Feeding with 4,4'-diaponeurosporene-producing Bacillus subtilis enhances the lactogenic immunity of sow.

Specific antibodies produced sow by oral porcine epidemic diarrhea virus (PEDV) vaccines would transfer to newborn piglets via colostrum, and it is an effective strategy to prevent porcine epidemic diarrhea (PED). However, there is a lag in the development of corresponding vaccines due to the rapid mutation of PEDV, which could increase the difficulty of PED prevention and control in pig farms. Hence, congenital lactogenic immunity was assessed by feeding 4,4'-diaponeurosporene-producing Bacillus subtilis (B.S-Dia) to sow on the 80th day of gestation in order to protect newborn piglets from PEDV infection. Firstly, we found that the quantities of T lymphocytes and monocytes in the blood and colostrum after oral administration of B.S-Dia were significantly increased as observed by flow cytometry, whereas the proliferative activity of T lymphocytes in colostrum was also markedly increased. Furthermore, enzyme-linked immunosorbent assay (ELISA) results revealed that levels of TGF (Transforming growth factor) -ß, Interleukin (IL) -6, lysozyme and lactoferrin were significantly increased. Finally, it was found in the piglets' challenge protection test that offspring pigs of the sows feeding B.S-Dia during pregnancy did not develop diarrhea symptoms and intestinal pathological changes at 48 h after infection with PEDV, and PEDV load in the jejunum and ileum was significantly reduced, but offspring pigs of the sows taking orally PBS during pregnancy developed pronounced diarrhea symptoms and extensive PEDV colonization was noted both in the jejunum and ileum. In summary, sow by oral administration of B.S-Dia substantially increased congenital lactogenic immunity, thereby preventing newborn piglets from being infected with PEDV.

Vascular response to the microcirculation in the fingertip by local vibration with varied amplitude.

Objectives: We investigated the effect of local vibration intensity on the vascular response to the microcirculation of the finger. Materials and methods: We performed hand-transmitted vibration experiments combined with laser Doppler flowmetry (LDF) to measure the blood perfusion signals of fingertips in the vibrated hand and the contralateral middle finger under the same frequency and different amplitude vibration, and to analyze the changes of microcirculatory blood perfusion levels in the fingers, and to investigate the effects of vibration stimulation on the endothelial, neural and myogenic regulatory frequency ranges of fingertips based on wavelet analysis. Furthermore, the transparent silicone films were fabricated and cultured with vascular endothelial cell (EC), which will undergo the local vibration with varied amplitude. And the expression of inflammatory factors was detected in the ECs. Results: Low-frequency vibration leads to a decreased blood flow in fingertip, and the degree of reduction in fingertip blood flow increases as the amplitude gradually increases, and the period required for blood flow to return to normal level after hand-transmitted vibration gradually increases. The decrease in blood flow is more pronounced in the vibrating hand than in the contralateral hand. In addition, nuclear factor-κB (NF-κB) expression increased significantly with the increase of vibration amplitude. Conclusion: High amplitude vibrations caused the inflammatory reaction of ECs which will lead to the altered endothelial regulatory activity. The endothelial regulatory activity is closely related to the blood perfusion in the microcirculation.

Comparison of the cross-protection of PPRSV sublineage 8.7 MLV vaccines against the recombinant NADC30-like strain.

The emergence of recombinant porcine reproductive and respiratory syndrome virus (PRRSV) has caused a substantial threat to the swine industry in recent years. However, the protective efficacy of different sublineage 8.7 PRRSV modified-live virus (MLV) vaccines against emerging strains were still obscure. In this study, a broad epidemiological investigation of PRRSV showed the prevalence of NADC30-like strain increased in Shandong Province, China from 2018 to 2020. Through piglet trial for vaccination and challenge with recombinant NADC30-like SDlz1601 strain, CH-1R MLV vaccine showed better protective effect than JXA1-R and TJM-F92 MLV vaccines in terms of clinical score and pathological observation. Moreover, all three MLV vaccines could reduce virus loads in the serum of piglets. This study provides valuable insights into the prevalence of the NADC30-like strain and the protective effect of PRRS MLV vaccines against recombinant NADC30-like strains, which could help to improve the prevention and control of PRRSV infections.

Upregulation of TLR4-Dependent ATP Production Is Critical for Glaesserella parasuis LPS-Mediated Inflammation.

