Levels Of Viral Antigen Research Articles

Detection of Double-Stranded RNA Intermediates During SARS-CoV-2 Infections of Syrian Golden Hamsters with Monoclonal Antibodies and Its Implications for Histopathological Evaluation of In Vivo Studies.

The SARS-CoV-2 pandemic has highlighted the challenges posed by the emergence and rapid global spread of previously unknown viruses. Early investigations on the pathogenesis of newly identified viruses are often hampered by a lack of appropriate sample material and conventional detection methods. In this study, viral replication within the lungs of SARS-CoV-2-infected Syrian golden hamsters was assessed by immunolabeling dsRNA intermediates with three different monoclonal antibodies in formalin-fixed, paraffin-embedded tissue samples. The presence of dsRNA was compared to viral antigen levels, viral titers, and genomic RNA replicates using three different variants of concern and an ancestral virus strain at a single time point and during the course of infection with an ancestral variant, and then validated using fluorescent 2-plex in situ hybridization. The results indicate that the detection of viral infection using anti-dsRNA antibodies is restricted to an early phase of infection with high viral replication activity. Additionally, the combined detection of dsRNA intermediates and viral antigens may help to bridge the interpretation gaps between viral antigen levels and viral titers at a single time point. Further testing in other viral infections or species is needed to assess the potential of dsRNA as an early marker for viral infections.

LBP-005 Efficacy, safety, tolerability, and immunogenicity of JNJ-0535 following a reduction of viral antigen levels through administration of siRNA JNJ-3989 in patients with chronic HBeAg negative hepatitis B-interim data of the OSPREY study

LBP-005 Efficacy, safety, tolerability, and immunogenicity of JNJ-0535 following a reduction of viral antigen levels through administration of siRNA JNJ-3989 in patients with chronic HBeAg negative hepatitis B-interim data of the OSPREY study

Kinetics of SARS-CoV-2 infection biomarkers in a household transmission study

SARS-CoV-2 is the causative agent of COVID-19. Timely and accurate diagnostic testing is vital to contain the spread of infection, reduce delays in treatment and care, and inform patient management. Optimal specimen type (e.g. nasal swabs or saliva), timing of sampling, viral marker assayed (RNA or antigen), and correlation with viral infectivity and COVID-19 symptoms severity remain incompletely defined. We conducted a field study to evaluate SARS-CoV-2 viral marker kinetics starting from very early times after infection. We measured RNA and antigen levels in nasal swabs and saliva, virus outgrowth in cell culture from nasal swabs, and antibody levels in blood in a cohort of 30 households. Nine household contacts (HHC) became infected with SARS-CoV-2 during the study. Viral RNA was detected in saliva specimens approximately 1–2 days before nasal swabs in six HHC. Detection of RNA was more sensitive than of antigen, but antigen detection was better correlated with culture positivity, a proxy for contagiousness. Anti-nucleocapsid antibodies peaked one to three weeks post-infection. Viral RNA and antigen levels were higher in specimens yielding replication competent virus in cell culture. This study provides important data that can inform how to optimally interpret SARS-CoV-2 diagnostic test results.

Multidrug-resistant mutations to antiviral and antibody therapy in an immunocompromised patient infected with SARS-CoV-2

