Porcine lung tissue slices: a culture model for PRCV infection and innate immune response investigations.

Respiratory coronaviruses (RCoVs) significantly threaten human health, necessitating the development of an ex vivo respiratory culture system for investigating RCoVs infection. Here, we successfully generated a porcine precision-cut lung slices (PCLSs) culture system, containing all resident lung cell types in their natural arrangement. Next, this culture system was inoculated with a porcine respiratory coronavirus (PRCV), exhibiting clinical features akin to humans who were infected by SARS-CoV-2. The results demonstrated that PRCV efficiently infected and replicated within PCLSs, targeting ciliated cells in the bronchioles, terminal bronchioles, respiratory bronchioles, and pulmonary alveoli. Additionally, through RNA-Seq analysis of the innate immune response in PCLSs following PRCV infection, expression levels of interferons, inflammatory cytokines and IFN stimulated genes were significantly upregulated. This ex vivo model may not only offer new insights into PRCV infection in the porcine respiratory tract but also serve as a valuable tool for studying human respiratory CoVs infection.

TLR9 mediates IgA production in the porcine small intestine during PEDV infection.

IgA plays a vital role in defending against the infectious pathogens. However, the specific regulatory pathways involved in IgA secretion in the context of PEDV infection have remained elusive. Therefore, in this study, we explore the molecular mechanisms underlying IgA secretion in response to infection, with a particular focus on PEDV, a devastating enteric virus affecting global swine production. Our investigation begins by examining changes in IgA concentrations in both serum and small intestinal contents following PEDV infection in 2- and 4-week-old pigs. Remarkably, a significant increase in IgA levels in these older pigs post-infection were observed. To delve deeper into the regulatory mechanisms governing IgA secretion in response to PEDV infection, isolated porcine intestinal B cells were co-cultured with monocytes derived DCs (Mo-DCs) in vitro. In the intestinal DC-B cell co-cultures, IgA secretion was found to increase significantly after PEDV infection, as well as upregulating the expression of AID, GLTα and PSTα reflecting isotype switching to IgA. In addition, the expression of TLR9 was upregulated in these cultures, as determined by RT-qPCR and western blotting. Moreover, our findings extend to in vivo observations, where we detected higher levels of TLR9 expression in the ileum of pig post PEDV infection. Collectively, our results highlight the ability of PEDV to stimulate the generation of IgA, particularly in elder pigs, and identify TLR9 as a critical mediator of IgA production within the porcine intestinal microenvironment during PEDV infection.

Construction of a g-C3N4/Bi(OH)3 Heterojunction for the Enhancement of Visible Light Photocatalytic Antibacterial Activity.

Photocatalytic technology has been recently conducted to remove microbial contamination due to its unique features of nontoxic by-products, low cost, negligible microbial resistance and broad-spectrum elimination capacity. Herein, a novel two dimensional (2D) g-C3N4/Bi(OH)3 (CNB) heterojunction was fabricated byincorporating Bi(OH)3 (BOH) nanoparticles with g-C3N4 (CN) nanosheets. This CNB heterojunction exhibited high photocatalytic antibacterial efficiency (99.3%) against Escherichia coli (E. coli) under visible light irradiation, which was 4.3 and 3.4 times that of BOH (23.0%) and CN (28.0%), respectively. The increase in specific surface area, ultra-thin layered structure, construction of a heterojunction and enhancement of visible light absorption were conducive to facilitating the separation and transfer of photoinduced charge carriers. Live/dead cell staining, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) assays and scanning electron microscopy (SEM) have been implemented to investigate the damage to the cell membrane and the leakage of the intracellular protein in the photocatalytic antibacterial process. The e-, h+ and O2•- were the active species involved in this process. This study proposed an appropriate photocatalyst for efficient treatment of bacterial contamination.

Ultrasound synergistic slightly acidic electrolyzed water treatment of grapes: Impacts on microbial loads, wettability, and postharvest storage quality.

