Project description:Molecular detection and characterization of antibiotic-resistant Escherichia coli isolated from urban and peri-urban gardening systems in Dhaka, Bangladesh
Project description:Molecular detection and characterization of antibiotic resistant Shigella flexneri isolated from urban and peri-urban gardening systems in Dhaka, Bangladesh
Project description:Molecular detection and characterization of antibiotic-resistant Citrobacter freundii isolated from urban and peri-urban gardening systems in Dhaka, Bangladesh
Project description:We applied an RNA capture and amplification technique, Selective Capture of Transcribed Sequences (SCOTS), and microarray hybridization to identify S. Typhi transcripts expressed in the blood of five humans infected with S. Typhi in Bangladesh. In total, we detected the expression of mRNAs for 2046 S. Typhi genes (44% of the S. Typhi genome) in human blood; expression of 912 genes was detected in all 5 patients, and expression of 1100 genes was detected in 4 or more patients. Identified transcripts were associated with the virulence-associated PhoP regulon, Salmonella pathogenicity islands, the use of alternative carbon and energy sources, synthesis and transport of iron, thiamine, and biotin, and resistance to antimicrobial peptides and oxidative stress. The most highly represented group were genes currently annotated as encoding proteins designated as hypothetical, unknown, or unclassified. Of the 2046 detected transcripts, 1320 (29% of the S. Typhi genome) had significantly different levels of detection in human blood compared to in vitro cultures; detection of 141 transcripts was significantly different in all 5 patients, and detection of 331 transcripts varied in at least 4 patients. These mRNAs encode proteins of unknown function, those involved in energy metabolism, transport and binding, cell envelope, cellular processes, and pathogenesis. We confirmed increased expression of a subset of identified mRNAs by quantitative-PCR. We compared transcriptional profiles of S. Typhi from the blood of infected humans to S. Typhi grown in vitro
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.