Decrease In Titer Research Articles

Humoral and Cellular Responses to COVID-19 Vaccination Indicate the Need for Post-Vaccination Testing in Frail Population.

Despite the high efficacy of the BNT162b2 vaccine in the general population, data on its immunogenicity among frail elderly individuals are limited. Recently, levels of anti-SARS-CoV-2 spike IgG antibodies and serum neutralization titers were confirmed as good immune markers of protection against the virus, with evidence showing a reverse correlation between these two parameters and susceptibility to infection. Here we analyzed sera from 138 nursing home residents (median age of 88.9 years) and 312 nursing home staff (median age of 50.7 years) to determine the humoral response to two doses of the BNT162b2 vaccine, and found markedly decreased serum anti-spike antibody levels and neutralization titers in the nursing home resident (NHR) group, with over 11% non-responders compared to only 1.3% among the controls. Moreover, three months post-vaccination, a significant decrease in antibody titers was observed in COVID-19-naive nursing home residents. Subsequent flow cytometry and interferon gamma secretion analyses indicated that antibody non-responders among NHRs also failed to mount cellular responses. The presented data emphasize that additional measures are needed in the population of frail elderly individuals. Given the high proportion of non-responders among NHRs, continued monitoring should be considered in this group.

Longitudinal analysis of antibody dynamics in COVID-19 convalescents reveals neutralizing responses up to 16 months after infection.

Elucidating the dynamics of the neutralizing antibody (nAb) response in coronavirus disease 2019 (COVID-19) convalescents is crucial in controlling the pandemic and informing vaccination strategies. Here we measured nAb titres across 411 sequential plasma samples collected during 1-480 d after illness onset or laboratory confirmation (d.a.o.) from 214 COVID-19 convalescents, covering the clinical spectrum of disease and without additional exposure history after recovery or vaccination against SARS-CoV-2, using authentic SARS-CoV-2 microneutralization (MN) assays. Forty-eight samples were also tested for neutralizing activities against the circulating variants using pseudotyped neutralization assay. Results showed that anti-RBD IgG and MN titres peaked at ~120 d.a.o. and subsequently declined, with significantly reduced nAb responses found in 91.67% of COVID-19 convalescents (≥50% decrease in current MN titres compared with the paired peak MN titres). Despite this decline, majority of the COVID-19 convalescents maintained detectable anti-RBD IgG and MN titres at 400-480 d.a.o., with undetectable neutralizing activity found in 14.41% (16/111) of the mild and 50% (5/10) of the asymptomatic infections at 330-480 d.a.o. Persistent antibody-dependent immunity could provide protection against circulating variants after one year, despite significantly decreased neutralizing activities against Beta, Delta and Mu variants. In conclusion, these data show that despite a marked decline in neutralizing activity over time, nAb responses persist for up to 480 d in most convalescents of symptomatic COVID-19, whereas a high rate of undetectable nAb responses was found in convalescents from asymptomatic infections.

In Vitro Inhibition of Replication of Dengue Virus Serotypes 1-4 by siRNAs Bound to Non-Toxic Liposomes.

Dengue virus is a ssRNA+ flavivirus, which produces the dengue disease in humans. Currently, no specific treatment exists. siRNAs regulate gene expression and have been used systematically to silence viral genomes; however, they require controlled release. Liposomes show favorable results encapsulating siRNA for gene silencing. The objective herein was to design and evaluate in vitro siRNAs bound to liposomes that inhibit DENV replication. siRNAs were designed against DENV1–4 from conserved regions using siDirect2.0 and Web-BLOCK-iT™ RNAiDesigner; the initial in vitro evaluation was carried out through transfection into HepG2 cells. siRNA with silencing capacity was encapsulated in liposomes composed of D-Lin-MC3-DMA, DSPC, Chol. Cytotoxicity, hemolysis, pro-inflammatory cytokine release and antiviral activity were evaluated using plaque assay and RT-qPCR. A working concentration of siRNA was established at 40 nM. siRNA1, siRNA2, siRNA3.1, and siRNA4 were encapsulated in liposomes, and their siRNA delivery through liposomes led to a statistically significant decrease in viral titers, yielded no cytotoxicity or hemolysis and did not stimulate release of pro-inflammatory cytokines. Finally, liposomes were designed with siRNA against DENV, which proved to be safe in vitro.

