Burkhard Malorny

The genus Arcanobacterium has expanded rapidly, with an increasing number of species described from domestic and wildlife hosts, underscoring the need for clearer delineation of species boundaries. In this study, 12 isolates initially presumed to belong to Arcanobacterium canis were examined using comparative phenotypic and genomic approaches to clarify their taxonomic position. The isolates showed partial phenotypic similarity to the type strain A. canis DSM 25104ᵀ, including consistent haemolysis, but differed in several enzymatic reactions and carbohydrate utilization patterns. Matrix-assisted laser desorption/ionization time-of-flight MS identified all isolates at the genus level, with only one displaying spectral similarity to A. canis . Phylogenetic analysis of five gene targets revealed that although the isolates shared sequence similarity with the type strain A. canis DSM 25104ᵀ, they consistently formed a separate clade. Core-genome phylogeny likewise placed the isolates on an independent branch. Pangenome analysis identified 2,178 gene clusters (58.8% core genes) and demonstrated that the cytolysin-like gene present in the type strain A. canis DSM 25104ᵀ is completely absent from all 12 isolates. Digital DNA–DNA hybridization and average nucleotide identity values fell below accepted species-level thresholds, indicating that the isolates represent a genomically distinct lineage within Arcanobacterium . Because chemotaxonomic analyses required for formal species description were not performed, the taxonomic status of this lineage remains unresolved. Nevertheless, the present findings refine species boundaries within Arcanobacterium and provide a robust foundation for future taxonomic and comparative genomic studies.

Background Bacteriophages are recognized modulators of microbiome composition and function, yet their role in the porcine upper respiratory tract, a primary gateway for pathogen colonization in the post-weaning period, remains unexplored. Unlike the porcine gut, no reference framework is available for respiratory sites. Furthermore, the low-biomass nature of nasopharyngeal specimens makes virome recovery highly sensitive to extraction strategy, but the extent to which workflow choice shapes ecological inference in this niche has not been evaluated. Results We profiled the nasopharyngeal phageome of post-weaning piglets across ten commercial farms (30 pen-level pools) using paired DNA-microbiome (DNA-m) and virus-like particle–enriched (VLP-e) short-read metagenomics (n = 60 libraries). Protocol choice strongly reshaped viral recovery (PERMANOVA R² = 0.448, p < 0.0001), with a contig overlap between workflows of n = 4,357), predicted replication strategy emerged as a dominant organizing axis: lifestyle explained up to 40.6% of compositional variation at family level. Host prediction linked phages to dominant upper-airway colonizers, including Streptococcaceae, Moraxellaceae, Pasteurellaceae, with a marked lifestyle–host polarization: virulent phages were preferentially linked to Bacteroidota (particularly Prevotella), whereas temperate phages were enriched in Streptococcaceae and Moraxellaceae. Integration of viral taxonomy and host affiliation resolved a modular architecture in which a few recurrent phage–host couplings (e.g., Suoliviridae–Bacteroidota, Peduoviridae–Pasteurellaceae, Aliceevansviridae–Streptococcaceae) were conserved but differentially weighted between virulent and temperate fractions. Conclusions This study establishes the first phageome catalogue and ecological framework for a respiratory site in livestock. The nasopharyngeal phageome is organized into recurrent, host-linked taxonomic modules jointly constrained by viral lineage, host affiliation and replication strategy, with lifestyle-dependent connections to key colonizers implicated in the porcine respiratory disease complex. This catalogue and its modular architecture provide a foundation for investigating phage-mediated modulation of bacterial dynamics during the post-weaning transition and for the selection of lytic phage candidates targeting respiratory pathogens.

Salmonella serotyping is shifting from slide agglutination toward whole-genome sequencing (WGS). While WGS allows for comprehensive analyses, phenotypic information about lipopolysaccharide-deficient (“LPS-rough”) isolates obtained from slide agglutination is lost. This discrepancy represents a challenge for Salmonella control in livestock because in the European Union, LPS-rough Salmonella isolated from food-producing animals that are untypable using slide agglutination alone are not subject to control measures, whereas isolates of certain serovars in certain matrices would be, when based on geno-serotyping. Here, we provide an account of the relevance of this phenotype in the context of routine diagnostics and food safety by characterizing the occurrence, diversity, and isolation matrices of LPS-rough isolates among non-human Salmonella enterica subsp. enterica isolates from Germany. Using available WGS data, we examined phylogenetic relationships and associations with certain genomic features. On average, 5% of isolates exhibited an LPS-rough phenotype across 46 serovars, but clonal distribution of this phenotype along the food chain was not evident. LPS-rough isolates were more commonly found in S . Choleraesuis isolated from wild boar and in S . Typhimurium isolated from pork products, compared with other matrices. We also found associations with two virulence factors, an AMR gene, and a plasmid marker. The present work lays the foundation for future research into the role of certain matrices, environments, or genomic factors in the development of LPS-rough Salmonella isolates and will facilitate the elucidation of the genomic basis of this phenotype, which may improve recommendations regarding risk management and control measures. IMPORTANCE: The present work highlights some of the challenges associated with the recent shift from serology- to sequence-based typing of Salmonella enterica serovars and provides a national perspective on the presence and relevance of the lipopolysaccharide-deficient (“LPS-rough”) phenotype in samples obtained from food, animals, and the environment, including considerations regarding the choice of typing method. We provide evidence that clonal distribution of isolates with this phenotype is unlikely, but that certain environments may favor its development, and certain genomic factors may increase survival rates of LPS-rough isolates when exposed to environmental stressors. These findings could have important implications for regulations regarding the surveillance and management of Salmonella isolated from food, feed, and animals in the future, in particular in the context of using different typing methods, and warrant further detailed research.