Antiviral and antibody therapies for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are being recommended for high-risk patients, but the potential for the development of multidrug-resistant mutations in immunocompromised patients is unclear. To investigate the treatment course in cases of prolonged viral shedding in an immunocompromised patient with SARS-CoV-2 infection, we conducted longitudinal measurements of laboratory tests, chest computed tomography (CT) image evaluations, antibody titers, and antigen levels in nasopharyngeal swabs. Furthermore, we performed whole-genome sequencing and digital PCR analysis to examine the mechanisms of drug resistance. We present a case of a 65-year-old man with a history of malignant lymphoma who was treated with multiple antiviral and antibody therapies, including sotrovimab, remdesivir, paxlovid (nirmatrelvir/ritonavir), and molnupiravir. Initially, viral antigen levels decreased after treatments. However, after the virus rebounded, the patient showed no virologic response. The viral genome analysis revealed a single Omicron subvariant (BA.1.1), which evolved within the host during the disease progression. The viruses had acquired multiple resistance mutations to nirmatrelvir (3 chymotrypsin-like protease [3CLpro] E166 A/V), sotrovimab (spike P337L and E340K), and remdesivir (RNA-dependent RNA polymerase [RdRp] V166L). Our results indicate that viruses with multidrug-resistant mutations and survival fitness persist in the infected subpopulation after drug selection pressure. This study was supported by the JSPS KAKENHI Early-Career Scientists 18K16292 (Y.H.), Grant-in-Aid for Scientific Research (B) 20H03668 and 23H02955 (Y.H.), the YASUDA Medical Foundation (Y.H.), the Uehara Memorial Foundation (Y.H.), the Takeda Science Foundation (Y.H.), and Kato Memorial Bioscience Foundation (Y.H.).

Lower pretreatment HBV DNA levels are associated with better off-treatment outcomes after nucleo(s)tide analogue withdrawal in patients with HBeAg-negative chronic hepatitis B: A multicentre cohort study

Background & AimsPretreatment predictors of finite nucleo(s)tide analogue (NUC) therapy remain elusive. We studied the association between pretreatment HBV DNA levels and outcomes after therapy cessation. MethodsPatients with chronic hepatitis B who were HBeAg negative at the start of NUC treatment were enrolled from sites in Asia and Europe. We studied the association between pretreatment HBV DNA levels and (1) clinical relapse (defined as HBV DNA >2,000 IU/ml + alanine aminotransferase >2 × the upper limit of normal or retreatment) and (2) HBsAg loss after NUC withdrawal. ResultsWe enrolled 757 patients, 88% Asian, 57% treated with entecavir, with a median duration of treatment of 159 (IQR 156–262) weeks. Mean pretreatment HBV DNA levels were 5.70 (SD 1.5) log IU/ml and were low (<20,000 IU/ml) in 150 (20%) and high (>20,000 IU/ml) in 607 (80%). The cumulative risk of clinical relapse at 144 weeks after therapy cessation was 22% among patients with pretreatment HBV DNA levels <20,000 IU/ml vs. 60% among patients with pretreatment HBV DNA levels >20,000 IU/ml, whereas the cumulative probabilities of HBsAg loss were 17.5% vs. 5% (p <0.001). In multivariable analysis, pretreatment HBV DNA levels <20,000 IU/ml were independently associated with a reduced likelihood of clinical relapse (adjusted hazard ratio 0.379, p <0.001) and with an increased chance of HBsAg loss (adjusted hazard ratio 2.872, p <0.001). ConclusionsLower pretreatment HBV DNA levels are associated with a lower risk of clinical relapse and a higher chance of HBsAg loss after cessation of NUC therapy, independent of end-of-treatment viral antigen levels. Further studies are needed to confirm these findings in non-Asian populations. Impact and ImplicationsA subgroup of patients with chronic hepatitis B may not require retreatment after stopping antiviral therapy. In this study, comprising 757 patients with chronic hepatitis B from Europe and Asia, we found that higher viral load before initiation of treatment was a risk factor for relapse after stopping treatment. Patients with a low HBV DNA level before starting antiviral therapy had the lowest risk of relapse, and a high chance of HBsAg loss, after stopping treatment. These findings can help select patients for treatment withdrawal and guide intensity of off-treatment monitoring.