Optimization of application schedule of camphecene, a novel anti-influenza compound, based on its pharmacokinetic characteristics.

Due to high variability and rapid life cycle, influenza virus is able to develop drug resistance against direct-acting antivirals. Development of novel virus-in113039hibiting drugs is therefore important goal. Previously, we identified camphor derivative, camphecene, as an effective anti-influenza compound. In the present study, we optimize the regimen of its application to avoid high sub-toxic concentrations. The protective activity of camphecene was assessed on the model of lethal pneumonia of mice caused by influenza viruses. Camphecene was administered either once a day or four times a day, alone or in combination with Tamiflu. Mortality and viral titer in the lungs were studied. Pharmacokinetics of camphecene was studied in rabbits. We have demonstrated that camphecene, being used every 6 h at a dose of 7.5mg/kg/day, results in antiviral effect that was statistically equal to the effect of 100 mg/kg/day once a day, that is, the same effect was achieved by 13 times lower daily dose of the drug. This effect was manifested in decrease of mortality and decrease of virus' titer in the lungs. The studies of pharmacokinetics of camphecene have demonstrated that it does not accumulate in blood plasma and that its m ultiple applications with dosage interval of 65 min are safe. In addition, the results of the study demonstrate also that camphecene possesses additive effect with Tamiflu, allowing to decrease the dose of the latter. The results suggest that due to safety and efficacy, camphecene can be further developed as potential anti-influenza remedy.

Long-term, infection-acquired immunity against the SARS-CoV-2 Delta variant in a hamster model.

The emergence of the SARS-CoV-2 Delta variant (B.1.617.2) raises concerns about potential reduced sensitivity of the virus to antibody neutralization and subsequent vaccine breakthrough infections. Here, we use a live virus neutralization assay with sera from Pfizer- and Moderna-vaccinated individuals to examine neutralizing antibody titers against SARS-CoV-2 and observe a 3.9- and 2.7-fold reduction, respectively, in neutralizing antibody titers against the Delta variant compared with an early isolate bearing only a D614G substitution in its spike protein. We observe similar reduced sensitivity with sera from hamsters that were previously infected with an early isolate of SARS-CoV-2. Despite this reduction in neutralizing antibody titers against the Delta variant, hamsters previously infected (up to 15 months earlier) with an early isolate are protected from infection with the Delta variant, suggesting that the immune response to the first infection is sufficient to provide protection against subsequent infection with the Delta variant.

Factors Influencing Pneumococcal Vaccination-Induced Total IgG, IgG2, and IgA Levels in Vaccinated Splenectomized Patients