Microbial contamination is the principal factor in the deterioration of postharvest storage quality in grapes. To mitigate this issue, we explored a synergistic treatment which combines ultrasound (US) and slightly acidic electrolyzed water (SAEW), and rigorously compared with conventional water cleaning (CW), exclusive US treatment, and standalone SAEW treatment. The US + SAEW treatment proved to be markedly superior in reducing total bacterial, mold & yeast counts on grapes. Specifically, it achieved reductions of 2.23 log CFU/g and 2.76 log CFU/g, respectively, exceeding the efficiencies of SAEW (0.78, 0.75), US (0.58, 0.65), and CW (0.24, 0.46). The efficacy of this synergistic treatment is attributed to the ultrasound removal of the wax layer on grape skins, which transitions the skin from hydrophobic to hydrophilic. This alteration increases the contact area between the grape surface and SAEW, thereby enhancing the antimicrobial efficacy of SAEW. From a physicochemical quality standpoint, the US + SAEW treatment exhibited multiple advantages. It not only minimized weight loss, color deviations, polyphenol oxidase activity and malondialdehyde synthesis in comparison to CW-treated samples but also preserved firmness, sugar-acid ratio and the activities of key enzymes including phenylalanine ammonia-lyase, superoxide dismutase and catalase, and thus maintaining high levels of total phenolics, total ascorbic acid, total anthocyanins, and antioxidants. Consequently, US + SAEW treatment put off the times of decay onset in grapes by 12 days, outperforming both SAEW (8) and US (4) in comparison to CW. These results highlight the potential of US + SAEW as an effective strategy for maintaining grape quality during their postharvest storage period.

Linoleic acid: a natural feed compound against porcine epidemic diarrhea disease.

IMPORTANCE: Porcine epidemic diarrhea virus (PEDV) is a pig coronavirus that causes severe diarrhea and high mortality in piglets, but as no effective drugs are available, this virus threatens the pig industry. Here, we found that the intestinal contents of specific pathogen-free pigs effectively blocked PEDV invasion. Through proteomic and metabolic analyses of the intestinal contents, we screened 10 metabolites to investigate their function and found that linoleic acid (LA) significantly inhibited PEDV replication. Further investigations revealed that LA inhibited viral replication and release mainly by binding with PEDV NSP5 to regulate the PI3K pathway and, in particular, inhibiting AKT phosphorylation. In vivo experiments illustrated that orally administered LA protected pigs from PEDV challenge and severe diarrhea. These findings provide strong support for exploring antiviral drugs for coronavirus treatment.

TGF-ß1 reduces the differentiation of porcine IgA-producing plasma cells by inducing IgM+ B cells apoptosis via Bax/Bcl2-Caspase3 pathway.

Transforming growth factor ß1 (TGF-ß1) performs a critical role in maintaining homeostasis of intestinal mucosa regulation and controls the survival, proliferation, and differentiation of many immune cells. In this study, we discovered that the infection of porcine epidemic diarrhea virus (PEDV), a coronavirus, upregulated TGF-ß1 expression via activating Tregs. Besides, recombinant porcine TGF-ß1 decreased the percentage of CD21+ B cells within the lymphocyte population in vitro. We further found that TGF-ß1 reduced the IgA-secreting B cell numbers and also inhibited plasma cell differentiation. Additional investigations revealed that TGF-ß1 induced the apoptosis of IgM+ B cells in both peyer's patches (PPs) and peripheral blood (PB) through the activation of the Bax/Bcl2-Caspase3 pathway. Conversely, the application of the TGF-ß1 signaling inhibitor SB431542 significantly antagonized the TGF-ß1-induced reduction of IgA secretion and B cell apoptosis and restored plasma cell differentiation. Collectively, TGF-ß1 plays an important role in regulating the survival and differentiation of porcine IgA-secreting B cells through the classical mitochondrial apoptosis pathway. These findings will facilitate future mucosal vaccine designs that target the regulation of TGF-ß1 for the control of enteric pathogens in the pig industry.

Clinical characteristics of infants hospitalized with early congenital syphilitic nephropathy: a single-center retrospective cross-sectional study in China.

