Portable Rice Disease Spores Capture and Detection Method Using Diffraction Fingerprints on Microfluidic Chip

Crop diseases cause great harm to food security, 90% of these are caused by fungal spores. This paper proposes a crop diseases spore detection method, based on the lensfree diffraction fingerprint and microfluidic chip. The spore diffraction images are obtained by a designed large field of view lensless diffraction detection platform which contains the spore enrichment microfluidic chip and lensless imaging module. By using the microfluidic chip to enrich and isolate spores in advance, the required particles can be captured in the chip enrichment area, and other impurities can be filtered to reduce the interference of impurities on spore detection. The light source emits partially coherent light and irradiates the target to generate diffraction fingerprints, which can be used to distinguish spores and impurities. According to the theoretical analysis, two parameters, Peak to Center ratio (PCR) and Peak to Valley ratio (PVR), are found to quantify these spores. The correlation coefficient between the detection results of rice blast spores by the constructed device and the results of microscopic artificial identification was up to 0.99, and the average error rate of the proposed device was only 5.91%. The size of the device is only 4 cm × 4 cm × 5 cm, and the cost is less than $150, which is one thousandth of the existing equipment. Therefore, it may be widely used as an early detection method for crop disease caused by spores.

Bacterial spore detection and analysis using hyperpolarized 129Xe chemical exchange saturation transfer (Hyper-CEST) NMR

Hyper-CEST 129Xe NMR spectroscopy was employed to detect Bacillus anthracis and Bacillus subtilis spores in solution, and interrogate the layers that comprise their structures.

Anthrax Spore Detection by a Luminex Assay Based on Monoclonal Antibodies That Recognize Anthrose-Containing Oligosaccharides

ABSTRACT The similarity of endospore surface antigens between bacteria of the Bacillus cereus group complicates the development of selective antibody-based anthrax detection systems. The surface of B. anthracis endospores exposes a tetrasaccharide containing the monosaccharide anthrose. Anti-tetrasaccharide monoclonal antibodies (MAbs) and anti-anthrose-rhamnose disaccharide MAbs were produced and tested for their fine specificities in a direct spore enzyme-linked immunosorbent assay (ELISA) with inactivated spores of a broad spectrum of B. anthracis strains and related species of the Bacillus genus. Although the two sets of MAbs had different fine specificities, all of them recognized the tested B. anthracis strains and showed only a limited cross-reactivity with two B. cereus strains. The MAbs were further tested for their ability to be implemented in a highly sensitive and specific bead-based Luminex assay. This assay detected spores from different B. anthracis strains and two cross-reactive B. cereus strains, correlating with the results obtained in direct spore ELISA. The Luminex assay (detection limit 103 to 104 spores per ml) was much more sensitive than the corresponding sandwich ELISA. Although not strictly specific for B. anthracis spores, the developed Luminex assay represents a useful first-line screening tool for the detection of B. anthracis spores.

Development of Antibodies against Anthrose Tetrasaccharide for Specific Detection of Bacillus anthracis Spores

ABSTRACT Methods for the immunological detection of Bacillus anthracis in various environmental samples and the discrimination of B. anthracis from other members of the B. cereus group are not yet well established. To generate specific discriminating antibodies, we immunized rabbits, mice, and chickens with inactivated B. anthracis spores and, additionally, immunized rabbits and mice with the tetrasaccharide β-Ant-(1→3)-α-l-Rhap-(1→3)-α-l-Rhap-(1→2)-l-Rhap. It is a constituent of the exosporium glycoprotein BclA and contains the newly discovered sugar anthrose 2-O-methyl-4-(3-hydroxy-3-methylbutamido)-4,6-dideoxy-β-d-glucose. The BclA protein is a major component of the exosporium of B. anthracis spores and is decorated by the tetrasaccharide indicated above. The anthrose-containing tetrasaccharide chain seems to be highly specific for B. anthracis, which makes it a key biomarker for the detection of these spores. The different immunizations led to anthrose-reactive polyclonal and monoclonal antibodies which were analyzed by various methods to characterize their ability to discriminate between B. anthracis and other Bacillus spp. Multiple applications, such as enzyme-linked immunosorbent assay, indirect immunofluorescence assay, and electron microscopy, revealed the specificities of the polyclonal and monoclonal antibodies generated for B. anthracis spore detection. All polyclonal antibodies were able to correctly identify the B. anthracis strains tested and showed only minimal cross-reactivities with other Bacillus strains. Moreover, the antibodies generated proved functional in a new capture assay for B. anthracis spores and could therefore be useful for the detection of spores in complex samples.