Splenectomy is closely associated with a lifetime risk of pneumococcal and other encapsulated bacterial infections. In this study, it was aimed to investigate the change of antibody levels after vaccination against Streptococcus pneumoniae according to age, gender, years after splenectomy and the possible effect of splenectomy on IgG avidity. In addition the education and awareness levels of the participants about post-splenectomy vaccination and infectious diseases were also analyzed. In the first of the three phases of this study, 32 individuals with splenectomy were enrolled. The awareness of the patients about the possible risks after splenectomy was investigated with a simple questionnaire. Routine laboratory test results were obtained and clinical examinations were performed. In the second stage, total Ig values of 29 splenectomy patients were determined. In the third phase, 14 splenectomy and 5 healthy volunteers were vaccinated according to the Vaccination Practices Advisory Committee (ACIP) guidelines. Pneumococcal-specific antibody levels and IgG avidity were detected by enzyme linked immunosorbent assay (ELISA). It was determined that 68.8% of the splenectomized patients were unaware of their vaccination status and 78.2% of them were unaware of the increased risk of infectious diseases in asplenic conditions. . According to the hospital information management system, all 31 (96.87%) patients, except one, were vaccinated with PPV23. As expected, vaccinated patients exhibited high levels of vaccine-specific antibody production with IgG, IgG2, and IgA antibody concentrations of 321 ± 76.68 mg/l, 73.07 ± 8.273 mg/l, and 117.8 ± 14.94 mg/l, respectively, but unvaccinated patients had very low antibody (IgG, IgG2 and IgA antibody concentrations were 11.5 mg/l, 1.3 mg/l and 1.2 mg/l, respectively) levels. Although there was no correlation between antibody titers and gender, age groups or presence of fever history, the decrease in total IgG, IgG2 and IgA titers were strongly correlated with the time since splenectomy. Antibody titers were found to be significantly lower in splenectomized patients vaccinated more than 10 years ago. Routine laboratory results were at normal levels except for low platelet count. On the other hand, both splenectomized and healthy control subjects displayed similar IgG avidity index values (%61.8 ve %64.4% inhibition in control and splenectomized subjects, respectively) after the vaccination schedule. It was shown that post-splenectomy vaccination with PPV23 induced high levels of pneumococcus-specific antibody production that can last for more than five years. It was determined that more efforts should be made to increase the level of knowledge about pneumococcal and other overwhelming post-splenectomy infections (OPSI) as the awareness of the patients about the risks of infection after splenectomy was poor. In particular, patients with splenectomy operation more than 10 years ago should be very careful about being asplenic as they were determined to have significantly lower level of vaccine-specific antibody production. Our study was also the first to show that splenectomy does not alter IgG avidity induced by pneumococcal vaccination.

Soft-sensors application for automated feeding control in high-throughput mammalian cell cultures.

The ever-increasing demand for biopharmaceuticals has created the need for improving the overall productivity of culture processes. One such operational concept that is considered is fed-batch operations as opposed to batch operations. However, optimal fed-batch operations require complete knowledge of the cell culture to optimize the culture conditions and the nutrients feeding. For example, when using high-throughput small-scale bioreactors to test multiple clones that do not behave the same, depletion or overfeeding of some key components can occur if the feeding strategy is not individually optimized. Over the recent years, various solutions for real-time measuring of the main cell culture metabolites have been proposed. Still, the complexity in the implementation of these techniques has limited their use. Soft-sensors present an opportunity to overcome these limitations by indirectly estimating these variables in real-time. This manuscript details the development of a new soft-sensor-based fed-batch strategy to maintain substrate concentration (glucose and glutamine) at optimal levels in small-scale multiparallel Chinese Hamster Ovary Cells cultures. Two alternatives to the standard feeding strategy were tested: an OUR soft-sensor-based strategy for glucose and glutamine (Strategy 1) and a dual OUR for glutamine and CO2 /alkali addition for glucose soft-sensor strategy (Strategy 2). The results demonstrated the applicability of the OUR soft-sensor-based strategy to optimize glucose and glutamine feedings, which yielded a 21% increase in final viable cell density (VCD) and a 31% in erythropoietin titer compared with the reference one. However, CO2 /alkali addition soft-sensor suffered from insufficient data to relate alkali addition with glucose consumption. As a result, the culture was overfed with glucose resulting in a 4% increase on final VCD, but a 9% decrease in final titer compared with the Reference Strategy.

P529 Evolution of COVID19 serology in a real-life population of IMID patients. Results of the BELCOMID study: BELgian Cohort study of COVID-19 in Immune Mediated Inflammatory Diseases (IMID)

