Background: A major bottleneck in the use of metagenome sequencing for human gut microbiome studies has been the lack of a comprehensive genome collection to be used as a reference database. Several recent efforts have been made to re-construct genomes from human gut metagenome data, resulting in a huge increase in the number of relevant genomes. In this work, we aimed to create a collection of the most prevalent healthy human gut prokaryotic genomes, to be used as a reference database, including both MAGs from the human gut and ordinary RefSeq genomes.Results: We screened > 5,700 healthy human gut metagenomes for the containment of > 490,000 publicly available prokaryotic genomes sourced from RefSeq and the recently announced UHGG collection. This resulted in a pool of > 381,000 genomes that were subsequently scored and ranked based on their prevalence in the healthy human metagenomes. The genomes were then clustered at a 97.5% sequence identity resolution, and cluster representatives (30,691 in total) were retained to comprise the HumGut collection. Using the Kraken2 software for classification, we find superior performance in the assignment of metagenomic reads, classifying on average 94.5% of the reads in a metagenome, as opposed to 86% with UHGG and 44% when using standard Kraken2 database. A coarser HumGut collection, consisting of genomes dereplicated at 95% sequence identity—similar to UHGG, classified 88.25% of the reads. HumGut, half the size of standard Kraken2 database and directly comparable to the UHGG size, outperforms them both.Conclusions: The HumGut collection contains > 30,000 genomes clustered at a 97.5% sequence identity resolution and ranked by human gut prevalence. We demonstrate how metagenomes from IBD-patients map equally well to this collection, indicating this reference is relevant also for studies well outside the metagenome reference set used to obtain HumGut. All data and metadata, as well as helpful code, are available athttp://arken.nmbu.no/~larssn/humgut/.

Purpose: Urinary Tract Infections (UTIs) are among the most prevalent infections globally. Every year, approximately 150 million people are diagnosed with UTIs worldwide. The current state-of-the-art diagnostic methods are culture-based and have a turnaround time of 2–4 days for pathogen identification and susceptibility testing. Methods: This study first establishes an optical density culture-based method for spiking healthy urine samples with the six most prevalent uropathogens. Urine samples were spiked at clinically significant concentrations of 10³-10⁵ CFU/ml. Three DNA extraction kits (BioStic, PowerFood, and Blood and Tissue) were investigated based on the DNA yield, average processing time, elution volume, and the average cost incurred per extraction. After DNA extraction, the samples were sequenced using MinION and Flongle flow cells. Results: The Blood and Tissue kit outperformed the other kits based on the investigated parameters. Using nanopore sequencing, all the pathogens and corresponding genes were only identified at a spike concentration of 10⁵ CFU/ml, achieved after 10 min and 3 hours of sequencing, respectively. However, some pathogens and antibiotic-resistance genes (ARG) could be identified from spikes at 10³ colony formation units (CFU/mL). The overall turnaround time was five hours, from sample preparation to sequencing-based identification of pathogen ID and antimicrobial resistance genes. Conclusion: This study demonstrates excellent promise in reducing the time required for informed antibiotic administration from 48 to 72 h to five hours, thereby reducing the number of empirical doses and increasing the chance of saving lives.

Cumulus cells (CCs) are pivotal during oocyte development. This study aimed to identify novel marker genes for porcine oocyte quality by examining the expression of selected genes in CCs and oocytes, employing the model of oocytes from prepubertal animals being of reduced quality compared to those from adult animals. Total RNA was extracted either directly after follicle aspiration or after in vitro maturation, followed by RT-qPCR. Immature gilt CCs accumulated BBOX1 transcripts, involved in L-carnitine biosynthesis, to a 14.8-fold higher level (p < 0.05) relative to sows, while for CPT2, participating in fatty acid oxidation, the level was 0.48 (p < 0.05). While showing no differences between gilt and sow CCs after maturation, CPT2 and BBOX1 levels in oocytes were higher in gilts at both time points. The apparent delayed lipid metabolism and reduced accumulation of ALDOA and G6PD transcripts in gilt CCs after maturation, implying downregulation of glycolysis and the pentose phosphate pathway, suggest gilt cumulus–oocyte complexes have inadequate ATP stores and oxidative stress balance compared to sows at the end of maturation. Reduced expression of BBOX1 and higher expression of CPT2 in CCs before maturation and higher expression of G6PD and ALDOA after maturation are new potential markers of oocyte quality.

