction took place. A further spread similar to the one in northern Italy can be assumed for the future due to similar climatic conditions

BayFlux: A Bayesian method to quantify metabolic Fluxes and their uncertainty at the genome scale.

Metabolic fluxes, the number of metabolites traversing each biochemical reaction in a cell per unit time, are crucial for assessing and understanding cell function. 13C Metabolic Flux Analysis (13C MFA) is considered to be the gold standard for measuring metabolic fluxes. 13C MFA typically works by leveraging extracellular exchange fluxes as well as data from 13C labeling experiments to calculate the flux profile which best fit the data for a small, central carbon, metabolic model. However, the nonlinear nature of the 13C MFA fitting procedure means that several flux profiles fit the experimental data within the experimental error, and traditional optimization methods offer only a partial or skewed picture, especially in non-gaussian situations where multiple very distinct flux regions fit the data equally well. Here, we present a method for flux space sampling through Bayesian inference (BayFlux), that identifies the full distribution of fluxes compatible with experimental data for a comprehensive genome-scale model. This Bayesian approach allows us to accurately quantify uncertainty in calculated fluxes. We also find that, surprisingly, the genome-scale model of metabolism produces narrower flux distributions (reduced uncertainty) than the small core metabolic models traditionally used in 13C MFA. The different results for some reactions when using genome-scale models vs core metabolic models advise caution in assuming strong inferences from 13C MFA since the results may depend significantly on the completeness of the model used. Based on BayFlux, we developed and evaluated novel methods (P-13C MOMA and P-13C ROOM) to predict the biological results of a gene knockout, that improve on the traditional MOMA and ROOM methods by quantifying prediction uncertainty