Rapid and effective detection of Macrobrachium rosenbergii nodavirus using a combination of nucleic acid sequence-based amplification test and immunochromatographic strip.

Nucleic acid sequence-based amplification (NASBA) provides a fast and convenient approach for nucleic acid amplification under isothermal conditions, and its combination with an immunoassay-based lateral flow dipstick (LFD) could produce a higher detection efficiency for M. rosenbergii nodavirus isolated from China (MrNV-chin). In this study, two specific primers and a labelled probe of the capsid protein gene of MrNV-chin were constructed. The process of this assay mainly included a single-step amplification at a temperature of 41 â„ƒ for 90 min, and hybridization with an FITC-labeled probe for 5 min, with the hybridization been required for visual identification during LFD assay. The test results indicated that, the NASBA-LFD assay showed sensitivity for 1.0 fg M. rosenbergii total RNA with MrNV-chin infection, which was 104 times that of the present RT-PCR approach for the detection of MrNV. In addition, no products were created for shrimps with infection of other kinds of either DNA or RNA virus, which indicated that the NASBA-LFD was specific for MrNV. Therefore, the combination of NASBA and LFD is a new alternative detection method for MrNV which is rapid, accurate, sensitive and specific without expensive equipment and specialised personnel. Early detection of this infectious disease among aquatic organisms will help implement efficient therapeutic strategy to prevent its spread, enhance animal health and limit loss of aquatic breeds in the event of an outbreak.

Geographical Origin Differentiation of Rice by LC-MS-Based Non-Targeted Metabolomics.

Many factors, such as soil, climate, and water source in the planting area, can affect rice taste and quality. Adulterated rice is common in the market, which seriously damages the production and sales of high-quality rice. Traceability analysis of rice has become one of the important research fields of food safety management. In this study, LC-MS-based non-targeted metabolomics technology was used to trace four rice samples from Heilongjiang and Jiangsu Provinces, namely, Daohuaxiang (DH), Huaidao No. 5 (HD), Songjing (SJ), and Changlixiang (CL). Results showed that the discrimination accuracy of the partial least squares discriminant analysis (PLS-DA) model was as high as 100% with satisfactory prediction ability. A total of 328 differential metabolites were screened, indicating significant differences in rice metabolites from different origins. Pathway enrichment analysis was carried out on the four rice samples based on the KEGG database to determine the three metabolic pathways with the highest enrichment degree. The main biochemical metabolic pathways and signal transduction pathways involved in differential metabolites in rice were obtained. This study provides theoretical support for the geographical origins of rice and elucidates the change mechanism of rice metabolic pathways, which can shed light on improving rice quality control.

Discrimination of Tetrastigma hemsleyanum according to geographical origin by near-infrared spectroscopy combined with a deep learning approach.

Recently, deep learning has presented as a powerful approach to overcome the deficiencies of the conventional biochemical approaches. In this study, a method for discriminating medicinal plant Tetrastigma hemsleyanum from different origins was proposed using near-infrared spectroscopy (NIRS) and deep learning models. Support vector machine (SVM), self-adaptive evolutionary extreme learning machine (SAE-ELM), and convolutional neural network (CNN) were used to process the near-infrared spectral data (4000-5600 cm-1). The results indicated that the average recognition accuracy of SVM on the test set samples (n = 60) reached 90%. The average recognition accuracy of SAE-ELM was 98.3%, while CNN correctly discriminated 100% of T. hemsleyanum from different origins. Notably, CNN avoids tedious redundant data preprocessing and is also able to save the trained model for the next call to achieve rapid detection. As above, this study provides an effective deep learning-based method for discriminating the geographical origins of T. hemsleyanum as well as providing a convenient and satisfactory approach to ensure the famous-region of other medicinal plants.

Neurotoxic Anatoxin-a Can Also Exert Immunotoxicity by the Induction of Apoptosis on Carassius auratus Lymphocytes in vitro When Exposed to Environmentally Relevant Concentrations.

