CRISPR/Cas12a-Responsive Smart DNA Hydrogel for Sensitive Electrochemiluminescence Detection of the Huanglongbing Outer Membrane Protein Gene.

Citrus Huanglongbing (HLB) is known as the cancer of citrus, where Candidatus Liberibacter asiaticus (CLas) is the most prevalent strain causing HLB. In this study, we report a novel electrochemiluminescence (ECL) biosensor for the highly sensitive detection of the CLas outer membrane protein (Omp) gene by coupling rolling circle amplification (RCA) with a CRISPR/Cas12a-responsive smart DNA hydrogel. In the presence of the target, a large number of amplicons are generated through RCA. The amplicons activate the trans-cleavage activity of CRISPR/Cas12a through hybridizing with crRNA, triggering the response of smart DNA hydrogel to release the encapsulated AuAg nanoclusters (AuAg NCs) on the electrode and therefore leading to a decreased ECL signal. The ECL intensity change (I0 - I) is positively correlated with the concentration of the target in the range 50 fM to 5 nM, with a limit of detection of 40 fM. The performance of the sensor has also been evaluated with 10 samples of live citrus leaves (five HLB negative and five HLB positive), and the result is in excellent agreement with the gold standard qPCR result. The sensing strategy has expanded the ECL versatility for detecting varying levels of dsDNA or ssDNA in plants with high sensitivity.

Effect of macrophage polarization on parasitic protection against type 1 diabetes mellitus.

Type 1 diabetes mellitus is a chronic disease caused by the destruction of pancreatic beta cells. Based on the hygiene hypothesis, a growing body of evidence suggests a negative association between parasitic infections and diabetes in humans and animal models. The mechanism of parasite-mediated prevention of type 1 diabetes mellitus may be related to the adaptive and innate immune systems. Macrophage polarization is a new paradigm for the treatment of type 1 diabetes mellitus, and different host macrophage subsets play various roles during parasite infection. Proinflammatory cytokines are released by M1 macrophages, which are important in the development of type 1 diabetes mellitus. Parasite-activated M2 macrophages prevent the development of type 1 diabetes mellitus and can influence the development of adaptive immune responses through several mechanisms, including Th2 cells and regulatory T cells. Here, we review the role and mechanism of macrophage polarization in parasitic protection against type 1 diabetes mellitus.

Immunization with recombinant schistosome adenylate kinase 1 partially protects mice against Schistosoma japonicum infection.

Highly effective and safe prophylactic vaccines are urgently needed to sustainably control schistosomiasis, one of the most serious endemic zoonoses in China. In this study, we characterized adenylate kinase 1 from Schistosoma japonicum (SjAK1), a phosphotransferase that regulates cellular energy and metabolism, and evaluated its potential as a recombinant vaccine. Based on real-time quantitative PCR, western blot, and immunolocalization, SjAK1 is active throughout the life of the worm, although its expression is higher in 21-day-old schistosomula, adult worms, and eggs deposited in the host liver. Further, the enzyme accumulates in the eggshell, intestinal epithelium, integument of adult worms and in the vitellaria tissue in female worms. A 594-bp full-length complementary DNA (cDNA) encoding SjAK1 was synthesized from total RNA of 3-day-old schistosomes, and immunization with recombinant SjAK1 reduced worm burden by 50%, decreased the density of eggs deposited in the host liver by 40%, and reduced the area of granulomas in the host liver by 56%. ELISA results showed that recombinant SjAK1 also stimulated Th1 cytokines such as IL-2 and IFN-γ, but not IL-5 and IL-4. Collectively, a recombinant form of the enzyme SjAK1 elicits partial protective immunity against Schistosoma japonicum infection and the induction of Th1 cytokines. Thus, the enzyme has potential as a component of a multivalent vaccine against schistosomiasis.