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.

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In Staphylococcus aureus , resistance to β-lactamase stable β-lactam antibiotics is mediated by the penicillinbinding protein 2a, encoded by mecA or by its homologues mecB or mecC. However, a substantial number of meticillin-resistant isolates lack known mec genes and, thus, are called meticillin resistant lacking mec (MRLM). This study aims to identify the genetic mechanisms underlying the MRLM phenotype. A total of 141 MRLM isolates and 142 meticillin-susceptible controls were included in this study. Oxacillin and cefoxitin minimum inhibitory concentrations were determined by broth microdilution and the presence of mec genes was excluded by PCR. Comparative genomics and a genome-wide association study (GWAS) approach were applied to identify genetic polymorphisms associated with the MRLM phenotype. The potential impact of such mutations on the expression of PBP4, as well as on cell morphology and biofilm formation, was investigated. GWAS revealed that mutations in gdpP were significantly associated with the MRLM phenotype. GdpP is a phosphodiesterase enzyme involved in the degradation of the second messenger cyclic-di-AMP in S. aureus . A total of 131 MRLM isolates carried truncations, insertions or deletions as well as amino acid substitutions, mainly located in the functional DHH-domain of GdpP. We experimentally verified the contribution of these gdpP mutations to the MRLM phenotype by heterologous complementation experiments. The mutations in gdpP had no effect on transcription levels of pbp4; however, cell sizes of MRLM strains were reduced. The impact on biofilm formation was highly strain dependent. We report mutations in gdpP as a clinically relevant mechanism for β-lactam resistance in MRLM isolates. This observation is of particular clinical relevance, since MRLM are easily misclassified as MSSA (meticillin-susceptible S. aureus ), which may lead to unnoticed spread of β-lactam-resistant isolates and subsequent treatment failure.

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Stocking Walleye Sander vitreus is a common management tool to augment populations where natural reproduction is limited. Some hatcheries have progressively raised larger fingerling Walleye to improve poststocking survival; however, little is known about the poststocking survival and behavior of large fingerling Walleye. We sought to evaluate the poststocking daily apparent survival, depth use, dispersal, and home range size of large fingerling Walleye (>200 mm TL) in three Iowa, USA, lakes. Walleye (209–265 mm; n = 15 per lake [45 fish total]) were implanted with radio tags, stocked on October 26–30, 2017, and tracked until May 30, 2018. Cormack–Jolly–Seber recapture models estimated that Walleye apparent survival increased with days poststocking and fish length, resulting in 76% (95% CI = 44–89%) cumulative survival by May. Walleye in Brushy Creek Lake were located in deeper water (mean ± SE = 5.1 ± 0.2 m) than those in Big Creek Lake (3.3 ± 0.2 m) or East Okoboji Lake (1.7 ± 0.1 m), but depth use did not vary with days poststocking. Walleye dispersed an average of 1,355 ± 234 m within 13 d across all lakes, with home range size being larger in Big Creek Lake (mean ± SE = 67.9 ± 21.7 ha) than in Brushy Creek Lake (15.5 ± 15.7 ha) or East Okoboji Lake (31.0 ± 14.0 ha). Our results indicate that Walleye poststocking survival is high overall, with most mortality occurring within 20 d as Walleye are dispersing, suggesting that managers should focus on improving survival during this critical period to improve stocking success.