An alternative SEM drying method using hexamethyldisilazane (HMDS) for microbial cell attachment studies on...

Area of Science:

• Microbiology
• Materials Science
• Microscopy

Background:

• Microbial colonization of sub-bituminous coal is of interest due to its role in biofragmentation, fermentation, and methane generation.
• Traditional critical point drying (CPD) for scanning electron microscopy (SEM) presents challenges with fragile coal samples and requires expensive equipment.

Purpose of the Study:

• To evaluate hexamethyldisilazane (HMDS) as a simpler, more affordable alternative to CPD for preparing bacterial-on-coal specimens for SEM imaging.
• To assess the effectiveness of HMDS in preserving bacterial cell morphology and attachment structures on sub-bituminous coal.

Main Methods:

• Specimen preparation for SEM using hexamethyldisilazane (HMDS) as a drying agent.
• Comparison of HMDS drying with critical point drying (CPD) for bacterial cells attached to sub-bituminous coal.
• Microscopic analysis of bacterial cell shape, aggregation, and attachment points to coal surfaces.

Main Results:

• HMDS effectively preserved the shape of individual and aggregated bacterial cells on coal, with minimal cell flattening.
• The method allowed for clear observation of bacterial attachment points to the coal surface, facilitated by EPS shrinkage.
• Positive results were observed for both short and long bacterial cells, even without glucose.

Conclusions:

• Hexamethyldisilazane (HMDS) is a suitable and advantageous drying agent for SEM preparation of bacterial-on-coal specimens.
• HMDS offers a safer, cheaper, and more practical alternative to CPD for studying microbial colonization on sub-bituminous coal.
• The technique facilitates the study of cell-coal and cell-cell interactions through EPS structure observation.