Prevalence of and risk factors for myopia among urban and rural children in Northeast China: protocol for a school-based cross-sectional study.

INTRODUCTION: The worldwide prevalence of myopia is high and continues to increase. In this study, a school screening programme for myopia will be implemented using the whole-process information method. The purpose of this study is to investigate the prevalence of myopia in urban and rural areas of Northeast China and to determine the factors related to myopia. METHODS AND ANALYSIS: This is a school-based cross-sectional study. Our study population will include 6000 school-aged children from 2 urban and 2 rural schools in Jinzhou, China. The study will be conducted using our self-developed internet-based intelligent data collection, transmission, storage and analysis system. Examination parameters include uncorrected distance visual acuity, presenting distance visual acuity, non-cycloplegic autorefraction, height, weight, waist circumference, hip circumference, spinal curvature and dental caries. The examination report will be automatically sent to parents, who will complete the questionnaire, and appropriate statistical analysis will be performed. The main outcome is the prevalence of myopia, defined as an equivalent spherical degree ≤-0.5 D. ETHICS AND DISSEMINATION: Ethical approval was obtained from the Third Affiliated Hospital of Jinzhou Medical University (number: JYDSY-KXYJ-IEC-2023-018). Findings will be published in a peer-reviewed journal. Subjects and their parents (or other authorised agents) give informed consent prior to study participation. TRIAL REGISTRATION NUMBER: ChiCTR2300072893.

Molecular characterization, expression and functional analysis of TAK1, TAB1 and TAB2 in Nile tilapia (Oreochromis niloticus).

The MAPK pathway is the common intersection of signal transduction pathways such as inflammation, differentiation and proliferation and plays an important role in the process of antiviral immunity. Streptococcus agalactiae will have a great impact on tilapia aquaculture, so it is necessary to study the immune response mechanism of tilapia to S. agalactiae. In this study, we isolated the cDNA sequences of TAK1, TAB1 and TAB2 from Nile tilapia (Oreochromis niloticus). The TAK1 gene was 3492 bp in length, contained an open reading frame (ORF) of 1809 bp and encoded a polypeptide of 602 amino acids. The cDNA sequence of the TAB1 gene was 4001 bp, and its ORF was 1491 bp, which encoded 497 amino acids. The cDNA sequence of the TAB2 gene was 4792 bp, and its ORF was 2217 bp, encoding 738 amino acids. TAK1 has an S_TKc domain and a coiled coil structure; the TAB1 protein structure contains a PP2C_SIG domain and a conserved PYVDXA/TXF sequence model; and TAB2 contains a CUE domain, a coiled coil domain and a Znf_RBZ domain. Homology analysis showed that TAK1 and TAB1 had the highest homology with Neolamprologus brichardi, and TAB2 had the highest homology with Simochromis diagramma (98.28 %). In the phylogenetic tree, TAK1, TAB1 and TAB2 formed a large branch with other scleractinian fishes. The tissue expression analysis showed that the expression of TAK1, TAB1 and TAB2 was highest in the muscle. The expression of TAK1, TAB1 and TAB2 was significantly induced in most of the tested tissues after stimulation with LPS, Poly I:C and S. agalactiae. The subcellular localization results showed that TAK1 was located in the cytoplasm, and TAB1 and TAB2 had certain distributions in the cytoplasm and nucleus. Coimmunoprecipitation (Co-IP) results showed that TRAF6 did not interact with the TAK1 protein but interacted with TAB2, while TAB1 did not interact with P38γ but interacted with TAK1. There was also an interaction between TAK1 and TAB2.

Performance of targeted next-generation sequencing in the detection of respiratory pathogens and antimicrobial resistance genes for children.