Glaesserella parasuis (G. parasuis), an important pathogenic bacterium, cause Glässer's disease, and has resulted in tremendous economic losses to the global swine industry. G. parasuis infection causes typical acute systemic inflammation. However, the molecular details of how the host modulates the acute inflammatory response induced by G. parasuis are largely unknown. In this study, we found that G. parasuis LZ and LPS both enhanced the mortality of PAM cells, and at the same time, the level of ATP was enhanced. LPS treatment significantly increased the expressions of IL-1ß, P2X7R, NLRP3, NF-κB, p-NF-κB, and GSDMD, leading to pyroptosis. Furthermore, these proteins' expression was enhanced following extracellular ATP further stimulation. When reduced the production of P2X7R, NF-κB-NLRP3-GSDMS inflammasome signaling pathway was inhibited, and the mortality of cells was reduced. MCC950 treatment repressed the formation of inflammasome and reduced mortality. Further exploration found that the knockdown of TLR4 significantly reduced ATP content and cell mortality, and inhibited the expression of p-NF-κB and NLRP3. These findings suggested upregulation of TLR4-dependent ATP production is critical for G. parasuis LPS-mediated inflammation, provided new insights into the molecular pathways underlying the inflammatory response induced by G. parasuis, and offered a fresh perspective on therapeutic strategies.

Novel p22 and p30 dual-proteins combination based indirect ELISA for detecting antibodies against African swine fever virus.

Introduction: African swine fever virus (ASFV) infection is one of the most complex and fatal hemorrhagic viral diseases, causing a devastating loss to the swine industry. Since no effective vaccine is available, prevention and control of ASFV heavily depends on early diagnostic detection. Methods: In this study, a novel indirect ELISA was established for detecting antibodies against ASFV using dual-proteins, p22 and p30. Recombinants p22 and p30 were expressed and purified from E.coli vector system by recombined plasmids pET-KP177R and pET-CP204L. p22 and p30 were mixed as antigens for developing the indirect ELISA. Results: Through optimizing coating concentrations of p30 and p22, coating ratio (p30: p22 = 1:3), and serum dilution (as 1:600), the established ELISA performed higher specificity, sensitivity, and repeatability against ASFV-positive serum. Furthermore, 184 clinical serum samples from suspected diseased pigs were verified the established ELISA in clinical diagnosis. The results showed that compared with two commercial ELISA kits, the established ELISA possessed higher sensitivity and almost uniform coincidence rate. Conclusion: The novel indirect ELISA based on dual-proteins p30 and p22 performed a valuable role in diagnostic detection of ASFV, providing a broad insight into serological diagnostic methods of ASFV.

Probing the Effect and Mechanism of Flue Gas on the Performance of Resorcinol/Hexamethylenetetramine-Based Polymer Gel in Flue Gas Flooding Reservoir.

Polymer gel plugging is an effective method for gas mobility control in flue gas flooding reservoirs. However, the effect and mechanism of flue gas on the performance of polymer gels have rarely been reported. In this study, a polymer gel was prepared by cross-linking hydrolyzed polyacrylamide (HPAM) and resorcinol/ hexamethylenetetramine (HMTA) to illuminate the influencing mechanism of flue gas composition on gel. The gel rheological testing results showed that flue gas promoted gelation performance, whereas it seriously threatened gel long-term stability, especially at high pressure conditions. The influence of CO2 on the polymer gel had the characteristic of multiplicity. The hydrodynamic radius (Rh) and the initial viscosity of HPAM solution decreased in the presence of CO2. Nonetheless, the dissolved CO2 expedited the decomposition rate of HMTA into formaldehyde, which promoted the cross-linking process of the HPAM, leading to a shorter gelation time. Oxidation-reduction potential (ORP) tests and Fourier transform infrared spectroscopy (FTIR) analysis indicated that O2 played a leading role in the oxidative degradation of HPAM compared to CO2 and threatened the gel long-term stability at elevated gas pressures. To address the adverse effects caused by flue gas, it is highly desirable to develop polymer gels by adding oxygen scavengers or strengthening additives.

A Novel Pathway for Porcine Epidemic Diarrhea Virus Transmission from Sows to Neonatal Piglets Mediated by Colostrum.

