Enhancing Security and Flexibility in the Industrial Internet of Things: Blockchain-Based Data Sharing and Privacy Protection.

To address the complexities, inflexibility, and security concerns in traditional data sharing models of the Industrial Internet of Things (IIoT), we propose a blockchain-based data sharing and privacy protection (BBDSPP) scheme for IIoT. Initially, we characterize and assign values to attributes, and employ a weighted threshold secret sharing scheme to refine the data sharing approach. This enables flexible combinations of permissions, ensuring the adaptability of data sharing. Subsequently, based on non-interactive zero-knowledge proof technology, we design a lightweight identity proof protocol using attribute values. This protocol pre-verifies the identity of data accessors, ensuring that only legitimate terminal members can access data within the system, while also protecting the privacy of the members. Finally, we utilize the InterPlanetary File System (IPFS) to store encrypted shared resources, effectively addressing the issue of low storage efficiency in traditional blockchain systems. Theoretical analysis and testing of the computational overhead of our scheme demonstrate that, while ensuring performance, our scheme has the smallest total computational load compared to the other five schemes. Experimental results indicate that our scheme effectively addresses the shortcomings of existing solutions in areas such as identity authentication, privacy protection, and flexible combination of permissions, demonstrating a good performance and strong feasibility.

Photochemically induced chloromethylation/cyclization of benzimidazole derivatives with CCl4/CHCl3.

Herein, it is reported that a series of trichloromethyl/dichloromethyl substituted benzimidazole derivatives have been synthesized by dechlorination of CCl4/CHCl3 to form polychloromethyl radicals and cyclization with an unactivated olefin under a purple LED lamp. The protocol features a wide substrate scope, high atom economy, and excellent regioselectivity, and is easy to scale up.

Mapping and characterization of the Rx3 gene for resistance to Xanthomonas euvesicatoria pv. euvesicatoria race T1 in tomato.

KEY MESSAGE: Rx3 encodes a typical CC-NBS-LRR resistance protein and confers the resistance to Xanthomonas euvesicatoria pv. euvesicatoria race T1 causing bacterial spot in tomato. Bacterial spot caused by at least four species of Xanthomonas is an epidemic disease severely affecting tomato production worldwide. The use of resistant cultivars is an economical and effective approach to control the disease. An unimproved tomato breeding line Hawaii 7988 has been considered as the most reliable source for resistance to X. euvesicatoria pv. euvesicatoria race T1, and the Rx3 locus located at a 4.53-Mb region on chromosome 5 (SL4.0) is the major locus for resistance to race T1 in this line. In the current study, the Rx3 locus was firstly located to a 1.05-Mb region based on comparisons of marker polymorphisms between the susceptible line Ohio 88119 and resistant lines Hawaii 7998, Ohio 9834 and FG02-7530. Using recombinant inbred lines (F5:6, F6:7, and F7:8) derived from a cross between Ohio 88119 and Ohio 9834, the Rx3 locus was finally mapped to a 64.3-kb interval between markers MG-Rx3-4 and MG-Rx3-A6. The Solyc05g053980 gene, designated as Rx3, encoding a coiled-coil nucleotide-binding leucine-rich repeat protein was considered as the candidate for the Rx3 locus. Expression of the gene could be induced by the infection of race T1 strain. Knockout of the Solyc05g053980 gene through CRISPR/Cas9 editing system in the resistant line FG02-7530 decreased resistance to race T1 strain. These results provide a close step for understanding the resistance mechanism to race T1 in Hawaii 7998 and guide tomato breeders accordingly to improve bacterial spot disease resistance in tomato.

Sesquiterpene Lactones from Sigesbeckia glabrescens Possessing Potent Anti-inflammatory Activity by Directly Binding to IKKα/ß.

