New Insights into the Genome Organization of Yeast Double-Stranded RNA LBC Viruses

The yeasts Torulaspora delbrueckii (Td) and Saccharomyces cerevisiae (Sc) may show a killer phenotype that is encoded in dsRNA M viruses (V-M), which require the helper activity of another dsRNA virus (V-LA or V-LBC) for replication. Recently, two TdV-LBCbarr genomes, which share sequence identity with ScV-LBC counterparts, were characterized by high-throughput sequencing (HTS). They also share some similar characteristics with Sc-LA viruses. This may explain why TdV-LBCbarr has helper capability to maintain M viruses, whereas ScV-LBC does not. We here analyze two stretches with low sequence identity (LIS I and LIS II) that were found in TdV-LBCbarr Gag-Pol proteins when comparing with the homologous regions of ScV-LBC. These stretches may result from successive nucleotide insertions or deletions (indels) that allow compensatory frameshift events required to maintain specific functions of the RNA-polymerase, while modifying other functions such as the ability to bind V-M (+)RNA for packaging. The presence of an additional frameshifting site in LIS I may ensure the synthesis of a certain amount of RNA-polymerase until the new compensatory indel appears. Additional 5′- and 3′-extra sequences were found beyond V-LBC canonical genomes. Most extra sequences showed high identity to some stretches of the canonical genomes and can form stem-loop structures. Further, the 3′-extra sequence of two ScV-LBC genomes contains rRNA stretches. The origin and possible functions of these extra sequences are here discussed.

Detection and characterisation of porcine circoviruses in wild boars in northeastern Serbia

The objective was to expand and update the knowledge on the presence and genotype diversity of porcine circoviruses 2 and 3 (PCV2 and PCV3) in the wild boar populations from the hunting grounds in northeastern Serbia. The presence of PCV3 was not determined, and PCV2 was confirmed in 40.32% of the organ samples from 124 wild boars hunted from 2018 to 2019, indicating their significance in virus circulation since traditional pig farms with irregular PCV2 vaccination strategies are widespread in this region. The most prevalent genotype was PCV2d, followed by PCV2b and PCV2a in 55.6%, 38.9%, and 5.5% of the examined samples, respectively. Nucleotide sequences of the detected strains were homogenous within the genotype and clustered within the subgroups PCV2d-2, PCV2b-1A/B, and PCV2a-2D with high identity to European, Chinese, and Serbian domestic pig sequences suggesting their origin. Wild boars presented with no clinical or pathological signs of infection, implying that these animals might be less susceptible to disease, particularly since the cofactors present in pig farming systems that support the disease development are absent in the wild. The high PCV2 detection frequency demonstrates the importance of wildlife monitoring to track virus population dynamics, especially in regions with free-range pig farming in order to plan adequate disease control strategies.

The Asterias Rubens Sea Star Igkappa Gene when Compared to Marthasterias Glacialis Sea Star Genome (Echinodermata)

The sea star IGKappa gene was cloned in 2014 by the use of primers. It was compared in the present work to Marthasterias glacialis sea star genome. A high identity was found with this last one.

A Predictive Energy Landscape Model of Metamorphic Protein Conformational Specificity

'Metamorphic' proteins challenge state-of-the-art structure prediction methods reliant on amino acid similarity. Unfortunately, this obviates a more effective thermodynamic approach necessary to properly evaluate the impact of amino acid changes on the stability of two different folds. A vital capability of such a thermodynamic approach would be the quantification of the free energy differences between 1) the energy landscape minima of each native fold, and 2) each fold and the denatured state. Here we develop an energetic framework for conformational specificity, based on an ensemble description of protein thermodynamics. This energetic framework was able to successfully recapitulate the structures of high-identity engineered sequences experimentally shown to adopt either Streptococcus protein GA or GB folds, demonstrating that this approach indeed reflected the energetic determinants of fold. Residue-level decomposition of the conformational specificity suggested several testable hypotheses, notably among them that fold-switching could be affected by local de-stabilization of the populated fold at positions sensitive to equilibrium perturbation. Since this ensemble-based compatibility framework is applicable to any structure and any sequence, it may be practically useful for the future targeted design, or large-scale proteomic detection, of novel metamorphic proteins.

