Michael Pietsch

Salmonella enterica subsp. enterica serovar Enteritidis ( S . Enteritidis) is the second most common serovar causing human salmonellosis in Aotearoa New Zealand (NZ). Sequence type (ST)183, which includes phage types 9a and 11, was the second most frequently isolated S . Enteritidis strain from human cases between 2020 and 2023. This ST is considered endemic in NZ as well as in mainland Europe and Great Britain, where the European hedgehog ( Erinaceus europaeus ) is a recognized wildlife reservoir. Hedgehogs were introduced to NZ in the late 19th century; however, their role in the ecology of ST183 in NZ has not been formally evaluated. The aim of this study was to investigate whether hedgehogs act as a reservoir for S . Enteritidis ST183 in NZ and to assess the evolutionary history and epidemiology of this strain across human, animal and environmental contexts. We analysed human, animal and environmental ST183 isolates, including hedgehog carcasses opportunistically sampled in NZ, using Bayesian phylogenetic methods integrated with national epidemiological data. Although S . Enteritidis ST183 was isolated from three of 45 hedgehog carcasses during our study, consistent with recent prior detections of ST183 in NZ hedgehogs, Bayesian phylogenetic analysis supports a most recent common ancestor for currently circulating ST183 strains in the late 20th century, ~100 years after the introduction of hedgehogs into NZ, providing no evidence that the strain was introduced concomitantly with hedgehogs. Epidemiological analysis revealed that, unlike in Europe, ST183 infections in NZ are more common in people aged over 60 years compared with non-ST183 S . Enteritidis infections, with rural residence and contact with farm animals identified as key risk factors. Together, these findings suggest that S . Enteritidis ST183 is established within the NZ rural environment, with evidence of interspecies transmission. While hedgehogs may contribute to the maintenance of ST183, they are unlikely to represent the original source of introduction, indicating a complex, multi-host ecology.

Notifications of Salmonella Strathcona infections increased in Europe in 2023 prompting a multi-country outbreak investigation. We aimed to describe the epidemiology of S . Strathcona infections in 17 European countries 2011–2024, investigate the genetic relatedness of S . Strathcona isolates and identify the vehicle. Cases were persons residing in the study area and with a laboratory-confirmed S . Strathcona infection 2011–2024. Confirmed cases had a S . Strathcona isolate clustering with the outbreak reference strain in core genome multilocus sequence typing (cgMLST) within 7 allelic differences (AD) and possible cases within 8–13 AD. Probable cases had an epidemiological link to a confirmed case and non-outbreak cases had an isolate > 13 AD from the outbreak reference strain. Since 2011, 662 S . Strathcona infections have been identified: 469 confirmed, 161 probable, 13 possible and 19 non-outbreak cases. Median age of the cases was 34 years (IQR: 19–58 years) and 306 (47.5%) were notified in 2023–2024. Most sequenced isolates (469/496; 94.5%) were highly genetically related (≤ 7 AD) over time and across countries, compatible with a common source. Epidemiological and traceback investigations identified small tomatoes from Sicily as the suspect food vehicle. Stringent control measures at the source are needed to stop the contamination and prevent future cases.

The enteric pathogen Salmonella enterica serovar Typhimurium relies on the activity of effector proteins to invade, replicate, and disseminate into host epithelial cells and other tissues, thereby causing disease. Secretion and injection of effector proteins into host cells is mediated by dedicated secretion systems, which hence represent major virulence determinants. Here, we report the identification of a synthetic small molecule with drug-like properties, C26, which suppresses the secretion of effector proteins and consequently hinders bacterial invasion of eukaryotic cells. C26 binds to and inhibits HilD, the transcriptional regulator of the major secretion systems. Although sharing the same binding pocket as the previously described long-chain fatty acid ligands, C26 inhibits HilD with a unique binding mode and a distinct mechanism. We provide evidence of intramacrophage activity and present analogs with improved potency and suitability as scaffolds to develop antivirulence agents against Salmonella infections in humans and animals.

Non-typhoidal Salmonella enterica is an important gastrointestinal pathogen causing a considerable burden of disease. Resistance to third generation cephalosporins poses a serious threat for treatment of severe infections. In this study occurrence, phylogenetic relationship, and mechanisms of third generation cephalosporin resistance were investigated for clinical non-typhoidal S. enterica isolates in Germany. From 2017 to 2019, we detected 168 unique clinical S. enterica isolates with phenotypic resistance to third generation cephalosporins in a nation-wide surveillance. Compared to previous years, we observed a significant (P=0.0002) and consistent increase in resistant isolates from 0.41 % in 2005 to 1.71 % in 2019. In total, 34 different serovars were identified, most often S. Infantis (n=41; 24.4 %), S. Typhimurium (n=27; 16.1 %), S. Kentucky (n=21; 12.5 %), and S. Derby (n=17; 10.1 %). Whole genome analyses revealed extended-spectrum β-lactamase (ESBL) genes as main cause for third generation cephalosporin resistance, and most prevalent were bla _CTX-M-1 (n=55), bla _CTX-M-14 (n=25), and bla _CTX-M-65 (n=23). There was no strict correlation between serovar, phylogenetic lineage, and ESBL type but some serovar/ESBL gene combinations were detected frequently, such as bla _CTX-M-1 and bla _CTX-M-65 in S. Infantis or bla _CTX-M-14b in S. Kentucky. The ESBL genes were mainly located on plasmids, including IncI, IncA/C variants, emerging pESI variants, and a novel bla _CTX-M-1harbouring plasmid. We conclude that third generation cephalosporin resistance is on the rise among clinical S. enterica isolates in Germany, and occurrence in various S. enterica serovars is most probably due to multiple acquisition events of plasmids.