Chromatographic fractionation of Sigesbeckia glabrescens led to the identification of 10 new sesquiterpene lactones, named siegesbeckialides I-O (1-7) and glabrescones A-C (8-10), along with 14 known analogues. An anti-inflammatory activity assay showed that siegesbeckialide I (1) most potently inhibited LPS-induced NO production in RAW264.7 murine macrophages. Furthermore, siegesbeckialide I suppressed the protein expression of iNOS and COX2, as well as the release of PGE2, IL-1ß, IL-6, and TNF-α in LPS-stimulated RAW264.7 cells. Mechanistically, siegesbeckialide I directly binds to inhibitors of IKKα/ß and suppresses their phosphorylation. This leads to the inhibition of IKKα/ß-mediated phosphorylation and degradation of inhibitor α of NF-κB (IκBα), as well as the activation of NF-κB signaling.

The calcium-dependent protein kinase ZmCDPK7 functions in heat-stress tolerance in maize.

Global warming poses a serious threat to crops. Calcium-dependent protein kinases (CDPKs)/CPKs play vital roles in plant stress responses, but their exact roles in plant thermotolerance remains elusive. Here, we explored the roles of heat-induced ZmCDPK7 in thermotolerance in maize. ZmCDPK7-overexpressing maize plants displayed higher thermotolerance, photosynthetic rates, and antioxidant enzyme activity but lower H2 O2 and malondialdehyde (MDA) contents than wild-type plants under heat stress. ZmCDPK7-knockdown plants displayed the opposite patterns. ZmCDPK7 is attached to the plasma membrane but can translocate to the cytosol under heat stress. ZmCDPK7 interacts with the small heat shock protein sHSP17.4, phosphorylates sHSP17.4 at Ser-44 and the respiratory burst oxidase homolog RBOHB at Ser-99, and upregulates their expression. Site-directed mutagenesis of sHSP17.4 to generate a Ser-44-Ala substitution reduced ZmCDPK7's enhancement of catalase activity but enhanced ZmCDPK7's suppression of MDA accumulation in heat-stressed maize protoplasts. sHSP17.4, ZmCDPK7, and RBOHB were less strongly upregulated in response to heat stress in the abscisic acid-deficient mutant vp5 versus the wild type. Pretreatment with an RBOH inhibitor suppressed sHSP17.4 and ZmCDPK7 expression. Therefore, abscisic acid-induced ZmCDPK7 functions both upstream and downstream of RBOH and participates in thermotolerance in maize by mediating the phosphorylation of sHSP17.4, which might be essential for its chaperone function.

Novel SCN9A missense mutations contribute to congenital insensitivity to pain: Unexpected correlation between electrophysiological characterization and clinical phenotype.

Congenital insensitivity to pain (OMIM 243000) is an extremely rare disorder caused by loss-of-function mutations in SCN9A encoding Nav1.7. Although the SCN9A mutations and phenotypes of painlessness and anosmia/hyposmia in patients are previously well documented, the complex relationship between genotype and phenotype of congenital insensitivity to pain remains unclear. Here, we report a congenital insensitivity to pain patient with novel SCN9A mutations. Functional significance of novel SCN9A mutations was assessed in HEK293 cells expressing Nav1.7, the results showed that p.Arg99His significantly decreased current density and reduced total Nav1.7 protein levels, whereas p.Trp917Gly almost abolished Nav1.7 sodium current without affecting its protein expression. These revealed that mutations in Nav1.7 in this congenital insensitivity to pain patient still retained partial channel function, but the patient showed completely painlessness, the unexpected genotypic-phenotypic relationship of SCN9A mutations in our patient may challenge the previous findings "Nav1.7 total loss-of-function leads to painlessness." Additionally, these findings are helpful for understanding the critical amino acid for maintaining function of Nav1.7, thus contributing to the development of Nav1.7-targeted analgesics.

CircRNAs as potential biomarkers for the clinicopathology and prognosis of glioma patients: a meta-analysis.