BACKGROUND: Early studies claimed that early congenital syphilitic (CS) nephropathy was rare, and systematic studies about this disease are absent, which may lead to poor awareness of early CS nephropathy in clinicians and result in misdiagnosis and poor patient prognosis. The present study systematically and comprehensively analyzes the clinical characteristics of infants with early CS nephropathy hospitalized in Beijing Ditan Hospital, an infectious disease hospital in China in order to improve the understanding and management of this disorder. METHODS: Data of the children with early CS from July 1, 2008, to December 31, 2021, were collected from the electronic medical record system of the hospital. Each patient's demographic characteristics, clinical history, mother's history of syphilitic infection, and laboratory values were extracted. The patients were enrolled to either the nephropathy group or the non-nephropathy group depending on diagnosis. Descriptive statistics was used to report basic demographics, clinical and laboratory test values, and variables were compared between the two groups by nonparametric tests, t test or χ2 tests. RESULTS: Of the 122 children with early CS enrolled, 24(19.7%) were diagnosed with early CS nephropathy. All of the children with CS nephropathy were young infants < 6 months old. A majority of them showed typical congenital syphilitic skin lesions, but a quarter of them did not have skin lesions. Compared with non-nephropathic children with early CS, those with nephropathy had higher frequency of hepatosplenomegaly, fever, edema, gastrointestinal (GI) symptoms, and anemia, as well as decreased C3 levels. Urinalysis results showed hematuria in all patients with early CS nephropathy, with proteinuria and renal function impairment in 91.7% and 12.5% of the patients, respectively. Nephritic-type nephrotic syndrome and glomerulonephritis were diagnosed in 45.8% and 54.2% of these patients, respectively. All infants with CS nephropathy were cured or improved after appropriate treatments. CONCLUSION: Infants with early CS nephropathy often presented with nephritic-type nephrotic syndrome or glomerulonephritis, and the typical skin lesions, fever, hepatosplenomegaly, and edema, etc., were its common clinical presentations, and these characteristics could help with the diagnosis. But for infants with nephropathy who did not have typical clinical presentations, CS should also be screened as an important etiology to avoid misdiagnosis.

PEDV promotes the differentiation of CD4+T cells towards Th1, Tfh, and Treg cells via CD103+DCs.

Porcine epidemic diarrhea virus (PEDV) can infect all ages of pigs, particularly newborn piglets with a mortality almost reaching to 80-100%, causing significant economic losses to the global pig industry. The mucosal immune response is crucial for PEDV prevention, in which specific dendritic cells (DCs) and differentiated T cells play vital roles. In this study, CD103+DCs were differentiated successfully with retinoic acid (RA) treatment in vitro. PEDV could not replicate efficiently in differentiated CD103+DCs but could promote maturation of CD103+DCs by up-regulating the expression of SLA-DR, CD1a, CD86, and cytokines of IL-1ß and IL-10. In addition, PEDV-infected CD103+DCs and CD4+T cells were co-cultured, and the results showed that the differentiation of CD4+T cells toward Th1, Tfh, and Treg, but not Th2. These results demonstrate that PEDV-infected CD103+DCs could promote the differentiation of CD4+T cells, which provided the basis for further study of mucosal response induced by PEDV via CD103+DCs.

Linoleic acid stimulation results in TGF-ß1 production and inhibition of PEDV infection in vitro.

Linoleic acid (LA) is recommended to improve pork quality. However, whether it affects the intestinal immune response in pigs is still unclear. Our ex vivo experiments demonstrated that LA stimulation resulted in increased frequencies of Tregs in PBMCs but not in Peyer's Patches (PPs). The results of RT-qPCR, flow cytometry, and ELISA indicated that LA increased the TGF-ß1 expression level in DCs isolated from PEDV infected pigs. Furthermore, RT-qPCR and flow cytometry results demonstrated that TGF-ß1 was associated with higher frequencies of Tregs both in PBMCs and PPs. Additional investigations showed that TGF-ß1 inhibited PEDV infection in vitro. Besides, knocking-out TGF-ß1 in IPEC-J2 cells resulted in higher viral load. Taken together, our results demonstrated that LA stimulation resulted in enhanced production of TGF-ß1 by DC, which resulted in higher frequencies of Tregs production and inhibition of PEDV infection.

Tofacitinib for the Treatment of Steroid-Induced Rosacea.

Purpose: Conventional treatment suffers from a dilemma of poor efficacy. The clinical use of Janus kinase (JAK) inhibitors in the treatment of steroid-induced rosacea has rarely been explored. Patients and Methods: We present a case of steroid-induced rosacea successfully treated with JAK inhibitor tofacitinib, with no adverse effects. Results: This case report of successful treatment shows a good clinical efficacy of JAK inhibitors tofacitinib in the treatment of SIR. Conclusion: JAK inhibitor tofacitinib may be a promising agent for the treatment with steroid-induced rosacea, especially for patients who have failed to conventional therapy.

