Degradation of ribosomal RNA during gametocytogenesis.

The life cycle of is characterized by complex regulatory changes that allow adaptation of the parasites to different environmental conditions, which are especially pronounced during transmission between the mammalian host and the insect vector. Previous studies have shown that uses three types of ribosomal RNAs (rRNA A-, S1- and S2-types) at different stages of its life cycle. We used Oxford Nanopore Technologies (ONT) direct RNA sequencing to investigate the dynamics of rRNA usage throughout the parasite's intraerythrocytic development, as well as in salivary gland sporozoites.

Duplexed direct RNA sequencing protocol using polyadenylation and polyuridylation.

Oxford Nanopore Technologies provides multiplexing options for DNA and cDNA sequencing, but not for direct RNA sequencing. Here we describe a duplexing approach and validate it by simultaneously sequencing the rRNA from wild type and knockout that have differential rRNA modifications, successfully demultiplexing the data using bioinformatics approaches.

Deciphering transcript architectural complexity in bacteria and archaea.

RNA transcripts are potential therapeutic targets, yet bacterial transcripts have uncharacterized biodiversity. We developed an algorithm for transcript prediction called tp.py using it to predict transcripts (mRNA and other RNAs) in K12 and E2348/69 strains (Bacteria:gamma-Proteobacteria), strains Scott A and RO15 (Bacteria:Firmicute), strains SG17M and NN2 strains (Bacteria:gamma-Proteobacteria), and (Archaea:Halobacteria).

Deciphering Bacterial and Archaeal Transcriptional Dark Matter and Its Architectural Complexity.

Transcripts are potential therapeutic targets, yet bacterial transcripts remain biological dark matter with uncharacterized biodiversity. We developed and applied an algorithm to predict transcripts for Escherichia coli K12 and E2348/69 strains (Bacteria:gamma-Proteobacteria) with newly generated ONT direct RNA sequencing data while predicting transcripts for Listeria monocytogenes strains Scott A and RO15 (Bacteria:Firmicute), Pseudomonas aeruginosa strains SG17M and NN2 strains (Bacteria:gamma-Proteobacteria), and Haloferax volcanii (Archaea:Halobacteria) using publicly available data. From >5 million E.

Common analysis of direct RNA sequencinG CUrrently leads to misidentification of mC at GCU motifs.

RNA modifications, such as methylation, can be detected with Oxford Nanopore Technologies direct RNA sequencing. One commonly used tool for detecting 5-methylcytosine (mC) modifications is Tombo, which uses an "Alternative Model" to detect putative modifications from a single sample. We examined direct RNA sequencing data from diverse taxa including viruses, bacteria, fungi, and animals.

Immunogenomic profile at baseline predicts host susceptibility to clinical malaria.

Introduction: Host gene and protein expression impact susceptibility to clinical malaria, but the balance of immune cell populations, cytokines and genes that contributes to protection, remains incompletely understood. Little is known about the determinants of host susceptibility to clinical malaria at a time when acquired immunity is developing.

Methods: We analyzed peripheral blood mononuclear cells (PBMCs) collected from children who differed in susceptibility to clinical malaria, all from a small town in Mali.

Common Analysis of Direct RNA SequencinG CUrrently Leads to Misidentification of 5-Methylcytosine Modifications at GCU Motifs.

RNA modifications, such as méthylation, can be detected with Oxford Nanopore Technologies direct RNA sequencing. One commonly used tool for detecting 5-methylcytosine (mC) modifications is Tombo, which uses an "Alternative Model" to detect putative modifications from a single sample. We examined direct RNA sequencing data from diverse taxa including virus, bacteria, fungi, and animals.

Accumulation of endosymbiont genomes in an insect autosome followed by endosymbiont replacement.

Eukaryotic genomes can acquire bacterial DNA via lateral gene transfer (LGT). A prominent source of LGT is Wolbachia, a widespread endosymbiont of arthropods and nematodes that is transmitted maternally through female germline cells. The DNA transfer from the Wolbachia endosymbiont wAna to Drosophila ananassae is extensive and has been localized to chromosome 4, contributing to chromosome expansion in this lineage.

X-treme loss of sequence diversity linked to neo-X chromosomes in filarial nematodes.

The sequence diversity of natural and laboratory populations of Brugia pahangi and Brugia malayi was assessed with Illumina resequencing followed by mapping in order to identify single nucleotide variants and insertions/deletions. In natural and laboratory Brugia populations, there is a lack of sequence diversity on chromosome X relative to the autosomes (πX/πA = 0.2), which is lower than the expected (πX/πA = 0.

Pim kinase inhibitor co-treatment decreases alternative non-homologous end-joining DNA repair and genomic instability induced by topoisomerase 2 inhibitors in cells with FLT3 internal tandem duplication.

Acute myeloid leukemia (AML) with fms-like tyrosine kinase 3 internal tandem duplication (FLT3-ITD) relapses with new chromosome abnormalities following chemotherapy, implicating genomic instability. Error-prone alternative non-homologous end-joining (Alt-NHEJ) DNA double-strand break (DSB) repair is upregulated in FLT3-ITD-expresssing cells, driven by c-Myc. The serine/threonine kinase Pim-1 is upregulated downstream of FLT3-ITD, and inhibiting Pim increases topoisomerase 2 (TOP2) inhibitor chemotherapy drug induction of DNA DSBs and apoptosis.

Comparison of long-read sequencing technologies in interrogating bacteria and fly genomes.

The newest generation of DNA sequencing technology is highlighted by the ability to generate sequence reads hundreds of kilobases in length. Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT) have pioneered competitive long read platforms, with more recent work focused on improving sequencing throughput and per-base accuracy. We used whole-genome sequencing data produced by three PacBio protocols (Sequel II CLR, Sequel II HiFi, RS II) and two ONT protocols (Rapid Sequencing and Ligation Sequencing) to compare assemblies of the bacteria Escherichia coli and the fruit fly Drosophila ananassae.