Hazardous anatoxin-a (ANTX-a) is produced by freshwater algal blooms worldwide, which greatly increases the risk of consumer exposure. Although ANTX-a shows widespread neurotoxicity in aquatic animals, little is known about its mechanism of action and biotransformation in biological systems, especially in immunobiological models. In this study, transmission electron microscopy results showed that ANTX-a can destroy lymphocytes of Carassius auratus in vitro by inducing cytoplasmic concentration, vacuolation, and swollen mitochondria. DNA fragmentations clearly showed a ladder pattern in agarose gel electrophoresis, which demonstrated that the apoptosis of fish lymphocytes was caused by exposure to ANTX-a. Flow cytometry results showed that the apoptotic percentage of fish lymphocytes exposed to 0.01, 0.1, 1, and 10 mg/L of ANTX-a for 12 h reached 18.89, 22.89, 39.23, and 35.58%, respectively. ANTX-a exposure induced a significant increase in reactive oxygen species (ROS) and malonaldehyde (MDA) in lymphocytes. The activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), and the glutathione (GSH) content of the 0.01 mg/L ANTX-a-treated group decreased significantly by about 41, 46, 67, and 54% compared with that of the control group (p < 0.01), respectively. Although these observations were dose-dependent, these results suggested that ANTX-a can induce lymphocyte apoptosis via intracellular oxidative stress and destroy the antioxidant system after a short exposure time of only 12 h. Besides neurotoxicity, ANTX-a may also be toxic to the immune system of fish, even when the fish are exposed to environmentally relevant concentrations, which clearly demonstrated that the potential health risks induced by ANTX-a in aquatic organisms requires attention.

Mechanism of immunosuppression in zebrafish (Danio rerio) spleen induced by environmentally relevant concentrations of perfluorooctanoic acid.

Perfluorooctanoic acid (PFOA) has been identified as a new persistent organic pollutant. This pollutant is ubiquitous in water and environments. Although PFOA is toxic to fishes, the precise immunotoxicological mechanism remains unclear. In this study, HPLC-MS analysis proved that PFOA can accumulate in the spleen of zebrafish. As comparison of 7-day and 14-day data, the cumulative content in the spleen significantly increased by 26% even in the 0.1 mg/L PFOA-treated group. Morphological observations revealed that PFOA can damage immune cells in zebrafish spleen by inducing vacuolization, lipofuscin granule production, and mitochondrial swelling. The Toll-like receptor 2 (TLR2)/myeloid differentiation factor 88 (myd88)/NF-κB (P65) pathway can mediate the mRNA expression levels of interferon (IFN) and B cell-activating factor (BAFF); immunoglobulin (Ig) secretion is further regulated. RT-PCR results indicated that the expression levels of P65 and IFN in the 1 mg/L group after PFOA exposure for 7 d increased by 4.03- and 3.28-fold, respectively, in a dose-dependent manner compared with those of the control group. The linear correlation coefficient (r2) was analyzed, and the results indicated that the Ig-mediated pathway can be affected by PFOA. For example, the r2 between IgD and P65 decreased from 0.641 (7 d) to 0.295 (14 d) after the cells were exposed to PFOA for a prolonged time; the r2 between IgD and IFN increased from 0.562 (7 d) to 0.808 (14 d). The triangle plot method strongly demonstrated that increased PFOA concentration and prolonged exposure to PFOA can inhibit Ig secretion. Therefore, immune organs, particularly the spleen, of zebrafish are vulnerable to PFOA. These results can help to improve the understanding of the possible noncarcinogenic risk mechanisms induced by PFOA.

Effects of Perfluorooctanoic Acid on the Associated Genes Expression of Autophagy Signaling Pathway of Carassius auratus Lymphocytes in vitro.

Perfluorooctanoic acid (PFOA) has been detected in various water bodies and caused harm to aquatic organisms. The aim of this study was to investigate the cytotoxicity and mechanism associated with autophagy and oxidative stress after exposure to PFOA (0, 1, 10, 100 µg/L) for 12 h on lymphocytes, which was isolated from the head kidney of Carassius auratus (C. auratus). Both of autophagy formation, cell activity, and intracellular reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), and superoxide dismutase (SOD) levels were measured. The relative expression of partial autophagy-related genes autophagy related 5 (Atg 5), autophagy related 7 (Atg 7), and Beclin 1 were also cloned and detected. Homologous relationships analysis showed high identities of genes in C. auratus and other fish by blast. C. auratus lymphocytes growth inhibition rates was increased induced by PFOA. Compared with the control group, the ROS generation and the MDA content were significantly increased in all of the PFOA-treated group. Besides, decreased SOD activity and decrease of GSH activity induced by PFOA further confirmed the occurrence of oxidative stress. The number of autophagosome formations was increased in a dose-dependent manner. Compared with the control group, Atg 7 and Beclin 1 mRNA expression was elevated significantly after PFOA exposed, showing a time-dependent manner, while mRNA expression of Atg 5 was increased remarkably in 100 µg/L PFOA-treated group. Our results indicated that PFOA caused oxidative damage to lymphocytes in C. auratus and caused various autophagy signaling pathway-associated genes imbalances in the lymphocytes. Autophagy signaling pathway-associated genes imbalance could weaken antioxidant capacity and involve in the mechanism of C. auratus lymphocytes oxidative injury caused by PFOA.