The mechanisms of colostrum-mediated virus transmission are difficult to elucidate because of the absence of experimental animal models and the difficulties in tissue sample collection from mothers in the peripartum period. Porcine epidemic diarrhea virus (PEDV) is a reemerging enteropathogenic coronavirus that has catastrophic impacts on the global pig industry. PEDV primarily infects neonatal piglets by multiple routes, especially 1- to 2-day-old neonatal piglets. Here, our epidemiological investigation and animal challenge experiments revealed that PEDV could be vertically transmitted from sows to neonatal piglets via colostrum, and CD3+ T cells in the colostrum play an important role in this process. The results showed that PEDV colonizing the intestinal epithelial cells (IECs) of orally immunized infected sows could be transferred to CD3+ T cells located just beneath the IECs. Next, PEDV-carrying CD3+ T cells, with the expression of integrin α4ß7 and CCR10, migrate from the intestine to the mammary gland through blood circulation. Arriving in the mammary gland, PEDV-carrying CD3+ T cells could be transported across mammary epithelial cells (MECs) into the lumen (colostrum), as illustrated by an autotransfusion assay and an MECs/T coculture system. The PEDV-carrying CD3+ T cells in colostrum could be interspersed between IECs of neonatal piglets, causing intestinal infection via cell-to-cell contact. Our study demonstrates for the first time that colostrum-derived CD3+ T cells comprise a potential route for the vertical transmission of PEDV. IMPORTANCE The colostrum represents an important infection route for many viruses. Here, we demonstrate the vertical transmission of porcine epidemic diarrhea virus (PEDV) from sows to neonatal piglets via colostrum. PEDV colonizing the intestinal epithelial cells could transfer the virus to CD3+ T cells located in the sow intestine. The PEDV-carrying CD3+ T cells in the sow intestine, with the expression of integrin α4ß7 and CCR10, arrive at the mammary gland through blood circulation and are transported across mammary epithelial cells into the lumen, finally leading to intestinal infection via cell-to-cell contact in neonatal piglets. Our study not only demonstrates an alternative route of PEDV infection but also provides an animal model of vertical transmission of human infectious disease.

Porcine Epidemic Diarrhea Virus Infection Disrupts the Nasal Endothelial Barrier To Favor Viral Dissemination.

Crossing the endothelium from the entry site and spreading in the bloodstream are crucial but obscure steps in the pathogenesis of many emerging viruses. Previous studies confirmed that porcine epidemic diarrhea virus (PEDV) caused intestinal infection by intranasal inoculation. However, the role of the nasal endothelial barrier in PEDV translocation remains unclear. Here, we demonstrated that PEDV infection causes nasal endothelial dysfunction to favor viral dissemination. Intranasal inoculation with PEDV compromised the integrity of endothelial cells (ECs) in nasal microvessels. The matrix metalloproteinase 7 (MMP-7) released from the PEDV-infected nasal epithelial cells (NECs) contributed to the destruction of endothelial integrity by degrading the tight junctions, rather than direct PEDV infection. Moreover, the proinflammatory cytokines released from PEDV-infected NECs activated ECs to upregulate ICAM-1 expression, which favored peripheral blood mononuclear cells (PBMCs) migration. PEDV could further exploit migrated cells to favor viral dissemination. Together, our results reveal the mechanism by which PEDV manipulates the endothelial dysfunction to favor viral dissemination and provide novel insights into how coronavirus interacts with the endothelium. IMPORTANCE The endothelial barrier is the last but vital defense against systemic viral transmission. Porcine epidemic diarrhea virus (PEDV) can cause severe atrophic enteritis and acute viremia. However, the mechanisms by which the virus crosses the endothelial barrier and causes viremia are poorly understood. In this study, we revealed the mechanisms of endothelial dysfunction in PEDV infection. The viral infection activates NECs and causes the upregulation of MMP-7 and proinflammatory cytokines. Using NECs, ECs, and PBMCs as in vitro models, we determined that the released MMP-7 contributed to the destruction of endothelial barrier, and the released proinflammatory cytokines activated ECs to facilitate PBMCs migration. Moreover, the virus further exploited the migrated cells to promote viral dissemination. Thus, our results provide new insights into the mechanisms underlying endothelial dysfunction induced by coronavirus infection.

Mechanism of transepithelial migration of lymphocytes into the milk in porcine mammary glands.

Lymphocytes in the colostrum play many important roles during lactation, including protecting newborn piglets against infections. The lymphocytes constantly enter the mammary gland from the mother's bloodstream before and during lactation. However, little is known about the mechanism of transport of maternal lymphocytes across the mammary glands into the milk (lumen). In this study, the maternal lymphocytes were detected in sow colostrum by immunofluorescent staining and fluorescence-activated cell sorting and lymphocytes were observed transmigrating into the breast acinar lumen. Furthermore, immunohistochemical staining revealed that CD3+ T, γδ+ T, and IgA+ B cells were primarily located at the base area of the mammary gland. Meanwhile, more lactating alveoli and blood capillaries were distributed in this area. Finally, a mammary epithelial cell (EpH4-Ev)/T cell co-culture system was established to explore the mechanism of lymphocyte transmigration across the mammary epithelial cells. The expression of CCL2 and CCL28 in EpH4-Ev cells, which facilitated the transmigration of lymphocytes, significantly increased in the presence of prolactin. Our results provide a better understanding of the concept of lactogenic immunity and pave the way for vaccination strategies for the induction of lactogenic immunity in pregnant swine.

