Lesson 1Antimicrobial susceptibility testing (AST) methods: disk diffusion, broth microdilution, automated AST systems — advantages, limitations, when to use eachThis section compares disk diffusion, broth microdilution, gradient strips, and automated AST systems, emphasizing test selection, setup, reading, quality control, and recognition of resistance mechanisms that may be missed by specific methods in our labs.
Disk diffusion setup, reading, and limitationsBroth microdilution and MIC determinationGradient diffusion strips: use and caveatsAutomated AST platforms and alert rulesChoosing AST methods for special organismsLesson 2Quality control, validation, and biosafety considerations for molecular testingThis section outlines quality control, validation, and biosafety for molecular testing, including verification of new assays, routine QC, contamination prevention, workflow separation, and compliance with regulatory standards in Zimbabwe.
Verification and validation of new PCR assaysInternal, external, and proficiency controlsPreventing amplicon contamination and carryoverUnidirectional workflow and lab zoningRegulatory and accreditation requirementsLesson 3Next-generation approaches: targeted amplicon sequencing, whole-genome sequencing (WGS) pipeline steps, assembly, annotation, resistance gene callingThis section introduces next-generation sequencing for resistance surveillance, outlining targeted amplicon workflows, WGS library preparation, sequencing, assembly, annotation, resistance gene calling, and basic quality control steps for local use.
Targeted amplicon design and library preparationWGS library prep, indexing, and run setupRead quality control and genome assembly choicesGenome annotation and resistance gene databasesReporting NGS findings and clinical relevanceLesson 4Specimen handling and culture: blood culture sets, incubation, subculture media and colony morphology cuesThis section explains best practices for specimen receipt, blood culture collection and incubation, subculture techniques, and interpretation of colony morphology, linking macroscopic features with likely pathogens in African hospitals.
Pre-analytical variables and specimen rejectionBlood culture set collection and incubation rulesSubculture media selection and inoculation patternsReading plates and recognizing mixed culturesColony morphology clues to common pathogensLesson 5Rapid phenotypic methods and immunochromatographic assays for common carbapenemasesThis section focuses on rapid phenotypic carbapenemase detection, including Carba NP variants and immunochromatographic lateral flow assays, with emphasis on performance, workflow integration, and result interpretation in busy clinical labs.
Principles of rapid phenotypic carbapenemase assaysCarba NP and derivatives: setup and readoutImmunochromatographic tests for KPC, NDM, OXA-48Sensitivity, specificity, and common interferencesAlgorithmic use with culture and molecular testsLesson 6Organism identification methods: biochemical panels, MALDI-TOF MS workflow and interpretationThis section reviews major organism identification strategies, including biochemical panels and MALDI-TOF MS, covering sample preparation, instrument workflow, spectral interpretation, troubleshooting, and integration with lab systems.
Design and interpretation of biochemical identification panelsMALDI-TOF sample prep for bacteria and yeastsAcquisition parameters and spectral quality controlDatabase matching, score cutoffs, and reportingCommon pitfalls and discordant ID resolutionLesson 7Phenotypic assays for resistance detection: ESBL confirmatory tests, modified Hodge test, Carba NP / mCIM / sCIM for carbapenemases, AmpC screeningThis section reviews phenotypic assays for resistance, including ESBL confirmatory tests, AmpC screening, and carbapenemase assays such as mCIM, sCIM, and Carba NP, with guidance on algorithms for complex patterns in our setting.
ESBL screening and confirmatory synergy testsPhenotypic detection of plasmid-mediated AmpCmCIM and sCIM workflows and interpretationCarba NP and related carbapenemase assaysIntegrating phenotypic and genotypic findingsLesson 8Molecular workflows: targeted PCR assays for blaCTX-M, blaKPC, blaNDM, blaOXA-48-like, qnr, 16S methyltransferasesThis section covers molecular workflows for key resistance genes, detailing assay design, controls, and interpretation for blaCTX-M, blaKPC, blaNDM, blaOXA-48-like, qnr, and 16S methyltransferases, integrated with phenotypic data.
Primer and probe design for resistance targetsDNA extraction methods and inhibition controlSingleplex vs multiplex PCR assay strategiesResult interpretation and limit of detectionReconciling PCR results with AST phenotypesLesson 9Interpreting MICs and breakpoints: CLSI vs EUCAST differences and how to select appropriate breakpointsThis section explains MIC interpretation using CLSI and EUCAST breakpoints, highlighting differences, updates, dose-specific breakpoints, and strategies for selecting standards in clinical laboratories across Zimbabwe.
MIC concepts, ECOFFs, and clinical breakpointsKey differences between CLSI and EUCASTDose-specific and infection-site breakpointsUpdating and version control of standardsDocumenting breakpoint choices in the lab
