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Tumor necrosis factor alpha (TNF-α) is an important proinflammatory cytokine and the only known cytokine that can directly kill tumor cells. Unlike mammalian counterparts, chicken TNF-α (chTNF-α) gene has not been identified until very recently due to its high GC content (∼70%) and long GC fragments. The biological functions of this newly-identified cytokine and its detection methods remain to be further investigated. In this study, the extracellular domain of chTNF-α was cloned into prokaryotic vector after codon optimization and recombinant chTNF-α protein was expressed. Subsequently, using recombinant chTNF-ɑ as immunogen, rabbit polyclonal antibody (pAb) and eight clones of mouse anti-chTNF-ɑ monoclonal antibodies (mAbs) were produced, respectively. Both the pAb and mAbs specifically recognized recombinant chTNF-ɑ expressed in E.coli and transfected COS-7 cells. Further mapping the antigenic region showed that all the mAbs recognized a region of amino acid residues 195-285 of chTNF-ɑ. Furthermore, an antigen-capture enzyme-linked immunosorbent assay for the detection of chTNF-ɑ was established using one mAb and the pAb. This assay showed no cross-reactivity with irrelevant Trx-fused antigens and could detect natural chTNF-ɑ expressed by mitogen-activated chicken splenocytes in a dose-dependent manner, with a detection limit of 1 ng/mL. Collectively, our results indicated that the mAbs and pAb against chTNF-α are specific and could be used for the study of the biological functions of chTNF-ɑ and the detection of chTNF-ɑ., Competing Interests: The authors declare no conflict of interest., (© 2022 The Author(s).)

The zoonotic H7N9 avian influenza virus emerged with the H9N2-origin internal gene cassette. Previous studies have reported that genetic reassortments with H9N2 were common in the first five human H7N9 epidemic waves. However, our latest work found that the circulating high pathogenicity H7N9 virus has established a dominant internal gene cassette and has decreased the frequency of reassortment with H9N2 since 2018. This dominant cassette of H7N9 was distinct from the cocirculating H9N2, although they shared a common ancestor. As a result, we suppose that this dominant cassette may benefit the viral population fitness and promote its continuous circulation in chickens., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Alzheimer's disease (AD) is a progressive neurodegenerative disease associated with aging, environmental, and genetic factors. Amyloid protein precursor ( APP ) is a known pathogenic gene for familial Alzheimer's disease (FAD), and now more than 70 APP mutations have been reported, but the genotype-phenotype correlation remains unclear. In this study, we collected clinical data from patients carrying APP mutations defined as pathogenic/likely pathogenic according to the American college of medical genetics and genomics (ACMG) guidelines. Then, we reanalyzed the clinical characteristics and identified genotype-phenotype correlations in APP mutations. Our results indicated that the clinical phenotypes of APP mutations are generally consistent with typical AD despite the fact that they show more non-demented symptoms and neurological symptoms. We also performed genotype-phenotype analysis according to the difference in APP processing caused by the mutations, and we found that there were indeed differences in onset age, behavioral and psychological disorders of dementia (BPSD) and myoclonus., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Liu, Xiao, Liu, Liao, Zhou, Weng, Zhou, Liu, Bi, Xu, Zhu, Yang, Zhang, Hao, Zhang, Wang, Jiao and Shen.)

"Carbon neutrality, carbon peaking" is China's national commitment to the whole world about its plans to manage global climate change. China faces many severe challenges in fulfilling its commitments to reduce emissions. China's digital economy is currently booming, and whether it can provide opportunities for reducing regional carbon emissions is worth exploring. This study constructed a comprehensive system to evaluate the development of its digital economy based on China's regional data and empirically tested the direct, indirect, and spatial effects of the comprehensive development of digital economy on regional carbon emissions. In addition, it examined the special stage characteristics using a Hansen threshold model. This study found the following: first, the digital economy significantly suppresses carbon emissions in general, notably with a spatial spillover effect to neighboring provinces. Secondly, an analysis of the mechanism shows that the comprehensive development of a digital economy can restrain regional carbon emissions through industrial progress and the optimization of energy consumption. Third, there are double thresholds, special driving trends and an "inverted N-type" relationship with development. Fourth, a spatial heterogeneity analysis revealed that significant "local" and "neighboring" impacts on the reduction of carbon emissions only exist in the central and eastern areas. This study has a reference value for releasing the dividend of digital economy development and reducing carbon emissions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Hao, Wen, Li, Xu and Xue.)

