Effects of baculovirus infection on intestinal microflora of BmNPV resistant and susceptible strain silkworm.

Bombyx mori L. (Lepidoptera: Bombycidae) nucleopolyhedrovirus (BmNPV) is a serious pathogen causing huge economic losses to sericulture. There is growing evidence that the gut microbiota of silkworms plays a critical role in shaping host responses and interactions with viral infection. However, little is known about the differences in the composition and diversity of intestinal microflora, especially with respect to silkworm strain differences and BmNPV infection-induced changes. Here, we aim to explore the differences between BmNPV-resistant strain A35 and susceptible strain P50 silkworm and the impact of BmNPV infection on intestinal microflora in different strains. The 16S rDNA sequencing analysis revealed that the fecal microbial populations were distinct between A35 and P50 and were significantly changed post BmNPV infection in both strains. Further analysis showed that the BmNPV-resistant strain silkworm possessed higher bacterial diversity than the susceptible strain, and BmNPV infection reduced the diversity of intestinal flora assessed by feces in both silkworm strains. In response to BmNPV infection, the abundance of Muribaculaceae increased in P50 and decreased in A35, while the abundance of Enterobacteriaceae decreased in P50 and increased in A35. These results indicated that BmNPV infection had various effects on the abundance of fecal microflora in different silkworm strains. Our findings not only broadened the understanding of host-pathogen interactions but also provided theoretical help for the breeding of resistant strains and healthy rearing of silkworms based on symbiotic bacteria.

Bmo-miR-3351 modulates glutathione content and inhibits BmNPV proliferation by targeting BmGSTe6 in Bombyx mori.

MicroRNAs (miRNAs) are small non-coding RNAs that play pivotal roles in the host response to invading pathogens. Among these pathogens, Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the main causes of substantial economic losses in sericulture, and there are relatively few studies on the specific functions of miRNAs in the B. mori-BmNPV interaction. Therefore, we conducted transcriptome sequencing to identify differentially expressed (DE) messenger RNAs (mRNAs) and miRNAs in the midgut of 2 B. mori strains (BmNPV-susceptible strain P50 and BmNPV-resistant strain A35) after BmNPV infection. Through correlation analysis of the miRNA and mRNA data, we identified a comprehensive set of 21 miRNAs and 37 predicted target mRNAs. Notably, miR-3351, which has high expression in A35, exhibited remarkable efficacy in suppressing BmNPV proliferation. Additionally, we confirmed that miR-3351 binds to the 3' untranslated region (3' UTR) of B. mori glutathione S-transferase epsilon 6 (BmGSTe6), resulting in its downregulation. Conversely, BmGSTe6 displayed an opposite expression pattern to miR-3351, effectively promoting BmNPV proliferation. Notably, BmGSTe6 levels were positively correlated with glutathione S-transferase activity, consequently influencing intracellular glutathione content in the infected samples. Furthermore, our investigation revealed the protective role of glutathione against BmNPV infection in BmN cells. In summary, miR-3351 modulates glutathione content by downregulating BmGSTe6 to inhibit BmNPV proliferation in B. mori. Our findings enriched the research on the role of B. mori miRNAs in the defense against BmNPV infection, and suggests that the antiviral molecule, glutathione, offers a novel perspective on preventing viral infection in sericulture.

Regulation of Bombyx mori ferritin heavy-chain homolog on ROS induces multiple effects on BmNPV replication.

Ferritin is an iron-binding protein composed of light-chain and heavy-chain homologs with a molecular weight of about 500 kDa. Free iron ions significantly affect reactive oxygen species (ROS) accumulation. Previous research has shown that Bombyx mori nucleopolyhedrosis virus (BmNPV) can increase ROS accumulation, activate autophagy, induce apoptosis, and upregulate the expression of B. mori ferritin heavy-chain homolog (BmFerHCH). However, the mechanism of mutual regulation between BmFerHCH and ROS-mediated autophagy and apoptosis induced by BmNPV remains unclear. In this study, we found that BmNPV induced the time-dependent accumulation of ROS in BmN cells, thereby promoting BmFerHCH expression. Interestingly, in BmFerHCH-overexpressed cells, BmNPV replication was inhibited in the first 18 h after infection but stimulated after 24 h. Further research on H2O2 or antioxidant-treated cells indicated that ROS-induced autophagy slightly increased in the early infection stage and increased BmNPV replication, while in the late stage, a large accumulation of ROS induced apoptosis and inhibited BmNPV replication. In this process, BmFerHCH inhibits BmNPV-induced ROS accumulation by chelating Fe2+. Taken together, BmFerHCH regulates ROS-mediated autophagy and apoptosis to achieve its various effects on BmNPV replication. These findings will help elucidate BmNPV-induced autophagy and apoptosis mediated by ROS and BmFerHCH, as well as the mutually fighting relationship between viruses and hosts.

