Accelerating Complete Phytoplasma Genome Assembly by Immunoprecipitation-Based Enrichment and MinION-Based DNA Sequencing for Comparative Analyses

Phytoplasmas are uncultivated plant-pathogenic bacteria with agricultural importance. Those belonging to the 16SrII group, represented by ‘Candidatus P. aurantifolia’, have a wide range of plant hosts and cause significant yield losses in valuable crops, such as pear, sweet potato, peanut, and soybean. In this study, a method that combines immunoprecipitation-based enrichment and MinION long-read DNA sequencing was developed to solve the challenge of phytoplasma genome studies. This approach produced long reads with high mapping rates and high genomic coverage that can be combined with Illumina reads to produce complete genome assemblies with high accuracy. We applied this method to strain NCHU2014 and determined its complete genome sequence, which consists of one circular chromosome with 635,584 bp and one plasmid with 4,224 bp. Although ‘Ca. P. aurantifolia’ NCHU2014 has a small chromosome with only 471 protein-coding genes, it contains 33 transporter genes and 27 putative effector genes, which may contribute to obtaining nutrients from hosts and manipulating host developments for their survival and multiplication. Two effectors, the homologs of SAP11 and SAP54/PHYL1 identified in ‘Ca. P. aurantifolia’ NCHU2014, have the biochemical activities in destabilizing host transcription factors, which can explain the disease symptoms observed in infected plants. Taken together, this study provides the first complete genome available for the 16SrII phytoplasmas and contributes to the understanding of phytoplasma pathogenicity.

Karyotype Variation in Eight Cultivars of Indian Dessert Banana (Musa acuminata L.) of Section Eumusa From Odisha, India

Banana (Musa spp.) cultivars especially dessert banana are important cash crop with high market demand all over the world as an integral part of the diet. The need for assessment of cytogenetic characters in Musa cultivars is inevitable as out of thousands of cultivars, cytogenetic characterization of most of them remains unresolved due to difficulties like small chromosome size, diversity in ploidy levels and high cultivar diversity which behave differently to standardized cytogenetic protocols. In this report, somatic chromosome number, detailed karyotype analysis including total chromosome length, volume, form percentage, Interphase Nuclear Volume (INV) were accessed on eight dessert type of Musa accessions from different places of Odisha. All the cultivars studied were found triploid (2n = 33) with a basic chromosome number of x=11. The karyotype formulae were assigned to each cultivar by grouping the chromosome according to their shared characteristics. The total chromosome length ranged from 54.95 µm in cv. Robusta to 81.5 µm in cv. Kathia with symmetric karyotype in all the studied cultivar. Karyotype formula revealed structural alteration of chromosome with Total Form percentage (TF%) variation from 35.65% in cv. Amritapani to 41.68% in cv. Patakpura that confirms more number of nearly median constricted chromosome as compared to sub-median chromosome. The total chromosome volume recorded from 10.78 µm3 in cv. Robusta to 15.99 µm3 in cv. Khatia and the INV varied from 1336.44 µm3 in cv. Dwarf Cavendish to 2048.37 µm3 in cv. Patakpura. The recorded structural variation might be due to differential genome specific condensation of chromosome. Chromosome length and volume found statistically significant among the cultivars.

Vibrio cholerae ParE2 toxin modulates its operon transcription by stabilization of an antitoxin DNA ruler

The parDE2 operon of Vibrio cholerae encodes a type II TA system, which is one of three loci in the superintegron of small chromosome II that show modest similarity to the parDE operon of plasmid RK2. ParE2, like plasmid RK2-encoded ParE, inhibits DNA gyrase, an essential topoisomerase that is also the target of quinolone antibacterial agents. Mechanistic understanding on ParE2 toxin inhibition by direct interaction with its cognate antitoxin and transcriptional autoregulation of the TA system are currently lacking. ParD2, the ribbon-helix-helix (RHH) antitoxin, auto-represses the parDE2 promoter. This repression is enhanced by ParE2, which therefore functions as a transcriptional co-repressor. Here we present protein-DNA interaction studies and high-resolution X-ray structures of the ParD2:ParE2 complex and isolated ParD2 antitoxin, revealing the basis of toxin inhibition and autoregulation of the TA operon by conditional cooperativity. Native mass spectrometry, SAXS and MALS studies confirm the presence of different oligomerization states of ParD2 in solution and the role of the DNA-binding hexameric ParD26:ParE22 assembly in transcriptional repression.