Introduction. Respiratory tract infection, which is associated with high morbidity and mortality, occurs frequently in children. At present, the main diagnostic method is culture. However, the low pathogen detection rate of the culture approach prevents timely and accurate diagnosis. Fortunately, next-generation sequencing (NGS) can compensate for the deficiency of culture, and its application in clinical diagnostics has become increasingly available.Gap Statement. Targeted NGS (tNGS) is a platform that can select and enrich specific regions before data enter the NGS pipeline. However, the performance of tNGS in the detection of respiratory pathogens and antimicrobial resistance genes (ARGs) in infections in children is unclear.Aim and methodology. In this study, we estimated the performance of tNGS in the detection of respiratory pathogens and ARGs in 47 bronchoalveolar lavage fluid (BALF) specimens from children using conventional culture and antimicrobial susceptibility testing (AST) as the gold standard.Results. RPIP (Respiratory Pathogen ID/AMR enrichment) sequencing generated almost 500 000 reads for each specimen. In the detection of pathogens, RPIP sequencing showed targeted superiority in detecting difficult-to-culture bacteria, including Mycoplasma pneumoniae. Compared with the results of culture, the sensitivity and specificity of RPIP were 84.4 % (confidence interval 70.5-93.5 %) and 97.7 % (95.9 -98.8%), respectively. Moreover, RPIP results showed that a single infection was detected in 10 of the 47 BALF specimens, and multiple infections were detected in 34, with the largest number of bacterial/viral coinfections. Nevertheless, there were also three specimens where no pathogen was detected. Furthermore, we analysed the drug resistance genes of specimens containing Streptococcus pneumoniae, which was detected in 25 out of 47 specimens in the study. A total of 58 ARGs associated with tetracycline, macrolide-lincosamide-streptogramin, beta-lactams, sulfonamide and aminoglycosides were identified by RPIP in 19 of 25 patients. Using the results of AST as a standard, the coincidence rates of erythromycin, tetracycline, penicillin and sulfonamides were 89.5, 79.0, 36.8 and 42.1 %, respectively.Conclusion. These results demonstrated the superiority of RPIP in pathogen detection, particularly for multiple and difficult-to-culture pathogens, as well as in predicting resistance to erythromycin and tetracycline, which has significance for the accurate diagnosis of pathogenic infection and in the guidance of clinical treatment.

Nile tilapia TBK1 interacts with STING and TRAF3 and is involved in the IFN-ß pathway in the immune response.

Nile tilapia (Oreochromis niloticus) occupies an important position in the culture of economic fish in China. However, the high mortality caused by streptococcal disease has had a significant impact on the tilapia farming industry. Therefore, it is necessary to clarify the immune mechanism of tilapia in response to Streptococcus agalactiae. As a hub in the natural immune signaling pathway, the junction molecule can help the organism defend against and clear pathogens and is crucial in the signaling pathway. In this study, the cDNA sequence of Nile tilapia TBK1 was cloned, and the expression profile was examined in normal fish and challenged fish. The cDNA sequence of the TBK1 gene was 3378 bp, and its open reading frame (ORF) was 2172 bp, encoding 723 amino acids. The deduced TBK1 protein contained an S_TKc domain, a coiled coil domain and a ubiquitin-like domain (ULD). TBK1 had the highest homology with zebra mbuna (Maylandia zebra) and Lake Malawi cichlid fish (Astatotilapia calliptera), both at 97.59%. In the phylogenetic tree, TBK1 forms a large branch with other scleractinian fish. TBK1 expression was highest in the brain and lowest in the liver. LPS, Poly I:C, and S. agalactiae challenge resulted in significant changes in TBK1 expression in the tissues examined. The subcellular localization showed that TBK1-GFP was distributed in the cytoplasm and could significantly increase IFN-ß activation. Pull-down results showed that there was an interaction between TBK1 and TRAF3 and an interaction between STING protein and TBK1 protein. The above results provide a basis for further investigation into the mechanism of TBK1 involvement in the signaling pathway.

Genomic organization and gene evolution of two warm temperature acclimation proteins (Wap65s) of Micropterus salmoides and their responses to temperature and bacterial/viral infections.

