Virulence factors in carbapenem-resistant hypervirulent Klebsiella pneumoniae.

Hypervirulence and carbapenem-resistant have emerged as two distinct evolutionary pathotypes of Klebsiella pneumoniae, with both reaching their epidemic success and posing a great threat to public health. However, as the boundaries separating these two pathotypes fade, we assist a worrisome convergence in certain high-risk clones, causing hospital outbreaks and challenging every therapeutic option available. To better understand the basic biology of these pathogens, this review aimed to describe the virulence factors and their distribution worldwide among carbapenem-resistant highly virulent or hypervirulent K. pneumoniae strains, as well as to understand the interplay of these virulence strains with the carbapenemase produced and the sequence type of such strains. As we witness a shift in healthcare settings where carbapenem-resistant highly virulent or hypervirulent K. pneumoniae are beginning to emerge and replace classical K. pneumoniae strains, a better understanding of these strains is urgently needed for immediate and appropriate response.

First Description of Ceftazidime/Avibactam Resistance in a ST13 KPC-70-Producing Klebsiella pneumoniae Strain from Portugal.

The combination of ceftazidime/avibactam (CZA) is a novel ß-lactam/ß-lactamase inhibitor with activity against Klebsiella pneumoniae carbapenemase (KPC)-producing Enterobacterales. Emerging cases caused by CZA-resistant strains that produce variants of KPC genes have already been reported worldwide. However, to the best of our knowledge, no CZA-resistant strains were reported in Portugal. In September 2019, a K. pneumoniae CZA-resistant strain was collected from ascitic fluid at a surgery ward of a tertiary University Hospital Center in Lisboa, Portugal. The strain was resistant to ceftazidime/avibactam, as well as to ceftazidime, cefoxitin, gentamicin, amoxicillin/clavulanic acid, and ertapenem, being susceptible to imipenem and tigecycline. A hypermucoviscosity phenotype was confirmed by string test. Whole-genome sequencing (WGS) analysis revealed the presence of an ST13 KPC70-producing K. pneumoniae, a KPC-3 variant, differing in two amino-acid substitutions (D179Y and T263A). The D179Y mutation in the KPC Ω-loop region is the most common amino-acid substitution in KPC-2 and KPC-3, further leading to CZA resistance. The second mutation causes a KPC-70 variant in which threonine replaces alanine (T263A). The CZA-resistant strain showed the capsular locus KL3 and antigen locus O1v2. Other important virulence factors were identified: fimbrial adhesins type 1 and type 3, as well as the cluster of iron uptake systems aerobactin, enterobactin, salmochelin, and yersiniabactin included in integrative conjugative element 10 (ICEKp10) with the genotoxin colibactin cluster. Herein, we report the molecular characterization of the first hypervirulent CZA-resistant ST13 KPC-70-producing K. pneumoniae strain in Portugal. The emergence of CZA-resistant strains might pose a serious threat to public health and suggests an urgent need for enhanced clinical awareness and epidemiologic surveillance.

First Outbreak of NDM-1-Producing Klebsiella pneumoniae ST11 in a Portuguese Hospital Centre during the COVID-19 Pandemic.

New Delhi metallo-ß-lactamase (NDM) carbapenemase has been considered a global threat due to its worldwide widespread in recent years. In Portugal, a very low number of infections with NDM-producing Enterobacterales has been reported. A total of 52 strains from 40 patients and 1 environmental sample isolated during COVID-19 pandemic were included in this study. Wholegenome sequencing (WGS) was performed on 20 carbapenemase-producing strains, including 17 NDM-1-producing Klebsiella pneumoniae ST11-KL105 lineage strains, one NDM-1-producing Escherichia coli ST58 strain and one KPC-3-producing K. pneumoniae ST147 strain, recovered from a total of 19 patients. Of interest, also one NDM-1-producing K. pneumoniae ST11-KL105 was collected from the hospital environment. Genome-wide phylogenetic analysis revealed an ongoing dissemination of NDM-1-producing K. pneumoniae ST11 strains (n = 18) with the same genetic features seen across multiple wards. Furthermore, the ST58 E. coli strain, collected from a patient rectal swab that was also colonised with a K. pneumoniae strain, also showed the IncFIA plasmid replicon and the blaNDM-1 gene (preceded by IS30 and followed by genes bleMBL, trpF, dsbC, cutA, groES and groEL). The blaNDM-1 is part of Tn125-like identical to those reported in Poland, Italy and India. The blaKPC-3 K. pneumoniae ST147-KL64 strain has the genetic environment Tn4401d isoform. In conclusion, herein we report the molecular epidemiology, resistome, virulome and mobilome of the first NDM-1 carbapenemase outbreak caused by K. pneumoniae ST11-KL105 lineage during the COVID-19 pandemic in Portugal. Moreover, the outbreak strains characterised included seventeen different patients (infected and colonised) and one environmental sample which also emphasises the role of commensal and hospital environment strains in the dissemination of the outbreak.

