Scientific Publications

The following scientific publications are related to Global Surveillance and the associated projects.

Setting a baseline for global urban virome surveillance in sewage
Nieuwenhuijse et al. 2020

The rapid development of megacities, and their growing connectedness across the world is becoming a distinct driver for emerging disease outbreaks. Early detection of unusual disease emergence and spread should therefore include such cities as part of risk-based surveillance. A catch-all metagenomic sequencing approach of urban sewage could potentially provide an unbiased insight into the dynamics of viral pathogens circulating in a community irrespective of access to care, a potential which already has been proven for the surveillance of poliovirus. Here, we present a detailed characterization of sewage viromes from a snapshot of 81 high density urban areas across the globe, including in-depth assessment of potential biases, as a proof of concept for catch-all viral pathogen surveillance. We show the ability to detect a wide range of viruses and geographical and seasonal differences for specific viral groups. Our findings offer a cross-sectional baseline for further research in viral surveillance from urban sewage samples and place previous studies in a global perspective.

DOI:    13-08-2020

ResFinder 4.0 for predictions of phenotypes from genotypes
Bortolaia et al. 2020

WGS-based antimicrobial susceptibility testing (AST) is as reliable as phenotypic AST for several antimicrobial/bacterial species combinations. However, routine use of WGS-based AST is hindered by the need for bioinformatics skills and knowledge of antimicrobial resistance (AMR) determinants to operate the vast majority of tools developed to date. By leveraging on ResFinder and PointFinder, two freely accessible tools that can also assist users without bioinformatics skills, we aimed at increasing their speed and providing an easily interpretable antibiogram as output.

The ResFinder code was re-written to process raw reads and use Kmer-based alignment. The existing ResFinder and PointFinder databases were revised and expanded. Additional databases were developed including a genotype-to-phenotype key associating each AMR determinant with a phenotype at the antimicrobial compound level, and species-specific panels for in silico antibiograms. ResFinder 4.0 was validated using Escherichia coli (n= 584),Salmonella spp. (n= 1081),Campylobacter jejuni (n= 239), Enterococcus faecium (n= 106), Enterococcus faecalis (n= 50) and Staphylococcus aureus (n= 163) exhibiting different AST profiles, and from different human and animal sources and geographical origins.

DOI:  11-08-2020

The gut microbiome but not the resistome is associated with urogenital schistosomiasis in preschool-aged children
Osakunor et al. 2020

Helminth parasites have been shown to have systemic effects in the host. Using shotgun metagenomic sequencing, we characterise the gut microbiome and resistome of 113 Zimbabwean preschool-aged children (1–5 years). We test the hypothesis that infection with the human helminth parasite, Schistosoma haematobium, is associated with changes in gut microbial and antimicrobial resistance gene abundance/diversity. Here, we show that bacteria phyla Bacteroidetes, Firmicutes, Proteobacteria, and fungi phyla Ascomycota, Microsporidia, Zoopagomycota dominate the microbiome. The abundance of Proteobacteria, Ascomycota, and Basidiomycota differ between schistosome-infected versus uninfected children. Specifically, infection is associated with increases in Pseudomonas, Stenotrophomonas, Derxia, Thalassospira, Aspergillus, Tricholoma, and Periglandula, with a decrease in Azospirillum. We find 262 AMR genes, from 12 functional drug classes, but no association with individual-specific data. To our knowledge, we describe a novel metagenomic dataset of Zimbabwean preschool-aged children, indicating an association between urogenital schistosome infection and changes in the gut microbiome.

DOI:     02-04-2020

Metaphylogenetic analysis of global sewage reveals that bacterial strains associated with human disease show less degree of geographic clustering
Ahrenfeldt et al. 2020

Knowledge about the difference in the global distribution of pathogens and non-pathogens is limited. Here, we investigate it using a multi-sample metagenomics phylogeny approach based on short-read metagenomic sequencing of sewage from 79 sites around the world. For each metagenomic sample, bacterial template genomes were identified in a non-redundant database of whole genome sequences. Reads were mapped to the templates identified in each sample. Phylogenetic trees were constructed for each template identified in multiple samples.