Plasma SARS-CoV-2 nucleocapsid antigen levels are associated with progression to severe disease in hospitalized COVID-19

BackgroundStudies quantifying SARS-CoV-2 have focused on upper respiratory tract or plasma viral RNA with inconsistent association with clinical outcomes. The association between plasma viral antigen levels and clinical outcomes has not been previously studied. Our aim was to investigate the relationship between plasma SARS-CoV-2 nucleocapsid antigen (N-antigen) concentration and both markers of host response and clinical outcomes.MethodsSARS-CoV-2 N-antigen concentrations were measured in the first study plasma sample (D0), collected within 72 h of hospital admission, from 256 subjects admitted between March 2020 and August 2021 in a prospective observational cohort of hospitalized patients with COVID-19. The rank correlations between plasma N-antigen and plasma biomarkers of tissue damage, coagulation, and inflammation were assessed. Multiple ordinal regression was used to test the association between enrollment N-antigen plasma concentration and the primary outcome of clinical deterioration at one week as measured by a modified World Health Organization (WHO) ordinal scale. Multiple logistic regression was used to test the association between enrollment plasma N-antigen concentration and the secondary outcomes of ICU admission, mechanical ventilation at 28 days, and death at 28 days. The prognostic discrimination of an externally derived “high antigen” cutoff of N-antigen ≥ 1000 pg/mL was also tested.ResultsN-antigen on D0 was detectable in 84% of study participants. Plasma N-antigen levels significantly correlated with RAGE (r = 0.61), IL-10 (r = 0.59), and IP-10 (r = 0.59, adjusted p = 0.01 for all correlations). For the primary outcome of clinical status at one week, each 500 pg/mL increase in plasma N-antigen level was associated with an adjusted OR of 1.05 (95% CI 1.03–1.08) for worse WHO ordinal status. D0 plasma N-antigen ≥ 1000 pg/mL was 77% sensitive and 59% specific (AUROC 0.68) with a positive predictive value of 23% and a negative predictive value of 93% for a worse WHO ordinal scale at day 7 compared to baseline. D0 N-antigen concentration was independently associated with ICU admission and 28-day mechanical ventilation, but not with death at 28 days.ConclusionsPlasma N-antigen levels are readily measured and provide important insight into the pathogenesis and prognosis of COVID-19. The measurement of N-antigen levels early in-hospital course may improve risk stratification, especially for identifying patients who are unlikely to progress to severe disease.

When and which patients should receive remdesivir?

When and which patients should receive remdesivir?

COVID-19 associated EBV reactivation and effects of ganciclovir treatment.

BackgroundSystemic reactivation of Epstein–Barr virus (EBV) may occur in novel coronavirus disease 2019 (COVID‐19) caused by the severe acute respiratory syndrome coronavirus‐2 (SARS‐CoV‐2). However, the clinical consequences of EBV reactivation remain uncertain.MethodsIn this retrospective study, we screened 1314 patients with confirmed COVID‐19 who died or were discharged between January 1, 2020 and March 12, 2020, in Wuhan Infectious Disease Hospital, Wuhan, China. Patients who had complete data for EBV serology and cytomegalovirus (CMV) serology were eligible. Serum levels of viral capsid antigen (VCA)‐immunoglobulin G (IgG), Epstein–Barr nuclear antigen‐IgG, VCA‐IgM, early antigen (EA)‐IgG, CMV‐IgG, and CMV‐IgM were compared between survivors and nonsurvivors. Dynamic changes of laboratory tests and outcomes were compared in patients with and without ganciclovir treatment. We used 1:1 matching based on age, gender, and illness severity to balance baseline characteristics.ResultsEBV reactivation was present in 55 of 217 patients. EBV reactivation was associated with age (57.91 [13.19] vs. 50.28 [12.66] years, p < .001), female gender (31 [56%] vs. 60 [37%], p = .02). Patients with EBV reactivation have statistically nonsignificant higher mortality rate (12 [22%] vs. 18 [11%], p = .08). EA‐IgG levels were significantly higher in nonsurvivors than in survivors (median difference: −0.00005, 95% confidence interval, CI [−3.10, 0.00], p = .05). As compared to patients with COVID‐19 who did not receive ganciclovir therapy, ganciclovir‐treated patients had improved survival rate (0.98, 95% CI [0.95, 1.00] vs. 0.88, 95% CI [0.81, 0.95], p = .01). Hemoglobin (p < .001) and prealbumin (p = .02) levels were significantly higher in ganciclovir‐treated patients.ConclusionA high proportion of COVID‐19 patients had EBV reactivation that may be associated with an increased risk of death. Whether treatment with ganciclovir may decrease the mortality of COVID‐19 patients complicated with EBV reactivation warrants to be addressed in a placebo‐controlled randomized trial in the future.

