MtlA gene sequencing based on 3rd -generation sequencing for Vibrio typing

Area of Science:

• Microbiology
• Genomics
• Bioinformatics

Background:

• Traditional methods for Vibrio parahaemolyticus strain typing, including serotyping and multilocus sequence typing (MLST), have limitations in resolution and accuracy, especially for closely related strains.
• The development of new sequencing technologies offers potential for improved strain characterization.

Purpose of the Study:

• To evaluate the utility of the Geneus G-seq500 nanopore sequencer for typing Vibrio parahaemolyticus strains.
• To assess the effectiveness of the mtlA gene as a novel typing marker for V. parahaemolyticus, offering advantages over existing methods.

Main Methods:

• Sequencing of the full-length mtlA gene (1,953 bp) from 33 V. parahaemolyticus strains using the Geneus G-seq500.
• Generation of long reads with high accuracy (mean quality Q35, sequence accuracy 99.88-99.93%) and development of a robust consensus sequence generation pipeline.
• Development of a novel naming rule for V. parahaemolyticus strains based on mtlA gene clustering, incorporating data from NCBI.

Main Results:

• The Geneus G-seq500 produced high-quality long reads, enabling accurate full-length mtlA gene sequencing.
• A new nomenclature system was established, designating the largest cluster of 2,321 strains as mtlA0001.
• The mtlA gene demonstrated high resolution and accuracy for V. parahaemolyticus strain typing, comparable to MLST but with improved cost-effectiveness and speed.

Conclusions:

• The Geneus G-seq500 exhibits high sequencing accuracy, making it suitable for microbial genomics applications.
• Utilizing the full-length mtlA gene is an effective strategy for V. parahaemolyticus strain typing and epidemiological surveillance.
• This approach provides a valuable tool for tracking and understanding the population structure of V. parahaemolyticus.