Warm temperature acclimation-related 65-kDa proteins (Wap65s) are fish plasma acute-phase glycoproteins homologous to hemopexin with high affinity and clearance for heme. The study characterized Mswap65-1 and Mswap65-2 genes in Micropterus salmoides. Structural analysis showed MsWap65s contained conserved heme-binding sites. MsWap65-1 had a chloride-binding site similar to hemopexin, while MsWap65-2 had an additional calcium-binding site. Phylogenetic and Ka/Ks analysis showed that fish Wap65s were evolutionarily conserved and underwent strong purifying selection. Functional divergence analysis indicated that fish Wap65-2 retained the putative function of ancestral Wap65, while Wap65-1 underwent neofunctional differentiation. QPCR showed Mswap65s were predominantly expressed in liver, but prolonged hyperthermy inhibited Mswap65-2 expression. Mswap65-2 expression was up-regulated in liver and spleen after Nocardia seriolae infection, while Mswap65-1 was down-regulated. MsWap65-2 may be associated with pathogenesis and play potential role in pathogen resistance. LMBV infection resulted in both significant downregulation of Mswap65s were both significantly down-regulated, with differences observed between sexes. We speculated the immune system might suppress expression after viral infection. Exogenous rMsWap65s were prepared, and injection of rMsWap65s alleviated phenylhydrazine-induced hemolysis and inhibited increases in heme, complement C3 and inflammatory symptoms. Our results contribute to an advanced understanding of the functions and mechanisms of MsWap65s in stress resistance.

Whole-genome resequencing reveals recent signatures of selection in five populations of largemouth bass ( Micropterus salmoides).

Largemouth bass ( Micropterus salmoides) is an economically important fish species in North America, Europe, and China. Various genetic improvement programs and domestication processes have modified its genome sequence through selective pressure, leaving nucleotide signals that can be detected at the genomic level. In this study, we sequenced 149 largemouth bass fish, including protospecies (imported from the US) and improved breeds (four domestic breeding populations from China). We detected genomic regions harboring certain genes associated with improved traits, which may be useful molecular markers for practical domestication, breeding, and selection. Subsequent analyses of genetic diversity and population structure revealed that the improved breeds have undergone more rigorous genetic changes. Through selective signal analysis, we identified hundreds of putative selective sweep regions in each largemouth bass line. Interestingly, we predicted 103 putative candidate genes potentially subjected to selection, including several associated with growth (p sst1 and grb10), early development ( klf9, sp4, and sp8), and immune traits ( pkn2, sept2, bcl6, and ripk2). These candidate genes represent potential genomic landmarks that could be used to improve important traits of biological and commercial interest. In summary, this study provides a genome-wide map of genetic variations and selection footprints in largemouth bass, which may benefit genetic studies and accelerate genetic improvement of this economically important fish.

Development of a Highly Permissive Mandarin Fish (Siniperca chuatsi) Kidney Cell Line for Mandarin Fish Ranavirus Using a Single-Cell Cloning Method.

Mandarin fish ranavirus (MRV) infection poses a substantial challenge to the mandarin fish culture industry as no effective preventive or therapeutic measures currently exist. The creation of a highly permissive cell line from a natural host is crucial for developing a vaccine for MRV and understanding its pathogenic mechanisms. In this research, the mandarin fish (Siniperca chuatsi) kidney cell line (SCK) was isolated from mandarin fish kidneys. Subsequently, SCK-a to SCK-g monoclonal cell lines were derived from the SCK cell population, distinguished by morphological variations. Notably, MRV infection induced an advanced cytopathic effect (CPE) in almost all cells of the SCK-f clone. Further tests showed that MRV achieved a peak viral titer of 1010.7 50% tissue culture infectious dose (TCID50)/mL and consistently exceeded 1010 TCID50/mL across nine passages in SCK-f cells. Electron microscopy verified the MRV virion integrity within SCK-f. In vivo experiments revealed that MRV infections led to cumulative mortality rates of 86.9% in mandarin fish and 88.9% in largemouth bass (Micropterus salmoides). Such results suggest that SCK-f is highly permissive to MRV. This study underscores the importance of cellular diversity in developing viral permissive cell lines. The SCK monoclonal cell line pool may offer potential for generating highly permissive cell lines for other mandarin fish viruses.

[Infrared Imaging Meibomian Gland Segmentation System Based on Deep Learning].

In order to better assist doctors in the diagnosis of dry eye and improve the ability of ophthalmologists to recognize the condition of meibomian gland, a meibomian gland image segmentation and enhancement method based on Mobile-U-Net network was proposed. Firstly, Mobile-Net is used as the coding part of U-Net for down sampling, and then features are extracted and fused with the features in decoder to guide image segmentation. Secondly, the segmentation of meibomian gland region is enhanced to assist doctors to judge the condition. Thirdly, a large number of meibomian gland images are collected to train and verify the semantic segmentation network, and the clarity evaluation index is used to verify the meibomian gland enhancement effect. The experimental results show that the similarity coefficient of the proposed method is stable at 92.71%, and the image clarity index is better than the similar dry eye detection instruments on the market.