BACKGROUND: An increasing number of studies have reported circular RNAs (circRNAs) as new potential biomarkers for the prognosis of gliomas. However, the overall prognostic value of circRNAs for glioma remains unclear. Therefore, this study is the first comprehensive evaluation of the clinicopathological and prognostic value of dysregulated circRNAs in the treatment of glioma patients. METHODS: We systematically reviewed the online databases of PubMed, Web of Science, EMBASE, and Cochrane Library to identify studies that explored the relationship between circRNA expression and clinicopathological and prognostic factors in glioma through April 11, 2020. The quality of the included studies was evaluated by the Newcastle-Ottawa Scale (NOS) checklists. Clinicopathological features were assessed by pooled odds ratios (ORs) and 95% confidence intervals (CIs), and overall survival (OS) was assessed by hazard ratios (HRs) and 95% CIs. RESULTS: Twenty-four eligible studies, including 22 studies of clinicopathological features, 1 diagnostic study, and 18 studies of prognosis, that included a total of 1390 patients were ultimately included in this study. Meta-analysis showed that highly expressed oncogenic circRNAs were significantly related to poor clinicopathological features (age: P = 0.026; tumor size: P ≤ 0.001; tumor grade: P ≤ 0.001; KPS: P = 0.012) and worse overall survival (OS) (HR = 2.01, 95% CI: 1.61-2.50, P ≤ 0.001). Moreover, we found that highly expressed tumor-suppressor circRNAs were related to better clinicopathological features (gender: P = 0.042; age: P = 0.014; tumor size: P = 0.022; tumor grade: P ≤ 0.001) and longer OS (HR = 2.70, 95% CI: 1.82-3.99, P ≤ 0.001). CONCLUSIONS: In conclusion, there is a significant correlation between the dysregulated expression of circRNAs and the clinicopathology and prognosis of glioma patients.

Production of a Functional Factor, p40, by Lactobacillus rhamnosus GG Is Promoted by Intestinal Epithelial Cell-Secreted Extracellular Vesicles.

The symbiotic relationship between the gut microbiome and the host provides a nutrient-rich environment for gut microbes and has beneficial effects on host health. Although the composition of the gut microbiome is known to be influenced by both host genetics and environmental factors, host effects on the activities and functions of the gut microbial communities remain poorly understood. Intestinal epithelial cells exert front-line responses to gut microbes and contribute to maintaining a healthy intestinal homeostasis. Here, seeking to elucidate whether intestinal epithelial cells modulate Lactobacillus rhamnosus GG (LGG) functions, we examined the production of p40, an LGG-derived secretory protein that protects intestinal epithelial cells against inflammation. We found that growth medium conditioned with colonic epithelial cell-derived components promotes p40 protein synthesis and secretion by LGG and enhances LGG-stimulated protective responses in intestinal epithelial cells. Furthermore, when LGG was cultured with the colonic luminal contents from healthy mice, p40 production was upregulated but was attenuated with luminal contents from mice with intestinal inflammation. Importantly, the colonic epithelial cell-derived components potentiated LGG-produced p40 levels in a mouse model of colitis and enhanced LGG-mediated amelioration of intestinal inflammation in this model. Notably, we found that colonic epithelial cell-secreted extracellular vesicles participate in communicating with LGG and that heat shock protein 90 (HSP90) in these vesicles might mediate the promotion of p40 production. These results reveal a previously unrecognized mechanism by which the anti-inflammatory effect of LGG is reinforced by intestinal epithelial cells and thereby maintains intestinal health.

Difficult intubation in a patient with acute epiglottitis and abscess complicated with cervical necrotizing fasciitis: A case report.