Cytogenetics of Four Species of the Green Clade Aplastodiscus Lutz, 1950 (Anura: Cophomantinae): New Insights into the Chromosomal Evolution of the Genus

The tree frog <i>Aplastodiscus</i> is a Neotropical taxon that encompasses 15 species in the Atlantic forest biome, with one isolated species in the Central Brazilian Cerrado. To date, only 8 species have been karyotyped, showing high levels of diploid number variation, which allowed clustering species in chromosome number groups: 2n = 24 (<i>Aplastodiscus perviridis</i> group), 2n = 22 (<i>Aplastodiscus albofrenatus</i> group), 2n = 20, and 2n = 18 (both within <i>Aplastodiscus albosignatus</i> group). This study aims to report karyotypic information on 4 species from the last 2 groups using classical and molecular cytogenetic techniques and hypothesize chromosomal evolutionary trends within the species groups. <i>Aplastodiscus weygoldti</i> showed 2n = 22; Ag-NOR and FISH 18S rDNA signals were located in the interstitial region of the short arms of chromosome pair 6. <i>Aplastodiscus cavicola, Aplastodiscus</i> sp. 4, and <i>Aplastodiscus</i> sp. 6 showed 2n = 18; Ag-NOR and FISH 18S rDNA bands were located in the terminal region of the long arm of chromosome pair 9. Our results support multiple and independent chromosome fusion events within <i>Aplastodiscus</i>, including a new chromosome fission event<i>.</i> Ag-NOR and FISH 18S rDNA patterns were restricted to the small chromosome pairs, similar to the other species within this genus, and confirm overall chromosome morphology conservation among the genera of Cophomantinae.

Karyotypic Characteristics and Genetic Relationships of Apricot Accessions from Different Ecological Groups

The present study aims to reveal the karyotypic characteristics and genetic relationships of apricot (Prunus armeniaca L.) accessions from different ecological groups. Fourteen, 9, and 30 accessions from the Central Asian ecological group, North China ecological group, and Dzhungar-Ili ecological group, respectively, were analyzed according to the conventional pressing plate method. The results showed that all the apricot accessions from the different ecological groups were diploid (2n = 2x = 16). The total haploid length of the chromosome set of the selected accessions ranged from 8.11 to 12.75 μm, which was a small chromosome, and no satellite chromosomes were detected. All accessions had different numbers of median-centromere chromosomes or sub-median-centromere chromosomes. The karyotypes of the selected accessions were classified as 1A or 2A. Principal component analysis revealed that the long-arm/short-arm ratio (0.968) and the karyotype symmetry index (−0.979) were the most valuable parameters, and cluster analysis revealed that the accessions from the Central Asian ecological group and Dzhungar-Ili ecological group clustered together. In terms of karyotypic characteristics, the accessions from the Dzhungar-Ili ecological group and Central Asian ecological group were closely related.

Chromosome-scale genome assembly for the duckweed Spirodela intermedia, integrating cytogenetic maps, PacBio and Oxford Nanopore libraries

Duckweeds are small, free-floating, morphologically highly reduced organisms belonging to the monocot order Alismatales. They display the most rapid growth among flowering plants, vary ~ 14-fold in genome size and comprise five genera. Spirodela is the phylogenetically oldest genus with only two mainly asexually propagating species: S. polyrhiza (2n = 40; 160 Mbp/1C) and S. intermedia (2n = 36; 160 Mbp/1C). This study combined comparative cytogenetics and de novo genome assembly based on PacBio, Illumina and Oxford Nanopore (ON) reads to obtain the first genome reference for S. intermedia and to compare its genomic features with those of the sister species S. polyrhiza. Both species’ genomes revealed little more than 20,000 putative protein-coding genes, very low rDNA copy numbers and a low amount of repetitive sequences, mainly Ty3/gypsy retroelements. The detection of a few new small chromosome rearrangements between both Spirodela species refined the karyotype and the chromosomal sequence assignment for S. intermedia.