Probability of HBsAg loss after nucleo(s)tide analogue withdrawal depends on HBV genotype and viral antigen levels

Nucleo(s)tide analogue (NUC) withdrawal may result in HBsAg clearance in a subset of patients. However, predictors of HBsAg loss after NUC withdrawal remain ill-defined. We studied predictors of HBsAg loss in a global cohort of HBeAg-negative patients with undetectable HBV DNA who discontinued long-term NUC therapy. Patients requiring retreatment after treatment cessation were considered non-responders. We enrolled 1,216 patients (991 with genotype data); 98 (8.1%) achieved HBsAg loss. The probability of HBsAg loss was higher in non-Asian patients (adjusted hazard ratio [aHR] 8.26, p <0.001), and in patients with lower HBsAg (aHR 0.243, p <0.001) and HBV core-related antigen (HBcrAg) (aHR 0.718, p= 0.001) levels. Combining HBsAg (<10, 10-100 or >100 IU/ml) and HBcrAg (<2log vs. ≥2 log) levels improved prediction of HBsAg loss, with extremely low rates observed in patients with HBsAg >100 IU/ml with detectable HBcrAg. HBsAg loss rates also varied with HBV genotype; the highest rates were observed for genotypes A and D, and none of the patients with HBV genotype E experienced HBsAg loss (p <0.001 for the overall comparison across genotypes; p <0.001 for genotypes A/D vs. genotypes B/C). HBV genotype C was independently associated with a higher probability of HBsAg loss when compared to genotype B among Asian patients (aHR 2.494; 95% CI 1.490-4.174, p= 0.001). The probability of HBsAg loss after NUC cessation varies according to patient ethnicity, HBV genotype and end-of-treatment viral antigen levels. Patients with low HBsAg (<100 IU/ml) and/or undetectable HBcrAg levels, particularly if non-Asian or infected with HBV genotype C, appear to be the best candidates for treatment withdrawal. A subset of patients may achieve clearance of hepatitis B surface antigen (HBsAg) - so-called functional cure - after withdrawal of nucleo(s)tide analogue therapy. In this multicentre study of 1,216 patients who discontinued antiviral therapy, we identified non-Asian ethnicity, HBV genotype C, and low hepatitis B surface antigen and hepatitis B core-related antigen levels as factors associated with an increased chance of HBsAg loss.

Characterization of the liver immune microenvironment in liver biopsies from patients with chronic HBV infection

Background & AimsWe aim to describe the liver immune microenvironment by analyzing liver biopsies from patients with chronic HBV infection (CHB). Host immune cell signatures and their corresponding localization were characterized by analyzing the intrahepatic transcriptome in combination with a custom multiplex immunofluorescence panel.MethodMatching FFPE and fresh frozen liver biopsies were collected from immune active patients within the open-label phase IV study GS-US-174-0149. RNA-Seq was conducted on 53 CHB liver biopsies from 46 patients. Twenty-eight of the 53 samples had matched FFPE biopsies and were stained with a 12-plex panel including cell segmentation, immune and viral biomarkers. Corresponding serum samples were screened using the MSD Human V-plex Screen Service to identify peripheral correlates for the immune microenvironment.ResultsUsing unsupervised clustering of the transcriptome, we reveal two unique liver immune signatures classified as immune high and immune low based on the quantification of the liver infiltrate gene signatures. Multiplex immunofluorescence analysis demonstrated large periportal lymphoid aggregates in immune high samples consisting of CD4 and CD8 T cells, B cells and macrophages. Differentiation of the high and low immune microenvironments was independent of HBeAg status and peripheral viral antigen levels. In addition, longitudinal analysis indicates that treatment and normalization of ALT correlates with a decrease in liver immune infiltrate and inflammation. Finally, we screened a panel of peripheral biomarkers and identified ICAM-1 and CXCL10 as biomarkers that strongly correlate with these unique immune microenvironments.ConclusionThese data provide a description of immune phenotypes in patients with CHB and show that immune responses are downregulated in the liver following nucleotide analogue treatment. This may have important implications for both the safety and efficacy of immune modulator programs aimed at HBV cure.Lay summaryLiver biopsies from patients with chronic hepatitis B were submitted to RNA-Seq and multiplex immunofluorescence and identified two different liver immune microenvironments: immune high and immune low. Immune high patients showed elevated immune pathways, including interferon signaling pathways, and increase presence of immune cells. Longitudinal analysis of biopsies from treatment experienced patients showed that treatment correlates with a marked decrease in inflammation and these findings may have important implications for both safety and efficacy of immune modulator programs for HBV cure.

