Highlights d Cities possess a consistent ''core'' set of non-human microbes d Urban microbiomes echo important features of cities and city-life d Antimicrobial resistance genes are widespread in cities d Cities contain many novel bacterial and viral species

One of the most widely used methods to detect an acute viral infection in clinical specimens is diagnostic real-time polymerase chain reaction. However, because of the COVID-19 pandemic, mass-spectrometry-based proteomics is currently being discussed as a potential diagnostic method for viral infections. Because proteomics is not yet applied in routine virus diagnostics, here we discuss its potential to detect viral infections. Apart from theoretical considerations, the current status and technical limitations are considered. Finally, the challenges that have to be overcome to establish proteomics in routine virus diagnostics are highlighted.

In shotgun proteomics, peptide and protein identification is most commonly conducted using database search engines, the method of choice when reference protein sequences are available. Despite its widespread use the database-driven approach is limited, mainly because of its static search space. In contrast, de novo sequencing derives peptide sequence information in an unbiased manner, using only the fragment ion information from the tandem mass spectra. In recent years, with the improvements in MS instrumentation, various new methods have been proposed for de novo sequencing. This review article provides an overview of existing de novo sequencing algorithms and software tools ranging from peptide sequencing to sequence-to-protein mapping. Various use cases are described for which de novo sequencing was successfully applied. Finally, limitations of current methods are highlighted and new directions are discussed for a wider acceptance of de novo sequencing in the community.

Campylobacter is the major bacterial agent of human gastroenteritis worldwide and represents a crucial global public health burden. Species differentiation of C. jejuni and C. coli and phylogenetic analysis is challenged by inter-species horizontal gene transfer. Routine real-time PCR on more than 4000 C. jejuni and C. coli field strains identified isolates with ambiguous PCR results for species differentiation, in particular, from the isolation source eggs. K-mer analysis of whole genome sequencing data indicated the presence of C. coli hybrid strains with huge amounts of C. jejuni introgression. Recombination events were distributed over the whole chromosome. MLST typing was impaired, since C. jejuni sequences were also found in six of the seven housekeeping genes. cgMLST suggested that the strains were phylogenetically unrelated. Intriguingly, the strains shared a stress response set of C. jejuni variant genes, with proposed roles in oxidative, osmotic and general stress defence, chromosome maintenance and repair, membrane transport, cell wall and capsular biosynthesis and chemotaxis. The results have practical impact on routine typing and on the understanding of the functional adaption to harsh environments, enabling successful spreading and persistence of Campylobacter. Since 2005, Campylobacter is the major zoonotic agent in the European Union, causing 250,161 confirmed campylobacteriosis cases in 2017 1. Around one third of the cases can be directly attributed to handling, preparation and consumption of broiler meat 2. Measures for Campylobacter reduction focus on virulence mechanisms and persistence factors, enabling the pathogen to successfully circulate within the food chain. Typing of Campylobacter by species differentiation methods and by multi-locus sequence typing (MLST) has become key tools for diagnostics and source attribution. Specific gene targets have proven stable and were, therefore, chosen for this purpose. Two of commonly used species differentiation markers 3-5 are mapA, a fitness factor in chicken colonization 6 and ceuE playing a role in iron acquisition 7. For MLST, central enzymatic functions, which are conserved in the genome were defined 8 and are commonly used for phylogenetic analysis. It was shown that high level of interspecies transfer of genetic material can occur between C. jejuni and C. coli 9. Adaptation to hosts can modulate the gene pool and allele variants and was suggested to be of more relevance than geographical location 10. Here we identified extensive interspecies gene transfer from C. jejuni to C. coli, impairing species differentiation and MLST analysis. Whole genome sequencing revealed that these hybrid strains shared C. jejuni gene variants, involved in stress response. Since the hybrids had predominantly been isolated from egg shells, we suggest that gene variations due to C. jejuni sequence introgression might have been a consequence of selection of survivors in a harsh environment.