Papers by Kristoffer Kiil
ASM Microbe Online, Abstracts, 2020
It has been hypothesized that, in aggregate, rare variants in coding regions of genes explain a s... more It has been hypothesized that, in aggregate, rare variants in coding regions of genes explain a substantial fraction of the heritability of common diseases. We sequenced the exomes of 1,000 Danish cases with common forms of type 2 diabetes (including body mass index > 27.5 kg/m 2 and hypertension) and 1,000 healthy controls to an average depth of 563. Our simulations suggest that our study had the statistical power to detect at least one causal gene (a gene containing causal mutations) if the heritability of these common diseases was explained by rare variants in the coding regions of a limited number of genes. We applied a series of gene-based tests to detect such susceptibility genes. However, no gene showed a significant association with disease risk after we corrected for the number of genes analyzed. Thus, we could reject a model for the genetic architecture of type 2 diabetes where rare nonsynonymous variants clustered in a modest number of genes (fewer than 20) are respons...
sequencing DTU Orbit (08/12/2018) A Danish Salmonella Bareilly outbreak investigated by the use o... more sequencing DTU Orbit (08/12/2018) A Danish Salmonella Bareilly outbreak investigated by the use of whole genome sequencing In 2012, we saw an increase of the Salmonella serotype Bareilly isolated from human infections. Bareilly is a rare serotype in Denmark, isolated from human infections between 2 and 9 times annually over the last 10 years. As a routine in rare serotypes, we use PFGE as the molecular method for cluster analysis, outbreak investigations, comparison with food and animal isolates as well as making international inquiries. In this case, we found Bareilly with the same PFGE profile from 8 patients. Seven of the cases could be traced back to an unknown food source served at a specific restaurant. At the same time four broiler flocks flocks were tested positive for Bareilly. Bareilly is also rare in the Danish food production, and it was the first time in more than 10 years that Bareilly was isolated in broiler flocks. PFGE was performed on these isolates as well and the...
Additional file 2: Figure S1. Maximum parsimony tree of 105 strains of Salmonella Infantis based ... more Additional file 2: Figure S1. Maximum parsimony tree of 105 strains of Salmonella Infantis based on 28,860 core-genome SNPs with Salmonella Infantis CVM44454 as the reference genome. Branches are labelled with the number of SNP differences. Strains belonging to E-Burst Group (eBG) 31 and 297 are marked in red circles. Figure S2. Q-plots based on probability values (Q) from STRUCTURE analysis of 2311 core-genome SNPs identified in 100 Salmonella Infantis strains with Salmonella Infantis CVM44454 as the reference genome. Genetic clusters are marked with curly brackets and cluster number. A: STRUCTURE analysis on main lineage with 85 strains B: STRUCTURE analysis on distant lineage with 15 strains. Figure S3. Mean evolutionary tree calculated from BEAST analysis with the best-fitted substitution model (GTR-BS-R) on 2311 core-genome SNPs. Branches are coloured according to clusters and branch length correlates with time in years. Figure S4. Maximum parsimony tree of 167 strains of Salmo...
Additional file 1: Table S1. Strain information on 105 strains of Salmonella Infantis. Table S2. ... more Additional file 1: Table S1. Strain information on 105 strains of Salmonella Infantis. Table S2. Q-values from STRUCTURE analysis of 100 strains of Salmonella Infantis.
2 ABSTRACT 1 We describe the design and evaluate the use of a high density oligonuclotide microar... more 2 ABSTRACT 1 We describe the design and evaluate the use of a high density oligonuclotide microarray covering seven 2 sequenced E. coli genomes in addition to several sequenced E. coli plasmids, bacteriophages, 3 pathogenicity islands and virulence genes. Its utility is demonstrated for comparative genomic profiling 4 of two unsequenced strains, O175:H16 D1 and O157:H7 3538 (∆stx2::cat) as well as two well-known 5 control strains, K-12 W3110 and O157:H7 EDL933. By using fluorescently labelled genomic DNA to 6 query the microarrays and subsequently analyse common virulence genes and phage elements, and 7 perform whole genome comparisons, we observed that O175:H16 D1 is a K-12 like strain and 8 confirmed that its φ3538 (∆stx2::cat) phage element originated from the E. coli 3538 (∆stx2::cat) strain 9 with which it shares a substantial proportion of phage elements. Moreover, a number of genes involved 10 in DNA transfer and recombination was identified in both new strains providing a li...