Hepatitis B Viral Markers in the Human Milk of HBsAg-Positive Mothers: An Observational Study

Quantification of viral antigens and viral loads in human milk samples from mothers infected with hepatitis B virus is largely unknown. The aim of the study was to quantitatively measure the levels of viral antigens and deoxyribonucleic acid of hepatitis B virus in human milk from mothers infected with hepatitis B virus. Fifty-five pairs of milk and serum samples from mothers with positive hepatitis B surface antigen, including 11 hepatitis B e antigen positive, were quantitatively tested to measure viral antigens by microparticle enzyme immunoassay and viral loads by real-time polymerase chain reaction assay. The median level of hepatitis B surface antigen in the human milk samples of mothers with positive or negative hepatitis B e antigen was each lower than that in the sera, respectively (1.10 vs. 4.32 log10 IU/ml, t = 10.693, p < .001; -0.77 vs. 2.53 log10 IU/ml, t = -25.135, p < .001). The titers of hepatitis B surface antigen or hepatitis B e antigen in the human milk samples were each correlated with that in maternal serum. The detectable level of deoxyribonucleic acid of hepatitis B virus in human milk ranged from 1.42-5.27 log10 IU/ml, whereas that in maternal sera was 1.44-8.66 log10 IU/ml. The viral level in human milk was not correlated with that in maternal circulation. The present study data illustrate the relatively low titers of viral markers in the milk of mothers with positive hepatitis B surface antigen.

Antiviral therapy after curative treatment of hepatocellular carcinoma in patients with chronic hepatitis B infection: Is tenofovir or entecavir preferred?

Antiviral therapy after curative treatment of hepatocellular carcinoma in patients with chronic hepatitis B infection: Is tenofovir or entecavir preferred?

Comparative analysis of antigen and molecular tests for the detection of Sars-CoV-2 and related variants: A study on 4266 samples

ObjectivesThe present study compared the performance of the Lumipulse G Sars-CoV-2 Ag kit with the TaqPath COVID-19 RT-PCR CE IVD kit. MethodsThe study was conducted on 4266 naso-oropharyngeal swabs. Samples were subjected to antigen RT-PCR tests for the detection of Sars-CoV-2 and related variants. Statistical analyses were conducted in R software. ResultsWe found 503 positives (including 138 H69-V70 deletion carriers) and 3763 negatives by RT-PCR, whereas 538 positives and 3728 negatives were obtained by antigen testing. We achieved empirical and binormal AU-ROCs of 0.920 and 0.990, accuracy of 0.960, sensitivity of 0.866, specificity of 0.973, positive and negative predictive values of 0.810 and 0.980. We obtained a positive correlation between viral loads and antigen levels (R2 = 0.81), finding a complete concordance for high viral loads (log10 copies/mL > 5.4). Antigen levels > 222 pg/mL were found to be reliable in assigning positive samples (p < 0.01). Concerning variant carriers, antigen test detected them with the same accuracy as other positive samples. ConclusionsMolecular and antigen tests should be evaluated regarding the prevalence of the area. In case of low prevalence, antigen testing can be employed as a first-line screening for the timely identification of affected individuals with high viral load, also if carriers of Sars-CoV-2 variants.

HTLV-1 targets human placental trophoblasts in seropositive pregnant women.

