Microbial Genomics and Infectious Disease¶
Chapter 126 | Harrison's 22e · Part 5 – Infectious Diseases: Bacterial · Chapter 126
Key Clinical Points¶
- Bacteriophages regulate pathogen abundance, influencing disease epidemiology.
- Genomic technologies enable rapid, sensitive diagnostics with unprecedented breadth of information.
- Traditional serologic methods have limited utility for real-time patient management and chronic infections.
- Metagenomic assembly workflows facilitate discovery of novel pathogens in complex samples.
- Phylogenetic analysis tracks transmission dynamics of viruses like Ebola and SARS-CoV-2.
- Genomics transforms infectious disease surveillance, diagnostics, and therapeutic development.
1. DEFINITION & OVERVIEW¶
Microbial genomics applies genomic technologies to study microbial pathogens and their interactions with hosts. This field transforms infectious disease diagnostics, surveillance, and therapeutic development through advanced sequencing methods and bioinformatics analysis.
1.1 Historical Context¶
• Traditional Microbiology: Relied on culture-based assays and biochemical testing for pathogen identification. • Serologic Limitations: Required acute- and convalescent-phase serum samples, limiting real-time utility for patient management. • Tuberculosis Detection: PPD skin testing showed poor sensitivity in immunocompromised populations.
1.2 Genomic Revolution¶
• Nucleic Acid Diagnostics: Offer improved speed, sensitivity, and specificity compared to culture methods. • Whole-Genome Sequencing (WGS): Enables identification of virulence factors and antibiotic resistance genes. • Metagenomics: Allows pathogen discovery in complex clinical samples containing mixed microbial communities.
1.3 Key Terms¶
• Contig: A DNA sequence representing a continuous fragment of a genome, assembled from overlapping sequences; relevant for de novo assembly of data not aligning to known genomes. • Horizontal Gene Transfer: The transfer of genes between organisms through mechanisms other than clonal descent (transformation, conjugation, or transduction). • Metagenome: Analysis of genetic material from multiple species directly from primary samples without requiring prior culture steps. • Microbial Genome-Wide Association Study (GWAS): An analytic framework to test statistical associations between microbial genotypes and phenotypes (e.g., antibiotic resistance, virulence). • Multilocus Sequence Typing (MLST): A method for typing organisms based on DNA sequence fragments from a prespecified set of genes. • Nucleic Acid Amplification Test (NAAT): A biochemical assay evaluating the presence of specific nucleic acids via amplification (e.g., PCR, ligase chain reactions). • Single-Nucleotide Polymorphism (SNP): Point mutations in DNA; the number of SNPs between isolates measures genetic distance. • Whole Genome Sequencing (WGS): A process determining the full DNA sequence of an organism's genome, facilitated by next-generation sequencing.
2. DIAGNOSTIC APPROACHES¶
Modern diagnostics integrate genomic technologies with traditional methods for comprehensive pathogen characterization.
2.1 Traditional Methods¶
• Phenotypic Classification: Based on morphology, growth requirements, and biochemical reactions. • Antibiotic Susceptibility Testing: Performed via agar diffusion or broth dilution methods. • Limitations: Slow turnaround time; inability to detect non-culturable pathogens.
2.2 Genomic Diagnostics¶
• PCR & RT-PCR: Rapid detection of viruses (HIV, HBV, HCV, SARS-CoV-2, influenza) and bacteria. • Metagenomics: Identification of novel pathogens via de novo assembly of unmapped reads. • Cell-Free DNA Sequencing: Unbiased amplification and sequencing of cell-free nucleic acid from blood; compares nonhuman sequences to known pathogen/contaminant genomes to identify circulating DNA. • 16S Ribosomal Gene Sequencing: Used for broader pathogen detection. • Microarray & Sequencing: Used for identifying viral pathogens (e.g., SARS-CoV-2, West Nile virus, MERS-CoV).
3. EPIDEMIOLOGIC APPLICATIONS¶
Genomic data transforms infectious disease surveillance and outbreak investigation.
3.1 Phylogenetic Analysis¶
• Transmission Tracking: Tracing transmission chains through genetic relatedness of pathogen isolates. • Outbreak Reconstruction: Estimating Time to Most Recent Common Ancestor (TMRCA). • Geographic Mapping: Identifying spread patterns in viral epidemics (e.g., Ebola, SARS-CoV-2).
3.2 Public Health Surveillance¶
• Antimicrobial Resistance (AMR): Real-time tracking of resistance gene dissemination. ◦ VRE: Detection of vanA or vanB genes. ◦ Carbapenemase: Detection of KPC, NDM, OXA-48, IMP, and VIM enzymes. • Disease Monitoring: Tracking vaccine-preventable diseases via genomic signatures. • Zoonotic Surveillance: Identifying zoonotic spillover events through comparative genomics. • Outbreak Tracking: Sequencing used for Klebsiella, S. aureus, M. tuberculosis, E. coli, Vibrio cholerae, Ebola, Zika, and influenza.