BackgroundImmunomodulators (IMM) and Targeted Immune-Modulating Therapies (TIMT) such as anti-TNF, anti-interleukins and Janus Kinase inhibitors, for treatment of Immune Mediated Inflammatory diseases (IMID) could theoretically interfere with the cytokine storm and humoral immune response against COVID19 infection and vaccination. We investigate seroprevalence and evolution of SARS-CoV2 antibodies in relation to previous vaccination and/or exposure to COVID19 and ongoing IMID-treatment in a Belgian, real-life population of IMID patients.MethodsA cross-disciplinary, prospective, observational cohort study was set up at two university hospitals. All patients with IMIDs of the gut (Crohn’s disease, ulcerative colitis), joints (rheumatoid, psoriatic or spondyloarthritis) and skin (psoriasis, hidradenitis suppurativa, atopic dermatitis) visiting the respective clinics were asked to participate. Patients had to fill out an electronic survey (REDCap®, based on WHO-ISARIC) and blood samples were drawn for serology testing (anti-Spike(S) and anti-viral Nucleocapsid(N) protein antibody IgG, Abbott). Results at baseline, prior to the national vaccination program and at 6 months follow-up are presented. R version 4.0.2 was used for statistical analyses.ResultsAt baseline 2165 IMID patients consented to take part. In 3.2% SARS-CoV2 anti-N seroconversion was confirmed. Of the anti-N seroconverted patients 72.9% reported a positive PCR test prior to inclusion.At 6-months follow-up, data of 1853 IMID patients was collected. Of these, 81.7% were fully and 14.4% partially vaccinated. Seroconversion for anti-N antibodies was confirmed in 2.5% of all participants and seroconversion for anti-S antibodies in 90.8%. In 5.1% (61/1483) of fully vaccinated IMID patients no seroconversion in anti-N nor anti-S antibodies was found.Chi Square analyses show, at 6-months follow-up, no significant association between anti-S seroconversion rate and treatment with systemic steroids (RiskRatio 1.22, 95%CI 0.38–3.9, P=0.99), TIMT (RiskRatio 0.57, 95% CI 0.3–1.1, P=0.12), IMM (RiskRatio 1.65, 95% CI 0.85–3.19, P=0.19) or combination treatment IMM/TIMT (RiskRatio 1.60, 95% CI 0.75–3.4, P=0.32).Appearance of COVID19 symptoms followed the epidemiological curve in Belgium (Fig1). ConclusionIn this real-life IMID cohort, the number of COVID19 cases confirmed by PCR prior to vaccination was low. Seroconversion rate for anti-N antibodies was lower at 6-months follow-up, suggesting decrease in antibody titre over time. Full COVID19 vaccination led to a high anti-S antibody seroconversion rate. Nonetheless, 5.1% of fully vaccinated patients showed no antibody seroconversion. So far, no significant association between anti-S antibody seroconversion and IMID treatment was noted.

Monitoring of SARS-CoV-2 Specific Antibodies after Vaccination

Vaccination is considered the most important measure to control the COVID-19 pandemic. Extensive follow-up studies with distinct vaccines and populations are able to promote robust and reliable data to better understand the effectiveness of this pharmacologic strategy. In this sense, we present data regarding binding and neutralizing (achieved by surrogate ELISA assay) antibodies throughout time, from vaccinated and previously infected (PI) health care workers (HCW) in Portugal. We analyzed serum samples of 132 HCW, who were vaccinated and with previous SARS-CoV-2 infection. Samples were collected before vaccination (baseline, M1), at second dose vaccine uptake (M2), and 25–70 days (M3) and 150–210 days (M4) after the second dose for vaccinated individuals. The IgG (anti-RBD/S) antibody geometric mean titers found on vaccinated HCW at M2 (GM = 116.1 BAU/mL; CI: 92.3–146.1) were significantly higher than those found on PI HCW at recruitment (M1) (GM = 35.9 BAU/mL; CI:15.4–83.4), and the neutralizing antibodies (nAb) were similar between these groups, of 93.2 UI/mL (95% CI 73.2–118.5) vs. 84.1 UI/mL (95% CI 40.4–155.9), respectively. We detected around 10-fold higher IgG (anti-RBD/S) antibodies titers in M3 when compared with M2, with a slight but significant decrease in titers from 36 days after the second dose vaccine uptake. The increase of nAb titers was correlated with IgG (anti-RBD/S) antibodies titers; however, in contrast to IgG (anti-RBD/S) antibodies titers, we did not detect a decrease in the nAb titer 36 days after a second vaccine dose uptake. At M4, a decrease of 8-fold in binding IgG (anti-RBD/S) and nAb was observed. No significant differences in antibody titers were observed by sex, age or chronic diseases. Our results suggest that IgG (anti-RBD/S) antibodies titers and nAb titers could be correlated, but an ongoing follow up of the cohort is required to better understand this correlation, and the duration of the immune response.

