Discovery and genetic characterization of a novel orthonairovirus in Ixodes ricinus ticks from Danube Delta.

Different arthropod species are vectors of a wide array of arboviruses (arthropod-borne viruses) and have likely been central to viral evolution. To better understand the extent of arthropod-borne pathogens, as well as their origin and evolutionary history, it is crucial to uncover the full range of microbial agents, including viruses associated with arthropods. In this study, a collection of ticks obtained in 2016 directly from mammal and bird hosts from several rural and natural sites of Danube Delta was subjected to transcriptome sequencing and amplification assays. Vector surveillance revealed the presence of a novel orthonairovirus species, designated Sulina virus, in Ixodes ricinus ticks. Phylogenetic clustering of each viral protein consistently placed the new virus in the Orthonairovirus genus as a new genogroup closely related to Tamdy orthonairovirus, a genogroup comprising both pathogenic and tick-associated orthonairoviruses. The serological testing of engorged ticks and blood of infected hosts, along with the inoculation of vertebrate cells and mice found no specific antibodies or viral replication, suggesting that Sulina virus is an orthonairovirus associated with the virome of Ixodes ricinus. Finally, the characterization of a novel orthonairovirus identified using high throughput sequencing will advance our knowledge of interactions between viruses and tick vectors, expanding our perspective on fundamental questions regarding orthonairovirus evolution, diversity, ecology and potential of emergence as pathogens.

New Phenotypes of Potato Co-induced by Mismatch Repair Deficiency and Somatic Hybridization.

As plants are sessile they need a very efficient system for repairing damage done by external or internal mutagens to their DNA. Mismatch repair (MMR) is one of the systems that maintain genome integrity and prevent homeologous recombination. In all eukaryotes mismatches are recognized by evolutionary conserved MSH proteins often acting as heterodimers, the constant component of which is MSH2. Changes affecting the function of MSH2 gene may induce a 'mutator' phenotype and microsatellite instability (MSI), as is demonstrated in MSH2 knock-out and silenced lines of Arabidopsis thaliana. The goal of this study was to screen for 'mutator' phenotypes in somatic hybrids between potato cvs. 'Delikat' and 'Désirée' and MMR deficient Solanum chacoense transformed using antisense (AS) or dominant negative mutant (DN) AtMSH2 genes. The results demonstrate that first generation fusion hybrids have a range of morphological abnormalities caused by uniparental MMR deficiency; these mutant phenotypes include: dwarf or gigantic plants; bushiness; curled, small, large or abnormal leaves; a deterioration in chloroplast structure; small deep-purple tubers and early dehiscent flowers. Forty percent of the viable somatic hybrids planted in a greenhouse, (10 out of 25 genotypes) had mutant phenotypes accompanied by MSI. The majority of the hybrids with 'mutator' phenotypes cultured on media containing kanamycin developed roots so sustaining the presence of selectable marker gene nptII, from the initial constructs. Here for the first time, MMR deficiency combined with somatic hybridization, are used to induce new phenotypes in plants, which supports the role of MMR deficiency in increasing introgressions between two related species.

Mismatch repair deficiency increases the transfer of antibiosis and antixenosis properties against Colorado potato beetle in somatic hybrids of Solanum tuberosum + S. chacoense.

BACKGROUND: Colorado potato beetle (CPB) has become the biggest enemy of cultivated potato worldwide. One of the most effective sources of resistance to CPB is Solanum chacoense, an accession with a high leptine glycoalkaloid content. The aim of our study was to assay the repellence and toxicity of S. chacoense, its somatic hybrids (SHs) and their backcross progenies (BC1 ) with potato for CPB adults and larvae. Transgenic S. chacoense, deficient in DNA mismatch repair (MMR), was also used to produce SHs, in order to increase homeologous recombination and hence introgression of wild-species DNA into the potato gene pool. RESULTS: Wild-type SH was highly resistant to CPB. Resistance to CPB of BC1 progenies showed a 1:3 inheritance pattern. MMR-deficient SHs performed better in the resistance analysis. Most MMR-deficient SHs had a similar toxicity as S. chacoense and an intensely repellent effect on CPB adults. Resistance of SHs and BC1 clones may be attributed to leptine biosynthesis, which was confirmed using a RAPD marker. CONCLUSION: This is the first report of SHs and their progenies exhibiting both antibiosis and antixenosis against CPB. Resistant SHs are an important step forward in combating this voracious pest of potato. © 2016 Society of Chemical Industry.

Identification of Hepatozoon erhardovae Krampitz, 1964 from bank voles (Myodes glareolus) and fleas in Southern Hungary.

In order to investigate the prevalence and life cycle of apicomplexan parasites, small mammals were live-trapped with modified Sherman traps in Southern Hungary between 2010 and 2012. Altogether, 528 rodents (Apodemus flavicollis Melchior, 1834, Apodemus agrarius Pallas, 1771, Myodes glareolus Schreber, 1780, Microtus agrestis Linnaeus, 1761, Mus musculus Linnaeus, 1758 and Micromys minutus Pallas, 1771) were collected and four shrews (Sorex spp.) were by-catched. Captured animals belonging to non-protected species were euthanized, and spleen samples were preserved for histological and molecular analyses. During the examination of spleen smears, Hepatozoon parasites were observed in eight out of 48 bank voles (M. glareolus). DNA was isolated from altogether 221 spleen samples, and 18S rDNA was amplified using two different PCR protocols. The eight bank vole samples were positive with PCR, but none of the other M. glareolus spleen samples or any of the tissue samples from other species were found to be infected. Sequenced amplicons were very similar to Hepatozoon spp. detected in M. glareolus in Spain and Poland. Ectoparasites were collected from the small mammal carcasses and from the vegetation. Hepatozoon DNA was not found in the 181 ticks removed from the small mammals or in the 162 ticks collected with flagging, but was detected in all three flea species (4/43 Megabothris turbidus Rothschild, 1909, 3/10 Ctenophthalmus assimilis Taschenberg, 1880 and 7/78 Ctenophthalmus agyrtes Heller, 1896). Based on gamont morphology, vertebrate and arthropod host species and DNA sequences, the parasites in our study can be identified as Hepatozoon erhardovae.

Culturable diversity of aerobic halophilic archaea (Fam. Halobacteriaceae) from hypersaline, meromictic Transylvanian lakes.

Perennially stratified salt lakes situated in the Transylvanian Basin (Central Romania) were surveyed for the diversity of culturable halophilic archaea (Fam. Halobacteriaceae). The physical and chemical characteristics of the waters indicated that all the investigated lakes were meromictic and neutral hypersaline. Samples collected from upper, intermediate, and deeper water layers and sediments were used for the isolation of halophilic strains followed by 16S rRNA gene-based identification and phenotypic characterization. The phylogenetic analysis of the 16S rRNA gene sequences revealed that all 191 isolates reported in this study and 43 strains previously isolated were affiliated with the family Halobacteriaceae and classified to 18 genera. Haloferax was the most frequently isolated genus (~47 %), followed by Halobacterium spp. (~12 %), and Halorubrum spp. (~11 %). Highest culturable diversity was detected in Brâncoveanu Lake, the oldest and saltiest of all studied lakes, while the opposite was observed in the most stable and least human-impacted Fara Fund Lake. One strain from Ursu Lake might possibly constitute a novel Halorubrum species as shown by phylogenetic analysis. Several haloarchaeal taxa recently described in Asian (i.e., Iran, China) saline systems were also identified as inhabiting the Transylvanian salt lakes thus expanding our knowledege on the geographic distribution of Halobacteriaceae.