Bovine leukemia virus (BLV) is widespread in global cattle populations, but the effects of its infection on milk quantity and quality have not been clearly elucidated in animal models. In this study, 30 healthy first-lactation cows were selected from ≈2,988 cows in a BLV-free farm with the same criteria of parity, age, lactation number, as well as milk yield, SCS, and composition (fat, protein, and lactose). Subsequently, these cows were randomly assigned to the intervention ( n = 15) or control ( n = 15) group, and reared in different cowsheds. Cows in the intervention group were inoculated with 1 × phosphate-buffered solution (PBS) resuspended in peripheral blood mononuclear cells (PBMC) from a BLV-positive cow, while the controls were inoculated with the inactivated PBMC from the same individual. From June 2016 to July 2021, milk weight (kg) was automatically recorded by milk sensors, and milk SCS and composition were originated from monthly performed dairy herd improvement (DHI) testing. Fluorescence resonance energy transfer (FRET)-qPCR and ELISA showed that cows in the intervention group were successfully infected with BLV, while cows in the control group were free of BLV for the entire period. At 45 days post-inoculation (DPI), the numbers of whole blood cells (WBCs) ( P = 0.010), lymphocytes (LYMs) ( P = 0.002), and monocytes (MNCs) ( P = 0.001) and the expression levels of IFN-γ ( P = 0.013), IL-10 ( P = 0.031), and IL-12p70 ( P = 0.008) increased significantly in the BLV infected cows compared to the non-infected. In lactation numbers 2-4, the intervention group had significantly higher overall milk yield ( P < 0.001), fat ( P = 0.031), and protein ( P = 0.050) than the control group, while milk SCS ( P = 0.038) and lactose ( P = 0.036) decreased significantly. Further analysis indicated that BLV infection was associated with increased milk yield at each lactation stage in lactation numbers 3-4 ( P = 0.021 or P < 0.001), but not with SCS and milk composition. Together, this 4-year longitudinal study revealed that artificial inoculation of BLV increased the milk yield in cows in this BLV challenge model., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Yang, Gong, Lu, Lu, Zhang, Meng, Peng, Chu, Cao, Hao, Sun, Wang, Qin, Wang, Shang and Yang.)

Interleukin-9 (IL-9) is a pleiotropic cytokine that acts on a variety of cells and tissues, and plays roles in inflammation and infection as well as tumor immunity. While mammalian IL-9s have been widely investigated, avian IL-9 has not yet been identified and characterized. In this study, we cloned chicken IL-9 (chIL-9) and performed a phylogenetic analysis, examined its tissue distribution, characterized the biological functions of recombinant chIL-9 (rchIL-9) and the expression form of natural chIL-9. Phylogenetic analysis showed that chIL-9 has less than 30% amino acid identity with mammalian IL-9s. The chIL-9 mRNA can be abundantly detected only in the testis and thymus, and are significantly up-regulated in peripheral blood mononuclear cells (PBMCs) upon mitogen stimulation. The rchIL-9 was produced by prokaryotic and eukaryotic expression systems and showed biological activity in activating monocytes/macrophages to produce inflammatory cytokines and promoting the proliferation of CD3 + T cells. In addition, four monoclonal antibodies (mAbs) and rabbit polyclonal antibody (pAb) against rchIL-9 were generated. Using anti-chIL-9 mAbs and pAb, natural chIL-9 expressed by the activated PBMCs of chickens with a molecular weight of 25kD was identified by Western-blotting. Collectively, our study reveals for the first time the presence of functional IL-9 in birds and lays the ground for further investigating the roles of chIL-9 in diseases and immunity., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 He, Chen, Hao, Yang and Shang.)

Whether structural alterations of intraretinal layers are indicators for the early diagnosis of Parkinson's disease (PD) remains unclear. We assessed the retinal layer thickness in different stages of PD and explored whether it can be an early diagnostic indicator for PD. In total, 397 [131, 146, and 120 with Hoehn-Yahr I (H-Y I), H-Y II, and H-Y III stages, respectively] patients with PD and 427 healthy controls (HCs) were enrolled. The peripapillary retinal nerve fiber layer (pRNFL), total macular retinal thickness (MRT), and macular volume (TMV) were measured by high-definition optical coherence tomography, and the macular intraretinal thickness was analyzed by the Iowa Reference Algorithms. As a result, the PD group had a significantly lower average, temporal quadrant pRNFL, MRT, and TMV than the HCs group (all p < 0.001). Moreover, the ganglion cell layer (GCL), inner plexiform layer (IPL), and outer nuclear layer were thinner in patients with PD with H-Y I, and significantly decreased as the H-Y stage increased. In addition, we observed that GCL and IPL thicknesses were both correlated with Movement Disorder Society-Unified Parkinson's Disease Rating Scale III (MDS-UPDRS III) scores and non-motor symptoms assessment scores. Furthermore, macular IPL thickness in the superior inner (SI) quadrant (IPL-SI) had the best diagnostic performance in patients with PD with H-Y I versus HCs, with a sensitivity and specificity of 75.06% and 81.67%, respectively. In conclusion, we confirmed the retinal structure was significantly altered in patients with PD in different clinical stages, and that GCL and IPL changes occurred during early PD disease and were correlated with MDS-UPDRS III scores and non-motor symptoms assessment scores. Furthermore, macular IPL-SI thickness might be performed as an early diagnostic indicator for PD., (© 2022. The Author(s).)