Microorganisms, 2021
Recurrent urinary tract infection (rUTI) remains a major problem for many women and therefore the... more Recurrent urinary tract infection (rUTI) remains a major problem for many women and therefore the pursuit for genomic and phenotypic traits which could define rUTI has been ongoing. The present study applied a genomic approach to investigate recurrent urinary tract infections by comparative analyses of recurrent and non-recurrent Escherichia coli isolates from general practice. From whole-genome sequencing data, phylogenetic clustering and genomic traits were studied on a collection of isolates which caused recurrent infection compared to non-recurrent isolates. In addition, genomic variation between the 1st and following infection was studied on a subset of the isolates. Evidence of limited adaptation between the recurrent infections based on single nucleotide polymorphism analyses with a range of 0–13 non-synonymous single nucleotide polymorphisms (SNPs) between the paired isolates. This included an overrepresentation of SNPs in metabolism genes. We identified several genes which ...
Clinical Infectious Diseases, 2016
Background. Listeriosis is a serious foodborne infection. Outbreaks of listeriosis occur rarely, ... more Background. Listeriosis is a serious foodborne infection. Outbreaks of listeriosis occur rarely, but have often proved difficult to solve. In June 2014, we detected and investigated a listeriosis outbreak in Denmark using patient interviews and whole-genome sequencing (WGS). Methods. We performed WGS on Listeria monocytogenes isolates from patients and available isolates from ready-to-eat foods and compared them using single-nucleotide polymorphism (SNP) analysis. Case patients had L. monocytogenes with ≤3 SNPs (the outbreak strain) isolated in September 2013−December 2014. Through interviews, we established case patients' food and clinical histories. Food production facilities were inspected and sampled, and we performed trace-back/trace-forward of food delivery chains. Results. In total, 41 cases were identified; 17 deaths occurred (41%). An isolate from a delicatessen meat (spiced meat roll) from company A was identical to the outbreak strain. Half of the patients were infected while hospitalized/institutionalized; institutions were supplied food by company A. The outbreak strain was repeatedly isolated from further samples taken within this company and within companies in its distribution chain. Products from company A were traced and recalled from >6000 food establishments, after which the outbreak ended. Conclusions. Ready-to-eat spiced meat roll from a single production facility caused this outbreak. The product, served sliced and cold, is popular among the elderly; serving it at hospitals probably contributed to the high case-fatality rate. WGS used for patient isolates and isolates from food control inspections, coupled with routine epidemiological follow-up, was instrumental in swiftly locating the source of infections, preventing further illnesses and deaths. Keywords. Listeria monocytogenes; outbreak investigation; foodborne diseases; bacterial genome; high-throughput nucleotide sequencing.
Figure 1 – Hits to Campylobacter found using BLAST (red) and Kraken (blue) Detection of Campyloba... more Figure 1 – Hits to Campylobacter found using BLAST (red) and Kraken (blue) Detection of Campylobacter in chicken faecal samples is possible from 106 CFU/g using BLAST (red bars) and from 104 CFU/g using Kraken (blue bars). For BLAST results hits are number of contigs matching Campylobacter in proportion to the total number of contigs. For Kraken results hits are number of reads assigned to Campylobacter in proportion to the total number of reads.
Scientific Reports
Despite phages’ ubiquitous presence and great importance in shaping microbial communities, little... more Despite phages’ ubiquitous presence and great importance in shaping microbial communities, little is known about the diversity of specific phages in different ecological niches. Here, we isolated, sequenced, and characterized 38 Escherichia coli-infecting phages (coliphages) from poultry faeces to gain a better understanding of the coliphage diversity in the poultry intestine. All phages belonged to either the Siphoviridae or Myoviridae family and their genomes ranged between 44,324 and 173,384 bp, with a G+C content between 35.5 and 46.4%. Phylogenetic analysis was performed based on single “marker” genes; the terminase large subunit, portal protein, and exonucleases, as well as the full draft genomes. Single gene analysis resulted in six distinct clusters. Only minor differences were observed between the different phylogenetic analyses, including branch lengths and additional duplicate or triplicate subclustering. Cluster formation was according to genome size, G+C content and pha...
