Combining Direct PCR Technology and Capillary Electrophoresis for an Easy-to-Operate and Highly Sensitive Infectious Disease Detection System for Shrimp.

Infectious diseases are considered the greatest threat to the modern high-density shrimp aquaculture industry. Specificity, rapidity, and sensitivity of molecular diagnostic methods for the detection of asymptomatic infected shrimp allows preventive measures to be taken before disease outbreaks. Routine molecular detection of pathogens in infected shrimp can be made easier with the use of a direct polymerase chain reaction (PCR). In this study, four direct PCR reagent brands were tested, and results showed that the detection signal of direct PCR in hepatopancreatic tissue was more severely affected. In addition, portable capillary electrophoresis was applied to improve sensitivity and specificity, resulting in a pathogen detection limit of 25 copies/PCR-reaction. Juvenile shrimp from five different aquaculture ponds were tested for white spot syndrome virus infection, and the results were consistent with the Organization for Animal Health's certified standard method. Furthermore, this methodology could be used to examine single post larvae shrimp. The overall detection time was reduced by more than 58.2%. Therefore, the combination of direct PCR and capillary electrophoresis for on-site examination is valuable and has potential as a suitable tool for diagnostic, epidemiological, and pathological studies of shrimp aquaculture.

How to evaluate the potential toxicity of therapeutic carbon nanomaterials? A comprehensive study of carbonized nanogels with multiple animal toxicity test models.

Carbon-based nanomaterials have great potential in medical applications, especially in the treatment of infectious diseases and even tumors. However, to safely execute the application of carbon nanomaterials in human treatments, conducting safety assessments and establishing suitable evaluation criteria are necessary. In this study, lysine-carbonized nanogels (Lys-CNGs) that display antibacterial and antiviral abilities were employed in a comprehensive evaluation of their toxicity profiles through assessments in different animal models and growth stages. It was observed that zebrafish at the embryo and eleutheroembryo stages experienced significant toxic effects at a concentration of 15-fold the recommended dosage (0.5 ppm), whereas adult zebrafish following long-term consumption of fodder containing Lys-CNGs presented no adverse effects. Further microbiota analysis indicated that Lys-CNGs did not cause significant changes in the composition of the intestinal bacteria. In contrast, in the toxicity assessments with mammalian animal models, the Lys-CNGs showed no adverse effects, such as weight loss, dermal irritation, and skin sensitization responses in rabbits and guinea pigs, even at a high dose of 2000 mg/kg body weight. Our study revealed that Lys-CNGs have different toxic effects on different growth stages of zebrafish. Researchers in this field should carefully consider the implications of these toxicity profiles during the development of therapeutic carbon-based nanomaterials and for comparison of studies.

Multifunctional carbonized nanogels to treat lethal acute hepatopancreatic necrosis disease.

BACKGROUND: Shrimp aquaculture has suffered huge economic losses over the past decade due to the outbreak of acute hepatopancreatic necrosis disease (AHPND), which is mainly caused by the bacteria Vibrio parahaemolyticus (V. parahaemolyticus) with the virulence pVA1 plasmid, which encodes a secretory photorhabdus insect-related (Pir) toxin composed of PirA and PirB proteins. The Pir toxin mainly attacks the hepatopancreas, a major metabolic organ in shrimp, thereby causing necrosis and loss of function. The pandemic of antibiotic-resistant strains makes the impact worse. METHODS: Mild pyrolysis of a mixture of polysaccharide dextran 70 and the crosslinker 1,8-diaminooctane at 180 â„ƒ for 3 h to form carbonized nanogels (DAO/DEX-CNGs) through controlled cross-linking and carbonization. The multifunctional therapeutic CNGs inherit nanogel-like structures and functional groups from their precursor molecules. RESULTS: DAO/DEX-CNGs manifest broad-spectrum antibacterial activity against Vibrio parahaemolyticus responsible for AHPND and even multiple drug-resistant strains. The polymer-like structures and functional groups on graphitic-carbon within the CNGs exhibit multiple treatment effects, including disruption of bacterial membranes, elevating bacterial oxidative stress, and neutralization of PirAB toxins. The inhibition of Vibrio in the midgut of infected shrimp, protection of hepatopancreas tissue from Pir toxin, and suppressing overstimulation of the immune system in severe V. parahaemolyticus infection, revealing that CNGs can effectively guard shrimp from Vibrio invasion. Moreover, shrimps fed with DAO/DEX-CNGs were carefully examined, such as the expression of the immune-related genes, hepatopancreas biopsy, and intestinal microbiota. Few adverse effects on shrimps were observed. CONCLUSION: Our work proposes brand-new applications of multifunctional carbon-based nanomaterials as efficient anti-Vibrio agents in the aquatic industry that hold great potential as feed additives to reduce antibiotic overuse in aquaculture.