New Delhi Metallo-Beta-Lactamase (NDM) plasmid predictions
The complete detection and genome sequencing of resistance plasmids is required to understand the co-occurrence of resistance genes on plasmids and their transmission, which is necessary to mitigate the spread of drug resistant bacterial infections. One group of multi-drug resistance plasmids of interest are the New Delhi Metallo-beta-lactamase (NDM) associ-ated plasmids. Bacteria pathogens with NDM genes are resistant to a broad range of beta-lactam antibiotics, including carbapenems, a mainstay for treating bacterial infections. We sequenced clinical isolates collected from patients that failed to respond to antibiotic treatments in the Hamilton-Niagara community using Next Generation Sequencing (NGS) technology, which is the standard technique for sequencing bacterial genomes. I analyzed their assembled genomes via the development of a new resistance plasmid prediction bioinformatics pipeline: RGI:Mobilome. RGI:Mobilome predicted NDM bearing plasmids in 15 isolates, along with other resistance genes on plasmids. However, the NDM plasmid predictions of all 15 isolates were fragmented due to incomplete genome assemblies, caused by repetitive sequences within bacterial genomes and use of short-read NGS technology. However, plasmid predictions greatly improved when we leveraged the high nucleotide accuracy of NGS reads and the structural resolving power of long reads generated with the Oxford Nanopore Technology (ONT) to produce ‘hybrid’ genome assemblies. From the 15 putative hybrid assembly NDM plasmids, one plasmid was a complete match to a known pKP-NDM1 plasmid, seven plasmids were “variants” of known and well characterized plasmids, and the remaining seven plasmids were “distant homologs” of known and well characterized plasmids, suggesting previously undescribed NDM-associated plasmids in our community. This thesis project reveals the diversity of NDM plasmid types within the Hamilton-Niagara community, which is valuable to epidemiologists and public health practitioners to devise actionable plans required to mitigate the spread of multi-drug resistance NDM plasmids and for clinicians to treat infections caused by NDM positive strains.
The growing challenge of microbial resistance emphasizes the importance of new antibiotics or reviving strategies for the use of old ones. Macrolide antibiotics are potent bacterial protein synthesis inhibitors with a formidable capacity to treat life-threatening bacterial infections, however, acquired and intrinsic resistance limits their clinical application. In the work presented here, we reveal that bicarbonate is a potent enhancer of the activity of macrolide antibiotics that overcomes both acquired and intrinsic resistance mechanisms. With a focus on azithromycin, a highly prescribed macrolide antibiotic, and using clinically relevant pathogens, we show that physiological concentrations of bicarbonate overcome drug resistance by increasing the intracellular concentration of azithromycin. We demonstrate the potential of bicarbonate as a formulation additive for topical use of azithromycin in treating a murine wound infection caused by Pseudomonas aeruginosa. Further, using a systemic murine model of methicillin-resistant Staphylococcus aureus infection, we demonstrate the potential role of physiological bicarbonate, naturally abundant in the host, to enhance the activity of azithromycin against macrolide-resistant MRSA. In all, our findings suggest that macrolide resistance, observed in the clinical microbiology laboratory using standard culturing techniques, is a poor predictor of efficacy in the clinic and that observed resistance should not necessarily hamper the use of macrolides. Whether as a formulation additive for topical use or as a natural component of host tissues, bicarbonate is a powerful potentiator of macrolides with the capacity to overcome drug resistance in life-threatening bacterial infections.
The McArthur lab welcomes back summer students Rachel Tran & Arman Edalatmand plus first timers Marcel Jansen & Emily Panousis! These four will be covering a lot of ground this summer, including AMR transmission dynamics, machine learning for automated bio curation, prediction of antibiotic production in Streptomycetes, algorithm development, and data harmonization.
Welcome Ashraf Bazan, who has joined the lab and the McMaster Biochemistry & Biomedical Sciences graduate program! A familiar face in the IIDR as he started if the Coombes lab, Ashraf joins us to lead an antimicrobial resistance (AMR) metagenomics investigation of azithromycin treatment of childhood diarrheal disease in Botswana, in collaboration with Dr. Jeffrey Pernica. Welcome Ashraf!