An analysis of the clinical features of children with early congenital Syphilis and Syphilitic Hepatitis.

BACKGROUND: The infection rate of congenital syphilis is gradually increasing, the clinical manifestations of some children with congenital syphilis are abnormal liver function, which is given the clinical diagnosis of syphilitic hepatitis. At present, there are few studies on the clinical features of children with early congenital syphilis combined with syphilitic hepatitis, so we set out to do such a study. We compared the liver function indicators before and after the treatment of syphilis to find the clinical features that can provide guidance for clinical diagnosis and treatment. METHODS: This study collected clinical data on 51 children with early congenital syphilis combined with syphilitic hepatitis in Beijing Ditan Hospital, affiliated with Capital Medical University, between April 2014 and October 2019. We observed their age, gender, clinical symptoms, and physical symptoms, as well as the pregnancy and childbirth history of their mothers. We also compared the liver function indicators before and after the treatment of the syphilis and analyzed the children's clinical features. RESULTS: The results of this study showed that the clinical manifestations in children with early congenital syphilis combined with syphilitic hepatitis were diverse. The most common clinical manifestation was anemia (56.9 %), followed by syphilitic rash (54.9 %), hands, feet, and whole-body peeling (35.3 %), and splenomegaly (29.4 %). Liver damage caused by a syphilis infection tends to result in elevated alanine aminotransferase, aspartate aminotransferase, and bilirubin, while albumin decreases. After the syphilis treatment, the liver function indexes were significantly improved compared with before treatment, and the difference was statistically significant (all p < 0.05). CONCLUSIONS: A child with abnormal liver function, especially with anemia, skin rash, peeling, abdominal distension, and hepatosplenomegaly should be highly suspected of having a syphilis infection. Once the diagnosis is made, the appropriate standard penicillin treatment should be started as soon as possible to improve the condition and prognosis of the child.

Analysis of clinical characteristics of severe pertussis in infants and children: a retrospective study.

BACKGROUND: The incidence of pertussis shows an increasing trend in recent years, but some clinicians often lack sufficient understanding of the clinical characteristics and risk factors for severe pertussis, and more effective measures should be taken to reduce the incidence and mortality of pertussis in young infants METHODS: A retrospective study was conducted, and 184 infants and children with pertussis who had been hospitalized in the Department of Pediatrics of Beijing Ditan Hospital affiliated with Capital Medical University from January 2016 to December 2017 were included. Clinical data of the patients were collected and the clinical characteristics were statistically analyzed RESULTS: Among the 184 patients, 41.85% were infants < 3 months of age, and 65.22% of the total patients were not vaccinated against pertussis. There were 22 critically ill children, among whom 4 died, and compared with mild cases, they had a higher proportion of children younger than 3 months of age and infants not vaccinated against pertussis (63.64% vs. 38.89% and 100% vs. 60.49%, respectively); a higher proportion of children with severe pneumonia (100% vs. 0%); higher leukocyte count(× 109/L , 35.80 ± 20.53 vs 19.41 ± 8.59); and a higher proportion of children with severe hyperleukocytosis (18.18% vs. 0%, respectively) (P<0.05) CONCLUSIONS: 1. Infants aged <3 months not vaccinated for pertussis appear more likely to become infected and have more severe disease. 2. Severe pneumonia and hyperleukocytosis are the main mechanisms underlying severe pertussis.

MyD88 Mediates Colitis- and RANKL-Induced Microfold Cell Differentiation.

Intestinal microfold (M) cells are critical for sampling antigens in the gut and initiating the intestinal mucosal immune response. In this study, we found that the oral administration of dextran sulfate sodium (DSS) and Salmonella infection induced colitis. In the process, the expression levels of M cell differentiation-related genes were synchronized with the kinetics of pro-inflammatory cytokines. Compared to wild-type (WT) mice, MyD88-/- mice exhibited significantly lower expression levels of M cell differentiation-related genes. However, DSS induced colitis in MyD88-/- mice but failed to promote the transcription of M cell differentiation related genes. Furthermore, the receptor activator of the Nuclear Factor-κB ligand (RANKL) upregulated the transcription of M cell differentiation related genes in murine intestinal organoids prepared from both WT and MyD88-/- mice. Meanwhile, fewer changes in M cell differentiation related genes were found in MyD88-/- mice as compared to WT mice. Hence, we concluded that myeloid differentiation factor 88 (MyD88) is an essential molecule for colitis- and RANKL-related differentiation of M cells.