Complete Mitochondrial Genome Sequence of .

The 13,647-bp complete mitochondrial genome of was sequenced and is syntenic to the mitochondrial genome of Phylogenetic analysis of the mitochondrial genome is consistent with the known phylogeny of ONC5 group filarial nematodes.

Complete Genome Sequence of Bp, the Endosymbiont of Brugia pahangi FR3.

Lymphatic filariasis is a devastating disease caused by filarial nematode roundworms, which contain obligate endosymbionts. Here, we assembled the genome of Bp, the endosymbiont of the filarial nematode , from Illumina, Pacific Biosciences, and Oxford Nanopore data. The complete, circular genome is 1,072,967 bp.

Nearly Complete Genome Sequence of Brugia pahangi FR3.

is a zoonotic parasite that is closely related to human-infecting filarial nematodes. Here, we report the nearly complete genome of , including assemblies of four autosomes and an X chromosome, with only seven gaps. The Y chromosome is still not completely assembled.

Drug Repurposing of Bromodomain Inhibitors as Potential Novel Therapeutic Leads for Lymphatic Filariasis Guided by Multispecies Transcriptomics.

To better understand the transcriptomic interplay of organisms associated with lymphatic filariasis, we conducted multispecies transcriptome sequencing (RNA-Seq) on the filarial nematode , its endosymbiont Bm, and its laboratory vector across the entire life cycle. In Bm, transcription of the noncoding 6S RNA suggests that it may be a regulator of bacterial cell growth, as its transcript levels correlate with bacterial replication rates. For , the transcriptional response reflects the stress that infection exerts on the mosquito with indicators of increased energy demand.

Complete Genome Sequence of Ana, the Wolbachia Endosymbiont of Drosophila ananassae.

Here, we present the complete genome sequence of the endosymbiont Ana, isolated from and derived from Oxford Nanopore and Illumina sequencing. We anticipate that this will aid in comparative genomics and the assembly of specifically in regions containing extensive lateral gene transfer events.

TwinBLAST: When Two Is Better than One.

Analysis of sequence read pairs can be essential for characterizing structural variation, including junction-spanning pairs of reads (JSPRs) suggesting recent lateral/horizontal gene transfer. TwinBLAST can be used to facilitate this analysis of JSPRs by enabling the visualization and curation of two BLAST reports side by side in a single interface.

Multispecies Transcriptomics Data Set of Brugia malayi, Its Endosymbiont Bm, and Aedes aegypti across the B. malayi Life Cycle.

Here, we present a comprehensive transcriptomics data set of Brugia malayi, its Wolbachia endosymbiont Bm, and its vector host. This study samples from 16 stages across the entire B. malayi life cycle, including stage 1 through 4 larvae, adult males and females, embryos, immature microfilariae, and mature microfilariae.

Targeted enrichment outperforms other enrichment techniques and enables more multi-species RNA-Seq analyses.

Enrichment methodologies enable the analysis of minor members in multi-species transcriptomic data. We compared the standard enrichment of bacterial and eukaryotic mRNA to a targeted enrichment using an Agilent SureSelect (AgSS) capture for Brugia malayi, Aspergillus fumigatus, and the Wolbachia endosymbiont of B. malayi (wBm).

Lateral gene transfer between prokaryotes and eukaryotes.

Lateral gene transfer (LGT) is an all-encompassing term for the movement of DNA between diverse organisms. LGT is synonymous with horizontal gene transfer, and the terms are used interchangeably throughout the scientific literature. While LGT has been recognized within the bacteria domain of life for decades, inter-domain LGTs are being increasingly described.

Extensive duplication of the Wolbachia DNA in chromosome four of Drosophila ananassae.

Background: Lateral gene transfer (LGT) from bacterial Wolbachia endosymbionts has been detected in ~20% of arthropod and nematode genome sequencing projects. Many of these transfers are large and contain a substantial part of the Wolbachia genome.

Results: Here, we re-sequenced three D.

Large-scale intersubspecific recombination in the plant-pathogenic bacterium Xylella fastidiosa is associated with the host shift to mulberry.

Homologous recombination plays an important role in the structuring of genetic variation of many bacteria; however, its importance in adaptive evolution is not well established. We investigated the association of intersubspecific homologous recombination (IHR) with the shift to a novel host (mulberry) by the plant-pathogenic bacterium Xylella fastidiosa. Mulberry leaf scorch was identified about 25 years ago in native red mulberry in the eastern United States and has spread to introduced white mulberry in California.

Detecting genetic introgression: high levels of intersubspecific recombination found in Xylella fastidiosa in Brazil.

Documenting the role of novel mutation versus homologous recombination in bacterial evolution, and especially in the invasion of new hosts, is central to understanding the long-term dynamics of pathogenic bacteria. We used multilocus sequence typing (MLST) to study this issue in Xylella fastidiosa subsp. pauca from Brazil, a bacterium causing citrus variegated chlorosis (CVC) and coffee leaf scorch (CLS).

Population genomic analysis of a bacterial plant pathogen: novel insight into the origin of Pierce's disease of grapevine in the U.S.

Invasive diseases present an increasing problem worldwide; however, genomic techniques are now available to investigate the timing and geographical origin of such introductions. We employed genomic techniques to demonstrate that the bacterial pathogen causing Pierce's disease of grapevine (PD) is not native to the US as previously assumed, but descended from a single genotype introduced from Central America. PD has posed a serious threat to the US wine industry ever since its first outbreak in Anaheim, California in the 1880s and continues to inhibit grape cultivation in a large area of the country.