Single-Cell Identification and Characterization of Viable but Nonculturable Campylobacter jejuni Using Raman Optical...

Area of Science:

• Microbiology
• Food Safety
• Spectroscopy

Background:

• Campylobacter jejuni is a major foodborne pathogen.
• The viable but nonculturable (VBNC) state allows C. jejuni to survive environmental stress.
• VBNC cells are difficult to detect using traditional methods.

Purpose of the Study:

• To develop a culture-independent method for identifying and characterizing VBNC C. jejuni cells.
• To analyze VBNC cells at the single-cell level for accurate detection.
• To understand the unique behavior and molecular characteristics of VBNC C. jejuni.

Main Methods:

• Induction of C. jejuni into the VBNC state using osmotic pressure and aerobic stress.
• Collection of single-cell Raman spectra using optical tweezers.
• Application of a convolutional neural network (CNN) for classification of VBNC and culturable cells.
• Utilizing gradient-weighted class activation mapping for spectral region analysis.

Main Results:

• VBNC C. jejuni cells were distinguished from culturable cells with approximately 92% accuracy using CNN.
• No significant spectral differences were observed between VBNC cells induced under different stressors or induction periods.
• Gradient-weighted class activation mapping identified key spectral regions associated with VBNC cell classification.

Conclusions:

• Single-cell Raman spectroscopy combined with machine learning provides an accurate method for detecting VBNC C. jejuni.
• The identified spectral regions correlate with known molecular changes in VBNC cells, offering insights into their characterization.
• This approach enhances the ability to detect and understand VBNC pathogens, improving food safety.