Utilization of Polyspecific Antiserum for Specific Radioimmunoassays: Radioimmunoassays for Rat Fetuin and Bikunin Were Developed by Using Antiserum Against Total Rat Serum Proteins

Polyspecific antiserum against total rat serum proteins was used to develop specific and sensitive radioimmunoassays for fetuin and bikunin, two minor protein components of rat plasma. The radioimmunoassays proved to be highly useful to trace bikunin and fetuin in the course of developing isolation procedures, since neither specific functional assays nor monospecific antisera were available. The two examples demonstrate that, in general, it will be possible to develop a specific and sensitive radioimmunoassay with antiserum raised against a crude antigen preparation, such as a body fluid or a tissue extract, provided that a minute amount of pure antigen is available for preparing the radioiodinated antigen

Fifth European Dirofilaria and Angiostrongylus Days (FiEDAD) 2016

Peer reviewe

Xenomonitoring of Mosquitoes (Diptera: Culicidae) for the Presence of Filarioid Helminths in Eastern Austria

Information on mosquito-borne filarioid helminths in Austria is scarce, but recent discoveries of Dirofilaria repens indicate autochthonous distribution of this parasite in Eastern Austria. In the current xenomonitoring study, more than 48,000 mosquitoes were collected in Eastern Austria between 2013 and 2015, using different sampling techniques and storage conditions, and were analysed in pools with molecular tools for the presence of filarioid helminth DNA. Overall, DNA of D. repens, Setaria tundra, and two unknown filarioid helminths were documented in twenty mosquito pools within the mitochondrial cox1 gene (barcode region). These results indicate that S. tundra, with roe deer as definite hosts, is common in Eastern Austria, with most occurrences in floodplain mosquitoes (e.g., Aedes vexans). Moreover, DNA of D. repens was found in an Anopheles plumbeus mosquito close to the Slovakian border, indicating that D. repens is endemic in low prevalence in Eastern Austria. This study shows that xenomonitoring is an adequate tool to analyse the presence of filarioid helminths, but results are influenced by mosquito sampling techniques, storage conditions, and molecular protocols

Avian Plasmodium in Eastern Austrian mosquitoes

Abstract Background Insect vectors, namely mosquitoes (Diptera: Culicidae), are compulsory for malaria parasites (Plasmodium spp.) to complete their life cycle. Despite this, little is known about vector competence of different mosquito species for the transmission of avian malaria parasites. Methods In this study, nested PCR was used to determine Plasmodium spp. occurrence in pools of whole individuals, as well as the diversity of mitochondrial cytochrome b gene sequences in wild-caught mosquitoes sampled across Eastern Austria in 2013–2015. Results A total of 45,749 mosquitoes in 2628 pools were collected, of which 169 pools (6.43%) comprising 9 mosquito species were positive for avian Plasmodium, with the majority of positives in mosquitoes of Culex pipiens s.l./Culex torrentium. Six different avian Plasmodium lineages were found, the most common were Plasmodium vaughani SYAT05, Plasmodium sp. Linn1 and Plasmodium relictum SGS1. In 2014, mosquitoes of the Culex pipiens complex were genetically identified and Culex pipiens f. pipiens presented with the highest number of avian Plasmodium positives (n = 37; 16.74%). Despite this, the minimum infection rate (MIR) was highest in Culex torrentium (5.36%) and Culex pipiens f. pipiens/f. molestus hybrids (5.26%). During 2014 and 2015, seasonal and annual changes in Plasmodium lineage distribution were also observed. In both years P. vaughani SYAT05 dominated at the beginning of the sampling period to be replaced later in the year by P. relictum SGS1 (2014) and Plasmodium sp. Linn1 (2015). Conclusions This is the first large-scale study of avian Plasmodium parasites in Austrian mosquitoes. These results are of special interest, because molecular identification of the taxa of the Cx. pipiens complex and Cx. torrentium enabled the determination of Plasmodium prevalence in the different mosquito taxa and hybrids of this complex. Since pools of whole insects were used, it is not possible to assert any vector competence in any of the examined mosquitoes, but the results are nonetheless valuable in providing an overview of avian Plasmodium species and lineages present in Austria

Number of mosquito pools positive for trypanosomatid DNA (trypanosome species, <i>T</i>. <i>avium</i>, <i>T</i>. <i>culicavium</i>, and <i>T</i>. <i>theileri</i>, are shown separately while <i>Crithidia</i> spp. infections are combined) according to the sampled months in 2014 and 2015 (Vienna and Eastern Austria).