Human T cell leukemia virus type 1 (HTLV-1) is mainly transmitted vertically through breast milk. The rate of mother-to-child transmission (MTCT) through formula feeding, although significantly lower than through breastfeeding, is approximately 2.4%-3.6%, suggesting the possibility of alternative transmission routes. MTCT of HTLV-1 might occur through the uterus, birth canal, or placental tissues; the latter is known as transplacental transmission. Here, we found that HTLV-1 proviral DNA was present in the placental villous tissues of the fetuses of nearly half of pregnant carriers and in a small number of cord blood samples. An RNA ISH assay showed that HTLV-1-expressing cells were present in nearly all subjects with HTLV-1-positive placental villous tissues, and their frequency was significantly higher in subjects with HTLV-1-positive cord blood samples. Furthermore, placental villous trophoblasts expressed HTLV-1 receptors and showed increased susceptibility to HTLV-1 infection. In addition, HTLV-1-infected trophoblasts expressed high levels of viral antigens and promoted the de novo infection of target T cells in a humanized mouse model. In summary, during pregnancy of HTLV-1 carriers, HTLV-1 was highly expressed in placental villous tissues, and villous trophoblasts showed high HTLV-1 sensitivity, suggesting that MTCT of HTLV-1 occurs through the placenta.

Comparison of automated SARS-CoV-2 antigen test for COVID-19 infection with quantitative RT-PCR using 313 nasopharyngeal swabs, including from seven serially followed patients

In routine clinical practice, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is determined by reverse-transcription PCR (RT-PCR). In the current pandemic, a more rapid and high-throughput method is in growing demand. Here, we validated the performance of a new antigen test (LUMIPULSE) based on chemiluminescence enzyme immunoassay. A total of 313 nasopharyngeal swabs (82 serial samples from 7 infected patients and 231 individual samples from 4 infected patients and 215 uninfected individuals) were analyzed for SARS-CoV-2 with quantitative RT-PCR (RT-qPCR) and then subjected to LUMIPULSE. We determined the cutoff value for antigen detection using receiver operating characteristic curve analysis and compared the performance of the antigen test with that of RT-qPCR. We also compared the viral loads and antigen levels in serial samples from seven infected patients. Using RT-qPCR as the reference, the antigen test exhibited 55.2% sensitivity and 99.6% specificity, with a 91.4% overall agreement rate (286/313). In specimens with > 100 viral copies and between 10 and 100 copies, the antigen test showed 100% and 85% concordance with RT-qPCR, respectively. This concordance declined with lower viral loads. In the serially followed patients, the antigen levels showed a steady decline, along with viral clearance. This gradual decline was in contrast with the abrupt positive-to-negative and negative-to-positive status changes observed with RT-qPCR, particularly in the late phase of infection. In summary, the LUMIPULSE antigen test can rapidly identify SARS-CoV-2-infected individuals with moderate to high viral loads and may be helpful for monitoring viral clearance in hospitalized patients.

Pilot study of tenofovir disoproxil fumarate and pegylated interferon-alpha 2a add-on therapy in Japanese patients with chronic hepatitis B.

A prospective pilot study of tenofovir disoproxil fumarate (TDF) and pegylated interferon alpha 2a (P-IFN) add-on therapy was conducted to evaluate its efficacy in reducing viral antigen levels in Japanese patients with chronic hepatitis B (UMIN 000020179). Patients with chronic hepatitis B receiving maintenance TDF therapy and exhibiting hepatitis B surface antigen (HBsAg) level > 800IU/ml were divided into two arms. P-IFN was added for 48weeks in the add-on arm (n = 32), while TDF monotherapy was maintained in the control arm (n = 51). Both groups were followed for 96weeks after baseline measurements. Almost all patients in the control arm displayed a slow and constant reduction in HBsAg during follow-up. In contrast, roughly half of the add-on arm exhibited a sharp decline in HBsAg during P-IFN administration, which disappeared after halting P-IFN. At 96weeks after baseline, 41% (13/32) of patients in the add-on arm had shown a rapid decrease in HBsAg, versus 2% (1/51) in the control arm (p < 0.001). Add-on therapy and increased cytotoxic T-cell response were significant factors associated with a rapid decrease in HBsAg according to multivariate analysis. In addition, higher HB core-related antigen (HBcrAg) level at baseline (p = 0.001) and add-on therapy (p = 0.036) were significant factors associated with a rapid reduction in HBcrAg. TDF and P-IFN add-on therapy in Japanese patients with chronic hepatitis B facilitated rapid decreases in HBsAg and HBcrAg. Further studies are needed to improve early HBsAg clearance rate.