Whole-Genome Sequencing Enables Molecular Characterization of Non-Clonal Group 258 High-Risk Clones (ST13, ST17, ST147 and ST307) among Carbapenem-Resistant Klebsiella pneumoniae from a Tertiary University Hospital Centre in Portugal.

The carbapenem-resistant Enterobacterales (CRE) strains have been identified by the World Health Organization as critical priority pathogens in research and development of diagnostics, treatments, and vaccines. However, recent molecular information about carbapenem-resistant K. pneumoniae (CRK) epidemiology in Portugal is still scarce. Thus, this study aimed to provide the molecular epidemiology, resistome, and virulome of CRK clinical strains recovered from a tertiary care hospital centre (2019-2021) using polymerase chain reaction (PCR) and the advanced molecular technique whole-genome sequencing (WGS). PCR amplification of carbapenemase genes was performed in 437 carbapenem-resistant K. pneumoniae strains. The most frequent carbapenemases were: KPC-3 (42%), followed by OXA-181 (20%), GES-5 (0.2%), and NDM-1 (0.2%). Additionally, 10 strains (2%) coproduced KPC-3 and OXA-181, and 1 strain coproduced KPC-3 and OXA-48 (0.2%). The genomic population structure of 68 strains characterized by WGS demonstrated the ongoing dissemination of four main high-risk clones: ST13, ST17, ST147, and ST307, while no clones belonging to the European predominant clonal groups (CG15 and CG258) were found. Moreover, we describe one K. pneumoniae ST39-KL62 that coproduced the NDM-1 carbapenemase and the extended-spectrum beta-lactamase CTX-M-15, and one K. pneumoniae ST29-KL54 GES-5 and BEL-1 coproducer. Furthermore, a high prevalence of iron siderophores were present in all CRK strains, with several strains presenting both colibactin and the hypermucoviscosity phenotype. Thus, the data presented here highlight an uncommon molecular epidemiology pattern in Portugal when compared with most European countries, further supporting the emergence and dissemination of nonclonal group 258 hypervirulent multidrug high-risk clones and the need to promote in-depth hospital molecular surveillance studies.

Luminescence Thermometry on the Route of the Mobile-Based Internet of Things (IoT): How Smart QR Codes Make It Real.

Quick Response (QR) codes are a gateway to the Internet of things (IoT) due to the growing use of smartphones/mobile devices and its properties like fast and easy reading, capacity to store more information than that found in conventional codes, and versatility associated to the rapid and simplified access to information. Challenges encompass the enhancement of storage capacity limits and the evolution to a smart label for mobile devices decryption applications. Organic-inorganic hybrids with europium (Eu3+) and terbium (Tb3+) ions are processed as luminescent QR codes that are able to simultaneously double the storage capacity and sense temperature in real time using a photo taken with the charge-coupled device of a smartphone. The methodology based on the intensity of the red and green pixels of the photo yields a maximum relative sensitivity and minimum temperature uncertainty of the QR code sensor (293 K) of 5.14% · K-1 and 0.194 K, respectively. As an added benefit, the intriguing performance results from energy transfer involving the thermal coupling between the Tb3+-excited level (5D4) and the low-lying triplet states of organic ligands, being the first example of an intramolecular primary thermometer. A mobile app is developed to materialize the concept of temperature reading through luminescent QR codes.