Trehalose hydrolysis and transport-related genes promote Bombyx mori nucleopolyhedrovirus proliferation through the phosphoinositide 3-kinase-Akt signalling pathway in BmN cell.

The reprogramming of host physiology has been considered an essential process for baculovirus propagation. Trehalose, the main sugar in insect blood, plays a crucial role as an instant energy source. Although the trehalose level is modulated following infection with Bombyx mori nucleopolyhedrovirus (BmNPV), the mechanism of trehalose metabolism in response to BmNPV infection is still unclear. In this study, we demonstrated that the trehalose level tended to be lower in BmNPV-infected hemolymph and higher in the midgut. The omics analysis revealed that two trehalose transporters, BmTret1-1 and BmTret1-2, and trehalase, BmTRE1 and BmTRE2, were differentially expressed in the midgut after BmNPV infection. BmTret1-1 and BmTret1-2 had the ability to transport trehalose into the cell and promoted cellular absorption of trehalose. Furthermore, the functions of BmTret1-1, BmTret1-2, BmTRE1 and BmTRE2 in BmNPV infection were analyzed. These genes were upregulated in the midgut after BmNPV infection. Virus amplification analysis revealed that these genes could promote BmNPV proliferation in BmN cells. In addition, these genes could promote the expression of BmPI3K, BmPDK1 and BmAkt and inhibit the expression of BmFoxO in the phosphoinositide 3-kinase (PI3K)-Akt signalling pathway. Similarly, the increased trehalose level in BmN cells could promote the expression of BmPI3K, BmPDK1 and BmAkt and inhibit the expression of BmFoxO. Taken together, BmNPV infection promote the expression of trehalose hydrolysis and transport-related genes. These changes affect the PI3K-Akt signalling pathway to facilitate BmNPV proliferation. These findings help clarify the relationship between trehalose metabolism and BmNPV infection.

Bombyx mori ferritin heavy-chain homolog facilitates BmNPV proliferation by inhibiting reactive oxygen species-mediated apoptosis.

Ferritin heavy-chain homolog (FerHCH), an iron-binding protein, plays an important role in the host defense against oxidative stress and pathogen infections. In our previous research, Bombyx mori native ferritin had an interaction with B. mori nucleopolyhedrovirus (BmNPV). However, the underlying molecular mechanism of single ferritin homolog responses to BmNPV infection remains unclear. In this study, we found that BmNPV titer and B. mori FerHCH (BmFerHCH) expression were positively correlated with the ferric iron concentration. We performed RNA interference (RNAi) and overexpression experiments to investigate the effects of BmFerHCH on BmNPV proliferation. BmFerHCH knockdown suppressed BmNPV proliferation in vivo and in vitro, whereas BmFerHCH overexpression facilitated BmNPV proliferation. In addition, the oxidative stress level was increased significantly in BmN cells after budded virus infection, while BmFerHCH could neutralize the increased ROS production induced by BmNPV. Of note, we found that ROS was involved in BmNPV-induced apoptosis. Through inhibiting ROS, apoptosis was suppressed by BmFerHCH, whereas BmFerHCH knockdown facilitated apoptosis. Therefore, we hypothesize that BmFerHCH-mediated inhibition of virus-induced apoptosis depends on suppressing ROS accumulation and, thereby, facilitates virus replication. These results suggest that BmFerHCH plays an important role in facilitating BmNPV proliferation and modulating BmFerHCH is potential strategy for studying host-pathogen interactions.