Phylogenetic Analyses of Rhizobia Isolated from Nodules of Lupinus angustifolius in Northern Tunisia Reveal Devosia sp. as a New Microsymbiont of Lupin Species

Thirty-two bacterial isolates were obtained from root nodules of Lupinus angustifolius growing in Northern Tunisia. Phylogenetic analyses based on recA and gyrB partial gene sequences grouped the strains into six clusters: four clusters belonged to the genus Bradyrhizobium (22 isolates), one to Microvirga (8 isolates) and one to Devosia (2 isolates), a genus that has not been previously reported to nodulate lupin. Representative strains of each group were further characterized. Multi-Locus Sequence Analysis (MLSA) based on recA and glnII gene sequences separated the strains within the genus Bradyrhizobium into four divergent clusters related to B. canariense, B. liaoningense, B. lupini, and B. algeriense, respectively. The latter might constitute a new Bradyrhizobium species. The strains in the Microvirga cluster showed high identity with M. tunisiensis. The Devosia isolates might also represent a new species within this genus. An additional phylogenetic analysis based on the symbiotic gene nodC affiliated the strains to symbiovars genistearum, mediterranense, and to a possibly new symbiovar. These results altogether contributed to the existing knowledge on the genetic diversity of lupin-nodulating microsymbionts and revealed a likely new, fast-growing, salt-tolerant rhizobial species within the genus Devosia as a potentially useful inoculant in agricultural practices or landscape restoration.

Molecular Identification of a Novel Iflavirus in Brown-Spotted Pitvipers (Protobothrops Mucrosquamatus)

Background: Iflaviridae is a family of small non-enveloped viruses with monopartite, positive-stranded RNA genomes, which are identified in arthropod hosts, primarily infecting insect species. Herein, we firstly identify the sequence of an iflavirus (YB-PMP20) found in brown-spotted pitvipers in China. Results: The sequence of YB-PMP20 showed high identity to the sequences of Hubei picorna-like virus (HUPV) (99.2% in nt), Vespa velutina-associated iflavirus like virus (VVAIV) (58.6% in nt) and Lygus lineolaris virus (LyIV-1) (46.6% in nt) in nucleotides encoding polyproteins. It contained a single large ORF (304–9291 nt) encoding 2996 amino acids. The deduced amino acid sequences were compared with those of iflavirus. Helicase, protease and the RdRp domain were found to be located at the 3´ end, and structural genes (VP1, VP2 and VP3) were found to be located at the 5´ end. Phylogenetic analysis indicated that YB-PMP20 belongs to the iflavirus cluster, and is similar to HUPV, LyIV-1 and VVAIV. Conclusion: The present study described the genetic characterization of a PmIFV strain in brown-spotted pitvipers. Our genomic data extend knowledge of the diversity of viruses in snakes.

Integrative Transcriptomic and Metabolomic Analysis at Organ Scale Reveals Gene Modules Involved in the Responses to Suboptimal Nitrogen Supply in Tomato

The development of high nitrogen use efficiency (NUE) cultivars under low N inputs is required for sustainable agriculture. To this end, in this study, we analyzed the impact of long-term suboptimal N conditions on the metabolome and transcriptome of tomato to identify specific molecular processes and regulators at the organ scale. Physiological and metabolic analysis revealed specific responses to maintain glutamate, asparagine, and sucrose synthesis in leaves for partition to sustain growth, while assimilated C surplus is stored in the roots. The transcriptomic analyses allowed us to identify root and leaf sets of genes whose expression depends on N availability. GO analyses of the identified genes revealed conserved biological functions involved in C and N metabolism and remobilization as well as other specifics such as the mitochondrial alternative respiration and chloroplastic cyclic electron flux. In addition, integrative analyses uncovered N regulated genes in root and leaf clusters, which are positively correlated with changes in the levels of different metabolites such as organic acids, amino acids, and formate. Interestingly, we identified transcription factors with high identity to TGA4, ARF8, HAT22, NF-YA5, and NLP9, which play key roles in N responses in Arabidopsis. Together, this study provides a set of nitrogen-responsive genes in tomato and new putative targets for tomato NUE and fruit quality improvement under limited N supply.