Role of intestinal extracellular matrix-related signaling in porcine epidemic diarrhea virus infection.

Porcine epidemic diarrhea virus (PEDV) is emerging as a major threat to the global swine industry. Clinical PEDV infection is associated with severe intestinal lesions, resulting in absorptive dysfunction and high mortality rates in suckling piglets. The extracellular matrix (ECM) is an important component of intestinal tissue, providing a structural framework and conveying tissue-specific signals to nearby enterocytes. In this study, we investigated the extensive ECM remodeling observed in intestinal epithelial cells infected with PEDV and elucidated the associated activated ECM receptor-related pathways. Protein-protein interaction network analysis revealed two significantly differentially expressed genes (cluster of differentiation 44 [CD44] and serpin family E member 1 [SERPINE1]) associated with the ECM. At the transcriptional level, both genes exhibited significant positive correlation with the extent of PEDV replication. Similarly, the expression of CD44 and PAI-1 (encoded by SERPINE1) was also increased in the intestines of piglets during viral infection. Furthermore, CD44 exhibited antiviral activity by enhancing the expression of antiviral cytokines (e.g., interleukin [IL]-6, IL-18, IL-11, and antimicrobial peptide beta-defensin 1) by activating nuclear factor-κB signaling. Conversely, PAI-1 was found to promote the release of progeny virions during PEDV infection, despite a decreased intracellular viral load. Nevertheless, the underlying mechanisms are still unclear. Taken together, our results highlighted the biological roles of specific ECM-regulated genes, i.e., CD44 and SERPINE1 in suppressing and promoting PEDV infection, thereby providing a theoretical foundation for the role of the ECM in intestinal infections and identifying potential therapeutic targets for PEDV.

Red blood cells serve as a vehicle for PEDV transmission.

As the most abundant cell type in the blood, red blood cells (RBCs) are serving for transporting oxygen. However, the mechanism by which RBCs binding virus remains largely unknown. Here, we demonstrated that porcine epidemic diarrhea virus (PEDV), a kind of coronavirus, could hijack RBCs and cause typical diarrhea in neonatal piglets. In an epidemiology investigation of PEDV, the RBCs samples from diarrheic pigs in several pig farms were found to be PEDV-positive. PEDV could bind to neonatal RBCs through CD71 and clathrin-mediated endocytosis, and its viability was maintained for 12 h. PEDV-loaded RBCs could transfer the virus to CD3+ T cells by conjugation and reach the intestine mucosa, where it caused infection. Finally, a further animal challenge revealed that transfusing with PEDV-loaded RBCs could cause intestinal epithelial cells (IECs) infection and typical diarrhea symptom. Therefore, our studies illustrated the mechanism by which PEDV could cause intestinal infection through hijacking RBCs, further providing a novel insight into the role of RBCs as potential cells for viral transmission in coronavirus pathogenesis.

Porcine reproductive and respiratory syndrome virus NADC30-like strain accelerates Streptococcus suis serotype 2 infection in vivo and in vitro.

Porcine reproductive and respiratory syndrome (PRRS), an economically significant pandemic disease, commonly results in increased impact of bacterial infections, including those by Streptococcus suis (S. suis). In recent years, PRRS virus (PRRSV) NADC30-like strain has emerged in different regions of China, and coinfected with S. suis and PRRSV has also gradually increased in clinical performance. However, the mechanisms involved in host innate responses towards S. suis and their implications of coinfection with NADC30-like strain remain unknown. Therefore, the pathogenicity of NADC30-like strain and S. suis serotype 2 (SS2) coinfection in vivo and in vitro was investigated in this study. The results showed that NADC30-like increased the invasion and proliferation of SS2 in blood and tissues, resulting in more severe pneumonia, myocarditis, and peritonitisas well as higher mortality rate in pigs. In vitro, NADC30-like strain increased the invasion and survival of SS2 in porcine alveolar macrophages (PAM) cells, causing more drastic expression of inflammatory cytokines and activation of NF-ĸB signalling. These results pave the way for understanding the interaction of S. suis with the swine immune system and their modulation in a viral coinfection.