Rachel Tran | Biochem 4T15 – Exploring the diversity of clinical multidrug resistance in Hamilton, Canada
Arman Edalatmand | Biochem 4T15 – Improving upon CARD*Shark and contextualizing antimicrobial resistance genes
Marcel Jansen | Biochem 4T15 – Translating the Comprehensive Antimicrobial Resistance Database to act as a stopgap for MEGARes
William Huynh | Biochem 4T15 – Expansion of Resistance Gene Identifier allows the identification of putative and novel frameshift mutations
Sohaib Syed | BiomedDC 4A15 – Expanding the scope of antimicrobial resistance surveillance of Mycobacterium tuberculosis in the Comprehensive Antibiotic Resistance Database
Thanks also goes to our 3rd year students Corie Niu (Biochem 3R06), Hamna Imtiaz (Biochem 3R06), Anna-Lisa Nguyen (HthSci 4D03), Sarah Yaqoob (Science 3RP3), & Hafsa Omer (LifeSci 3RP3)!
Quick update on the status of the McArthur Lab and the Comprehensive Antibiotic Resistance Database. While our home institution McMaster University is closed to on-site research and undergraduate/graduate teaching, all McArthur Lab members and CARD staff are working from home. However, like many genomics labs in Canada we are directly helping our clinical colleagues in analysis of SARS-CoV-2 sequences, lending processing power, staff time, and expertise. Response time for CARD might be a bit slow as we are stretched a bit thin. But on May 1st, 2020 we will be joined by William Huynh as Junior CARD Curator & Help Desk manager. We are excited to have William join us and expect to come back strong supporting the AMR research community in May.
PURPOSE: Antimicrobial resistance (AMR), especially multidrug resistance, is one of the most serious global threats facing public health. We performed a proof of concept study assessing the suitability of shotgun proteomics as a complementary approach to whole-genome sequencing (WGS) for detecting AMR determinants.
EXPERIMENTAL DESIGN: We used previously published shotgun proteomics and WGS data on four isolates of Campylobacter jejuni to perform AMR detection by searching the Comprehensive Antibiotic Resistance Database, and we assessed their detection ability relative to genomics screening and traditional phenotypic testing measured by minimum inhibitory concentration.
RESULTS: Both genomic and proteomic approaches identified the wild type and variant molecular determinants responsible for resistance to tetracycline and ciprofloxacin, in agreement with phenotypic testing. In contrast, the genomic method identified the presence of the β-lactamase gene, blaOXA-61 , in three isolates. However, its corresponding protein product was detected in only a single isolate, consistent with results obtained from phenotypic testing.
Alcock BP, Raphenya AR, Lau TTY, Tsang KK, Bouchard M, Edalatmand A, Huynh W, Nguyen A-LV, Cheng AA, Liu S, Min SY, Miroshnichenko A, Tran H-K, Werfalli RE, Nasir JA, Oloni M, Speicher DJ, Florescu A, Singh B, Faltyn M, Hernandez-Koutoucheva A, Sharma AN, Bordeleau E, Pawlowski AC, Zubyk HL, Dooley D, Griffiths E, Maguire F, Winsor GL, Beiko RG, Brinkman FSL, Hsiao WWL, Van Domselaar G, McArthur AG.
The Comprehensive Antibiotic Resistance Database (CARD; https://card.mcmaster.ca) is a curated resource providing reference DNA and protein sequences, detection models and bioinformatics tools on the molecular basis of bacterial antimicrobial resistance (AMR). CARD focuses on providing high-quality reference data and molecular sequences within a controlled vocabulary, the Antibiotic Resistance Ontology (ARO), designed by the CARD biocuration team to integrate with software development efforts for resistome analysis and prediction, such as CARD’s Resistance Gene Identifier (RGI) software. Since 2017, CARD has expanded through extensive curation of reference sequences, revision of the ontological structure, curation of over 500 new AMR detection models, development of a new classification paradigm and expansion of analytical tools. Most notably, a new Resistomes & Variants module provides analysis and statistical summary of in silico predicted resistance variants from 82 pathogens and over 100 000 genomes. By adding these resistance variants to CARD, we are able to summarize predicted resistance using the information included in CARD, identify trends in AMR mobility and determine previously undescribed and novel resistance variants. Here, we describe updates and recent expansions to CARD and its biocuration process, including new resources for community biocuration of AMR molecular reference data.