The Immune-Specific E3 Ubiquitin Ligase MARCH1 Is Upregulated during Human Cytomegalovirus Infection to Regulate Iron Levels.

Human cytomegalovirus (HCMV) modulates numerous cellular pathways to facilitate infection. Iron is essential to many cellular processes and is often incorporated into proteins and enzymes involved in oxidative phosphorylation and DNA synthesis and repair, among others. Despite its prominent role in the cell, little is known about the regulation of iron metabolism during HCMV infection. Herein, we observe modulation of the transferrin receptor (TfR) during infection and a corresponding change in the cellular labile iron pool. TfR and the iron pool are increased early during infection and then return to mock levels at the late stages of infection. We identified the cellular ubiquitin ligase MARCH1 as an important regulator of TfR. MARCH1 plays a proviral role during infection, as its knockdown leads to a decrease in infectious titers. Knockdown of MARCH1 also leads to an increase in ROS, lipid peroxidation, and mitochondrial dysfunction. Inhibiting an early increase in TfR expression during infection also decreases virus production. These findings indicate the importance of tightly regulating iron metabolism during HCMV infection to facilitate efficient virus production. IMPORTANCE Iron is essential for cells, playing important roles in energy generation, DNA replication, and gene expression. During infection, HCMV alters many cellular processes to aid its replication. We found that iron levels are tightly regulated during infection and that dysregulation of iron levels alters the ability to produce infectious virions. We also found that HCMV inactivates many of the cellular safeguards put in place to deal with excess iron. Thus, infected cells become more susceptible to variations in iron levels, which could be exploited as a therapeutic strategy for dealing with HCMV infections.

Outer Membrane Vesicles (OMVs) of Pseudomonas aeruginosa Provide Passive Resistance but Not Sensitization to LPS-Specific Phages.

Outer membrane vesicles (OMVs) released from gram-negative bacteria are key elements in bacterial physiology, pathogenesis, and defence. In this study, we investigated the role of Pseudomonas aeruginosa OMVs in the anti-phage defence as well as in the potential sensitization to LPS-specific phages. Using transmission electron microscopy, virion infectivity, and neutralization assays, we have shown that both phages efficiently absorb on free vesicles and are unable to infect P. aeruginosa host. Nevertheless, the accompanying decrease in PFU titre (neutralization) was only observed for myovirus KT28 but not podovirus LUZ7. Next, we verified whether OMVs derived from wild-type PAO1 strain can sensitize the LPS-deficient mutant (Δwbpl PAO1) resistant to tested phages. The flow cytometry experiments proved a quite effective and comparable association of OMVs to Δwbpl PAO1 and wild-type PAO1; however, the growth kinetic curves and one-step growth assay revealed no sensitization event of the OMV-associated phage-resistant P. aeruginosa deletant to LPS-specific phages. Our findings for the first time identify naturally formed OMVs as important players in passive resistance (protection) of P. aeruginosa population to phages, but we disproved the hypothesis of transferring phage receptors to make resistant strains susceptible to LPS-dependent phages.