Emergence of Hybrid Resistance and Virulence Plasmids Harboring New Delhi Metallo-β-Lactamase in Klebsiella pneumoniae in Russia

The emergence of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKp) is a new threat to healthcare. In this study, we analyzed nine CR-hvKp isolates of different sequence-types (ST) recovered from patients with nosocomial infections in two hospitals in Saint Petersburg. Whole-genome sequencing showed that eight of them harbored large mosaic plasmids carrying resistance to carbapenems and hypervirulence simultaneously, and four different types of hybrid plasmids were identified. BLAST analysis showed a high identity with two hybrid plasmids originating in the UK and Czech Republic. We demonstrated that hybrid plasmids emerged due to the acquisition of resistance genes by virulent plasmids. Moreover, one of the hybrid plasmids carried a novel New Delhi metallo-beta-lactamase (NDM) variant, differing from NDM-1 by one amino acid substitution (D130N), which did not provide significant evolutionary advantages compared to NDM-1. The discovery of structurally similar plasmids in geographically distant regions suggests that the actual distribution of hybrid plasmids carrying virulence and resistance genes is much wider than expected.

A Subtype of Human Bocavirus Detected in Rattus Norvegicus Feces in China

Background Bocavirus is a typical zoonotic pathogen with a wide range of hosts. Here, we report the epidemiology of human bocavirus (HBoV) detected in Rattus norvegicus. Methods Between May 2015 and May 2017, 357 R. norvegicus were captured in four Chinese provinces. Polymerase chain reaction was used to investigate the prevalence of HBoV in fecal samples. Phylogenetic analysis and sequencing of the entire viral genome were undertaken. Results HBoV was detected in 0.84% (3/357) of samples. Phylogenetic analysis based on the partial VP1 region and near-full-sequence regions showed that HBoV obtained in R. norvegicus was genetically closely related to HBoV-2. One near-full‐length HBoV genome (named “GZ533”) was acquired, and phylogenetic analysis of the three positive sequences revealed that they shared very high identity in nucleotides and amino acids in the VP1 region (96.0%–99.1%). Comparison of GZ533 and other HBoVs revealed ~100% identity of amino acids in the VP1 region, whereas only 37.5% identity of amino acids when compared with R. norvegicus bocavirus.Conclusion HBoV-2 was detected in R. norvegicus in China. R. norvegicus may be a carrier of HBoV infection, and its impact on public health merits attention.

Application of retrotransposon-based Inter-SINE Amplified Polymorphism (ISAP) markers for the differentiation of common poplar genotypes

On the basis of retrotransposon-insertion polymorphisms, molecular markers were developed for the identification and differentiation of poplar (<i>Populus</i> spp.) genotypes. For this purpose, short interspersed nuclear elements (SINEs) in the genome sequence of <i>Populus tremula</i> were identified and assigned to different SINE-families. For families with high copy number and high identity values, primers were derived to amplify Inter- SINE Amplified Polymorphisms (ISAPs) with polymerase chain reaction (PCR). The resulting fragments produce genotype-specific fingerprints. This molecular approach utilizes standard laboratory equipment and is therefore easy to use for the verification of plant material. We demonstrate the functionality of three distinct ISAP primer combinations by comparison to ten simple sequence repeat (SSR) markers to differentiate 23 poplar genotypes. Already by using a single ISAP primer combination all genotypes can be clearly discriminated. Furthermore, the cluster analysis based on three primer combinations divides clones according to their genetic background into two subclusters (by a bootstrap value of 98). Our results clearly demonstrate the usability of ISAP markers to differentiate genotypes and trace progenies of poplar trees.