2′-Fucosyllactose (2′-FL), a major component of fucosylated human milk oligosaccharides, is beneficial to human health in various ways like prebiotic effect, protection from pathogens, anti-inflammatory activity and reduction of the risk of neurodegeneration. Here, a whole-cell fluorescence biosensor for 2′-FL was developed. Escherichia coli (E. coli) was engineered to catalyse the cleavage of 2′-FL into l-fucose and lactose by constitutively expressing α-l-fucosidase. Escherichia coli ∆L YA, in which lacZ is deleted and lacY is retained, was employed to disable lactose consumption. E. coli ∆L YA constitutively co-expressing α-l-fucosidase and a red fluorescence protein (RFP) exhibited increased fluorescence intensity in media containing 2′-FL. However, the presence of 50 g/L lactose reduced the RFP intensity due to lactose-induced cytotoxicity. Preadaptation of bacterial strains to fucose alleviated growth hindrance by lactose and partially recovered the fluorescence intensity. The fluorescence intensity of the cell was linearly proportional to 1-5 g/L 2′-FL. The whole-cell sensor will be versatile in developing a 2′-FL detection system. Human milk oligosaccharides (HMOs) are present in human breast milk and are closely associated with health benefits. HMOs act as decoys for pathogens (e.g., virus, bacteria, and protozoa) by inhibiting their ability to bind to the surface of epithelial cells 1-4. Fucosylated oligosaccharides which are not found in bovine milk account for 50% of total HMOs. The HMOs 2′-fucosyllactose (2′-FL; Fuc-α1,2-Gal-β1,4-Glc) and 3-fucosyllactose (3-FL; Gal-β1,4-Fuc-α1,3-Glc), with a concentration range of 0.5-2 g/L, account for the largest portion of fucosylated HMOs 5,6. Fucosylated HMOs circulate systemically, affecting the host immune response, the regulation of tumour metastasis, and resistance to bacteria, fungi, and other pathogens 2,7,8. The concentration of 2′-FL in breast milk affects the ability to protect against vital systemic infections in nursing infant 8. While low content of 2′-FL in breast milk has been associated with a higher rate of diarrhoea during lactation 9,10 , about 20% of human milk do not contain 2′-FL 11. For these reasons, 2′-FL and 3-FL are spotlighted as nutraceutical and pharmaceutical ingredients; thus, a simple and visually measurable method is indispensable in evaluating their level in breast milk. Various methods have been developed for the production of 2′-FL, including whole-cell biocatalysis 12-20 , enzymatic synthesis 21,22 , and chemical synthesis 23,24. Regardless of the method used, a simple detection and quantification method is indispensable for the development of 2′-FL production. Quantification of 2′-FL has been mostly done using high-performance liquid chromatography (HPLC), high-pH anion exchange chromatography, and liquid chromatography-mass spectrometry (LC-MS) 15,25-28. However, those types of equipment are time-consuming, labour-intensive, and expensive. Therefore, those methods are unlikely to be used in a high-throughput manner or for ...
