Rapid detection of bacteriophages in starter culture using water-in-oil-in-water emulsion microdroplets

Abstract

Bacteriophage contamination of starter culture and raw material poses a major problem in the fermentation industry. In this study, a rapid detection of lytic phage contamination in starter culture using water-in-oil-in-water (W/O/W) emulsion microdroplets was described. A model bacteria with varying concentrations of lytic phages were encapsulated in W/O/W emulsion microdroplets using a simple needle-in-tube setup. The detection of lytic phage contamination was accomplished in 1 h using the propidium iodide labeling of the phage-infected bacteria inside the W/O/W emulsion microdroplets. Using this approach, a detection limit of 10(2) PFU/mL of phages was achieved quantitatively, while 10(4) PFU/mL of phages could be detected qualitatively based on visual comparison of the fluorescence images. Given the simplicity and sensitivity of this approach, it is anticipated that this method can be adapted to any strains of bacteria and lytic phages that are commonly used for fermentation, and has potential for a rapid detection of lytic phage contamination in the fermentation industry.

Understanding and controlling the release mechanism of Escherichia coli in double W1/O/W2emulsion globules in the presence of NaCl in the W2phase

“…10 A uorescence-based method for rapid detection of bacteriophages was developed by encapsulating E. coli and lytic phages in W 1 /O/W 2 emulsion. 11 More recently W 1 /O/W 2 emulsions have been used as 3D microenvironments for the containment and growth of bacterial biolms. 12 However, there is a lack of understanding of the mechanism by which bacteria are released from the W 1 phase of W 1 /O/W 2 emulsions.…”

Fast phage detection and quantification: An optical density-based approach

“…The growing interest in phage monitoring in these fields has prompted the development of a new generation of agile and sensitive methods able to overcome the limitations derived from DAL. These methods are based either on the direct detection of viral particles by PCR [21], qPCR [22, 23], Raman spectroscopy [24], immunoassay [25, 26], MALDI-TOF [27, 28], or on the lysis of the host organism by flow cytometry [29], fluorescence microscopy [30], enzyme release [13, 31, 32], surface plasmon resonance (SPR) [33, 34] or impedance measurements [35]. Sophisticated as they are, many of these methods do not match the sensitivity and precision of the DAL method.…”

Multiplexed and continuous microfluidic sensors using dynamic complex droplets

“…[1][2][3][4] In the context of sensing applications, these all-liquid materials are extremely sensitive to changes in interfacial tensions and are able to dynamically and reversibly transform their physical morphology and optical properties in the presence of chemical analytes. [5][6][7][8][9][10][11] For example, they have previously been demonstrated as effective detection platforms for biological analytes-such as E. coli, 12,13 Listeria monocytogenes, 14,15 SARS-CoV-2, 16,17 and bacteriophages 18 -as well as environmental contaminants. 19,20 While these examples served as successful proofs-of-concept, crucial limitations exist that prevent the development of an on-site sensing platform for real-world samples and situations.…”