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\nKlebsiella pneumoniae is a common pathogen of healthcare-associated infections expressing a plethora of antimicrobial resistance loci, including ADP-ribosyltransferase coding genes (arr), able to mediate rifampicin resistance. The latter has activity against a broad range of microorganisms by inhibiting DNA-dependent RNA polymerases. This study aims to characterise the arr distribution and genetic context in 138 clinical isolates of K. pneumoniae and correlate these with rifampicin resistance. All isolates were subjected to whole-genome sequencing for species identification, typing and AMR genes identification, along with the determination of the minimum inhibitory concentration (MIC) of rifampicin. Molecular detection of arr genes and class 1 integrons was performed for rifampicin-resistant isolates. Efflux activity was investigated as a possible determinant of rifampicin resistance in isolates devoid of known genetic determinants. Twelve isolates exhibited high rifampicin MICs (≥ 64 mg/L), 124 showed intermediate MICs (16-32 mg/L) and two displayed low (8 mg/L) MICs. Two arr allelic variants, arr-2 and arr-3, were found across one and nine K. pneumoniae isolates, respectively, all within class 1 integrons, including a newly described integron, and all associated with high rifampicin MICs  (≥ 64 mg/L). Elevated resistance levels were additionally linked to increased arr-2/3 expression and closer proximity to the promoter. No arr gene or rpoB mutations were found across the remaining two isolates and no correlation between efflux activity and high-level rifampicin resistance was found for both isolates. In conclusion, this study demonstrates that arr genes confer high levels of rifampicin resistance in K. pneumoniae highlighting its widespread dissemination within class 1 integrons., Competing Interests: Declarations. Competing interests: The authors declare no competing interests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Klebsiella pneumoniae (Kp) bacteria are an increasing threat to public health and represent one of the most concerning pathogens involved in life-threatening infections and antimicrobial resistance (AMR). To understand the epidemiology of AMR of Kp in Portugal, we analysed whole genome sequencing, susceptibility testing and other meta data on 509 isolates collected nationwide from 16 hospitals and environmental settings between years 1980 and 2019. Predominant sequence types (STs) included ST15 (n = 161, 32%), ST147 (n = 36, 7%), ST14 (n = 26, 5%) or ST13 (n = 26, 5%), while 31% of isolates belonged to STs with fewer than 10 isolates. AMR testing revealed widespread resistance to aminoglycosides, fluoroquinolones, cephalosporins and carbapenems. The most common carbapenemase gene was bla KPC-3 . Whilst the distribution of AMR linked plasmids appears uncorrelated with ST, their frequency has changed over time. Before year 2010, the dominant plasmid group was associated with the extended spectrum beta-lactamase gene bla CTX-M-15 , but this group appears to have been displaced by another carrying the bla KPC-3 gene. Co-carriage of bla CTX-M and bla KPC-3 was uncommon. Our results from the largest genomics study of Kp in Portugal highlight the active transmission of strains with AMR genes and provide a baseline set of variants for future resistance monitoring and epidemiological studies., (© 2022. The Author(s).)

The Beijing genotype comprises a highly disseminated strain type that is frequently associated with multidrug resistant (MDR) tuberculosis (TB) and increased transmissibility but, countries such as Portugal and Guinea-Bissau fall outside the regions phylogeographically associated with this specific genotype. Nevertheless, recent data shows that this genotype might be gradually emerging in these two countries as an underlying cause of primary MDR-TB. Here, we describe the emergence of Mycobacterium tuberculosis Beijing strains associated with MDR-TB in Portugal and Guinea-Bissau demonstrating the presence of the well described superclusters 100-32 and 94-32 in Portugal and Guinea-Bissau, respectively. Genome-wide analysis and comparison with a global genomic dataset of M. tuberculosis Beijing strains, revealed the presence of two genomic clusters encompassing isolates from Portugal and Guinea-Bissau, GC1 ( n  = 121) and GC2 ( n  = 39), both of which bore SNP signatures compatible with the 100-32/B0/W148 and 94-32/Central Asia Outbreak clades, respectively. Moreover, GC2 encompasses a cross-border cluster between Portugal, Guinea-Bissau and Brazil thus supporting migration-associated introduction of MDR-TB and subsequent clonal expansion at the community-level. The comparison with global Beijing datasets demonstrates the global reach of the disease and its complex dissemination across multiple countries while in parallel there are clear microevolutionary trajectories towards extensively drug resistant TB.