Chapter 12

Microbial Genomics

Introduction to Microbial Genomics

Microbial genomics is the comprehensive study of the genetic material of microorganisms. It involves sequencing, analyzing, and comparing the genomes of bacteria, archaea, viruses, and microbial eukaryotes to understand their structure, function, and evolution.

• Genome: The complete set of genetic material (DNA) in an organism.

• Gene: A segment of DNA that encodes a functional product, usually a protein.

• Genome size: Measured in base pairs (bp), kilobase pairs (kbp), or megabase pairs (Mbp).

Example: Comparing a bacterial genome of 10 Mbp to a eukaryotic genome of 10 Mbp, eukaryotes typically have more non-coding DNA, so bacteria may have more coding genes per Mbp.

Gene Content in Microbial Genomes

• Gene density: Bacteria generally have higher gene density (more genes per Mbp) than eukaryotes due to less non-coding DNA.

• Non-coding DNA: Eukaryotes have a larger proportion of non-coding DNA, including introns and regulatory sequences.

• Metagenomics: The study of genetic material recovered directly from environmental samples, allowing the analysis of entire microbial communities (microbiomes).

Example: Metagenomic sequencing can reveal the diversity and function of microbes in soil, water, or the human gut.

Proteomics

Introduction to Proteomics

Proteomics is the large-scale study of the structure, function, and interactions of proteins produced by an organism. It provides insights into cellular processes and how proteins respond to environmental changes.

• Proteome: The entire set of proteins expressed by a genome, cell, tissue, or organism at a certain time.

• Proteomic analysis: Used to identify proteins, study post-translational modifications, and understand protein-protein interactions.

Techniques in Proteomics

• Mass spectrometry (MS): A key technique for identifying and quantifying proteins by measuring the mass-to-charge ratio of peptide fragments.

• Post-translational modifications: Chemical changes to proteins after synthesis, such as phosphorylation or glycosylation, which can be detected by MS.

Example: Mass spectrometry can be used to detect disease biomarkers in blood samples.

Metabolomics

Introduction to Metabolomics

Metabolomics is the study of the complete set of small-molecule metabolites (such as sugars, amino acids, and lipids) produced by an organism. It provides a snapshot of the metabolic state and can reveal responses to environmental changes.

• Metabolome: The full complement of metabolites found within a cell or organism.

• Metabolic pathways: Series of enzymatic reactions that transform substrates into products.

• Mass spectrometry in metabolomics: Used for accurate identification and quantification of metabolites, even at low concentrations.

Example: Metabolomics can be used to study how bacteria respond to antibiotics or environmental stressors.

Single-Cell Genomics

Introduction to Single-Cell Genomics

Single-cell genomics involves sequencing the DNA from individual cells, allowing researchers to study genetic diversity and function at the single-cell level.

• Amplification: Techniques such as multiple displacement amplification (MDA) are used to amplify small amounts of DNA from single cells.

• Applications: Identifying rare microbes, studying cell-to-cell variation, and linking specific microbes to metabolic functions.

Example: Single-cell genomics can identify uncultured bacteria in environmental samples.

Biotechnology & Synthetic Biology

Introduction to Genetic Engineering

Biotechnology uses living organisms or their components to develop useful products. Synthetic biology involves designing and constructing new biological parts or systems.

• Molecular cloning: Inserting a gene of interest into a vector (such as a plasmid) and introducing it into a host organism for replication or expression.

• Vectors: DNA molecules (plasmids, viruses) used to carry foreign genetic material into a cell.

• Recombinant DNA: DNA molecules formed by laboratory methods to bring together genetic material from multiple sources.

Example: Production of human insulin in bacteria using recombinant DNA technology.

Applications of Genetic Engineering

• Transgenic organisms: Organisms that have been genetically modified to carry genes from other species.

• Agrobacterium tumefaciens and Ti plasmid: Used to transfer genes into plants, creating genetically modified crops.

• Bovine somatotropin: A recombinant hormone used to increase milk production in cows.

Example: Genetically modified crops with improved resistance to pests or environmental stress.

Genome Editing: CRISPR-Cas9

Introduction to CRISPR

CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) is a natural defense mechanism in bacteria that has been adapted as a powerful genome editing tool.

• CRISPR-Cas9 system: Uses a synthetic guide RNA (sgRNA) to direct the Cas9 nuclease to a specific DNA sequence, where it makes a double-stranded break.

• Insertion: A DNA fragment with homology to the cut site can be inserted using the cell's repair machinery.

• Deletion: Two cut sites can be created, and the intervening DNA is removed during repair.

Example: CRISPR can be used to knock out genes in bacteria to study their function or to correct genetic mutations in higher organisms.

Applications of CRISPR

• Gene therapy: Correcting genetic defects in humans.

• Crop improvement: Creating disease-resistant or higher-yielding plants.

• Sterile insect technique: Releasing genetically modified sterile insects to control pest populations.

Summary Table: Omics Technologies in Microbiology

Omics Field	Definition	Main Techniques	Applications
Genomics	Study of the complete DNA sequence of an organism	DNA sequencing, PCR, metagenomics	Microbial diversity, evolution, gene discovery
Proteomics	Study of all proteins expressed by a genome	Mass spectrometry, 2D gel electrophoresis	Protein function, disease biomarkers
Metabolomics	Study of all metabolites in a cell or organism	Mass spectrometry, NMR spectroscopy	Metabolic pathways, stress responses
Single-cell genomics	Sequencing DNA from individual cells	MDA, single-cell sequencing	Microbial ecology, rare species detection