Previous case reports indicate Arcanobacterium ’s opportunistic pathogenic potential. However, the true diversity of the genus remains understudied. Here, we present the complete genome of Arcanobacterium wilhelmae isolated from a diseased rhinoceros, suspected to play a role in its condition. These genomic data may enable future advancements in understanding Arcanobacterium pathogenicity.

Many species of the genus Arcanobacterium are known as opportunistic pathogens and have been isolated in association with infectious diseases in humans and animals. Here, we present the complete genome sequence of another opportunistic pathogenic representative, namely Arcanobacterium canis , isolated from the otitis externa of an English bulldog.

For successful elucidation of a food-borne infection chain, the availability of high-quality sequencing data from suspected microbial contaminants is a prerequisite. Commonly, those investigations are a joint effort undertaken by different laboratories and institutes. To analyze the extent of variability introduced by differing wet-lab procedures on the quality of the sequence data we conducted an interlaboratory study, involving four bacterial pathogens, which account for the majority of food-related bacterial infections: Campylobacter spp., Shiga toxin-producing Escherichia coli, Listeria monocytogenes, and Salmonella enterica. The participants, ranging from German federal research institutes, federal state laboratories to universities and companies, were asked to follow their routine in-house protocols for short-read sequencing of 10 cultures and one isolated bacterial DNA per species. Sequence and assembly quality were then analyzed centrally. Variations within isolate samples were detected with SNP and cgMLST calling. Overall, we found that the quality of Illumina raw sequence data was high with little overall variability, with one exception, attributed to a specific library preparation kit. The variability of Ion Torrent data was higher, independent of the investigated species. For cgMLST and SNP analysis results, we found that technological sequencing artefacts could be reduced by the use of filters, and that SNP analysis was more suited than cgMLST to compare data of different contributors. Regarding the four species, a minority of Campylobacter isolate data showed the in comparison highest divergence with regard to sequence type and cgMLST analysis. We additionally compared the assembler SPAdes and SKESA for their performance on the Illumina data sets of the different species and library preparation methods and found overall similar assembly quality metrics and cgMLST statistics.

The characterization of Shiga toxin-producing Escherichia coli (STEC) is necessary to assess their pathogenic potential, but isolation of the strain from complex matrices such as milk remains challenging. In previous work, we have shown the potential of long-read metagenomics to characterize eae-positive STEC from artificially contaminated raw milk without isolating the strain. The presence of multiple E. coli strains in the sample was shown to potentially hinder the correct characterization of the STEC strain. Here, we aimed at determining the STEC:commensal ratio that would prevent the characterization of the STEC. We artificially contaminated pasteurized milk with different ratios of an eae-positive STEC and a commensal E. coli and applied the method previously developed. Results showed that the STEC strain growth was better than the commensal E. coli after enrichment in acriflavine-supplemented BPW. The STEC was successfully characterized in all samples with at least 10 times more STEC post-enrichment compared to the commensal E. coli. However, the presence of equivalent proportions of STEC and commensal E. coli prevented the full characterization of the STEC strain. This study confirms the potential of long-read metagenomics for STEC characterization in an isolation-free manner while refining its limit regarding the presence of background E. coli strains.

The genus Arcanobacterium is constantly growing as novel species are identified. In particular, harbor seals have proven to be a common reservoir for bacteria of this genus. Here, we announce the complete genome sequence of another Arcanobacterium species—namely, Arcanobacterium pinnipediorum strain DSM 28752, isolated from a harbor seal.

This study provides the first integral genomic characterization of the resistome and mobilome of sentinel microorganisms for antimicrobial resistance (AMR) surveillance from two different swine production systems. Relevant differences were observed among taxa in the resistomes and mobilomes they harbored, revealing their distinctive risk in AMR dissemination and spread.

Background: We benchmarked sequencing technology and assembly strategies for short-read, long-read, and hybrid assemblers in respect to correctness, contiguity, and completeness of assemblies in genomes of Francisella tularensis. Benchmarking allowed in-depth analyses of genomic structures of the Francisella pathogenicity islands and insertion sequences. Five major high-throughput sequencing technologies were applied, including next-generation “short-read” and third-generation “long-read” sequencing methods. Results: We focused on short-read assemblers, hybrid assemblers, and analysis of the genomic structure with particular emphasis on insertion sequences and the Francisella pathogenicity island. The A5-miseq pipeline performed best for MiSeq data, Mira for Ion Torrent data, and ABySS for HiSeq data from eight short-read assembly methods. Two approaches were applied to benchmark long-read and hybrid assembly strategies: long-read-first assembly followed by correction with short reads (Canu/Pilon, Flye/Pilon) and short-read-first assembly along with scaffolding based on long reads (Unicyler, SPAdes). Hybrid assembly can resolve large repetitive regions best with a “long-read first” approach. Conclusions: Genomic structures of the Francisella pathogenicity islands frequently showed misassembly. Insertion sequences (IS) could be used to perform an evolutionary conservation analysis. A phylogenetic structure of insertion sequences and the evolution within the clades elucidated the clade structure of the highly conservative F. tularensis.