Deleting ORF71L of infectious spleen and kidney necrosis virus (ISKNV) resulted in virulence attenuation in Mandarin fish.

Infectious spleen and kidney necrosis virus (ISKNV), the type species of the genus Megalocytivirus, infects a variety of teleost fish species and causes substantial losses in the aquaculture industry worldwide. ISKNV ORF71L is 1611 bp in length, encodes a 537-amino-acid peptide and was previously identified as a viral structural protein in the ISKNV virion. In this study, the ORF71L deletion mutant virus strain ISKNV-Δ71 was obtained through a homologous recombination approach. The multistep growth curves showed that ISKNV-Δ71 replication was faster than ISKNV-WT replication in mandarin fish fry cells (MFF-1 cells) before 48 h post-infection (hpi). The cumulative mortality of ISKNV-Δ71-infected mandarin fish (Siniperca chuatsi) was lower than that of fish infected with ISKNV-WT. The copy numbers of viral genome equivalents (GEs) in ISKNV-Δ71-infected mandarin fish spleens were also lower than those in ISKNV-WT-infected spleens. Deletion of ORF71L resulted in ISKNV virulence attenuation in mandarin fish. Furthermore, we found that the number of melanomacrophage centers (MMCs) in ISKNV-Δ71-infected mandarin fish spleens was higher than that in ISKNV-WT-infected mandarin fish spleens. Transcriptomic analysis showed that the cytokine-cytokine receptor interaction pathway had the most significant change between ISKNV-Δ71- and ISKNV-WT-infected MFF-1 cells. These results indicated ORF71L is a virulence-related gene of ISKNV. ORF71L could be considered as a potential target for the development of engineered attenuated live vaccines via multigene deletion or as a potential insertion site for exogenous protein expression.

Enhanced Shielding Performance of Layered Carbon Fiber Composites Filled with Carbonyl Iron and Carbon Nanotubes in the Koch Curve Fractal Method.

Layered carbon fiber composites (CFC) with enhanced shielding effectiveness (SE) were prepared with mixed fillers of carbon nanotubes (CNTs) and carbonyl iron powders (CIPs) in the form of a Koch curve fractal. In the layered composite structure, glass fiber (GF) cloth was used in the wave-transmissive layer (WTL), and the carbon fiber (CF) cloth was used in the supporting layer (SL). Between WTL and SL, CNTs and CIPs were distributed in epoxy resin in the form of a Koch curve fractal to act as an absorbing layer (AL), and copper foil was used as a reflective layer (RL) and bonded at the bottom of the whole composites. The layered structure design and excellent interlayer interface integration obviously improved the SE performance of the CFC. The SE of different samples was investigated, and the results show that, with the increase in the number (n) of Koch curve fractals, the SE of the samples enhanced in the low frequency scope (1-5 GHz). The sample with n = 2 has the highest SE value of 73.8 dB at 2.3 GHz. The shielding performance of the fractal sample filled by CNTs and CIPs simultaneously has a comprehensive improvement in the whole scope of 1-18 GHz, especially for the sample with n = 2. The cumulative bandwidth value of the SE exceeding 55 dB is about 14.3 GHz, accounting for 85% of the whole frequency scope, indicating the composite fabricated in this paper is an electromagnetic shielding material with great prospect.

Identification and expression analysis of cellular and viral microRNAs in CyHV3-infected KCF-1 cells.

MicroRNAs (miRNAs) are small non-coding RNAs with approximately 22 nucleotides (nt) that are encoded by a diverse range of metazoan eukaryotes, plants and viruses. CyHV-3 (cyprinid herpesvirus-3) is a member of the Alloherpesviridae virus family and has caused severe economic losses for the common carp and koi carp fishery industries. In this study, a total of 15,987,652 clean reads were generated from a cDNA library of CyHV-3-infected KCF-1 (koi caudal fin) cells using high-throughput sequencing technology. Following annotation and secondary structure prediction, 28 miRNAs were identified as novel candidate miRNAs encoded by common carp (Cyprinus carpio), and seven miRNAs were shown to be encoded by CyHV-3. Next, 19 host miRNAs and seven viral miRNAs were validated by stem-loop real-time PCR. Northern blot analysis confirmed the presence of 14 host miRNAs and five CyHV-3-encoded novel miRNAs. The results of this study expand the knowledge of common carp and CyHV-3 microRNAs and provide a useful theoretical foundation for further study of CyHV-3.

