Multi-Locus Sequence Typing

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What is the MLST Approach

MLST is a sophisticated molecular technique essential for the accurate identification and differentiation of bacterial strains. It involves the sequencing of internal fragments from multiple housekeeping genes—commonly seven—each spanning 400-600 base pairs. These housekeeping genes are crucial for fundamental cellular functions and exhibit minimal genetic variation within a species. Using primers specific to these genes, MLST performs PCR amplification and sequencing to determine allele numbers at designated loci. This process generates unique allele profiles or sequence types (STs) for each strain. By analyzing these profiles through methods like pairwise difference matrices and constructing phylogenetic trees, MLST enables a detailed understanding of genetic relationships and evolutionary trajectories among bacterial strains.

Key Features of MLST

• Standardized Framework: MLST uses a structured method to sequence housekeeping genes, assigning unique identifiers to alleles for consistent and comparable bacterial typing.
• Sequence-Based Typing: It relies on nucleotide sequences rather than fragment lengths, offering precise and reproducible strain identification.
• Allelic Profiles: The Sequence Type (ST) represents a strain's allelic profile across multiple loci, facilitating evolutionary and epidemiological analyses.

Why is MLST Used

The utilization of MLST provides several essential benefits:

• High Resolution and Precision: MLST provides detailed strain differentiation, essential for pinpointing outbreaks and mapping bacterial evolution with fine genetic distinctions.
• Reproducibility: By adhering to uniform protocols and leveraging centralized databases, MLST delivers reliable and comparable results across various research settings.
• Application Flexibility: MLST's adaptability to numerous bacterial species makes it a valuable tool for diverse research applications, from clinical diagnostics to environmental studies.

Differences Between MLST and MLSA

While both MLST and Multi-Locus Sequence Analysis (MLSA) revolve around the examination of multiple gene loci, their objectives and applications diverge significantly:

Purpose and Scope:

• MLST: This method is principally employed for the typing and differentiation of bacterial strains within a species. By analyzing sequence variations in predefined housekeeping genes, MLST can accurately distinguish between different strains.
• MLSA: Conversely, MLSA is predominantly utilized for constructing phylogenetic relationships and examining evolutionary lineages among diverse species or higher taxonomic groups.

Gene Selection:

• MLST: Typically focuses on a fixed set of housekeeping genes, which are conserved and essential across various strains, serving fundamental cellular functions.
• MLSA: Encompasses a more extensive array of genes, which may include both highly conserved and more variable genes, depending on the phylogenetic questions being addressed.

Application:

• MLST: Predominantly applied in epidemiological studies, including outbreak investigations and detailed strain differentiation. Its high resolution makes it an invaluable tool for understanding the transmission and spread of bacterial pathogens.
• MLSA: More suited for in-depth phylogenetic studies, MLSA facilitates the reconstruction of evolutionary histories and the mapping of genetic relationships among broader taxonomic categories.

Applications of MLST

The applications of MLST include, but are not limited to, the following areas:

• Epidemiology: Tracking and monitoring bacterial outbreaks, identifying sources of infections, and assessing the spread of pathogenic strains.
• Microbial Ecology: Studying the distribution and diversity of microbial populations in environmental samples.
• Pharmacology: Investigating resistance patterns and susceptibility to antibiotics among different bacterial strains.