European Journal of Clinical Microbiology & Infectious Diseases
Campylobacter jejuni is recognised as the leading cause of bacterial gastroenteritis in industria... more Campylobacter jejuni is recognised as the leading cause of bacterial gastroenteritis in industrialised countries. Although the majority of Campylobacter infections are self-limiting, antimicrobial treatment is necessary in severe cases. Therefore, the development of antimicrobial resistance (AMR) in Campylobacter is a growing public health challenge and surveillance of AMR is important for bacterial disease control. The aim of this study was to predict antimicrobial resistance in C. jejuni from whole-genome sequencing data. A total of 516 clinical C. jejuni isolates collected between 2014 and 2017 were subjected to WGS. Resistance phenotypes were determined by standard broth dilution, categorising isolates as either susceptible or resistant based on epidemiological cutoffs for six antimicrobials: ciprofloxacin, nalidixic acid, erythromycin, gentamicin, streptomycin, and tetracycline. Resistance genotypes were identified using an in-house database containing reference genes with know...
Genes
In microbial food safety, molecular methods such as quantitative PCR (qPCR) and next-generation s... more In microbial food safety, molecular methods such as quantitative PCR (qPCR) and next-generation sequencing (NGS) of bacterial isolates can potentially be replaced by diagnostic shotgun metagenomics. However, the methods for pre-analytical sample preparation are often optimized for qPCR, and do not necessarily perform equally well for qPCR and sequencing. The present study investigates, through screening of methods, whether qPCR can be used as an indicator for the optimization of sample preparation for NGS-based shotgun metagenomics with a diagnostic focus. This was used on human fecal samples spiked with 10 3 or 10 6 colony-forming units (CFU)/g Campylobacter jejuni, as well as porcine fecal samples spiked with 10 3 or 10 6 CFU/g Salmonella typhimurium. DNA was extracted from the samples using variations of two widely used kits. The following quality parameters were measured: DNA concentration, qPCR, DNA fragmentation during library preparation, amount of DNA available for sequencing, amount of sequencing data, distribution of data between samples in a batch, and data insert size; none showed any correlation with the target ratio of the spiking organism detected in sequencing data. Surprisingly, diagnostic metagenomics can have better detection sensitivity than qPCR for samples spiked with 10 3 CFU/g C. jejuni. The study also showed that qPCR and sequencing results may be different due to inhibition in one of the methods. In conclusion, qPCR cannot uncritically be used as an indicator for the optimization of sample preparation for diagnostic metagenomics.
Emerging Infectious Diseases
C ampylobacter jejuni is the most frequent cause of bacterial gastroenteritis in industrialized c... more C ampylobacter jejuni is the most frequent cause of bacterial gastroenteritis in industrialized countries worldwide (1,2), including in Denmark, where ≈4,000 Campylobacter infections are reported annually (3). Despite the high notification rates, Campylobacter infections are believed to be highly underdiagnosed (4,5). Denmark's national surveillance system for Campylobacter is based on observation of the number of notified human cases (3); a substantial number of Campylobacter outbreaks may be overlooked because of a lack of routine microbiological typing of isolates. This hypothesis is supported by evidence from a recent study, in which whole-genome sequencing (WGS)-based typing of selected clinical Campylobacter isolates from patients in Denmark identified numerous small outbreak-like clusters (6). This finding suggests that more outbreaks occur in Denmark than the few typically large outbreaks associated with a single event that are detected by the current surveillance system (7-10). To achieve a national surveillance system that will detect ongoing Campylobacter outbreaks in real time, highly discriminatory subtyping of isolates is needed. Thus, we more comprehensively evaluated the frequency of Campylobacter outbreaks among humans in Denmark by using WGS-based typing. To match the clinical isolates with their sources, we compared them with isolates from food and animals, covering the main putative sources of human Campylobacter infections (i.e., contact with or consumption of animals or animal products, primarily contaminated poultry meat). Although Campylobacter infections are primarily foodborne, a recent case-control study in Denmark found that contact with animals and the environment might account for a substantial proportion of domestic infections (11). Several other reported sources of infection include unpasteurized milk, drinking water, bathing water, vegetables, and fruits (1,12-14). WGS offers high-resolution discriminatory subtyping and has been successfully implemented for