INTRODUCTION: Acute epiglottitis is not uncommon and it can cause high mortality due to airway obstruction. Acute epiglottitis complicated with cervical necrotizing fasciitis has rarely been reported, and it is also a life-threatening disease with a fatality rate of 7% to 50%. PATIENT CONCERNS: A 64-year-old woman presented to our hospital with chief complaints of sore throat and cervical swelling, long with foreign body sensation and hoarseness. Endoscopic laryngoscopy showed erythematous and swollen epiglottis with purulent secretions on the surface. Computed tomography (CT) scan showed swollen epiglottis and swelling of the neck with air- and fluid-containing necrotizing tissue. DIAGNOSES: The diagnosis was acute epiglottitis and abscess complicated with cervical necrotizing fasciitis. INTERVENTIONS: With the patient in awake condition, airway access was established by performing intubation with adjunctive use of gum elastic bougie, followed by surgical debridement under general anesthesia; a flap was used for skin coverage and intravenous piperacillin-tazobactam was administered. OUTCOMES: The patient was discharged without complications. CONCLUSION: Gum elastic bougie is a usable tool in difficult intubation. Adequate pre-anesthesia evaluation, patient sedation, and gentle manipulation assured the intubation success in this case.

Standardized training nurses's humanistic care practice ability: A cross-sectional survey in western China during COVID-19.

AIM: This study aims to investigate the current status and determinants of humanistic care practice abilities among standardized training nurses in China during the post-COVID-19 era, as well as to propose management strategies. DESIGN: A cross-sectional study. METHODS: Deliberately chosen were 517 standardized training nursing students from a provincial-level training facility in western China. RESULTS: The respondents had a mean ± SD age of 21.23 ± 1.34 years, and 92.0% of them voluntarily opted for the nursing profession. Almost all (99.8%) respondents had at least a college degree. The standardized training nurses scored an average of (130.31 ± 14.18) on humanistic care ability, which was significantly related to some sociodemographic variables. The average scores for the five dimensions of nursing communication ability, psychological adjustment ability, moral and legal application ability, nursing aesthetic ability, and care practice ability were 30.78, 17.61, 32.23, 18 and 31.67, respectively. All these dimensions showed positive correlations with the overall score of humanistic care practice ability.

Emerging Drug Delivery Vectors: Engineering of Plant-Derived Nanovesicles and Their Applications in Biomedicine.

Extracellular vesicles can transmit intercellular information and transport biomolecules to recipient cells during various pathophysiological processes in the organism. Animal cell exosomes have been identified as potential nanodrugs delivery vehicles, yet they have some shortcomings such as high immunogenicity, high cytotoxicity, and complicated preparation procedures. In addition to exosomes, plant-derived extracellular vesicles (PDVs), which carry a variety of active substances, are another promising nano-transport vehicles emerging in recent years due to their stable physicochemical properties, wide source, and low cost. This work briefly introduces the collection and characterization of PDVs, then focuses on the application of PDVs as natural or engineered drug carriers in biomedicine, and finally discusses the development and challenges of PDVs in future applications.

Sucralose triggers insulin resistance leading to follicular dysplasia in mice.

Sucralose, the extensively utilized sweetener, might lead to metabolic disorders with prolonged consumption, but it remains uncertain if sucralose has any impact on female reproductive health. We incorporated sucralose into drinking water and observed food intake, body weight, estrous cycle, follicular development, serum hormones, and insulin sensitivity of mice. The mice did not experience any changes in their food intake or body weight after consuming sucralose. However, they displayed irregularities in the estrous cycle, marked by a reduced count of primordial, primary, and secondary follicles, coupled with a significant increase in the number of antral follicles. There was a decline in follicle-stimulating hormone (FSH), estradiol (E2), and progesterone (P4) levels, while testosterone (T) and luteinizing hormone (LH) levels surged, leading to a notable elevation in the LH / FSH ratio. Sucralose also induced insulin resistance, as evidenced by elevated insulin levels and impaired insulin tolerance, which responded to an increase in bacterial-derived serum endotoxin. By eliminating insulin resistance with rosiglitazone (RSG), eradicating intestinal flora-derived endotoxins with neomycin (NEO), or enhancing intestinal barrier function with indole-3-carbinol (I3C), the abnormalities in estrous cycle, disruptions in follicular development, hormonal imbalances and elevation in serum endotoxins induced by sucralose were successfully reversed. The present study indicates that sucralose-induced follicular dysplasia in mice is probably related to impaired intestinal permeability, infiltration of endotoxins, initiation of systemic inflammation, and insulin resistance.