4. CLINICAL IMPACT¶
Genomic approaches improve patient management and therapeutic strategies.
4.1 Personalized Medicine¶
• Targeted Therapy: Tailoring antibiotic therapy based on specific resistance gene profiling. • Prognostic Prediction: Identifying virulence factors to predict disease severity. • Immunotherapy: Guiding development of immunotherapies for infectious diseases.
4.2 Therapeutic Innovation¶
• Vaccine Design: Developing vaccines targeting conserved genomic regions. • CRISPR Technology: Development of CRISPR-based diagnostics and therapeutics. • Novel Antimicrobials: Discovery of new antimicrobial peptides through genomic screening.
5. CLINICAL SUMMARY & WORKFLOWS¶
Key procedural pathways for clinical application.
Metagenomic Assembly Workflow (Figure 126-3)¶
- Clinical Sample → 2. DNA Extraction & Sequencing → 3. Mapping to Reference Genomes → 4. De novo Assembly of 'unmapped' reads into contigs → 5. Phylogenetic Comparison to known genomes → 6. Identification of Novel/Known Pathogens.
Antibiotic Resistance Detection¶
• Vancomycin Resistance (VRE): PCR detection of vanA or vanB. • Carbapenemase Detection: PCR and targeted sequencing for KPC, NDM, OXA-48, IMP, and VIM.
Reference Tables¶
TABLE 126-1 Glossary of Selected Terms in Genomics TERM Contig¶
Harrison's 22e, p.976
| TERM | DEFINITION |
|---|---|
| Contig | A DNA sequence representing a continuous fragment of a genome, assembled from overlapping sequences; relevant for de novo assembly of sequence data that do not align to previously sequenced genomes |
| Horizontal gene transfer |
The transfer of genes between organisms through mechanisms other than by clonal descent, such as through transformation, conjugation, or transduction |
| Metagenomics | Analysis of genetic material from multiple species directly from primary samples without requiring prior culture steps |
| Microbial genome- wide association study (GWAS) |
An analytic framework to test statistical associations between microbial genotypes and phenotypes of interest, such as antibiotic resistance and virulence |
| Multilocus sequence typing |
A method for typing organisms based on DNA sequence fragments from a prespecified set of genes |
| Nucleic acid amplification test (NAAT) |
A biochemical assay that evaluates for the presence of a particular string of nucleic acids through amplification by one of several methods, including polymerase and ligase chain reactions |
| Single-nucleotide polymorphism (SNP) |
Point mutations in DNA, the number of which in different microbial isolates is a measure of their genetic distance from one another |
| Whole genome sequencing (WGS) |
A process that determines the full DNA sequence of an organism’s genome; has been greatly facilitated by next- generation sequencing technology |
TABLE 126-2 Selected Clinical Applications of Infectious Disease Genomics APPLICATION Organism Identification Viral…¶
Harrison's 22e, p.977
| APPLICATION | TECHNOLOGY | NOTES/EXAMPLES |
|---|---|---|
| Organism Identification | ||
| Viral detection | PCR, RT-PCR | Identification of HIV, HBV, HCV, respiratory viruses including SARS-CoV-2 and influenza, and others for diagnosis and response to therapy |
| PCR | ||
| Pathogen detection | PCR, RT-PCR, NAAT | Multiplexed identification of dozens of viruses, bacteria, yeasts, and parasites from a variety of clinical specimens |
| 16S ribosomal gene sequencing |
||
| Pathogen detection | Cell-free DNA sequencing |
Unbiased amplification and sequencing of cell-free nucleic acid from blood, with analytical comparison of resulting nonhuman sequences to genomes of known pathogens and contaminants, in order to identify circulating pathogen DNA; anecdotal clinical use to establish etiology of systemic or focal infection, though clinical utility and optimal use cases still evolving |
| Pathogen Discovery | ||
| Sequencing, metagenomic assembly |
||
| Viral pathogens | Microarray, sequencing | Hybridization of clinical samples to microarrays from phylogenetically diverse known viruses identified the first SARS coronavirus and others. Direct sequencing has identified SARS-CoV-2, West Nile virus, and MERS-CoV, among others. Use is primarily in research. |
| Antibiotic Resistance | ||
| PCR | ||
| VRE detection | PCR | Detection of the vanA or vanB gene, the main genotypic causes of vancomycin resistance in Enterococcus |
| PCR, NAAT | ||
| Carbapenemase detection | PCR | Detection of genes encoding one of several types of enzymes (KPC, NDM, OXA-48, IMP, VIM) that hydrolyze carbapenems, accounting for much but not all carbapenemase resistance in Enterobacteriaceae |
| Targeted sequencing | ||
| Epidemiology | ||
| Outbreak and epidemic tracking |
Sequencing | Application to tracking outbreaks and epidemics on local and international scales, including spread of carbapenemase-producing Klebsiella, S. aureus, M. tuberculosis, E. coli, Vibrio cholerae, Ebola virus, Zika virus, and influenza virus |
| Sequencing |