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BackgroundExtended spectrum beta-lactamases-producing (ESBL-P) and/or carbapenem-resistant (CR) Klebsiella pneumoniae have severely restricted available treatment options in healthcare settings in Vietnam. Understanding the diversity and transmission mechanisms of ESBL- and carbapenemase-encoding K. pneumoniae is important in both hospital and community settings for patient management.MethodsWe conducted a 6-month prospective cohort study of 69 Intensive care unit (ICU) patients from two hospitals in Hanoi, Vietnam. Longitudinally collected samples from patients and the ICU environment were cultured on selective media, and 357 K. pneumoniae colonies were whole genome sequenced. We performed phylogenetic analyses, and correlated phenotypic antimicrobial susceptibility testing with genotypic features of K. pneumoniae isolates. We constructed transmission networks of patient samples, relating ICU admission times and locations with genetic similarity of infecting K. pneumoniae.FindingsDespite being geographically and clinically separated, the two hospitals shared closely related strains carrying the same array of antimicrobial resistance genes. Many patients carried the same resistant K. pneumoniae clone from admission to discharge. 45.9% of total isolates carried both ESBL- and carbapenemase-encoding genes, with high minimum inhibitory concentrations. We found a novel co-occurrence of blaKPC-2 and blaNDM-1 in 46. 6% of samples from the globally successful ST15 lineage.InterpretationThese results highlight the high prevalence of ESBL-positive carbapenem-resistant K. pneumoniae in Vietnamese ICUs. Through studying K. pneumoniae ST15 in detail, we illustrated how important resistance genes are coalescing in stains carried broadly by patients entering the two hospitals directly or through referral.FundingThis study was supported by the Medical Research Council Newton Fund, United Kingdom (grant MR/N029399/1); the Ministry of Science and Technology, Vietnam (grant HNQT/SPÐP/04.16); This research was funded in whole by the Wellcome Trust (grant 206194). For the purpose of Open Access, the author has applied a CC BY public copyright licence to any Author Accepted Manuscript version arising from this submission.

Here we describe the development and evaluation of a novel an air-dried high-resolution melt (HRM) assay to detect eight major extended spectrum beta-Lactamase (ESBL) (SHV and CTXM groups 1 and 9) and Carbapenemase (NDM, IMP, KPC, VIM and OXA-48) genes that cause antimicrobial resistance. The assay was evaluated using 440 DNA samples extracted from bacterial isolates from Nepal, Malawi and UK and 390 clinical Enterobacteriaceae isolates with known resistance phenotypes from Nepal. The sensitivity and specificity for detecting the ESBL and Carbapenemase genes in comparison to the reference gel-base PCR and sequencing was 94.7% (95%CI: 92.5%-96.5%) and 99.2% (95%CI: 98.8%-99.5%) and 98.5% (95%CI: 97.0%-99.4%) and 98.5% (95%CI: 98.0%-98.9%) when compared to the original wet format. The overall phenotypic agreement was 91.1% (95%CI: 90.0%-92.9%) on predicting resistance to cefotaxime and carbapenems. We observed good inter-machine reproducibility of the air-dried HRM assay using the Rotor-Gene Q, QuantStudio™ 5, CFX96, LightCycler® 480 and MIC. Assay stability upon storage in the fridge (6.2°C ± 0.9), room temperature (20.35°C ± 0.7) and oven (29.7°C ± 1.4) were assessed at six time points for eight months and no loss of sensitivity occurred under all conditions. We present here a ready-to-use air-dried HRM-PCR assay that offers an easy, thermostable, fast and accurate tool for the detection of ESBL and Carbapenamase genes to improve AMR diagnosis and treatment.

Carriage dynamics of drug-resistant bacteria, especially within households, are poorly understood. This limits the ability to develop effective interventions for controlling the spread of antimicrobial resistance in the community. Two groups consisting of: (i) patients with urinary tract infection requiring antimicrobial treatment; and (ii) patients who were not prescribed antimicrobial treatment were prospectively recruited at three European sites: Antwerp (Belgium), Geneva (Switzerland) and Lodz (Poland). Each index patient and up to three additional household members provided faecal samples at baseline, completion of antimicrobial therapy (or 7-10 days after the first sample for the non-exposed) and 28 days after the second sample. We analysed household-level and individual-level fluoroquinolone resistant Enterobacteriaceae (FQR-E) acquisition and carriage data using Bayesian multi-state Markov models. At the individual level, we estimated a median baseline FQR-E acquisition rate of 0.006 (95%CrI = [0.004, 0.01]) per day, and a median duration of carriage of 24.4 days (95% CrI=[15.23,41.38]). Nitrofurantoin exposure was associated with a reduced rate of FQR-E acquisition (HR=0.28, 95%CrI=[0.14,0.56]), while fluoroquinolone exposure had no clear association with rates of FQR-E acquisition (HR=1.43, 95% CrI=[0.81,2.53]) at individual level. There was evidence that rates of FQR-E acquisition varied by site, and coming from Lodz was associated with a higher acquisition rate (HR=3.56, 95% CrI=[1.92, 6.34]). Prolonged duration of carriage was associated with exposure to fluoroquinolone or nitrofurantoin during the study, use of any antimicrobial agent in the prior 12 months and travel to endemic regions. At household level, we found strong evidence of positive association between FQR-E acquisition and fluoroquinolone exposure (HR=3.43, 95% CrI=[1.51,7.74]). There was weak evidence of negative association between FQR-E acquisition and nitrofurantoin exposure (HR=0.42, 95%CrI=[0.12, 1.24]. Similar to the individual level, carriage duration was also associated with antimicrobial exposure at the household level. Our study has identified within household contacts as an important route for FQR-E transmission and highlights the need for prioritising household focused interventions to control FQR-E spread.

A suspected outbreak of multi-drug resistant (MDR) Klebsiella pneumoniae in a Malawian neonatal unit was investigated using whole-genome sequencing. Strain-types, virulence and resistance genes of K. pneumoniae isolated from patients from the hospital over a four-year period were identified. A MDR ST340 clone was implicated as the likely outbreak cause.