DOI:     20-02-2020

Clades of huge phages from across Earth’s ecosystems
Basem Al-Shayeb et al. 2020

Bacteriophages typically have small genomes and depend on their bacterial hosts for replication. Here we sequenced DNA from diverse ecosystems and found hundreds of phage genomes with lengths of more than 200 kilobases (kb), including a genome of 735 kb, which is—to our knowledge—the largest phage genome to be described to date. Thirty-five genomes were manually curated to completion (circular and no gaps). Expanded genetic repertoires include diverse and previously undescribed CRISPR–Cas systems, transfer RNAs (tRNAs), tRNA synthetases, tRNA-modification enzymes, translation-initiation and elongation factors, and ribosomal proteins.

DOI:   12-02-2020

Using sewage for surveillance of antimicrobial resistance
Frank M. Aarestrup and Mark E. J. Woolhouse

Antimicrobial resistance (AMR), a cross-cutting and increasing threat to global health, is a complex problem with multiple and interconnected drivers. Reliable surveillance data that accurately describe and characterize the global occurrence and distribution of AMR are essential for tracking changes in resistance over time, setting national and global priorities, assessing the impacts of interventions, identifying new kinds of resistance, and supporting investigation of (international) outbreaks of resistant pathogens. AMR surveillance data can also inform development of treatment guidelines. Yet it has proven difficult to achieve these objectives on a global scale, and especially in low- and middle-income countries (LMICs), largely because current surveillance systems deliver data that are extremely variable in quality and quantity and highly heterogeneous in terms of which population is sampled (usually a category of hospital patients) and what drug-bug combinations are included. Here, we outline a plan for a global AMR surveillance system based on applying next-generation sequencing (NGS) to human sewage that will be especially helpful for community AMR surveillance, which is difficult to achieve in other ways, and will provide an affordable surveillance option in resource-poor settings.

DOI:  10.1126/science.aba3432     07-02-2020

Prediction of Acquired Antimicrobial Resistance for Multiple Bacterial Species Using Neural Networks
Aytan-Aktug et al. 2020

Machine learning has proven to be a powerful method to predict antimicrobial resistance (AMR) without using prior knowledge for selected bacterial species-antimicrobial combinations. To date, only species-specific machine learning models have been developed, and to the best of our knowledge, the inclusion of information from multiple species has not been attempted. The aim of this study was to determine the feasibility of including information from multiple bacterial species to predict AMR for an individual species, since this may make it easier to train and update resistance predictions for multiple species and may lead to improved predictions.

DOI:  21-01-2020

Using Genomics to Track Global Antimicrobial Resistance
Hendriksen et al. 2019

The recent advancements in rapid and affordable DNA sequencing technologies have revolutionized diagnostic microbiology and microbial surveillance. The availability of bioinformatics tools and online accessible databases has been a prerequisite for this. We conducted a scientific literature review and here we present a description of examples of available tools and databases for antimicrobial resistance (AMR) detection and provide future perspectives and recommendations. At least 47 freely accessible bioinformatics resources for detection of AMR determinants in DNA or amino acid sequence data have been developed to date. These include, among others but not limited to, ARG-ANNOT, CARD, SRST2, MEGARes, Genefinder, ARIBA, KmerResistance, AMRFinder, and ResFinder.

DOI:   04-09-2019

Worldwide human mitochondrial haplogroup distribution from urban sewage
Pipek et al. 2019

Community level genetic information can be essential to direct health measures and study demographic tendencies but is subject to considerable ethical and legal challenges. These concerns become less pronounced when analyzing urban sewage samples, which are ab ovo anonymous by their pooled nature. We were able to detect traces of the human mitochondrial DNA (mtDNA) in urban sewage samples and to estimate the distribution of human mtDNA haplogroups. An expectation maximization approach was used to determine mtDNA haplogroup mixture proportions for samples collected at each different geographic location. Our results show reasonable agreement with both previous studies of ancient evolution or migration and current US census data; and are also readily reproducible and highly robust. Our approach presents a promising alternative for sample collection in studies focusing on the ethnic and genetic composition of populations or diseases associated with different mtDNA haplogroups and genotypes.