<p>Number of mosquito pools positive for trypanosomatid DNA (trypanosome species, <i>T</i>. <i>avium</i>, <i>T</i>. <i>culicavium</i>, and <i>T</i>. <i>theileri</i>, are shown separately while <i>Crithidia</i> spp. infections are combined) according to the sampled months in 2014 and 2015 (Vienna and Eastern Austria).</p

An SSU rDNA-based Bayesian phylogenetic tree representing the most likely new <i>Herpetomonas</i> species obtained from a mosquito collected in Eastern Austria.

<p>Bootstrap values from Bayesian posterior probabilities (5 million generations) and bootstrap percentages for maximum-likelihood (PhyML) analysis (1,000 replicates) are shown at the nodes; dashes indicate <50% bootstrap support or different topology; asterisks mark branches with maximal statistical support; double-crossed branches are 50% of the original length. The tree was rooted with five sequences of <i>Phytomonas</i> spp., the closest relative of the genus <i>Herpetomonas</i>. Parasite names, names of strains or GenBank accession numbers are given; the branch lengths are drawn proportionally to the amount of changes (scale bar).</p

Trypanosomatid parasites in Austrian mosquitoes

<div><p>Trypanosomatid flagellates have not been studied in Austria in any detail. In this study, specific nested PCR, targeted on the ribosomal small subunit, was used to determine the occurrence and diversity of trypanosomatids in wild-caught mosquitoes sampled across Eastern Austria in the years 2014−2015. We collected a total of 29,975 mosquitoes of 19 species divided in 1680 pools. Of these, 298 (17.7%), representing 12 different mosquito species, were positive for trypanosomatid DNA. In total, seven trypanosomatid spp. were identified (three <i>Trypanosoma</i>, three <i>Crithidia</i> and one <i>Herpetomonas</i> species), with the highest parasite species diversity found in the mosquito host <i>Coquillettidia richiardii</i>. The most frequent parasite species belonged to the mammalian <i>Trypanosoma theileri</i>/<i>cervi</i> species complex (found in 105 pools; 6.3%). The avian species <i>T</i>. <i>culicavium</i> (found in 69 pools; 4.1%) was only detected in mosquitoes of the genus <i>Culex</i>, which corresponds to their preference for avian hosts. Monoxenous trypanosomatids of the genus <i>Crithidia</i> and <i>Herpetomonas</i> were found in 20 (1.3%) mosquito pools. One third (n = 98) of the trypanosomatid positive mosquito pools carried more than one parasite species. This is the first large scale study of trypanosomatid parasites in Austrian mosquitoes and our results are valuable in providing an overview of the diversity of these parasites in Austria.</p></div

Number of mosquito pools positive for trypanosomatid DNA (trypanosome species, <i>T</i>. <i>avium</i>, <i>T</i>. <i>culicavium</i>, and <i>T</i>. <i>theileri</i>, are shown separately while <i>Crithidia</i> spp. infections are combined) according to the sampled months in 2014 and 2015 (Vienna and Eastern Austria).

<p>Number of mosquito pools positive for trypanosomatid DNA (trypanosome species, <i>T</i>. <i>avium</i>, <i>T</i>. <i>culicavium</i>, and <i>T</i>. <i>theileri</i>, are shown separately while <i>Crithidia</i> spp. infections are combined) according to the sampled months in 2014 and 2015 (Vienna and Eastern Austria).</p