Transcriptomic analysis of koi (Cyprinus carpio) spleen tissue upon cyprinid herpesvirus 3 (CyHV3) infection using next generation sequencing.

Cyprinid Herpesvirus 3 (CyHV-3) can infect and specifically cause a huge economic loss in both common carp (Cyprinus carpio) and its ornamental koi variety. The molecular mechanisms underlying CyHV-3 infection are not well understood. In this study, koi spleen tissues of both mock and CyHV-3 infection groups were collected, and high-throughput sequencing technology was used to analyze the differentially expressed genes (DEGs) at the transcriptome level. A total of 105,356,188 clean reads from two libraries were obtained. After the de novo assembly of the transcripts, 129,314 unigenes were generated. Of these unigenes, 70,655 unigenes were matched to the known proteins in the database, while 2190 unigenes were predicted by ESTScan software. Comparing the infection group to the mock group, a total of 23,029 significantly differentially expressed unigenes were identified, including 10,493 up-regulated DEGs and 12,536 down-regulated DEGs. GO (Gene Ontology) annotation and functional enrichment analysis indicated that all of the DEGs were annotated into GO terms in three main GO categories: biological process, cellular component and molecular function. KEGG (Kyoto Encyclopedia of Genes and Genomes) analysis of the DEGs showed that a total of 12,002 DEG unigenes were annotated into 256 pathways classified into 6 main categories. Additionally, 20 differentially expressed genes were validated by quantitative real-time PCR. As the first report of a transcriptome analysis of koi carp with CyHV-3 infection, the data presented here provide knowledge of the innate immune response against CyHV-3 in koi carp and useful data for further research of the molecular mechanism of CyHV-3 infection.

Ion Torrent next-generation sequencing reveals the complete mitochondrial genome of koi carp (Cyprinus carpio, Cyprinidae).

The complete mitochondrial genome of the koi carp (Cyprinus carpio, Cyprinidae) was sequenced in the present study by using Ion Torrent Personal Genome Machine (PGM) platform for the first time. The mitochondrial genome sequence is 16 581 bp in size and consists of 13 protein-coding genes, 22 tRNA genes, two rRNA genes and one control region. The gene order and organization were similar to most of the other teleost. The nucleotide compositions of the light strand are 24.82% of A, 31.92% of T, 27.53% of G and 15.73% of C. With the exception of eight tRNA genes and the NADH dehydrogenase subunit 6 (ND6), all other mitochondrial genes are encoded on the heavy strand. The phylogenetic tree constructed using a maximum-likelihood model showed sister relationship of koi carp to other Cyprinidae fishes.

Extracellular virion proteins of two Chinese CyHV-3/KHV isolates, and identification of two novel envelope proteins.

Proteins in extracellular virions of two Chinese cyprinid herpesvirus-3/koi herpesvirus (CyHV-3/KHV) isolates were identified through one-dimensional gel-based matrix-assisted laser desorption/ionization tandem time of flight mass spectrometry in combination with liquid chromatography tandem mass spectrometry (LC ESI-MS/MS). A total of 43 viral proteins were identified, seven of which (pORF62, 68, 43, 51, 92, 84, and 72) were characterized as marked characteristic viral proteins in CyHV-3/KHV virions and six of which (pORF11, 27, 83, 91, 106 and 116) were not detected previously. Of the newly identified proteins, pORF83 was identified as a novel viral minor envelope protein by Western blot analysis and indirect immunofluorescence assay. Another 27 cellular proteins were validated by LC ESI-MS/MS to be involved in purified extracellular CyHV-3/KHV virions, among which a highly abundant, virus-inducible stress protein was shown and characterized as a cell-derived envelope protein in CyHV-3/KHV virions. Our study extends the number of known CyHV-3/KHV virion-associated viral proteins from 40 to 46 and identifies a novel viral envelope protein as well as a cellular envelope protein. The present findings provide some new insights for better understanding CyHV-3/KHV.