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MobiSeq: De novo SNP discovery in model and non-model species through sequencing the flanking region of transposable elements

Rey-Iglesia A, Gopalakrishan S, Carøe C, Alquezar-Planas DE, Ahlmann Nielsen A, Röder T, et al.

Mol Ecol Resour, 2019 Mar;19(2):512-525.
PMID: 30575257 DOI: 10.1111/1755-0998.12984

In recent years, the availability of reduced representation library (RRL) methods has catalysed an expansion of genome-scale studies to characterize both model and non-model organisms. Most of these methods rely on the use of restriction enzymes to obtain DNA sequences at a genome-wide level. These approaches have been widely used to sequence thousands of markers across individuals for many organisms at a reasonable cost, revolutionizing the field of population genomics. However, there are still some limitations associated with these methods, in particular the high molecular weight DNA required as starting material, the reduced number of common loci among investigated samples, and the short length of the sequenced site-associated DNA. Here, we present MobiSeq, a RRL protocol exploiting simple laboratory techniques, that generates genomic data based on PCR targeted enrichment of transposable elements and the sequencing of the associated flanking region. We validate its performance across 103 DNA extracts derived from three mammalian species: grey wolf (Canis lupus), red deer complex (Cervus sp.) and brown rat (Rattus norvegicus). MobiSeq enables the sequencing of hundreds of thousands loci across the genome and performs SNP discovery with relatively low rates of clonality. Given the ease and flexibility of MobiSeq protocol, the method has the potential to be implemented for marker discovery and population genomics across a wide range of organisms-enabling the exploration of diverse evolutionary and conservation questions.
Fulltext High diversity of picornaviruses in rats from different continents revealed by deep sequencing

Hansen TA, Mollerup S, Nguyen NP, White NE, Coghlan M, Alquezar-Planas DE, et al.

Emerg Microbes Infect, 2016 Aug 17;5(8):e90.
PMID: 27530749 DOI: 10.1038/emi.2016.90

Outbreaks of zoonotic diseases in humans and livestock are not uncommon, and an important component in containment of such emerging viral diseases is rapid and reliable diagnostics. Such methods are often PCR-based and hence require the availability of sequence data from the pathogen. Rattus norvegicus (R. norvegicus) is a known reservoir for important zoonotic pathogens. Transmission may be direct via contact with the animal, for example, through exposure to its faecal matter, or indirectly mediated by arthropod vectors. Here we investigated the viral content in rat faecal matter (n=29) collected from two continents by analyzing 2.2 billion next-generation sequencing reads derived from both DNA and RNA. Among other virus families, we found sequences from members of the Picornaviridae to be abundant in the microbiome of all the samples. Here we describe the diversity of the picornavirus-like contigs including near-full-length genomes closely related to the Boone cardiovirus and Theiler's encephalomyelitis virus. From this study, we conclude that picornaviruses within R. norvegicus are more diverse than previously recognized. The virome of R. norvegicus should be investigated further to assess the full potential for zoonotic virus transmission.
Fulltext Contaminating viral sequences in high-throughput sequencing viromics: a linkage study of 700 sequencing libraries

Asplund M, Kjartansdóttir KR, Mollerup S, Vinner L, Fridholm H, Herrera JAR, et al.

Clin Microbiol Infect, 2019 Oct;25(10):1277-1285.
PMID: 31059795 DOI: 10.1016/j.cmi.2019.04.028

OBJECTIVES: Sample preparation for high-throughput sequencing (HTS) includes treatment with various laboratory components, potentially carrying viral nucleic acids, the extent of which has not been thoroughly investigated. Our aim was to systematically examine a diverse repertoire of laboratory components used to prepare samples for HTS in order to identify contaminating viral sequences.
METHODS: A total of 322 samples of mainly human origin were analysed using eight protocols, applying a wide variety of laboratory components. Several samples (60% of human specimens) were processed using different protocols. In total, 712 sequencing libraries were investigated for viral sequence contamination.
RESULTS: Among sequences showing similarity to viruses, 493 were significantly associated with the use of laboratory components. Each of these viral sequences had sporadic appearance, only being identified in a subset of the samples treated with the linked laboratory component, and some were not identified in the non-template control samples. Remarkably, more than 65% of all viral sequences identified were within viral clusters linked to the use of laboratory components.
CONCLUSIONS: We show that high prevalence of contaminating viral sequences can be expected in HTS-based virome data and provide an extensive list of novel contaminating viral sequences that can be used for evaluation of viral findings in future virome and metagenome studies. Moreover, we show that detection can be problematic due to stochastic appearance and limited non-template controls. Although the exact origin of these viral sequences requires further research, our results support laboratory-component-linked viral sequence contamination of both biological and synthetic origin.
Fulltext High-Throughput Sequencing-Based Investigation of Viruses in Human Cancers by Multienrichment Approach

Mollerup S, Asplund M, Friis-Nielsen J, Kjartansdóttir KR, Fridholm H, Hansen TA, et al.

J Infect Dis, 2019 09 13;220(8):1312-1324.
PMID: 31253993 DOI: 10.1093/infdis/jiz318

BACKGROUND: Viruses and other infectious agents cause more than 15% of human cancer cases. High-throughput sequencing-based studies of virus-cancer associations have mainly focused on cancer transcriptome data.
METHODS: In this study, we applied a diverse selection of presequencing enrichment methods targeting all major viral groups, to characterize the viruses present in 197 samples from 18 sample types of cancerous origin. Using high-throughput sequencing, we generated 710 datasets constituting 57 billion sequencing reads.
RESULTS: Detailed in silico investigation of the viral content, including exclusion of viral artefacts, from de novo assembled contigs and individual sequencing reads yielded a map of the viruses detected. Our data reveal a virome dominated by papillomaviruses, anelloviruses, herpesviruses, and parvoviruses. More than half of the included samples contained 1 or more viruses; however, no link between specific viruses and cancer types were found.
CONCLUSIONS: Our study sheds light on viral presence in cancers and provides highly relevant virome data for future reference.
Fulltext Genomic analysis of the causative agents of coccidiosis in domestic chickens

Reid AJ, Blake DP, Ansari HR, Billington K, Browne HP, Bryant J, et al.

Genome Res, 2014 Oct;24(10):1676-85.
PMID: 25015382 DOI: 10.1101/gr.168955.113

Global production of chickens has trebled in the past two decades and they are now the most important source of dietary animal protein worldwide. Chickens are subject to many infectious diseases that reduce their performance and productivity. Coccidiosis, caused by apicomplexan protozoa of the genus Eimeria, is one of the most important poultry diseases. Understanding the biology of Eimeria parasites underpins development of new drugs and vaccines needed to improve global food security. We have produced annotated genome sequences of all seven species of Eimeria that infect domestic chickens, which reveal the full extent of previously described repeat-rich and repeat-poor regions and show that these parasites possess the most repeat-rich proteomes ever described. Furthermore, while no other apicomplexan has been found to possess retrotransposons, Eimeria is home to a family of chromoviruses. Analysis of Eimeria genes involved in basic biology and host-parasite interaction highlights adaptations to a relatively simple developmental life cycle and a complex array of co-expressed surface proteins involved in host cell binding.