PathoFact: a pipeline for the prediction of virulence factors and antimicrobial resistance genes in metagenomic data

Microbiome. 2021 Feb 17;9(1):49. doi: 10.1186/s40168-020-00993-9.


Background: Pathogenic microorganisms cause disease by invading, colonizing, and damaging their host. Virulence factors including bacterial toxins contribute to pathogenicity. Additionally, antimicrobial resistance genes allow pathogens to evade otherwise curative treatments. To understand causal relationships between microbiome compositions, functioning, and disease, it is essential to identify virulence factors and antimicrobial resistance genes in situ. At present, there is a clear lack of computational approaches to simultaneously identify these factors in metagenomic datasets.

Results: Here, we present PathoFact, a tool for the contextualized prediction of virulence factors, bacterial toxins, and antimicrobial resistance genes with high accuracy (0.921, 0.832 and 0.979, respectively) and specificity (0.957, 0.989 and 0.994). We evaluate the performance of PathoFact on simulated metagenomic datasets and perform a comparison to two other general workflows for the analysis of metagenomic data. PathoFact outperforms all existing workflows in predicting virulence factors and toxin genes. It performs comparably to one pipeline regarding the prediction of antimicrobial resistance while outperforming the others. We further demonstrate the performance of PathoFact on three publicly available case-control metagenomic datasets representing an actual infection as well as chronic diseases in which either pathogenic potential or bacterial toxins are hypothesized to play a role. In each case, we identify virulence factors and AMR genes which differentiated between the case and control groups, thereby revealing novel gene associations with the studied diseases.

Conclusion: PathoFact is an easy-to-use, modular, and reproducible pipeline for the identification of virulence factors, bacterial toxins, and antimicrobial resistance genes in metagenomic data. Additionally, our tool combines the prediction of these pathogenicity factors with the identification of mobile genetic elements. This provides further depth to the analysis by considering the genomic context of the pertinent genes. Furthermore, PathoFact's modules for virulence factors, toxins, and antimicrobial resistance genes can be applied independently, thereby making it a flexible and versatile tool. PathoFact, its models, and databases are freely available at . Video abstract.

Keywords: Antimicrobial resistance; Bacterial toxins; Bioinformatics; Metagenomics; Microbiome; Mobile genetic elements; Virulence factors.

Publication types

  • Research Support, Non-U.S. Gov't
  • Video-Audio Media

MeSH terms

  • Anti-Bacterial Agents / pharmacology
  • Anti-Infective Agents / pharmacology*
  • Drug Resistance, Bacterial / drug effects
  • Drug Resistance, Bacterial / genetics*
  • Metagenomics*
  • Software*
  • Virulence Factors / genetics*


  • Anti-Bacterial Agents
  • Anti-Infective Agents
  • Virulence Factors