Immune stimulatory and anti-HIV-1 potential of extracts derived from marine brown algae Padina tetrastromatica.

Background Marine brown algae are biologically diverse and their medicinal value has been explored limited. We assessed whether Padina tetrastromatica Hauck will possess the immune stimulatory and human immunodeficiency virus (HIV-1) inhibitory activity. Materials and Methods Aqueous and methanolic extracts were tested for the Th1/Th2 cytokines using PBMC. Subsequently, leukotriene B4 (LTB4), nitric oxide (NO) and anti-oxidant effect were analyzed using RAW264.7 cells. In addition, Padina extracts were tested for the HIV-1 clade C & A by measuring the levels of viral p24 antigen in infected peripheral blood mononuclear cells (PBMCs) and against reverse transcriptase (RT). Results At 100 μg/mL, aqueous and methanolic extracts produced a significant amount of IL-10 and IFN-γ at 24 h and 72 h post-stimulation by PBMCs. It also produced a significant amount of LTB4, NO and had an antioxidant effect on RAW264.7 cell, suggesting the immune stimulating potential of P. tetrastromatica. Upon infection of PBMCs with 100 TCID50, aqueous and methanolic extracts of P. tetrastromatica inhibited HIV-1 C (>90%) and HIV-1 A (>50%) showed a significant reduction in HIV-1 p24 levels and HIV-1 RT inhibition (>50%). GC-MS study revealed a relative abundance of tetradecanoic and oleic acid in the methanolic extract of P. tetrastromatica, which might be responsible for immune stimulation and anti-HIV-1 activity. Conclusion At lower concentrations (100 mg/mL), the aqueous and methanolic extracts of P. tetrastromatica showed the strong immune stimulation and greatest anti-HIV-1 potential in vitro. This study demonstrates the therapeutic potential of these brown algae P. tetrastromatica for the benefit of mankind.

The characteristics of hDPP4 transgenic mice subjected to aerosol MERS coronavirus infection via an animal nose-only exposure device.

BackgroundMiddle East respiratory syndrome coronavirus (MERS‐CoV), which is not fully understood in regard to certain transmission routes and pathogenesis and lacks specific therapeutics and vaccines, poses a global threat to public health.MethodsTo simulate the clinical aerosol transmission route, hDPP4 transgenic mice were infected with MERS‐CoV by an animal nose‐only exposure device and compared with instillation‐inoculated mice. The challenged mice were observed for 14 consecutive days and necropsied on days 3, 5, 7, and 9 to analyze viral load, histopathology, viral antigen distribution, and cytokines in tissues.ResultsMERS‐CoV aerosol‐infected mice with an incubation period of 5‐7 days showed weight loss on days 7‐11, obvious lung lesions on day 7, high viral loads in the lungs on days 3‐9 and in the brain on days 7‐9, and 60% survival. MERS‐CoV instillation‐inoculated mice exhibited clinical signs on day 1, obvious lung lesions on days 3‐5, continuous weight loss, 0% survival by day 5, and high viral loads in the lungs and brain on days 3‐5. Viral antigen and high levels of proinflammatory cytokines and chemokines were detected in the aerosol and instillation groups. Disease, lung lesion, and viral replication progressions were slower in the MERS‐CoV aerosol‐infected mice than in the MERS‐CoV instillation‐inoculated mice.ConclusionhDPP4 transgenic mice were successfully infected with MERS‐CoV aerosols via an animal nose‐only exposure device, and aerosol‐ and instillation‐infected mice simulated the clinical symptoms of moderate diffuse interstitial pneumonia. However, the transgenic mice exposed to aerosol MERS‐CoV developed disease and lung pathology progressions that more closely resembled those observed in humans.