Identification and mapping of new genes for resistance to downy mildew in lettuce

Key message Eleven new major resistance genes for lettuce downy mildew were introgressed from wild Lactuca species and mapped to small regions in the lettuce genome. Abstract Downy mildew, caused by the oomycete pathogen Bremia lactucae Regel, is the most important disease of lettuce (Lactuca sativa L.). The most effective method to control this disease is by using resistant cultivars expressing dominant resistance genes (Dm genes). In order to counter changes in pathogen virulence, multiple resistance genes have been introgressed from wild species by repeated backcrosses to cultivated lettuce, resulting in numerous near-isogenic lines (NILs) only differing for small chromosome regions that are associated with resistance. Low-pass, whole genome sequencing of 11 NILs was used to identify the chromosome segments introgressed from the wild donor species. This located the candidate chromosomal positions for resistance genes as well as additional segments. F2 segregating populations derived from these NILs were used to genetically map the resistance genes to one or two loci in the lettuce reference genome. Precise knowledge of the location of new Dm genes provides the foundation for marker-assisted selection to breed cultivars with multiple genes for resistance to downy mildew.

Seed Composition Survey of a Peanut CSSL Population Reveals Introgression Lines with Elevated Oleic/Linoleic Profiles

ABSTRACT The peanut CSSL population represents one of the ways that interspecific hybridization has been used to introduce genetic variation into cultivated peanut. The lines were developed by crossing Fleur 11, a farmer preferred spanish cultivar from West Africa with a synthetic allotetraploid. The latter was developed by crossing A. duranensis to A. ipaensis and tetraploidizing the resultant hybrid. Subsequent selection with genetic markers resulted in a population comprising lines with small chromosome segments from the wild in a cultivated peanut background. The objective of this study was to characterize the protein, total oil, fatty acid and sugar profiles of the population. The results indicated that the values of Fleur 11 for all the traits analyzed were within the normal range expected in peanut. Since the population had a uniform genetic background derived from Fleur 11, the profiles for a majority of the lines were comparable to Fleur 11. However, three lines (CSSL 84, CSSL 100 and CSSL 111) were found to have elevated oleic acid and reduced linoleic and palmitic acid relative to Fleur 11. The oleic to linoleic acid ratios (O/L) for these lines were 118, 104 and 97% greater than that of Fleur 11, respectively. While the increased values are still considered to be within the normal oleic acid range, the effect of introgressions on these lines represent the possibility of discovering new sources of high O/L polymorphisms. Such polymorphisms have the potential for use in further improving peanut oil quality.

Identification of Vibrio cholerae Strains of the «Haitian» Group by PCR Based on INDEL-Typing

The aim of the work was to find a genetic INDEL-marker of the Haitian group of Vibrio cholerae strains, what allow carrying out their identification by means PCR.Materials and methods. For searching INDEL-markers we used the data from GenBank database on complete genomic sequences of V. cholerae strains El Тor isolated in different continents in different years. For the analysis we used the author's software written in the Java programming language. The NextGIS system was used for mapping.Results and discussion. We found that the deletion of 8 nucleotides in the gene VCA1095 located on a small chromosome and encoding chemotaxis protein СheA is a characteristic genetic feature of the «Haitian» group strains. Primers have been developed to detect this deletion in PCR.Conclusion. The method of detection of strains of cholera vibrions «Haitian group» on the basis of INDELmarkers was developed and the distribution of such strains in the world before and after 2010 was shown.

Imprints of tumor mutation burden on chromosomes and relation to cancer risk in humans: A pan-cancer analysis

Cancers, resulting in uncontrolled cell proliferation, are driven by accumulation of somatic mutations. Genome-wide sequencing has produced a catalogue of millions of somatic mutations, which contain the evolutionary history of the cancers. However, the connection between the mutation accumulation and disease development and risks is poorly understood. Here, we analyzed more than 1,200,000 mutations from 5,000 cancer patients and revealed two novel signatures for 16 cancer types in The Cancer Genome Atlas (TCGA) database. A strong correlation between Tumor Mutation Burden (TMB) and the Patient Age at Diagnosis (PAD) is observed for cancers with low TMB (mean value less than 3 mutations per million base pairs) but is absent in cancers with high TMB. Surprisingly, the differences in cancer risk between the sexes are also mainly driven by the disparity in mutation burden. The TMB variations, imprinted at the chromosome level, also reflect accumulation of mutation clusters within small chromosome segments in high TMB cancers. A model, relating mutation rates to PAD and latency period for cancer detection, explains the key findings.