BMC Genomics
Background: Salmonella Infantis (S. Infantis) is one of the most frequent Salmonella serovars iso... more Background: Salmonella Infantis (S. Infantis) is one of the most frequent Salmonella serovars isolated from human cases of salmonellosis and the most detected serovar from animal and food sources in Europe. The serovar is commonly associated with poultry and there is increasing concern over multidrug resistant clones spreading worldwide, as the dominating clones are characterized by presence of large plasmids carrying multiple resistance genes. Increasing the knowledge of the S. Infantis population and evolution is important for understanding and preventing further spread. In this study, we analysed a collection of strains representing different decades, sources and geographic locations. We analysed the population structure and the accessory genome, in particular we identified prophages with a view to understand the role of prophages in relation to the evolution of this serovar. Results: We sequenced a global collection of 100 S. Infantis strains. A core-genome SNP analysis separated five strains in e-Burst Group (eBG) 297 with a long branch. The remaining strains, all in eBG31, were divided into three lineages that were estimated to have separated approximately 150 years ago. One lineage contained the vast majority of strains. In five of six clusters, no obvious correlation with source or geographical locations was seen. However, one cluster contained mostly strains from human and avian sources, indicating a clone with preference for these sources. The majority of strains within this cluster harboured a pESI-like plasmid with multiple resistance genes. Another lineage contained three genetic clusters with more rarely isolated strains of mainly animal origin, possibly less sampled or less infectious clones. Conserved prophages were identified in all strains, likely representing bacteriophages which integrated into the chromosome of a common ancestor to S. Infantis. We also saw that some prophages were specific to clusters and were probably introduced when the clusters were formed. Conclusions: This study analysed a global S. Infantis population and described its genetic structure. We hypothesize that the population has evolved in three separate lineages, with one more successfully emerging lineage. We furthermore detected conserved prophages present in the entire population and cluster specific prophages, which probably shaped the population structure.
We present CleanRecomb, a tool to quickly filter a SNP matrix for likely recombination events.Met... more We present CleanRecomb, a tool to quickly filter a SNP matrix for likely recombination events.MethodThe method evaluates segments with identical SNP profiles over the genome, based on the assumption that SNPs in the absense of recombination events are uniformly distributed across the genome. The method is evaluated on a set of 9 ST200 E. coli genome sequences.ResultsThe detected recombination events coincide with regions of elevated SNP density.
Whole-genome sequencing is becoming the method of choice but provides redundant data for many tas... more Whole-genome sequencing is becoming the method of choice but provides redundant data for many tasks. ReadFilter (https://github.com/ssi-dk/serum_readfilter) is offered as a way to improve run time of these tasks by rapidly filtering reads against user-specified sequences in order to work with a small fraction of original reads while maintaining accuracy. This can noticeably reduce mapping time and substantially reduce de novo assembly time.
Microbial Genomics
We present the LiSEQ (Listeria SEQuencing) project, funded by the European Food Safety Agency (EF... more We present the LiSEQ (Listeria SEQuencing) project, funded by the European Food Safety Agency (EFSA) to compare Listeria monocytogenes isolates collected in the European Union from ready-to-eat foods, compartments along the food chain (e.g. food-producing animals, food-processing environments) and humans. In this article, we report the molecular characterization of a selection of this data set employing whole-genome sequencing analysis. We present an overview of the strain diversity observed in different sampled sources, and characterize the isolates based on their virulence and resistance profile. We integrate into our analysis the global L. monocytogenes genome collection described by Moura and colleagues in 2016 to assess the representativeness of the LiSEQ collection in the context of known L. monocytogenes strain diversity.
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Papers by Kristoffer Kiil