Positive modulation of mGluR5 attenuates seizures and reduces TNF-α+ macrophages and microglia in the brain in a murine model of virus-induced temporal lobe epilepsy

Viral encephalitis markedly increases the risk for the development of epilepsy. The Theiler's murine encephalomyelitis virus (TMEV)-induced model of seizures/epilepsy is a murine model of both viral-induced seizures/epilepsy and human Temporal Lobe Epilepsy. The inflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α have been shown to play a role in seizure development in the TMEV-induced model of seizures/epilepsy, and infiltrating macrophages along with microglia have been shown to be major producers of these cytokines. The metabotropic glutamate receptor 5 (mGluR5) is a G-protein coupled receptor that has been shown to reduce IL-6 and TNF-α and to provide neuroprotection in other disease models. Therefore, we hypothesized that stimulation of mGluR5 would not only reduce seizures but attenuate IL-6 and TNF-α production in microglia and macrophages in the TMEV model. We found that pharmacological stimulation of mGluR5 with the selective positive allosteric modulator VU0360172 not only reduced acute seizure outcomes, but also reduced the percent of microglia and macrophages producing TNF-α 3 days post infection. Furthermore, treatment with VU0360172 did not alter the level of viral antigen, compared to controls, showing that this treatment does not compromise viral clearance. These results establish that mGluR5 may represent a therapeutic target in the TMEV-induced model of seizures/epilepsy.

Outcome of Antiviral Immunity in the Liver Is Shaped by the Level of Antigen Expressed in Infected Hepatocytes.

The liver bears unique immune properties that support both immune tolerance and immunity, but the mechanisms responsible for clearance versus persistence of virus‐infected hepatocytes remain unclear. Here, we dissect the factors determining the outcome of antiviral immunity using recombinant adenoviruses that reflect the hepatropism and hepatrophism of hepatitis viruses. We generated replication‐deficient adenoviruses with equimolar expression of ovalbumin, luciferase, and green fluorescent protein driven by a strong ubiquitous cytomegalovirus (CMV) promoter (Ad‐CMV‐GOL) or by 100‐fold weaker, yet hepatocyte‐specific, transthyretin (TTR) promoter (Ad‐TTR‐GOL). Using in vivo bioluminescence to quantitatively and dynamically image luciferase activity, we demonstrated that Ad‐TTR‐GOL infection always persists, whereas Ad‐CMV‐GOL infection is always cleared, independent of the number of infected hepatocytes. Failure to clear Ad‐TTR‐GOL infection involved mechanisms acting during initiation as well as execution of antigen‐specific immunity. First, hepatocyte‐restricted antigen expression led to delayed and curtailed T‐cell expansion—10,000‐fold after Ad‐CMV‐GOL versus 150‐fold after Ad‐TTR‐GOL‐infection. Second, CD8 T‐cells primed toward antigens selectively expressed by hepatocytes showed high PD‐1/Tim‐3/LAG‐3/CTLA‐4/CD160 expression levels similar to that seen in chronic hepatitis B. Third, Ad‐TTR‐GOL but not Ad‐CMV‐GOL‐infected hepatocytes escaped being killed by effector T‐cells while still inducing high PD‐1/Tim‐3/LAG‐3/CTLA‐4/CD160 expression, indicating different thresholds of T‐cell receptor signaling relevant for triggering effector functions compared with exhaustion. Conclusion: Our study identifies deficits in the generation of CD8 T‐cell immunity toward hepatocyte‐expressed antigens and escape of infected hepatocytes expressing low viral antigen levels from effector T‐cell killing as independent factors promoting viral persistence. This highlights the importance of addressing both the restauration of CD8 T‐cell dysfunction and overcoming local hurdles of effector T‐cell function to eliminate virus‐infected hepatocytes.