Project description:Antimicrobial resistance (AMR) is an increasing challenge for therapy and management of bacterial infections. Currently, antimicrobial resistance detection relies on phenotypic assays, which are performed independently of species identification. On the contrary, phenotypic prediction from molecular data using genomics is gaining interest in clinical microbiology and might become a serious alternative in the future. Although, in general protein analysis should be superior to genomics for phenotypic prediction, no untargeted proteomics workflow specifically related to AMR detection has been proposed so far. In this study, we present a universal proteomics workflow to detect the bacterial species and antimicrobial resistance related proteins in the absence of secondary antibiotic cultivation in less than 4 h from a primary culture. The method was validated using a sample cohort of 7 bacterial species and 11 AMR determinants represented by 13 protein isoforms which resulted in a sensitivity of 92 % (100 % with vancomycin inference) and a specificity of 100 % with respect to AMR determinants. This proof-of concept study demonstrates the high potential of untargeted proteomics for clinical microbiology.
Project description:Antimicrobial resistance (AMR) is a major public health threat of global proportions, which has the potential to lead to approximately ten million deaths per year by 2050. Pressured by this wicked problem, in 2014, the World Health Organization launched a call for member states to share AMR data through the implementation of the Global Antimicrobial Resistance Surveillance System (GLASS), to appropriately scale and monitor the general situation world-widely. In 2017, Brazil joined GLASS and, in 2018, started its own national antimicrobial surveillance program (BR-GLASS) to understand the impact of resistance in the country. We compiled data obtained from the complete routine of three hospitals' microbiology labs during the year of 2018. This pilot data sums up to 200,874 antimicrobial susceptibility test results from 11,347 isolates. It represents 119 different microorganisms recovered from 44 distinct types of clinical samples. Specimens came from patients originating from 301 Brazilian cities, with 4,950 of these isolates from presumed Healthcare-Associated Infections (HAIs) and the other 6,397 community-acquired cases. The female population offered 58% of the collected samples, while the other 42% were of male origin. The urinary tract was the most common topography (6,372/11,347 isolates), followed by blood samples (2,072/11,347). Gram-negative predominated the bacterial isolates: Escherichia coli was the most prevalent in general, representing 4,030 isolates (89.0% of these from the urinary tract). Coagulase-negative Staphylococci were the most prevalent bacteria in blood samples. Besides these two species, the ESKAPE group have consolidated their prevalence. Regarding drug susceptibility results, 141,648 (70.5%) were susceptible, 9,950 (4.9%) intermediate, and 49,276 (24.5%) resistant. Acinetobacter baumannii was the most worrisome microorganism, with 65.3% of the overall antimicrobial susceptibility tests showing resistance, followed by ESBL-producing Klebsiella pneumoniae, with a global resistance rate of 59%. Although this is a pilot project (still limited to one state), this database shows the importance of a nation-wide surveillance program,[153mm][-12mm] Q14 especially considering it already had patients coming from 301 distinct counties and 18 different states. The BR-GLASS Program is an ongoing project that intends to encompass at least 95 hospitals distributed in all five geographical regions in Brazil within the next 5 years.
Project description:Antimicrobial resistance is a global health concern; Enterobacterales resistant to third-generation cephalosporins (3GCs) and carbapenems are of the highest priority. Here, we conducted genome sequencing and standardized quantitative antimicrobial susceptibility testing of 4,195 isolates of Escherichia coli and Klebsiella pneumoniae resistant to 3GCs and Enterobacterales with reduced meropenem susceptibility collected across Japan. Our analyses provided a complete classification of 3GC resistance mechanisms. Analyses with complete reference plasmids revealed that among the blaCTX-M extended-spectrum β-lactamase genes, blaCTX-M-8 was typically encoded in highly similar plasmids. The two major AmpC β-lactamase genes were blaCMY-2 and blaDHA-1. Long-read sequencing of representative plasmids revealed that approximately 60% and 40% of blaCMY-2 and blaDHA-1 were encoded by such plasmids, respectively. Our analyses identified strains positive for carbapenemase genes but phenotypically susceptible to carbapenems and undetectable by standard antimicrobial susceptibility testing. Systematic long-read sequencing enabled reconstruction of 183 complete plasmid sequences encoding three major carbapenemase genes and elucidation of their geographical distribution stratified by replicon types and species carrying the plasmids and potential plasmid transfer events. Overall, we provide a blueprint for a national genomic surveillance study that integrates standardized quantitative antimicrobial susceptibility testing and characterizes resistance determinants.
Project description:National networks of laboratory-based surveillance of antimicrobial resistance (AMR) monitor resistance trends and disseminate these data to AMR stakeholders. Whole-genome sequencing (WGS) can support surveillance by pinpointing resistance mechanisms and uncovering transmission patterns. However, genomic surveillance is rare in low- and middle-income countries. Here, we implement WGS within the established Antimicrobial Resistance Surveillance Program of the Philippines via a binational collaboration. In parallel, we characterize bacterial populations of key bug-drug combinations via a retrospective sequencing survey. By linking the resistance phenotypes to genomic data, we reveal the interplay of genetic lineages (strains), AMR mechanisms, and AMR vehicles underlying the expansion of specific resistance phenotypes that coincide with the growing carbapenem resistance rates observed since 2010. Our results enhance our understanding of the drivers of carbapenem resistance in the Philippines, while also serving as the genetic background to contextualize ongoing local prospective surveillance.