The aim of this study was to gain an overview of the genetic diversity of Salmonella found in wildlife in Germany. We were particularly interested in exploring whether wildlife acts as a reservoir of certain serovars/subtypes or antimicrobial resistance (AMR) genes. Moreover, we wanted to explore the potential of Salmonella in spreading from wildlife to livestock and humans. To answer these questions, we sequenced 260 Salmonella enterica subsp. enterica isolates sampled between 2002 and 2020 from wildlife across Germany, using short-read whole genome sequencing. We found, consistent with previous findings, that some Salmonella sequence types are associated with certain animal species, such as S. Choleraesuis ST145 with wild boar and S. Enteritidis ST183 with hedgehogs. Antibiotic resistance was detected in 14.2% of all isolates, with resistance against important WATCH group antibiotics present in a small number of isolates. We further found that wildlife isolates do not form separate phylogenetic clusters distant to isolates from domestic animals and foodstuff, thus indicating frequent transmission events between these reservoirs. Overall, our study shows that Salmonella in German wildlife are diverse, with a low AMR burden and close links to Salmonella populations of farm and food-production environments.

(1) Background: Resistance plasmids are under selective conditions beneficial for the bacterial host, but in the absence of selective pressure, this carriage may cause fitness costs. Compensation of this fitness burden is important to obtain competitive ability under antibiotic-free conditions. In this study, we investigated fitness effects after a conjugative transfer of plasmids containing various beta-lactamase genes transferred into Escherichia coli. (2) Methods: Fourteen beta-lactamase-encoding plasmids were transferred from clinical donor strains to E. coli J53. Growth rates were compared for all transconjugants and the recipient. Selected transconjugants were challenged in long-term growth experiments. Growth rates were assessed at different time points during growth for 500 generations. Whole-genome sequencing (WGS) of initial and evolved transconjugants was determined. Results: Most plasmid acquisitions resulted in growth differences, ranging from −4.5% to 7.2%. Transfer of a single bla_CMY-16-carrying plasmid resulted in a growth burden and a growth benefit in independent mating. Long-term growth led to a compensation of fitness burdens and benefits. Analyzing WGS revealed genomic changes caused by Single Nucleotide Polymorphisms (SNPs) and insertion sequences over time. Conclusions: Fitness effects associated with plasmid acquisitions were variable. Potential compensatory mutations identified in transconjugants’ genomes after 500 generations give interesting insights into aspects of plasmid–host adaptations.

One of the most demanding challenges in infection control is the worldwide dissemination of multidrug-resistant (MDR) bacteria in clinical settings. Especially the increasing prevalence of carbapenemase producing Gram-negative pathogens poses an urgent threat to public health, as these enzymes confer resistance to almost all β-lactam antibiotics including carbapenems. In this study, we report a prolonged nosocomial outbreak of various NDM-1-producing Enterobacterales species due to clonal spread and cross-species exchange of plasmids and possibly transposons. Between July 2015 and September 2017, a total of 51 carbapenemase-positive isolates were collected from 38 patients and three environmental sources in a single German hospital. Combining molecular typing methods and whole genome sequencing, the metallo-β-lactamase gene bla_NDM–1 was found to be present in 35 isolates of which seven additionally carried the carbapenemase gene bla_KPC–2. Core genome MLST (cgMLST) revealed different clusters of closely related isolates of Escherichia coli, Klebsiella pneumoniae, Citrobacter freundii, Morganella morganii or Enterobacter cloacae indicating clonal spread. The detailed reconstruction of the plasmid sequences revealed that in all outbreak-associated isolates bla_NDM–1 was located on similar composite transposons, which were also very similar to Tn125 previously described for Acinetobacter baumannii. In contrast to Tn125, these structures were flanked by IS26 elements, which could facilitate horizontal gene transfer. Moreover, the identical plasmid was found to be shared by E. coli and M. morganii isolates. Our results highlight the importance of detailed genome-based analyses for complex nosocomial outbreaks, allowing the identification of causal genetic determinants and providing insights into potential mechanisms involved in the dissemination of antibiotic resistances between different bacterial species.