Epidemiological and clinical characteristics of 35 children with COVID-19 in Beijing, China.

IMPORTANCE: Within the coronavirus disease 2019 (COVID-19) global pandemic, more attention is warranted for whether this new infectious disease has unique manifestations in children. OBJECTIVE: To retrospectively determine the epidemiological and clinical characteristics of 35 children with COVID-19 in Beijing, China. METHODS: We collected data for 35 children diagnosed with COVID-19 who were admitted to Beijing Ditan Hospital from January 2020 to June 2020, and analyzed their epidemiological characteristics, clinical manifestations, laboratory examinations, chest imaging findings, treatments, and outcomes. RESULTS: The children comprised 18 boys (51.4%) and 17 girls (48.6%) aged 6 months to 15 years. All patients had clear epidemiological history, with family clusters accounting for 28 cases (80.0%) and clear tracing of exposure to high epidemic areas in the remaining 7 cases (20.0%). Four (11.4%) patients were classified as asymptomatic, 17 (48.6%) as acute upper respiratory infection, and 14 (40.0%) as mild pneumonia, with no severe or critical cases. Clinical manifestations were mild, including fever in 18 (51.4%), cough in 14 (40.0%), and nausea and diarrhea in 7 (20.0%) patients. White blood cell count was mostly normal (26 cases, 74.3%) or decreased (7 cases, 20.0%); lymphocyte percentage was increased in 24 (68.7%); neutrophil percentage was decreased in 25 (71.4%); alanine aminotransferase was increased in 3 (8.6%); and serum potassium was decreased in 4 (11.4%). Time to negative viral nucleic acid testing was 2-42 days (mean: 14.0 ± 9.4 days). Chest imaging examination revealed that 20 patients (57.1%) had different forms of lung inflammation. Treatment was mainly isolation and nutritional support. Eleven patients were treated with interferon atomization inhalation. No patients required oxygen therapy. All 35 children were cured and discharged. Length of hospital stay was 9-54 days (mean: 25.4 ± 13.8 days). During regular follow-up after discharge, 5 children showed positivity again in the viral nucleic acid test and were re-hospitalized for observation and treatment. The mean length of re-hospitalization stay was 10.8 days. INTERPRETATION: Children with COVID-19 mainly become infected within their family, and children of all ages are generally susceptible. The disease in children is mostly mild and the prognosis is good. Nucleic acid tests in some patients become positive again after discharge, suggesting that it is of great significance to carry out centralized isolation medical observations and active nucleic acid tests in close contacts for early detection of patients and routine epidemic prevention and control.

Corrigendum: Emergence of Getah Virus Infection in Horse With Fever in China, 2018.

[This corrects the article DOI: 10.3389/fmicb.2019.01416.].

Emergence of Getah Virus Infection in Horse With Fever in China, 2018.

Getah virus (GETV) is a mosquito-borne virus that was first determined in Malaysia in 1955, and can infect humans and multiple other mammals. GETV infection in horses has been reported in Japan and India, and causes great economic losses. In China, GETV has been identified in mosquitoes, pigs, foxes, and cattle with a wide geographical distribution, but has not been detected in horses. In August 2018, a sudden onset of fever was observed in racehorse in an equestrian training center in Guangdong Province in southern China. Blood samples were collected from the sick horse, and PCR/RT-PCR analysis was performed to screen for equine viral pathogens associated with fever. The results indicated that the samples were GETV RNA positive. After RT-PCR, sequencing, and assembly, the genome of the first Chinese horse-derived GETV strain, GZ201808, was obtained. Compared with the genome sequences of other GETV strains, twelve unique nucleotide substitutions were observed in GZ201808. The genome of GZ201808 had the highest genetic identity (99.6%) with AH9192, which was detected in pigs in China in 2017. Phylogenetic analysis indicated that GZ201808 clustered in Group III, and was located in an independent branch distant from other horse-derived GETV strains, indicating a unique evolutionary pattern of GZ201808. This study first determined and described the disease course of horse infected with GETV in China, sequenced and characterized the genome of the field horse-derived GETV strain, and therefore presented an unequivocal report of GETV infection in horses in China.

