Background: Advancements in DNA sequencing technology have transformed the field of bacterial genomics, allowing for faster and more cost effective chromosome level assemblies compared to a decade ago. However, transforming raw reads into a complete genome model is a significant computational challenge due to the varying quality and quantity of data obtained from different sequencing instruments, as well as intrinsic characteristics of the genome and desired analyses. To address this issue, we have developed a set of container-based pipelines using Nextflow, offering both common workflows for inexperienced users and high levels of customization for experienced ones. Their processing strategies are adaptable based on the sequencing data type, and their modularity enables the incorporation of new components to address the community’s evolving needs. Methods: These pipelines consist of three parts: quality control, de novo genome assembly, and bacterial genome annotation. In particular, the genome annotation pipeline provides a comprehensive overview of the genome, including standard gene prediction and functional inference, as well as predictions relevant to clinical applications such as virulence and resistance gene annotation, secondary metabolite detection, prophage and plasmid prediction, and more. Results: The annotation results are presented in reports, genome browsers, and a web-based application that enables users to explore and interact with the genome annotation results. Conclusions: Overall, our user-friendly pipelines offer a seamless integration of computational tools to facilitate routine bacterial genomics research. The effectiveness of these is illustrated by examining the sequencing data of a clinical sample of Klebsiella pneumoniae.

The Amazonian rainforest is a hyper-diverse ecosystem in the number of species and the myriad of intertaxon relationships that are mostly understudied. In order to characterize a dominant and economically important Amazonian species, the Brazil nut tree (Bertholletia excelsa Bonpl.), at the genome level, wegenerated high-coverage long-read sequencing data from the leaves of a single individual. The genome assembly revealed an unexpected discovery: two circular contigs that could be assigned to the chromosome and a plasmid of a Pantoea stewartii strain. Comparative genomics revealed that this strain belongs to the indologenes subspecies and displays high synteny with other strains isolated from diseased leaves of the neotropical palm Bactris gasipaes Kunth. Investigation of pathogenicity-related genes revealed the absence of the entire type III secretion system gene cluster in the plasmid, which was otherwise highly similar to a plasmid from an isolate known to cause disease in Dracaena sanderiana Mast. In contrast, several genes associated with plant-growth promoting traits were detected, including genes involved in indole-3-acetic acid (IAA) production, phosphate solubilization, and biosynthesis of siderophores. In summary, we report the genome of an uncultivated P. stewartii subsp. indologenes strain associated with the Brazil nut tree and potentially a plant growth-promoting bacteria.

Antimicrobial resistance (AMR) is an increasing and urgent issue for human health worldwide, as it leads to the reduction of available antibiotics to treat bacterial infections, in turn increasing hospital stays and lethality. Therefore, the study and genomic surveillance of bacterial carriers of resistance in and outside of clinical settings is of utter importance. A colony of multidrug resistant (MDR) bacteria identified as Klebsiella spp., by 16S rDNA amplicon sequencing, has been isolated from an urban lake in Brazil, during a drug-degrading bacterial prospection. Genomic analyses revealed the bacteria as Klebsiella pneumoniae species. Furthermore, the in silico Multilocus Sequence Typing (MLST) identified the genome as a new sequence type, ST5236. The search for antimicrobial resistance genes (ARGs) detected the presence of genes against beta-lactams, fosfomycin, acriflavine and efflux pumps, as well as genes for heavy metal resistance. Of particular note, an extended-spectrum beta-lactamase gene (blaCTX-M-15) has been detected in close proximity to siphoviridae genes, while a carbapenemase gene (KPC-2) has been found in an extrachromosomal contig, within a novel non-Tn4401 genetic element (NTE_KPC). An extrachromosomal contig found in the V3 isolate is identical to a contig of a K. pneumoniae isolate from a nearby hospital, which indicates a putative gene flow from the hospital network into Paranoá lake. The discovery of a MDR isolate in this lake is worrisome, as the region has recently undergone periods of water scarcity causing the lake, which receives treated wastewater effluent, and is already used for recreational purposes, to be used as an environmental buffer for drinking water reuse. Altogether, our results indicate an underrepresentation of environmental K. pneumoniae among available genomes, which may hamper the understanding of the population dynamics of the species in the environment and its consequences in the spread of ARGs and virulence genes.

Summary: Single-cell RNA sequencing (scRNA-seq) has become a popular approach for studying the transcriptome, providing a powerful tool for discovering and characterizing cell types and their developmental trajectories. However, scRNA-seq analysis is complex, requiring a continuous, iterative process to refine the data processing and uncover relevant biological information. We present Asc-Seurat, a feature rich workbench, providing a user-friendly and easy-to-install web application encapsulating the necessary tools for an all-encompassing and fluid scRNA-seq data analysis. Availability and implementation: Asc-Seurat is available at https://github.com/KirstLab/asc_seurat/ and released under GNU 3 license. Contact: mkirst@ufl.edu Supplementary information: Supplementary data are available at Bioinformatics online.