Detection of white spot syndrome virus in seafood samples using a magnetosome-based impedimetric biosensor.

White spot syndrome virus (WSSV) is a significant threat to the aquaculture sector, causing mortality among crabs and shrimps. Currently available diagnostic tests for WSSV are not rapid or cost-effective, and a new detection method is therefore needed. This study demonstrates the development of a biosensor by functionalization of magnetosomes with VP28-specific antibodies to detect WSSV in seafood. The magnetosomes (1 and 2 mg/ml) were conjugated with VP28 antibody (0.025-10 ng/µl), as confirmed by spectroscopy. The magnetosome-antibody conjugate was used to detect the VP28 antigen. The binding of antigen to the magnetosome-antibody complex resulted in a change in absorbance. The magnetosome-antibody-antigen complex was then concentrated and brought near a screen-printed carbon electrode by applying an external magnetic field, and the antigen concentration was determined using impedance measurements. The VP28 antigen (0.025 ng/µl) bound more efficiently to the magnetosome-VP28 antibody complex (0.025 ng/µl) than to the VP28 antibody (0.1 ng/µl) alone. The same assay was repeated to detect the VP28 antigen (0.01 ng/µl) in WSSV-infected seafood samples using the magnetosome-VP28 antibody complex (0.025 ng/µl). The WSSV in the seafood sample was also drawn toward the electrode due to the action of magnetosomes controlled by the external magnetic field and detected using impedance measurement. The presence of WSSV in seafood samples was verified by Western blot and RT-PCR. Cross-reactivity assays with other viruses confirmed the specificity of the magnetosome-based biosensor. The results indicate that the use of the magnetosome-based biosensor is a sensitive, specific, and rapid way to detect WSSV in seafood samples.

Magnetosome-anti-Salmonella antibody complex based biosensor for the detection of Salmonella typhimurium.

Epidemic Salmonellosis contracted through the consumption of contaminated food substances is a global concern. Thus, simple and effective diagnostic methods are needed. Magnetosome-based biosensors are gaining attention because of their promising features. Here, we developed a biosensor employing a magnetosome-anti-Salmonella antibody complex to detect lipopolysaccharide (somatic "O" antigen) and Salmonella typhimurium in real samples. Magnetosome was extracted from Magnetospirillum sp. RJS1 and characterized by microscopy. The magnetosome samples (1 and 2 mg/mL) were directly conjugated to anti-Salmonella antibody (0.8-200 µg/mL) and confirmed by spectroscopy and zeta potential. The concentrations of magnetosome, antibody and lipopolysaccharide were optimized by ELISA. The 2 mg/mL-0.8 µg/mL magnetosome-antibody complex was optimal for detecting lipopolysaccharide (0.001 µg/mL). Our assay is a cost-effective (60%) and sensitive (50%) method in detection of lipopolysaccharide. The optimized magnetosome-antibody complex was applied to an electrode surface and stabilized using an external magnetic field. Increased resistance confirmed the detection of lipopolysaccharide (at 0.001-0.1 µg/mL) using impedance spectroscopy. Significantly, the R2 value was 0.960. Then, the developed prototype biosensor was applied to food and water samples. ELISA confirmed the presence of lipopolysaccharide in homogenized infected samples and cross reactivity assays confirmed the specificity of the biosensor. Further, the biosensor showed low detection limit (101 CFU/mL) in water and milk sample demonstrating its sensitivity. Regression coefficient of 0.974 in water and 0.982 in milk was obtained. The magnetosome-antibody complex captured 90% of the S. typhimurium in real samples which was also confirmed in FE-SEM. Thus, the developed biosensor is selective, specific, rapid and sensitive for detection of S. typhimurium.

Development of magnetosomes-based biosensor for the detection of Listeria monocytogenes from food sample.

Listeriosis through contaminated food is one of the leading causes of premature deaths in pregnant women and new born babies. Here, the authors have developed a magnetosomes-based biosensor for the rapid, sensitive, specific and cost-effective detection of Listeria monocytogenes from food sample. Magnetosomes were extracted from Magnetospirillum sp. RJS1 and then directly bound to anti-Listeriolysin antibody (0.25-1 µg/ml), confirmed in spectroscopy. Listeriolysin (LLO) protein (0.01-7 µg/ml) was optimised in enzyme-linked immunosorbent assay. Magnetosomes was conjugated with LLO antibody (0.25 µg/ml) in optimum concentration to detect LLO protein (0.01 µg/ml). Magnetosomes-LLO antibody complex was 25% cost effective. The magnetosomes-LLO antibody complex was directly stabilised on screen printed electrode using external magnet. The significant increase in resistance (RCT value) on the electrode surface with increase in concentration of LLO protein was confirmed in impedance spectroscopy. The L. monocytogenes contaminated milk and water sample were processed and extracted LLO protein was detected in the biosensor. The specificity of the biosensor was confirmed in cross-reactivity assay with other food pathogens. The detection limit of 101 Cfu/ml in both water and milk sample manifests the sensitive nature of the biosensor. The capture efficiency and field emission scanning electron microscopy confirmed positive interaction of Listeria cells with magnetosomes-antibody complex.