Rift Valley fever virus Gn V5-epitope tagged virus enables identification of UBR4 as a Gn interacting protein that facilitates Rift Valley fever virus production

Rift Valley fever virus (RVFV) is an arbovirus that was first reported in the Rift Valley of Kenya which causes significant disease in humans and livestock. RVFV is a tri-segmented, negative-sense RNA virus consisting of a L, M, and S segments with the M segment encoding the glycoproteins Gn and Gc. Host factors that interact with Gn are largely unknown. To this end, two viruses containing an epitope tag (V5) on the Gn protein in position 105 or 229 (V5Gn105 and V5Gn229) were generated using the RVFV MP-12 vaccine strain as a backbone. The V5-tag insertion minimally impacted Gn functionality as measured by replication kinetics, Gn localization, and antibody neutralization assays. A proteomics-based approach was used to identify novel Gn-binding host proteins, including the E3 ubiquitin-protein ligase, UBR4. Depletion of UBR4 resulted in a significant decrease in RVFV titers and a reduction in viral RNA production.

Engineering a Dynamic Controllable Infectivity Switch in Bacteriophage T7.

Transcriptional repressors play an important role in regulating phage life cycle. Here, we examine how synthetic transcription repressors can be used in bacteriophage T7 to create a dynamic, controllable infectivity switch. We engineered T7 phage by replacing a large region of the early phage genome with different combinations of ligand-responsive promoters and ribosome binding sites (RBS) designed to control the phage RNA polymerase, gp1. Phages with engineered infectivity switch are fully viable at levels comparable to wildtype T7, when not repressed, indicating the phage can be engineered without loss of fitness. The most effective switch used a TetR-responsive promoter and an attenuated RBS, resulting in a 2-fold increase in latent period and a 10-fold decrease in phage titer when repressed. Phage activity can be further tuned using different inducer concentrations. Our study provides a proof of concept for how a simple synthetic circuit introduced into the phage genome enables user control over phage infectivity.

The induction of virus-neutralizing antibodies in influenza A-infected mice treated with Oseltamivir phosphate: effect of dosage and scheduling.

Oseltamivir phosphate (OS) is currently the most frequently used influenza antiviral drug. It moderates the course of influenza virus type A(IAV) infection, however, its impact on the induction of virus-neutralizing antibodies (VNAbs) is not understood in details. Here, we examined the influence of low (10mg/kg) or high (60mg/kg) doses of OS on the viral titer in lungs of BALB/c mice infected with 0.5LD50 of IAV and on the level of VNAbs. Prophylactic application of OS (6hbefore the infection) delayed the increase of viral titer in lungs with alower peak in comparison to non-treated control mice. After therapeutic OS application (44hafter the infection), maximum of virus titer did not significantly change. However, the induction of VNAbs strongly decreased, to 16.7%-18.1% of the control, after preventive application of high OS dose. Aminimal decrease of VNAbs titers was observed in groups of mice treated with low dose of OS applied therapeutically. They lowered to 91.1% / 14 or to 94.1% / 21 days post infection (p.i.) of VNAbs titers of non-treated control mice. In all other groups, levels of VNAbs titers dropped to 26.5-53.7% of those of non-treated mice. It should be noted that VNAbs titers were in direct proportion to maximal virus titers in mouse lungs of corresponding groups. In summary, after OS application the clinical symptoms of the disease were milder or non-observable in all OS-treated groups, but the lowering of VNAbs titers was dependent on the OS dose and interval between drug app-lication and the start of infection. Keywords: influenza Avirus; Oseltamivir; prophylactic treatment; therapeutic treatment; virus-neutralizing antibodies.

Inactivation of coronaviruses under irradiation by UVA-range light-emitting diodes

We report the results of the development of an experimental stand based on UVA light-emitting diodes (UVA LEDs) with radiation wavelengths of 385 and 395 nm for studying experimentally the inactivation of viruses of the coronavirus family, including SARS-CoV-2. Methodological grounds are presented for determining the inactivation dose that provides a predetermined decrease in the virus titre under the impact of UVA radiation. The effect of the diode radiation divergence on the virus photoinactivation process is investigated. It is shown that UVA LEDs can be used to reduce the virus titre by 4 orders of magnitude.

Serological Screening of Immunoglobulin G against SARS-CoV-2 Nucleocapsid and Spike Protein before and after Two Vaccine Doses among Healthcare Workers in Japan.