Robust Vascular Segmentation for Raw Complex Images of Laser Speckle Contrast Based on Weakly Supervised Learning.

Laser speckle contrast imaging (LSCI) is widely used for in vivo real-time detection and analysis of local blood flow microcirculation due to its non-invasive ability and excellent spatial and temporal resolution. However, vascular segmentation of LSCI images still faces a lot of difficulties due to numerous specific noises caused by the complexity of blood microcirculation's structure and irregular vascular aberrations in diseased regions. In addition, the difficulties of LSCI image data annotation have hindered the application of deep learning methods based on supervised learning in the field of LSCI image vascular segmentation. To tackle these difficulties, we propose a robust weakly supervised learning method, which selects the threshold combinations and processing flows instead of labor-intensive annotation work to construct the ground truth of the dataset, and design a deep neural network, FURNet, based on UNet++ and ResNeXt. The model obtained from training achieves high-quality vascular segmentation and captures multi-scene vascular features on both constructed and unknown datasets with good generalization. Furthermore, we intravital verified the availability of this method on a tumor before and after embolization treatment. This work provides a new approach for realizing LSCI vascular segmentation and also makes a new application-level advance in the field of artificial intelligence-assisted disease diagnosis.

Unpaired fundus image enhancement based on constrained generative adversarial networks.

Fundus photography (FP) is a crucial technique for diagnosing the progression of ocular and systemic diseases in clinical studies, with wide applications in early clinical screening and diagnosis. However, due to the nonuniform illumination and imbalanced intensity caused by various reasons, the quality of fundus images is often severely weakened, brings challenges for automated screening, analysis, and diagnosis of diseases. To resolve this problem, we developed strongly constrained generative adversarial networks (SCGAN). The results demonstrate that the quality of various datasets were more significantly enhanced based on SCGAN, simultaneously more effectively retaining tissue and vascular information under various experimental conditions. Furthermore, the clinical effectiveness and robustness of this model were validated by showing its improved ability in vascular segmentation as well as disease diagnosis. Our study provides a new comprehensive approach for FP and also possesses the potential capacity to advance artificial intelligence-assisted ophthalmic examination.

Neuroprotective effects of anti-TRAIL-ICG nanoagent and its multimodal imaging evaluation in cerebral ischemia-reperfusion injury.

Cerebral ischemia-reperfusion injury (CIRI) is a major challenge to neuronal survival in acute ischemic stroke (AIS). However, effective neuroprotective agents remain to be developed for the treatment of CIRI. In this work, we have developed an Anti-TRAIL protein-modified and indocyanine green (ICG)-responsive nanoagent (Anti-TRAIL-ICG) to target ischemic areas and then reduce CIRI and rescue the ischemic penumbra. In vitro and in vivo experiments have demonstrated that the carrier-free nanoagent can enhance drug transport across the blood-brain barrier (BBB) in stroke mice, exhibiting high targeting ability and good biocompatibility. Anti-TRAIL-ICG nanoagent played a better neuroprotective role by reducing apoptosis and ferroptosis, and significantly improved ischemia-reperfusion injury. Moreover, the multimodal imaging platform enables the dynamic in vivo examination of multiple morphofunctional information, so that the dynamic molecular events of nanoagent can be detected continuously and in real time for early treatment in transient middle cerebral artery occlusion (tMCAO) models. Furthermore, it has been found that Anti-TRAIL-ICG has great potential in the functional reconstruction of neurovascular networks through optical coherence tomography angiography (OCTA). Taken together, our work effectively alleviates CIRI after stoke by blocking multiple cell death pathways, which offers an innovative strategy for harnessing the apoptosis and ferroptosis against CIRI.

