Single-Cell Sequencing of a Bile Sample From an Acute Cholecystitis Patient

Area of Science:

• Genomics
• Microbiology
• Clinical Diagnostics

Background:

• Single-cell sequencing is emerging for clinical samples, but its application in bacterial infections remains limited.
• Acute cholecystitis is a common gallbladder inflammation, often caused by bacterial infections.

Purpose of the Study:

• To investigate the utility of single-cell genome sequencing for analyzing bacterial infections in a clinical sample.
• To characterize the bacterial species present in a bile sample from a patient with acute cholecystitis.

Main Methods:

• Whole genome sequencing of four *Escherichia coli* isolates from a bile sample using the single-cell amplified genome (SAG) sequencing technique.
• Comparative genomic analysis using average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) against type strains.
• 16S metagenome analysis to identify predominant bacterial genomes.

Main Results:

• The four *E. coli* isolates were genetically identical (ANI 99.98-100%, dDDH 100%).
• Genomic analysis indicated these strains were more closely related to *Shigella sonnei* (ANI 98.65-98.66%) than *E. coli* (ANI 96.79-96.80%), despite lacking key *Shigella* virulence genes.
• 16S metagenome analysis identified *E. coli* as the predominant species (93.15%), but SAG sequencing coverage was limited (9.45-42.88%), resulting in sequence gaps.

Conclusions:

• Single-cell genome sequencing, despite current limitations in coverage and data gaps, can offer novel insights into bacterial infections.
• The study highlights potential discrepancies between standard identification and whole-genome-based phylogenetic placement for clinical isolates.
• Further refinement of SAG sequencing protocols is necessary to improve genome coverage and data quality for clinical applications.