This study sought to evaluate the effects of two vaccine doses and the extent of SARS-CoV-2 infection among healthcare workers. We measured immunoglobulin G antibody titers against SARS-CoV-2 nucleocapsid and spike protein among healthcare workers at Gunma University Hospital. In March 2021, prior to BNT-162b2 vaccination, two of 771 participants were seropositive for nucleocapsid and spike protein, whereas 768 were seronegative. The remaining one participant was seropositive for nucleocapsid protein but seronegative for spike protein. A total of 769 participants were seropositive for spike protein after two vaccination doses. The two seropositive participants prior to vaccination showed the highest antibody titers after the second vaccination. They were probably infected with SARS-CoV-2 without clinical symptoms before March 2021. Four weeks after the second vaccination, a younger age was associated with higher antibody titers against SARS-CoV-2 spike protein. Thirty-two weeks after the second vaccination, blood samples were collected from 342 of 769 participants. Antibody titers at 32 weeks after the second vaccination significantly decreased compared with those at 4 weeks after the second vaccination among all age groups. The rate of decrease in antibody titers between 4 and 32 weeks after the second vaccination was greater in the female participants. No sex differences were observed in the antibody titers within each age group. BNT-162b2 vaccination thus induced seroconversion in an age-dependent manner. Serological screening could further establish the likelihood of subclinical SARS-CoV-2 infection.

Validation of mailed via postal service dried blood spot cards on commercially available HIV testing systems.

The demand for HIV testing using dried blood spots (DBS) has increased recently. However, DBS is not an approved sample for HIV testing in Japan. This study examined the validation of HIV testing with DBS, prepared at the laboratory or remotely and mailed via postal service to the laboratory. DBS were punched out from a 5.5 mm diameter circle on filter paper, then eluted with 600 μL of phosphate buffered saline overnight at 4℃, and analyzed by Lumipulse S HIVAg/Ab (LUM). The mean LUM count of DBS was 237.4-times diluted compared to titrated plasma. Repeated sample testing showed that although LUM count of DBS decreased slightly with increase in sample storage time (up to one month), it did not affect the result of HIV testing with DBS. Based on testing of 50 HIV+ confirmed cases and 50 HIV- persons, the estimated sensitivity was 98% (49/50) with a specificity of 100% when the cut-off value is 0.5. The single false negative case was a patient with undetectable viral load over the last 10 years, resulting in a decrease of antibody titer below the cut-off level. In conclusion, although DBS cannot completely replace plasma in HIV testing because the sensitivity was a little lower than that of plasma, it can be potentially useful for a screening test by self-finger-prick and postal service use. This will allow people to receive HIV testing without visiting public health centers.

Adenovirus-mediated RNA interference against herpes simplex virus infection in vitro.

Herpetic keratitis caused by the herpes simplex virus (HSV) is the most common form of ocular herpes that causes corneal blindness. Although treatments for herpes keratitis have improved in recent years. there is still considerable room for new treatments against viral infection that shows great promise. The aim of the study was to evaluate the effect of RNA interference on HSV Type 1 (HSV1) infection in vitro, first prophylactically then therapeutically. The highly conserved glycoproteins D (gD) and E (gE) were chosen as targets for this study. Different small interfering RNA (siRNA) duplexes that target gD and gE were designed and chemically synthesized. The recombinant adenovirus type 5 was developed and used as the vehicle with which we delivered the siRNA into the Vero cells infected with the HSV1 KOS strain. Evaluation of the efficacy of siRNA-mediated inhibition was performed either before virus inoculation (prophylactically) or after virus inoculation at the first appearance of lesions (therapeutically). The expression of messenger RNA encoding gD and gE was detected using a real-time polymerase chain reaction (qPCR). We analyzed HSV replication in Vero cells, cytotoxicity of HSV, and cell viability. When used prophylactically, the siRNA-targeting gD and gE created a more marked decrease in viral titer than when used therapeutically. The transfection of cells with recombinant adenovirus containing the siRNA expression cassette was associated with very low cytotoxicity. Adenovirus-mediated siRNA-targeting gD and gE genes effectively inhibit the replication of the HSV in Vero cells. In addition, these findings indicate that the prophylactic use of siRNA is far more effective at inhibiting HSV replication than the therapeutic use.

