Testing assembly strategies of Francisella tularensis genomes to infer an evolutionary conservation analysis of genomic structures

Neubert, Kerstin; Zuchantke, Eric; Leidenfrost, Robert Maximilian; Wuenschiers, Roebbe; Grützke, Josephine; Malorny, Burkhard; Brendebach, Holger; Dahouk, Sascha Al; Homeier, Timo; Hotzel, Helmut; Reinert, Knut; Tomaso, Herbert; Busch, Anne

doi:10.1186/s12864-021-08115-x

Cited by 5 publications

(2 citation statements)

References 148 publications

(107 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Any correlation between sequencing depth or evenness with assembly quality in plastid genomes may, therefore, be compounded by the idiosyncrasies of specific NGS platforms and assembly software tools. Given that most NGS sequencing platforms exhibit idiosyncratic error rates between 0.1% and 1% of the generated nucleotides Fox et al [33], a non-trivial number of the sequence reads underlying a genome assembly may represent erroneous data, possibly causing a distinct variation in sequencing depth or evenness [11,34]. Similarly, many of the various software tools for de novo plastid genome assembly [35,36] cannot assemble a complete plastid genome directly, but generate long sequence contigs that must still be post-processed and concatenated to obtain a complete plastome assembly [37].…”

Section: Introductionmentioning

confidence: 99%

Depth and evenness of sequencing coverage are associated with assembly quality, genome structure, and choice of sequencing platform in archived plastid genomes

Jenke,

Smith,

Gruenstaeudl

2024

Preprint

View full text Add to dashboard Cite

Background: Depth and evenness of sequencing coverage are considered important indicators of assembly quality. In plastid genomics, where new data generation has outstripped the development of suitable assembly quality indicators, these coverage metrics could provide valuable insights into the quality of plastomes of different size, structure, and evolutionary origin. However, the typical variation of sequencing depth and evenness among archived plastid genomes, the differences of these metrics among individual plastome sections, and any association with methodological factors have yet to be evaluated. Methods: This investigation aims to explore the variation of sequencing depth and sequencing evenness across publicly accessible plastid genomes as well as the potential associations that these metrics have with plastome structure, assembly quality, and methodological provenance. Specifically, we assess if sequencing evenness and reduced sequencing depth have significant correlations with, or significant differences among, individual genome sections, assembly quality indicators, the sequencing platforms employed, and the assembly software tools used. To that end, we retrieve published plastid genomes, their sequence reads, and their genome metadata from public sequence databases, measure sequencing depth and evenness across genome partitions and full-length genomes, and test several hypotheses regarding genome structure, assembly quality, and methodological provenance through non-parametric statistical tests. Results: The results of our analyses indicate significant differences in sequencing depth across the four structural partitions as well as between the coding and non-coding sections of the plastid genomes, a significant correlation between sequencing evenness and the number of ambiguous nucleotides per genome, and significant differences in sequencing evenness across various sequencing platforms. Conclusions: Based on these results, we conclude that sequencing depth may exhibit a variation that is associated with the quadripartite structure and the assembly quality of plastid genomes, and that sequencing evenness in plastid genomes is likely influenced by the type of sequencing platform employed.

show abstract

Section: Introductionmentioning

confidence: 99%

Depth and evenness of sequencing coverage are associated with assembly quality, genome structure, and choice of sequencing platform in archived plastid genomes

Jenke,

Smith,

Gruenstaeudl

2024

Preprint

View full text Add to dashboard Cite

show abstract

“…Following publication of the original article [ 1 ], it was reported that Röbbe Wünschiers’ name was misspelled as Roebbe Wuenschiers.…”

mentioning

confidence: 99%

Correction to: Testing assembly strategies of Francisella tularensis genomes to infer an evolutionary conservation analysis of genomic structures

et al. 2021

Self Cite

View full text Add to dashboard Cite

Benchmarking short and long read polishing tools for nanopore assemblies: achieving near-perfect genomes for outbreak isolates

Luan,

Commichaux,

Hoffmann

et al. 2024

BMC Genomics

View full text Add to dashboard Cite

Background Oxford Nanopore provides high throughput sequencing platforms able to reconstruct complete bacterial genomes with 99.95% accuracy. However, even small levels of error can obscure the phylogenetic relationships between closely related isolates. Polishing tools have been developed to correct these errors, but it is uncertain if they obtain the accuracy needed for the high-resolution source tracking of foodborne illness outbreaks. Results We tested 132 combinations of assembly and short- and long-read polishing tools to assess their accuracy for reconstructing the genome sequences of 15 highly similar Salmonella enterica serovar Newport isolates from a 2020 onion outbreak. While long-read polishing alone improved accuracy, near perfect accuracy (99.9999% accuracy or ~ 5 nucleotide errors across the 4.8 Mbp genome, excluding low confidence regions) was only obtained by pipelines that combined both long- and short-read polishing tools. Notably, medaka was a more accurate and efficient long-read polisher than Racon. Among short-read polishers, NextPolish showed the highest accuracy, but Pilon, Polypolish, and POLCA performed similarly. Among the 5 best performing pipelines, polishing with medaka followed by NextPolish was the most common combination. Importantly, the order of polishing tools mattered i.e., using less accurate tools after more accurate ones introduced errors. Indels in homopolymers and repetitive regions, where the short reads could not be uniquely mapped, remained the most challenging errors to correct. Conclusions Short reads are still needed to correct errors in nanopore sequenced assemblies to obtain the accuracy required for source tracking investigations. Our granular assessment of the performance of the polishing pipelines allowed us to suggest best practices for tool users and areas for improvement for tool developers.

show abstract

Testing assembly strategies of Francisella tularensis genomes to infer an evolutionary conservation analysis of genomic structures

Cited by 5 publications

References 148 publications

Depth and evenness of sequencing coverage are associated with assembly quality, genome structure, and choice of sequencing platform in archived plastid genomes

Depth and evenness of sequencing coverage are associated with assembly quality, genome structure, and choice of sequencing platform in archived plastid genomes

Correction to: Testing assembly strategies of Francisella tularensis genomes to infer an evolutionary conservation analysis of genomic structures

Benchmarking short and long read polishing tools for nanopore assemblies: achieving near-perfect genomes for outbreak isolates

Contact Info

Product

Resources

About