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A requirement for flow to enable the development of Ureaplasma parvum biofilms in vitro
R S Rowlands1, K Kragh2, S Sahu1,3
1Microbiology and Infection Research Group, Department of Biomedical Science, Cardiff School of Sport and Health Sciences, Cardiff Metropolitan University, Cardiff, UK.
Aims:
To use a flow-based method to establish, quantify and visualize biofilms of Ureaplasma parvum.
Methods And Results:
Absorbance readings of a U. parvum HPA5 culture were taken at 550 nm every 3 h for 30 h in order to establish a growth curve, with viability determined by the number of colour changing units (CCUs). Biofilms were established using the DTU flow-cell with a flow rate of 0·01 ml min-1 and compared to the static control. Titres of bacteria were determined by CCU and biofilm biomass was quantified by Syto9 staining and COMSTAT analysis. High-resolution images were obtained by scanning electron microscopy (SEM). Flow resulted in significantly more biofilm and higher cell titre (0·599 µm3 /µm2 ± 0·152 and 4 × 108 CCU per ml, respectively) compared with static conditions (0·008 µm3 /µm2 ± 0·010 and no recoverable cells, respectively). SEM revealed pleomorphic cells, with signs of budding and possible membrane vesicle formation.
Conclusions:
Flow is an essential requirement for the establishment of U. parvum biofilms.
Significance And Impact Of The Study:
This is the first quantification of biofilm biomass formed by U. parvum. It is now possible to establish viable biofilms of U. parvum which will allow for future testing of antimicrobial agents and understanding of virulence-associated with adhesion.
Insights
Flow-based methods are essential for establishing Ureaplasma parvum biofilms. This study quantifies biofilm biomass, enabling future antimicrobial testing and virulence studies.
Area of Science:
- Microbiology
- Biofilm formation
- Ureaplasma parvum
Background:
- Ureaplasma parvum is a common commensal bacterium that can cause opportunistic infections.
- Biofilm formation is a critical factor in bacterial pathogenesis and antimicrobial resistance.
- Current methods for studying Ureaplasma parvum biofilms are limited.
Purpose of the Study:
- To develop and validate a flow-based method for establishing, quantifying, and visualizing Ureaplasma parvum biofilms.
- To compare biofilm formation under flow versus static conditions.
- To provide a foundation for future studies on Ureaplasma parvum virulence and antimicrobial susceptibility.
Main Methods:
- Ureaplasma parvum HPA5 culture growth curve established via absorbance readings (550 nm) and viability assessed by colony-forming units (CFUs).
- Biofilms formed using a DTU flow-cell at 0.01 ml/min flow rate, compared to static controls.
- Biofilm biomass quantified using Syto9 staining and COMSTAT analysis; bacterial titres determined by CFUs.
- High-resolution imaging performed using scanning electron microscopy (SEM).
Main Results:
- Flow conditions significantly enhanced Ureaplasma parvum biofilm formation (0.599 µm³/µm²) compared to static conditions (0.008 µm³/µm²).
- Higher bacterial titres (4 × 10⁸ CFU/ml) were observed in flow conditions versus no recoverable cells in static conditions.
- SEM revealed pleomorphic Ureaplasma parvum cells with budding and potential membrane vesicle formation.
Conclusions:
- Flow is a critical requirement for the establishment of viable Ureaplasma parvum biofilms.
- This study presents the first quantification of Ureaplasma parvum biofilm biomass.
- The developed method facilitates future testing of antimicrobial agents and understanding of Ureaplasma parvum adhesion-associated virulence.