Knowledge about the dispersal and gene flow patterns in wild animals are important for our understanding of population ecology and the connectedness of populations. It is also important for management relating to disease control and the transmission of new and emerging diseases. Our study aimed to evaluate the genetic structuring among comparative samples of red foxes in a small part of Scandinavia and to estimate the gene flow and potential directionality in the movements of foxes using an optimized set of microsatellite markers. We compared genetic samples of red foxes (Vulpes vulpes) from two areas in Sweden and two areas in Norway, including red fox samples from areas where the occurrence of the cyclophyllic tapeworm Echinococcus multilocularis has been documented, and areas without known occurrence of the parasite. Our results show a high level of gene flow over considerable distances and substantiates migration from areas affected with E. multilocularis into Norway where the parasite is not yet detected. The results allow us to better understand the gene flow and directionality in the movement patterns of red foxes, which is important for wildlife management authorities regarding the spread of E. multilocularis.

Identifying and characterizing cold responsive genes in Fragaria vesca associated with or responsible for low temperature tolerance is a vital part of strawberry cultivar development. In this study we have investigated the transcript levels of eight genes, two dehydrin genes, three putative ABA-regulated genes, two cold–inducible CBF genes and the alcohol dehydrogenase gene, extracted from leaf and crown tissues of three F. vesca genotypes that vary in cold tolerance. Transcript levels of the CBF/DREB1 transcription factor FvCBF1E exhibited stronger cold up-regulation in comparison to FvCBF1B.1 in all genotypes. Transcripts of FvADH were highly up-regulated in both crown and leaf tissues from all three genotypes. In the ‘ALTA’ genotype, FvADH transcripts were significantly higher in leaf than crown tissues and more than 10 to 20-fold greater than in the less cold-tolerant ‘NCGR1363’ and ‘FDP817’ genotypes. FvGEM, containing the conserved ABRE promoter element, transcript was found to be cold-regulated in crowns. Direct comparison of the kinetics of transcript and protein accumulation of dehydrins was scrutinized. In all genotypes and organs, the changes of XERO2 transcript levels generally preceded protein changes, while levels of COR47 protein accumulation preceded the increases in COR47 RNA in ‘ALTA’ crowns.

Background: A major bottleneck in the use of metagenome sequencing for human gut microbiome studies has been the lack of a comprehensive genome collection to be used as a reference database. Several recent efforts have been made to re-construct genomes from human gut metagenome data, resulting in a huge increase in the number of relevant genomes. In this work, we aimed to create a collection of the most prevalent healthy human gut prokaryotic genomes, to be used as a reference database, including both MAGs from the human gut and ordinary RefSeq genomes. Results: We screened > 5,700 healthy human gut metagenomes for the containment of > 490,000 publicly available prokaryotic genomes sourced from RefSeq and the recently announced UHGG collection. This resulted in a pool of > 379,000 genomes that were subsequently scored and ranked based on their prevalence in the healthy human metagenomes. The genomes were then clustered at subspecies resolution, and cluster representatives were retained to comprise the HumGut collection. Using the Kraken2 software for classification, we find superior performance in the assignment of metagenomic reads, classifying on average 94.5% of the reads in a metagenome, as opposed to 86% with UHGG and 44% when using standard Kraken2 database. HumGut, half the size of standard Kraken2 database and directly comparable to the UHGG size, outperforms them both. Conclusions: The HumGut collection contains > 30,000 genomes clustered at subspecies resolution and ranked by human gut prevalence. We demonstrate how metagenomes from IBD-patients map equally well to this collection, indicating this reference is relevant also for studies well outside the metagenome reference set used to obtain HumGut. We believe this is a valuable resource in a field in dire need of method standardization. All data and metadata, as well as helpful code, are available at http://arken.nmbu.no/~larssn/humgut/ .