Aircraft route recovery based on distributed integer programming method.

In order to further promote the application and development of unmanned aviation in the manned field, and reduce the difficulty that airlines cannot avoid due to unexpected factors such as bad weather, aircraft failure, and so on, the problem of restoring aircraft routes has been studied. To reduce the economic losses caused by flight interruption, this paper divides the repair problem of aircraft operation plans into two sub problems, namely, the generation of flight routes and the reallocation of aircraft. Firstly, the existing fixed-point iteration method proposed by Dang is used to solve the feasible route generation model based on integer programming. To calculate quickly and efficiently, a segmentation method that divides the solution space into mutually independent segments is proposed as the premise of distributed computing. The feasible route is then allocated to the available aircraft to repair the flight plan. The experimental results of two examples of aircraft fault grounding and airport closure show that the method proposed in this paper is effective for aircraft route restoration.

Novel Mechanisms Underlying Rubber Accumulation and Programmed Cell Death in Laticiferous Canals of Decaisnea insignis Fruits: Cytological and Transcriptomic Analyses.

Natural rubber is one of the most important industrial raw materials, and its biosynthesis is still a fascinating process that is still largely unknown. In this research, we studied Decaisnea insignis, a unique rubber-producing plant that is different from other rubber-producing species due to the presence of lactiferous canals in its pericarp. The present study aims to provide novel insights into the mechanisms underlying rubber accumulation and PCD by subjecting the Decaisnea insignis laticiferous canals to light microscopy, TUNEL assay, and DAPI staining, as well as viability analysis, cellular ultrastructure analysis, and molecular analysis using light microscopy, scanning electron microscopy, immunofluorescence labeling, transmission electron microscopy, and transcriptome sequencing. At the cellular level, the origin of small rubber particles in the laticiferous canals had no morphological correlation with other organelles, and these particles were freely produced in the cytosol. The volume of the rubber particles increased at the sunken and expanding stage, which were identified as having the characteristics of programmed cell death (PCD); meanwhile, plenty of the rubber precursors or rubber particles were engulfed by the vacuoles, indicating a vacuole-mediated autophagy process. The accumulation of rubber particles occurred after the degeneration of protoplasts, suggesting a close association between rubber biosynthesis and PCD. The molecular analysis revealed the expression patterns of key genes involved in rubber biosynthesis. The upstream genes DiIPP, DiFPP, and DiGGPPS showed a decreasing trend during fruit ripening, while DiHRT, which is responsible for rubber particle extension, exhibited the highest expression level during the rubber particle formation. Moreover, the transcription factors related to PCD, DiLSD1, and DiLOL2 showed a negative correlation with the expression pattern of DiHRT, thus exhibiting strict rules of sequential expression during rubber biosynthesis. Additionally, the expression trends of DiXCP1 and DiCEP1, which act as proteases during PCD, were positively correlated with DiGGPPS expression. In conclusion, the findings suggest that the autophagic PCD may play a crucial role in rubber accumulation in D. insignis. Further research is still needed to fully understand the complex regulatory network underlying rubber biosynthesis in plants.

Weight bearing training alleviates muscle atrophy and pyroptosis of middle-aged rats.