Macular pigment and serum zeaxanthin levels with Goji berry supplement in early age-related macular degeneration.

AIM: To evaluate the efficacy of Goji berry supplementation on improving macular pigment, serum zeaxanthin levels and visual acuity in patients with early age-related macular degeneration (AMD). METHODS: A total of 114 patients (aged from 51 to 92y, mean age 69.53±8.41y) with early AMD were enrolled in our prospective, randomized controlled study. The included patients were assigned randomly to the Goji group (n=57) with 25 g of Goji berries supplementation per day for 90d and the control group (n=57) with their normal diet for 90d. Macular pigment optical density (MPOD) was measured using heterochromatic flicker photometry (HFP). The levels of serum lutein (L)/zeaxanthin (Z) were analyzed using high-performance liquid chromatography (HPLC). MPOD, serum L/Z levels and best corrected visual acuity (BCVA) were recorded at baseline and 90d. RESULTS: In the Goji group, there were no statistically significant differences in the serum L levels between the baseline (0.199±0.149 µmol/mL) and 90d (0.203±0.181 µmol/mL) (t=-0.186, P=0.850); however the serum Z levels were increased at 90d (0.101±0.087 µmol/mL) compared with those at the baseline (0.029±0.032 µmol/mL) (t=6.412, P<0.001). Patients treated with Goji berry for 90d showed an elevated MPOD (0.877±0.202 DU) from the baseline (0.731±0.205 DU) (t=-4.741, P=0.000). In contrast to the control group, the serum Z levels and MPOD were higher in the Goji group at 90d (both P<0.05). At 90d, patients with Goji berry supplementation had a relative decrease in BCVA (0.21±0.18 logMAR) compared with the baseline (0.27±0.20) (t=2.397, P=0.020). CONCLUSION: Overall, daily supplementation with Goji berry for 90d improves MPOD by increasing serum Z levels rather than serum L levels in early AMD patients. Goji berry may be an effective therapeutic intervention for preventing the progression of early AMD.

Publisher Correction: Emergence of Different Recombinant Porcine Reproductive and Respiratory Syndrome Viruses, China.

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Emergence of Different Recombinant Porcine Reproductive and Respiratory Syndrome Viruses, China.

Epidemiological investigations were conducted on recently emerging porcine reproductive and respiratory syndrome virus (PRRSV) strains in Shandong province in 2014-2015. The proportion of the NADC30 strain identified by ORF7 sequence alignment has been gradually increasing. Three emerging PRRSV strains were successfully isolated, and the complete genomic sequences were determined. Our results indicate the importance of recombinant strains in Shandong province, China. There was a varied degree of recombination of two or three strains (classical, HP-PRRSV and/or NADC30). Moreover, the recombination strains affected the pathogenicity of newly emerged strains.

A simple synthetic route to polyoxovanadate-based organic-inorganic hybrids using EEDQ as an ester coupling agent.

A reaction strategy for the post-functionalization of hexavanadate derivatives is presented herein. In this study, five polyoxovanadate-based organic-inorganic hybrids TBA2[V6O13{(OCH2)3CCOO(CH2)15CH3}2] (2), TBA2[V6O13{(OCH2)3CCOO(CH2)8CH[double bond, length as m-dash]CH2}2] (3), TBA2[V6O13{(OCH2)3CCOOCH2CF3}2] (4), TBA2[V6O13{(OCH2)3CCOO(CH2CH2O)3CH3}2] (5), and TBA2[V6O13{(OCH2)3CCOO(CH2)12OH}2]·2CH3CH2OH (6) were successfully synthesized via esterification of carboxylic acid groups-containing TBA2[V6O13{(OCH2)3CCOOH}2] (1) with five alcohols possessing different functional groups. These hybrids were characterized by single crystal X-ray diffraction, IR, ESI-MS, 1H and 13C-NMR spectroscopies, and elemental analysis. Moreover, the formation of large vesicles was observed in a mixed solution of compound 2 due to its surfactant-like structure.

The BALB/c mouse infection model for improving the Haemophilus parasuis serotyping scheme.

The use of BALB/c mouse as an alternative model to study Haemophilus parasuis (HPS) infections was evaluated, supplying the serotyping scheme by comparing the pathogenicity of different serovar HPS in pigs and mice challenge using statistical analysis. Results showed that the pathogenicity of different serovar HPS in mouse was consistent with in pigs, proving that this model is a viable alternative to pigs. It provides a convenient methodology for determining the virulence of HPS strains.