MICROBIAL TECHNOLOGY
is now the method of choice within the reference setting, replacing phenotypic methods as the first line for antimicrobial susceptibility determination. Whole-genome sequencing can simultaneously give information regarding the genetic relatedness of different clinical isolates. Characterisation of mycobacterial isolates using WGS works particularly well for Mycobacterium tuberculosis strains but is also effective for other well- described non-tuberculosis mycobacteria. Given the slow genomic evolution of M. tuberculosis, varying only around one single nucleotide polymorphism (SNP) every two years, the data derived from WGS can be used as an effective indicator to identify clusters of related cases. The advantages of WGS as an
Natasha Weston provided an overview of WGS of M. tuberculosis and other mycobacterial species.
mechanism is not clear and requires further investigation. These agar-based methods, along with the gold-standard of broth-microdilution, have the additional difficulty of interpreting and reporting the susceptibility results using increasingly complex breakpoint tables. Ensuring the results tested within the microbiology laboratory translate to understandable results for users is critical for it to impact patient care.
Molecular methods offer a more rapid, gene specific mechanism that can be particularly useful in organisms with infection, prevention and control implications such as methicillin-resistant Staphylococcus aureus (MRSA) or carbapenemase-producing organisms (CPOs). Detection of the gene in a diagnostic setting can be used to rule-in patients as having a resistant organism and there are multiple commercial assays available for doing so, albeit this is more complex for Gram-negative than Gram- positive bacteria.
This captivating session showed the advantages of the amount of data gained when applying whole-genome sequencing (WGS) in antimicrobial resistance and how this can be used for epidemiological purposes, outbreak investigations and to examine biomarkers in organisms. While the genotypic methods are unable to allow understanding of the interactions of more than one resistance mechanism within one organism, and do not always predict therapeutic failure, they have significant public health and outbreak management implications.
Dr Hopkins discussed the literature and 16
evidence body surrounding phenotypic and genotypic antimicrobial susceptibility testing, and while the volume of evidence is there, the drawback is the tendency for studies to focus on individual organisms, reducing its applicability into a diagnostic laboratory.
The need for most laboratories to use a combination of methods requires a sound knowledge of the methods, their limitations and the most appropriate application. This makes antimicrobial resistance (AMR) a key area of microbiology where multidisciplinary working of biomedical scientists, clinical scientists, pharmacists, medical microbiologists and infection control is crucial, and brings together the science with the clinical applications, for the patient.
Tuberculosis and whole-genome sequencing Dr Natasha Weston, who was previously a Senior Clinical Fellow at the National Mycobacterial Reference Service in Birmingham, concluded the morning session with a look at how WGS is being applied to the diagnosis of TB and other mycobacterial infections. While PCR is playing an increasing diagnostic role, it can be costly and provides quite limited antimicrobial susceptibility information. Dr Weston demonstrated that WGS has advantages in diagnosis, treatment and epidemiologically. By applying WGS techniques, a reliable rapid identification can be obtained with an indication of multiple resistance markers potentially weeks before that possible by traditional techniques. Whole-genome sequencing
outbreak investigation tool were vividly illustrated with a series of case studies showing how socially distinct cases can be linked using WGS. Data can now be available in less than a week from initial positive cultures, which allows a much more rapid and accurate picture of TB outbreaks and transmission events to be determined. This means that public health interventions, contact tracing and treatment can be targeted much more precisely and effectively than previously possible. Transmission events that are transient or short lived can be difficult to connect but WGS has allowed the recognition of clusters with little or no social overlap. Dr Weston clearly demonstrated within these case studies the real benefits that more rapid availability of typing data can provide, where it is even possible to identify laboratory cross-contamination events.
External quality assessment: the challenges The increasing complexity of the technologies that have been introduced has meant a greatly increased requirement for QC in the context of wider quality assurance. During this continual period of progression, the complexity of the analysers has increased markedly from detection of single gene targets to multiplex assays capable of detecting multiple targets from a range of different pathogens simultaneously. Conversely, the time taken to generate a result has markedly decreased, with some platforms providing an actionable result within 30 minutes. Nowadays, over 80% of microbiology laboratories in the UK report that they use molecular assays for virology diagnostic testing. It was with this background in mind that Dr Elaine McCulloch, Technical Project Manager, QCMD, Glasgow discussed the need for quality assurance in molecular diagnostics
SEPTEMBER 2022
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