Background: Age-related muscle atrophy and adipose accumulation begin to occur in young and middle-aged individuals, and exercise at an early age improves body composition. Pyroptosis may play an essential role in age-related low-grade inflammation. This study aimed to explore the alleviation of muscle atrophy by weight-bearing training with increasing age via inhibition of pyroptosis. Methods: Ninety 8-month-old male SD rats were randomly divided into three groups: (1) normal baseline group (N group, n = 10), sacrificed after adaptive feeding; control group (C group, n = 40); and weight-bearing running group (R group, n = 40). Blood samples, adipose tissue (AT), and extensor digitorum longus (EDL) were collected after 8, 16, 24, and 32-weeks intervention. Results: The body weight, muscle mass, fat mass, plasma lipid, AT wet weight, adipocyte cross-sectional area (CSA), and apoptosis rates of AT and EDL were increased, while the muscle mass, wet weight, and fiber CSA of EDL were decreased by aging, which were reversed by exercise. Weight-bearing training promoted protein synthesis in EDL, inhibited protein degradation in EDL, and expression of pyroptotic key proteins in EDL and AT in rats. Conclusion: Weight-bearing training improves body composition and alleviates age-related muscle atrophy in rats, and its mechanism may be related to the inhibition of pyroptosis in the EDL and AT and the improvement of muscle protein metabolism.

Developmental Programmed Cell Death Involved in Ontogenesis of Dictamnus dasycarpus Capitate Glandular Hairs.

Plant glandular trichomes have received much attention due to their commercial and biological value. Recent studies have focused on the development of various glands in plants, suggesting that programmed cell death (PCD) may play an important role during the development of plant secretory structures. However, the development processes and cytological characteristics in different types of plant secretory structures differed significantly. This study aims to provide new data on the developmental PCD of the capitate glandular hairs in Dictamnus dasycarpus. Light, scanning, immunofluorescence labeling, and transmission electron microscopy were used to determine the different developmental processes of the capitate glandular hairs from a cytological perspective. Morphologically, the capitate glandular hair originates from one initial epidermal cell and differentiates into a multicellular trichome characterized by two basal cells, two lines of stalk cells, and a multicellular head. It is also histochemically detected by essential oils. TUNEL-positive reactions identified nuclei with diffused fluorescence or an irregular figure by DAPI, and Evans blue staining showed that the head and stalk cells lost their viability. Ultrastructural evidence revealed the developmental process by two possible modes of PCD. Non-autolytic PCD was characterized by buckling cell walls and degenerated nuclei, mitochondria, plastids, multivesicular body (MVB), and end-expanded endoplasmic reticulum in the condensed cytoplasm, which were mainly observed in the head cells. The MVB was detected in the degraded vacuole, a degraded nucleus with condensed chromatin and diffused membrane, and eventual loss of the vacuole membrane integrity exhibited typical evidence of vacuole-mediated autolytic PCD in the stalk cells. Furthermore, protoplasm degeneration coupled with dark oil droplets and numerous micro-dark osmiophilic substances was observed during late stages. The secretion mode of essential oils is also described in this paper.

Transgenic Microalgae Expressing Double-Stranded RNA as Potential Feed Supplements for Controlling White Spot Syndrome in Shrimp Aquaculture.

Viral infection of farmed fish and shellfish represents a major issue within the aquaculture industry. One potential control strategy involves RNA interference of viral gene expression through the oral delivery of specific double-stranded RNA (dsRNA). In previous work, we have shown that recombinant dsRNA can be produced in the chloroplast of the edible microalga Chlamydomonas reinhardtii and used to control disease in shrimp. Here, we report a significant improvement in antiviral dsRNA production and its use to protect shrimp against white spot syndrome virus (WSSV). A new strategy for dsRNA synthesis was developed that uses two convergent copies of the endogenous rrnS promoter to drive high-level transcription of both strands of the WSSV gene element in the chloroplast. Quantitative RT-PCR indicated that ~119 ng dsRNA was produced per liter of culture of the transgenic microalga. This represents an ~10-fold increase in dsRNA relative to our previous report. The engineered alga was assessed for its ability to prevent WSSV infection when fed to shrimp larvae prior to a challenge with the virus. The survival of shrimp given feed supplemented with dried alga containing the dsRNA was significantly enhanced (~69% survival) relative to a negative control (<10% survival). The findings suggest that this new dsRNA production platform could be employed as a low-cost, low-tech control method for aquaculture.