DOI:     12-08-2019

Global phylogeography and ancient evolution of the widespread human gut virus crAssphage
Edwards et al. 2019

Microbiomes are vast communities of microorganisms and viruses that populate all natural ecosystems. Viruses have been considered to be the most variable component of microbiomes, as supported by virome surveys and examples of high genomic mosaicism. However, recent evidence suggests that the human gut virome is remarkably stable compared with that of other environments. Here, we investigate the origin, evolution and epidemiology of crAssphage, a widespread human gut virus. Through a global collaboration, we obtained DNA sequences of crAssphage from more than one-third of the world’s countries and showed that the phylogeography of crAssphage is locally clustered within countries, cities and individuals.

DOI:   08-07-2019

Global monitoring of antimicrobial resistance based on metagenomics analyses of urban sewage
Hendriksen et al. 2019

Antimicrobial resistance (AMR) is a serious threat to global public health, but obtaining representative data on AMR for healthy human populations is difficult. Here, we use metagenomic analysis of untreated sewage to characterize the bacterial resistome from 79 sites in 60 countries. We find systematic differences in abundance and diversity of AMR genes between Europe/North-America/Oceania and Africa/Asia/South-America. Antimicrobial use data and bacterial taxonomy only explains a minor part of the AMR variation that we observe. We find no evidence for cross-selection between antimicrobial classes, or for effect of air travel between sites. However, AMR gene abundance strongly correlates with socio-economic, health and environmental factors, which we use to predict AMR gene abundances in all countries in the world. Our findings suggest that global AMR gene diversity and abundance vary by region, and that improving sanitation and health could potentially limit the global burden of AMR.

DOI:   08-03-2019

Abundance and diversity of the faecal resistome in slaughter pigs and broilers in nine European countries
Munk et al. 2018

Antimicrobial resistance (AMR) in bacteria and associated human morbidity and mortality is increasing. The use of antimicrobials in livestock selects for AMR that can subsequently be transferred to humans. This flow of AMR between reservoirs demands surveillance in livestock and in humans. We quantified and characterized the acquired resistance gene pools (resistomes) of 181 pig and 178 poultry farms from nine European countries, sequencing more than 5,000 Gb of DNA using shotgun metagenomics. We quantified acquired AMR using the ResFinder database and a second database constructed for this study, consisting of AMR genes identified through screening environmental DNA. The pig and poultry resistomes were very different in abundance and composition. There was a significant country effect on the resistomes, more so in pigs than in poultry. We found higher AMR loads in pigs, whereas poultry resistomes were more diverse. We detected several recently described, critical AMR genes, including mcr-1 and optrA, the abundance of which differed both between host species and between countries. We found that the total acquired AMR level was associated with the overall country-specific antimicrobial usage in livestock and that countries with comparable usage patterns had similar resistomes. However, functionally determined AMR genes were not associated with total drug use.

DOI:      23-07-2018

Meta-genomic analysis of toilet waste from long distance flights; a step towards global surveillance of infectious diseases and antimicrobial resistance
Petersen et al. 2015

Human populations worldwide are increasingly confronted with infectious diseases and antimicrobial resistance spreading faster and appearing more frequently. Knowledge regarding their occurrence and worldwide transmission is important to control outbreaks and prevent epidemics. Here, we performed shotgun sequencing of toilet waste from 18 international airplanes arriving in Copenhagen, Denmark, from nine cities in three world regions. An average of 18.6 Gb (14.8 to 25.7 Gb) of raw Illumina paired end sequence data was generated, cleaned, trimmed and mapped against reference sequence databases for bacteria and antimicrobial resistance genes. An average of 106,839 (0.06%) reads were assigned to resistance genes with genes encoding resistance to tetracycline, macrolide and beta-lactam resistance genes as the most abundant in all samples. We found significantly higher abundance and diversity of genes encoding antimicrobial resistance, including critical important resistance (e.g. blaCTX-M) carried on airplanes from South Asia compared to North America. Presence of Salmonella enterica and norovirus were also detected in higher amounts from South Asia, whereas Clostridium difficile was most abundant in samples from North America. Our study provides a first step towards a potential novel strategy for global surveillance enabling simultaneous detection of multiple human health threatening genetic elements, infectious agents and resistance genes.

DOI:       10-07-2015
29 JANUARY 2022