Newcastle disease virus selectively infects dividing cells and promotes viral proliferation.

Newcastle disease virus (NDV) can select cells to infect, but the mechanism of its cell selectivity has not been comprehensively investigated. Here, we use HeLa cells to establish that NDV can selectively infect cells at the single-cell level. We labeled proliferating cells with 5'-bromo-2-deoxyuridine (BrdU) and examined the colocalization of BrdU with NDV in cells to clarify the relationships between NDV infection and cell proliferation. Receptors at the plasma membrane mediate NDV entry into host cells. We labeled sialic acid receptor isoforms, compared their densities between different cell types and measured the sialic acid receptor densities in different cell phases. Our results suggest that NDV displays host tropism to HeLa cells compared to BHK cells and that the differences in the receptor isoform expression patterns between cell types contribute to the selection of HeLa by NDV. At the single-cell level, the dynamics of receptor expression changes during different cell phases contributing to the selection of cells in S/G2 phase for NDV infection. Furthermore, cell proliferation benefits viral replication, and enhanced virus replication leads to increased damage to cells. The elucidation of the mechanisms underlying host cell selection by NDV may help in the screening and characterizing of additional candidate oncolytic virus strains.

Clinical research on therapeutic effect of combined application of lobaplatin and irinotecan in treating recurrant small cell lung cancer.

To observe and analyze the therapeutic effect of combinated application of lobaplatin and irinotecan in treating recurrant small cell lung cancer. The 140 patients who were treated in our hospital for recurrant small cell lung cancer were selected as research objects. All selected patients were subjected to combined application of obaplatin and irinotecan, the total therapeutic effect was observed, and the adverse reactions occurring during treatment were recorded. Through observing the total treatment effective ratio of 140 patients recurrant small cell lung cancer, the number of complete remission cases, number of partial remission cases, number of stable disease cases, and number of disease progression cases were 40(28.57%), 28(20.00%), 30(21.43%), 42(30.00%), respectively, with total effective ratio of 48.57%. The average time to progression (TTP) was (4.5±0.8) months, average overall survival (OS) was (7.6±1.2) months. The toxic and adverse effects mainly included hematological toxicity and gastrointestinal adverse reaction, such as leukocyte reduction, neutrophil reduction, thrombocytopenia, decreased hemoglobin, nausea and vomiting, diarrhea. No toxicity-related death occurred. In treatment of patients with recurrant small cell lung cancer, the combined application of lobaplatin and irinotecan can achieve great results, which is a safe and reliable way of treatment.

Newcastle disease virus V protein inhibits apoptosis in DF-1 cells by downregulating TXNL1.

Many viral proteins are related to suppressing apoptosis in target cells and are hence beneficial to viral replication. The V protein of Newcastle disease virus (NDV) is one such protein that plays an important role in inhibiting apoptosis in a species-specific manner. However, to date, there have been no reports clarifying the antiapoptotic mechanisms of the V protein. The present study was undertaken to determine the apoptotic potential of the V protein in a chicken embryo fibroblast cell line (DF-1 cell) and to elucidate its molecular mechanisms of action. Here, a yeast two-hybrid system was used to screen the host proteins that interact with the V protein and identified thioredoxin-like protein 1 (TXNL1) as a potential binding partner. Immuno-colocalization of V protein and TXNL1 protein in DF-1 cells further verified the interaction of the two proteins. Through the overexpression of TXNL1 protein and knockdown of TXNL1 protein in DF-1 cells, the effects of NDV replication and cell apoptosis were examined. Cell apoptosis was detected by flow cytometry. The mRNA and protein expression levels of Bax, Bcl-2 and Caspase-3 were detected by quantitative real-time PCR (Q-PCR) and Western blotting. NDV expression was detected by Q-PCR and plaque assay. The results revealed that the TXNL1 protein induced apoptosis and inhibited NDV replication in DF-1 cells. Furthermore, the Western blot and Q-PCR results suggested that TXNL1 induced cell apoptosis through a pathway involving Bcl-2\Bax and Caspase-3. Finally, this work provides insight into the mechanism by which the V protein inhibits apoptosis.