Bacterial resistance to antibiotics have become one of the most severe threats in global public health, so the development of new-style antimicrobial agents is urgent. In this work, quaternized carbon quantum dots (qCQDs) with broad-spectrum antibacterial activity were synthesized by a simple green "one-pot" method using dimethyl diallyl ammonium chloride and glucose as reaction precursors. The qCQDs displayed satisfactory antibacterial activity against both Gram-positive and gram-negative bacteria. In rat models of wounds infected with mixed bacteria, qCQDs obviously restored the weight of rats, significantly reduced the death of rats from severe infection, and promoted the recovery and healing of infected wounds. Biosafety tests confirmed that qCQDs had no obvious toxic and side effects during the testing stage. The analysis of quantitative proteomics revealed that qCQDs mainly acted on ribosomal proteins in Staphylococcus aureus (Gram-positive bacteria) and significantly down-regulated proteins associated with citrate cycle in Escherichia coli (Gram-negative bacteria). Meanwhile, real-time quantitative PCR confirmed that the variation trend of genes corresponding to the proteins associated with ribosome and citrate cycle was consistent with the proteomic results after treatment of qCQDs, suggesting that qCQDs has a new antibacterial mechanism which is different from the reported carbon quantum dots with antibacterial action. STATEMENT OF SIGNIFICANCE: With the development of the research on carbon quantum dots, the application of carbon quantum dots in the field of medicine has attracted extensive attention. In this paper, quaternized carbon quantum dots (qCQDs) with antimicrobial activity prepared by specific methods were studied, including antimicrobial spectrum, antimicrobial mechanism and in vivo antimicrobial application. The antimicrobial mechanism of qCQDs was studied by proteomics and RT-qRCR, and the different mechanisms of qCQDs against Gram-positive and Gram-negative bacteria were also found. This study provides a research foundation for the application of carbon quantum dots in antimicrobial field, and also expands the application range of carbon quantum dots in medicine field., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier Ltd.)

Urban flooding can be predicted by using different modeling approaches. This study considered different methods of modeling the dynamic flow interactions between pipe systems and surface flooding in urban areas. These approaches can be divided into two categories based on surface runoff collection units. This paper introduces a new hydrodynamic model that couples the storm water management model and the 2D overland model. The model's efficiency was validated based on the aforementioned experimental dataset; agreement was verified by correlation values above 0.6. Additionally, this study used different approaches and compared their accuracy in predicting flooding patterns. The results show that the use of sub-catchments to model the collection of surface runoff was not predictive of the inundation process, indicating a lower goodness of fit with the recorded values than that of adopting cells. Moreover, to determine which method of adopting cells to collect runoff could better predict rainstorm-induced inundation, an error and correlation analysis was conducted. The analysis found low error and high correlation, suggesting that inundation can be effectively predicted by the new approaches. Ultimately, this study contributes to existing work on numerical analysis of the interaction methods of urban flooding.

A solid-phase extraction (SPE)-micellar electrokinetic chromatography (MEKC) method was developed for the simultaneous determination of four beta-lactam antibiotics (ampicillin, amoxicillin, penicillin V and cephalexin) in spiked milk samples. The milk samples were extracted by SPE after protein precipitation. The analysis was performed by MEKC at 18 kV with phosphate buffer containing sodium dodecyl sulfate (SDS) and the detection was carried out at 210 nm. The four antibiotics could be baseline-separated less than 9 min. The linearities in 0.5 - 20 mg/L could be constructed with the correlation coefficients between 0.994 3 - 0.997 6, and the limits of detection were 0.16 - 0.20 mg/L. The average recoveries of antibiotics from milk were over 70% except amoxicillin. The method is sensitive, specific and easy for the determination of beta-lactam antibiotic residues in milk.