Persistence of Antibodies Against Spike Glycoprotein of SARS-CoV-2 in Healthcare Workers Post Double Dose of BBV-152 and AZD1222 Vaccines.

Purpose: We investigated the persistence of the vaccine-induced immunoglobulin G (IgG) antibodies against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) among healthcare workers (HCWs) in Odisha who received a complete dose of either Covaxin or Covishield vaccine.Methods: This 24-week longitudinal cohort study was conducted from January to July 2021 with participants from 6 healthcare and research facilities of Odisha to understand the dynamicity of the vaccine-induced IgG antibodies against SARS-CoV-2 after the complete dose of vaccines.Results: Serum samples were collected from 614 participants during each follow-up and were tested in two chemiluminescent microparticle immunoassay (CLIA)-based platforms to detect SARS-CoV-2 antibodies both qualitatively and quantitatively. Among these participants, 308 (50.2%) participants were Covishield recipients and the rest 306 (49.8%) participants took Covaxin. A total of 81 breakthrough cases were recorded and the rest 533 HCWs without any history of postvaccination infection showed significant antibody waning either from T3 (Covaxin recipient) or T4 (Covishield recipient). The production of vaccine-induced IgG antibodies is significantly higher (p < 0.001) in Covishield compared with Covaxin. Covishield recipients produced higher median anti-S IgG titer than Covaxin. No statistically significant differences in antibody titers were observed based on age, gender, comorbidities, and blood groups.Conclusion: This 6-month follow-up study documents a 2-fold and 4-fold decrease in spike antibody titer among Covishield and Covaxin recipients, respectively. The clinical implications of antibody waning after vaccination are not well understood. It also highlights the need for further data to understand the long-term persistence of vaccine-induced antibody and threshold antibody titer required for protection against reinfection.

Immunogenicity after 6 months of BNT162b2 vaccination in frail or disabled nursing home residents: The COVID-A Study.

There is incomplete information regarding evolution of antibody titers against SARS-CoV-2 after a two-dose strategy vaccination with BNT162b2 in older adults in long-term care facilities (LTCFs) with frailty, disability, or cognitive impairment. We aimed to determine IgG antibody titer loss in older adults in LTCFs. This is a multicenter longitudinal cohort study including 127 residents (90 females and 37 males) with a mean age of 82.7 years (range 65-99) with different frailty and disability profiles in two LTCFs in Albacete, Spain. Residents received two doses of BNT162b2 as per label, and antibody levels were determined 1 and 6months after the second dose. Age, sex, previous history of coronavirus disease 2019 (COVID-19), comorbidity (Charlson Index), performance in activities of daily living (Barthel Index), frailty (FRAIL instrument), and cognitive status were assessed. The mean antibody titers 1 and 6months after the second vaccine dose were 32,145 AU/ml (SD 41,206) and 6182 AU/ml (SD 13,316), respectively. Across all participants, the median antibody titer loss measured 77.6% (interquartile range [IQR] 23.8%). Notably, the decline of titers in individuals with pre-vaccination COVID-19 infection was significantly lower than in those without a history of SARS-CoV-2 infection (72.2% vs. 85.3%; p < 0.001). The median titer decrease per follow-up day was 0.47% (IQR 0.14%) and only pre-vaccination COVID-19 was associated with lower rate of antibody decline at 6months (hazard ratio 0.17; 95% confidence interval 0.07-0.41; p < 0.001). Frailty, disability, older age, cognitive impairment, or comorbidity were not associated with the extent of antibody loss. Older adults in LTCFs experience a rapid loss of antibodies over the first 6 months after the second dose of BNT162b2 vaccine. Only pre-vaccination COVID-19 is associated with a slower rate of antibody decrease. Our data support immunization with a third dose in this vulnerable, high-risk population.