An LSTM-Based Prediction Method for Lower Limb Intention Perception by Integrative Analysis of Kinect Visual Signal.

Recently, computer vision and deep learning technology has been applied in various gait rehabilitation researches. Considering the long short-term memory (LSTM) network has been proved an excellent performance in learn sequence feature representations, we proposed a lower limb joint trajectory prediction method based on LSTM for conducting active rehabilitation on a rehabilitation robotic system. Our approach based on synergy theory exploits that the follow-up lower limb joint trajectory, i.e. limb intention, could be generated by joint angles of the previous swing process of upper limb which were acquired from Kinect platform, an advanced computer vision platform for motion tracking. A customize Kinect-Treadmill data acquisition platform was built for this study. With this platform, data acquisition on ten healthy subjects is processed in four different walking speeds to acquire the joint angles calculated by Kinect visual signals of upper and lower limb swing. Then, the angles of hip and knee in one side which were presented as lower limb intentions are predicted by the fore angles of the elbow and shoulder on the opposite side via a trained LSTM model. The results indicate that the trained LSTM model has a better estimation of predicting the lower limb intentions, and the feasibility of Kinect visual signals has been validated as well.

Using Eye Aspect Ratio to Enhance Fast and Objective Assessment of Facial Paralysis.

A rapid and objective assessment of the severity of facial paralysis allows rehabilitation physicians to choose the optimal rehabilitation treatment regimen for their patients. In this study, patients with facial paralysis were enrolled as study objects, and the eye aspect ratio (EAR) index was proposed for the eye region. The correlation between EAR and the facial nerve grading system 2.0 (FNGS 2.0) score was analyzed to verify the ability of EAR to enhance FNGS 2.0 for the rapid and objective assessment of the severity of the facial paralysis. Firstly, in order to accurately calculate the EAR, we constructed a landmark detection model based on the face images of facial paralysis patients (FP-FLDM). Evaluation results showed that the error rate of facial feature point detection in patients with facial paralysis of FP-FLDM is 17.1%, which was significantly superior to the landmark detection model based on normal face images (NF-FLDM). Secondly, in this study, the Fréchet distance was used to calculate the difference in bilateral EAR of facial paralysis patients and to verify the correlation between this difference and the corresponding FNGS 2.0 score. The results showed that the higher the FNGS 2.0 score , the greater the difference in bilateral EAR. The correlation coefficient between the bilateral EAR difference and the corresponding FNGS 2.0 score was 0.9673, indicating a high correlation. Finally, through a 10-fold crossvalidation, we can know that the accuracy of scoring the eyes of patients with facial paralysis using EAR was 85.7%, which can be used to enhance the objective and rapid assessment of the severity of facial paralysis by FNGS 2.0.

An Unobtrusive Computerized Assessment Framework for Unilateral Peripheral Facial Paralysis.

Unilateral peripheral facial paralysis (UPFP) is a form of facial nerve paralysis and clinically classified according to conditions of facial symmetry. Prompt and precise assessment is crucial to neural rehabilitation of UPFP. The prevalent House-Brackmann (HB) grading system relies on subjective judgments with significant interobservation variation. Therefore, to explore an objective method for the UPFP assessment, clinical image sequences are captured using a web camera setup while 5 healthy and 27 UPFP subjects perform a group of predefined actions, including keeping expressionless, raising brows, closing eyes, bulging cheek, and showing teeth in turn. First, facial region is decided using Haar cascade classifier, and then landmark points are acquired by a supervised descent method. Second, these landmark points are used to generate a group of features reflecting the structural parameters of regions of eyebrows, eyes, nose, and mouth, respectively. Third, correlation coefficients are computed between the raw features HB scores. To reduce feature dimensions, only those with correlation coefficients larger than an empirically selected value, 0.35, are input into a support vector machine to generate a classifier. With the classifier, exact match (discrepancy = 0 between result from proposed method and HB scores) rate at 49.9%, and loose match (discrepancy = 1) rate at 87.97% are achieved on the experiment data. After sample augmentation, the final rate is increased to 90.01%, outperformed previous reports. In conclusion, it is demonstrated with an unobtrusive web camera setup, encouraging results have been generated with the proposed framework in this exploratory study.